Methods and systems for inducing aggregation of fungal proteins and fungal food products made thereby

Abstract

In particular, methods are disclosed for inducing aggregation of fungal proteins in liquid dispersions of filamentous fungal particles by adjusting pH, adding functional ingredients, and / or adding salt. Also disclosed are fungal food products, such as cheese and cheese curd-like food products, and tofu-like food products, that can be produced by such methods.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Patent Application No. 63 / 436,413, filed December 30, 2022, the entire contents of which are incorporated herein by reference.

[0002] This disclosure relates generally to food products made from edible filamentous fungi and methods of making the same, and in particular to solid and / or colloidal fungal food products (e.g., cheese and cheese curd-like food products, tofu-like food products, edible gels, etc.) made by aggregation of filamentous fungal proteins in a liquid dispersion of the filamentous fungal proteins. [Background technology]

[0003] Many traditional food products, such as cheese, cheese curds, and tofu, are formed by a process in which proteins (e.g., dairy proteins in the case of cheese and cheese curds, soy proteins in the case of tofu) are coagulated from a liquid dispersion of the protein (e.g., milk, soy milk), and these coagulated proteins can then be further processed and converted into food products. With increasing commercial interest in vegan and / or hypoallergenic alternatives to such food products, many attempts have been made to create analogs of these traditional food products by coagulating other types of proteins, such as fungal proteins, from their liquid dispersions. However, for reasons that are not always well understood, it can be more difficult to induce coagulation of fungal proteins from a liquid dispersion than it is to induce coagulation of dairy or plant proteins. Furthermore, even if fungal proteins can be coagulated from a liquid dispersion, it may be more difficult or costly to process the resulting coagulated protein into a suitable food product; by way of non-limiting example, fungal protein coagulates may be smaller and / or more brittle than dairy and / or soy protein coagulates and therefore may be more difficult to process into a food product (e.g., a tofu analog) with effective structural integrity and texture.

[0004] Many previous efforts to process coagulated fungal proteins into food products have attempted to overcome these drawbacks by combining the coagulated fungal proteins with binders or gelling agents such as alginate, carrageenan, and / or egg whites. However, these ingredients can be expensive, allergenic, and / or non-vegan, and / or can adversely affect the desired sensory properties (e.g., color, taste, etc.) of the resulting food product.

[0005] Thus, there is a need in the art for methods and systems for providing solid and / or colloidal fungal protein compositions with limited or no addition of traditional non-fungal binders and gelling agents. Furthermore, such methods and systems would utilize relatively small amounts of readily available and inexpensive components, such as common food-safe acids and / or bases, non-fungal proteins, and / or oligosaccharides and / or polysaccharides, and / or salts, which would be advantageously included in the resulting fungal protein compositions. Summary of the Invention

[0006] In one aspect of the present disclosure, a method for making a solid and / or colloidal fungal food material comprises inducing aggregation of fungal proteins in a liquid dispersion of filamentous fungal particles.

[0007] In embodiments, the triggering step may include at least one of (i) adjusting the pH of the liquid dispersion, (ii) adding one or more functional ingredients to the liquid dispersion, and (iii) adding one or more salts to the liquid dispersion. The triggering step may, but need not, include (i). The triggering step may, but need not include (ii). The triggering step may, but need not include (iii). The triggering step may, but need not include (i) and (ii). The triggering step may, but need not include (i) and (iii). The triggering step may, but need not include (ii) and (iii). The triggering step may, but need not include (i), (ii), and (iii).

[0008] In embodiments, the liquid dispersion of filamentous fungus particles may include oil and / or solid fat. The method may, but need not, include a step of combining a liquid phase, filamentous fungus particles, and oil and / or solid fat to form a liquid dispersion before the inducing step. The combining step may, but need not, include blending the liquid phase and filamentous fungus particles with oil and / or solid fat. The blending may, but need not, include high-speed shearing. The high-speed shearing may, but need not, include shearing the liquid phase, filamentous fungus particles, and oil and / or solid fat at a rotational speed of at least about 10,000 rpm for at least about 2 minutes. The combining step may, but need not, include adding an emulsifier. The emulsifier may, but need not, be selected from the group consisting of carboxymethylcellulose, carrageenan, cellulose, guar gum, lecithin, mono- and diglycerides of fatty acids, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, polysorbates, stearoyl lactylate, sorbitan esters, sucrose esters, sucroglycerides, xanthan gum, and combinations thereof. The oil and / or solid fat may, but need not, comprise an oil selected from the group consisting of acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof. The oil and / or solid fat may, but need not, comprise a solid fat selected from the group consisting of blubber, butter, chicken fat, clarified butter, cocoa butter, dripping, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof. The oil content of the liquid dispersion may, but need not, be from about 1% to about 5% by weight.

[0009] In embodiments, the inducing step may include (ii) and the one or more functional ingredients may include a non-fungal protein, which may, but need not, be selected from the group consisting of bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof.

[0010] In embodiments, the inducing step may include (ii) and the one or more functional ingredients may include one or more enzymes, which may, but need not, be selected from the group consisting of catalase, chymosin, lactase, lipase, transglutaminase, and combinations thereof.

[0011] In embodiments, the inducing step may include (i), where the inducing step may involve lowering the pH of the liquid dispersion. The pH of the liquid dispersion may, but need not, be lowered by adding an acid to the liquid dispersion. The acid may, but need not, be selected from the group consisting of sorbic acid, benzoic acid, formic acid, acetic acid, dehydroacetic acid, lactic acid, propionic acid, boric acid, malic acid, fumaric acid, ascorbic acid, erythorbic acid, citric acid, tartaric acid, phosphoric acid, metatartaric acid, adipic acid, succinic acid, thiodipropionic acid, phytic acid, alginic acid, hydrochloric acid, sulfuric acid, gluconic acid, glutamic acid, guanylic acid, inosinic acid, cyclamic acid, cholic acid, and combinations thereof. The pH of the liquid dispersion may, but need not, be lowered by adding an acidifying microbial culture to the liquid dispersion. The inducing step may, but need not, further include heating the liquid dispersion. The liquid dispersion can, but need not, be heated to a temperature of about 150°F to about 180°F (about 65.5°C to about 83°C). The method can, but need not, include the step of further heating the liquid dispersion to a temperature of about 180°F to about 200°F (about 83°C to about 94°C) after the inducing step.

[0012] In embodiments, the liquid dispersion may include at least one salt of calcium or magnesium, and / or the method may include (iii), wherein the one or more salts include at least one salt of calcium or magnesium. The at least one salt of calcium or magnesium may be selected from the group consisting of calcium carbonate, calcium sorbate, calcium benzoate, calcium sulfite, calcium bisulfite, calcium formate, calcium acetate, calcium propionate, calcium ascorbate, calcium lactate, monocalcium citrate, dicalcium citrate, tricalcium citrate, monocalcium phosphate, dicalcium phosphate, tricalcium phosphate, calcium malate, calcium hydrogen malate, calcium tartrate, calcium fumarate, calcium glyceryl phosphate, disodium calcium ethylenediaminetetraacetate, calcium lactobionate, calcium alginate, dicalcium diphosphate, dihydrogen calcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, The additive may, but need not, be selected from the group consisting of calcium ferrocyanide, dicalcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium silicate, calcium aluminosilicate, calcium stearate, calcium gluconate, synthetic calcium aluminate, calcium diglutamate, calcium guanylate, calcium inosinate, calcium 5'-ribonucleotide, calcium iodate, calcium bromate, calcium peroxide, calcium cyclamate, calcium saccharinate, magnesium lactate, monomagnesium phosphate, dimagnesium phosphate, magnesium citrate, magnesium salts of fatty acids, magnesium carbonate, magnesium bicarbonate, magnesium chloride, magnesium sulfate, magnesium oxide, magnesium silicate, magnesium trisilicate, magnesium stearate, magnesium gluconate, magnesium diglutamate, and combinations thereof.

[0013] In embodiments, the liquid dispersion may further comprise at least one of a flavoring agent, a taste modifier, and a plant masking agent.

[0014] In an embodiment, at least a portion of the filamentous fungal particles may be produced by size reduction of a cohesive filamentous fungal mycelial biomass, which may be, but need not be, produced by liquid surface fermentation or solid-state fermentation.

[0015] In an embodiment, at least a portion of the filamentous fungal particles may be produced by submerged fermentation.

[0016] In embodiments, the filamentous fungal particles may be in the form of a powder having a particle size of about 30 μm to about 400 μm.

[0017] In embodiments, the filamentous fungal particles may consist essentially of fungal mycelium.

[0018] In embodiments, the filamentous fungal particles may comprise at least about 50% by weight of fungal mycelia, may comprise at least about 75% by weight of fungal mycelia, or may comprise at least about 95% by weight of fungal mycelia.

[0019] In embodiments, the solids content of the liquid dispersion may be from about 4% to about 7% by weight.

[0020] In embodiments, the weight ratio of filamentous fungal particles to liquid in the liquid dispersion may be from about 1:10 to about 10:1.

[0021] In embodiments, the liquid dispersion may be stable at room temperature for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

[0022] In embodiments, the liquid dispersion may be stable at refrigerated temperatures for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

[0023] In an embodiment, the liquid dispersion may be a mixed-type mycelial biomass composition comprising a first mycelial biomass type and a second mycelial biomass type, wherein the first and second mycelial biomass types are different from each other. The first mycelial biomass type may be, but need not be, a clumped mycelial biomass type, and the second mycelial biomass type may be, but need not be, a submerged mycelial biomass type. The first mycelial biomass type may be, but need not be, selected from the group consisting of biomat pieces, biomat powder, biomat dispersion, and spray-dried biomat powder. The second mycelial biomass type may be, but need not be, selected from the group consisting of submerged slurry, submerged dough, submerged powder, submerged dispersion, and submerged spray-dried powder. The first and second mycelial biomass types may each be, but need not be, a submerged mycelial biomass type. The first and second mycelial biomass forms may, but need not, be selected from the group consisting of a soaked liquid biomass, a soaked dough, a soaked powder, a soaked dispersion, and a soaked spray-dried powder. The liquid dispersion may, but need not, be a combined liquid dispersion, and the method may, but need not, include, prior to the inducing step, blending a mixture of the first mycelial biomass form and a first liquid to form a first liquid dispersion, blending a mixture of the second mycelial biomass form and a second liquid to form a second liquid dispersion, and combining the first and second liquid dispersions to form a combined liquid dispersion.

[0024] In embodiments, the gel may be formed in an inducing step, which may, but need not, include adjusting the pH of the liquid dispersion to a gelling pH of about 4 or less. The gelling pH may, but need not, be about 3.5.

[0025] In embodiments, the mycelium curd can be formed in a triggering step, and the method can further include separating at least a portion of the liquid phase of the liquid dispersion from the mycelium curd. In the separating step, the at least a portion of the liquid phase can be, but need not be, at least about 90% by weight of the liquid phase. The triggering step can include, but need not be, adjusting the pH of the liquid dispersion to a pH of about 2 to about 4. In the triggering step, the pH can be, but need not be, adjusted to a pH of about 3.5. The separating step can include, but need not be, pressing the mycelium curd through a mesh filter. The mesh filter can, but need not be, comprise a cloth. The cloth can, but need not be, cheesecloth. The mesh filter can, but need not be, comprise a fine mesh sieve. The method can, but need not be, comprise forming the mycelium curd into a block.

[0026] In another aspect of the present disclosure, the food material comprises aggregated filamentous fungal mycelial biomass.

[0027] In embodiments, the food material may further comprise oil and / or solid fat. The oil and / or solid fat may, but need not, comprise an oil selected from the group consisting of acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof. The oil and / or solid fat may, but need not, comprise a solid fat selected from the group consisting of marine tallow, butter, chicken fat, clarified butter, cocoa butter, drippings, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof.

[0028] In embodiments, the food material may further comprise a non-fungal protein, which may, but need not, be selected from the group consisting of bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof.

[0029] In an embodiment, at least a portion of the filamentous fungal particles may be produced by size reduction of a cohesive filamentous fungal mycelial biomass, which may be, but need not be, produced by liquid surface fermentation or solid-state fermentation.

[0030] In an embodiment, at least a portion of the filamentous fungal particles may be produced by submerged fermentation.

[0031] In embodiments, the filamentous fungal particles may consist essentially of fungal mycelium.

[0032] In embodiments, the filamentous fungal particles may comprise at least about 50% by weight of fungal mycelia, may comprise at least about 75% by weight of fungal mycelia, or may comprise at least about 95% by weight of fungal mycelia.

[0033] In an embodiment, the fungal card may be in the form of a block.

[0034] In embodiments, the food material may be free of non-fungal gelling agents.

[0035] In an embodiment, the food material may consist essentially of aggregated filamentous fungal mycelial biomass.

[0036] In embodiments, the food material may consist of filamentous fungal mycelial biomass and at least one acid or base.

[0037] In embodiments, the food material may consist of filamentous fungal mycelial biomass and at least one functional ingredient.

[0038] In embodiments, the food material may consist of filamentous fungal mycelial biomass and at least one salt.

[0039] In embodiments, the food material may further comprise a microbial food culture.

[0040] In embodiments, the food material may have a hardness of from about 1N to about 50N.

[0041] In embodiments, the food material may have an adhesive strength of from about 0.001 N·mm to about 60 N·mm.

[0042] In embodiments, the food material may have a cohesive strength of from about 0.001 to about 4.

[0043] In an embodiment, the food material may be a fungal curd made by the method for making a fungal curd as disclosed herein.

[0044] In another aspect of the present disclosure, the mixed-type mycelial biomass composition comprises a first mycelial biomass type and a second mycelial biomass type, wherein the first mycelial biomass type and the second mycelial biomass type are different mycelial biomass types.

[0045] In an embodiment, the composition may be a food material. The food material may be, but is not required to be, selected from the group consisting of a flour, a plurality of solid particles other than a flour, a liquid dispersion, an emulsion, a foam, a gel, a sol, and a solid foam. The food material may be, but is not required to be, a flour, and the flour may, but is not required to be, a filamentous fungus particle having a particle size of about 30 μm to about 400 μm. The food material may be, but is not required to be, a plurality of solid particles other than a flour, and the plurality of solid particles may, but is not required to be, a filamentous fungus particle having a particle length of about 0.05 mm to about 500 mm, a particle width of about 0.03 mm to about 7 mm, and a particle height of about 0.03 mm to about 1.0 mm. The food material may be, but is not required to be, a liquid dispersion or sol, and the mass ratio of filamentous fungus particles to liquid in the liquid dispersion or sol may, but is not required to be, about 1:10 to about 10:1. The food material can be, but need not be, a liquid dispersion or sol, and the liquid dispersion or sol can be, but need not be, stable for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.The food material may have a foam stability of at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, or at least about 95%, such as for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, The foam may, but need not, have a duration of at least about 1 day, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

[0046] In an embodiment, the food product may include a food ingredient.

[0047] In embodiments, the first mycelial biomass format is a clumped mycelial biomass format and the second mycelial biomass format is a soaked mycelial biomass format. The first mycelial biomass format may, but need not, be selected from the group consisting of biomat pieces, biomat powder, biomat dispersion, and spray-dried biomat powder. The second mycelial biomass format may, but need not, be selected from the group consisting of soaked slurries, soaked doughs, soaked powders, soaked dispersions, and soaked spray-dried powders. The composition may, but need not, be a food ingredient. The composition may, but need not be a gel. The composition may, but need not be a food product selected from the group consisting of a blancmange-like food product, a butter-like food product, a custard-like food product, a jam-like food product, a jelly-like food product, a margarine-like food product, and a yogurt-like food product. The mass ratio of the first mycelial biomass form to the second mycelial biomass form can be, but need not be, about 1:10 to about 10:1.

[0048] In embodiments, the first mycelial biomass form and the second mycelial biomass form may each be a soaked mycelial biomass form. The first mycelial biomass form and the second mycelial biomass form may each be, but need not be, selected from the group consisting of a soaked slurry, a soaked dough, a soaked powder, a soaked dispersion, and a soaked spray-dried powder. The composition may, but need not be, a food ingredient. The composition may, but need not be, a gel. The composition may, but need not be, a food product selected from the group consisting of a blancmange-like food product, a butter-like food product, a custard-like food product, a jam-like food product, a jelly-like food product, a margarine-like food product, and a yogurt-like food product. The mass ratio of the first mycelial biomass form to the second mycelial biomass form may, but need not be, about 1:10 to about 10:1.

[0049] In another aspect of the present disclosure, a method for producing a fungal gel includes at least one of the steps of (i) adjusting the pH of a liquid dispersion, (ii) adding one or more functional ingredients to the liquid dispersion, and (iii) adding one or more salts to the liquid dispersion, wherein the liquid dispersion is a mixed-type mycelial biomass composition comprising a first mycelial biomass type and a second mycelial biomass type, and the first mycelial biomass type and the second mycelial biomass type are different mycelial biomass types.

[0050] In an embodiment, the fungal gel may be a food product, which may, but need not, be selected from the group consisting of a blancmange like food product, a butter like food product, a custard like food product, a jam like food product, a jelly like food product, a margarine like food product, and a yogurt like food product.

[0051] In embodiments, the mass ratio of the first mycelial biomass form to the second mycelial biomass form may be from about 1:10 to about 10:1.

[0052] In embodiments, the first mycelial biomass format may be a cohesive mycelial biomass format selected from the group consisting of biomat pieces, biomat flour, biomat dispersion, and spray-dried biomat powder, and the second mycelial biomass format may be a soaked mycelial biomass format selected from the group consisting of soaked slurry, soaked dough, soaked powder, soaked dispersion, and soaked spray-dried powder. The first mycelial biomass format may, but need not, be selected from the group consisting of biomat pieces, biomat flour, and spray-dried biomat powder, and the second mycelial biomass format may, but need not be selected from the group consisting of soaked dough and soaked powder.

[0053] In embodiments, the first and second mycelial biomass formats may each be a soaked mycelial biomass format selected from the group consisting of a soaked paste, a soaked powder, a soaked liquid dispersion, and a soaked spray-dried powder. The first and second mycelial biomass formats may, but need not, be selected from the group consisting of a soaked dough, a soaked powder, and a soaked spray-dried powder.

[0054] In an embodiment, a mixed-form mycelial biomass composition can be produced by a method comprising blending a mixture of a first mycelial biomass form and a first liquid to form a first liquid dispersion, blending a mixture of a second mycelial biomass form and a second liquid to form a second liquid dispersion, and combining the first liquid dispersion and the second liquid dispersion to form the mixed-form mycelial biomass composition.

[0055] In embodiments, the inducing step includes (i), and in the inducing step, the pH of the liquid dispersion can be adjusted to a gelling pH of about 4 or less. The gelling pH can be, but need not be, about 3.5.

[0056] In another aspect of the present disclosure, a method for making a fungal tofu analog food product includes inducing aggregation of fungal proteins in a liquid dispersion of filamentous fungal particles to form a fungal curd, wherein the inducing step includes at least one of (i) adjusting the pH of the liquid dispersion, (ii) adding one or more functional ingredients to the liquid dispersion, and (iii) adding one or more salts to the liquid dispersion, and compressing the fungal curd to form the fungal tofu analog food product.

[0057] In embodiments, the method may further comprise the step of separating the fungal curd from the liquid phase of the liquid dispersion after the inducing step.

[0058] In embodiments, the liquid dispersion may include oil and / or solid fat. The method may, but need not, further include combining the liquid phase, the filamentous fungus particles, and the oil and / or solid fat to form a liquid dispersion before the inducing step. The combining step may, but need not, include blending the liquid phase and the filamentous fungus particles with the oil and / or solid fat. The blending may, but need not, include high-speed shearing. The high-speed shearing may, but need not, include shearing the liquid phase, the filamentous fungus particles, and the oil and / or solid fat at a rotational speed of at least about 10,000 rpm for at least about 2 minutes. The combining step may, but need not, include adding an emulsifier. The emulsifier may, but need not, be selected from the group consisting of carboxymethylcellulose, carrageenan, cellulose, guar gum, lecithin, mono- and diglycerides of fatty acids, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, polysorbates, stearoyl lactylate, sorbitan esters, sucrose esters, sucroglycerides, xanthan gum, and combinations thereof. The oil and / or solid fat may, but need not, comprise an oil selected from the group consisting of acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof. The oil and / or solid fat may, but need not, comprise a solid fat selected from the group consisting of marine tallow, butter, chicken fat, clarified butter, cocoa butter, dripping, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof. The oil content of the liquid dispersion can be, but need not be, from about 1% to about 5% by weight.

[0059] In embodiments, the inducing step may include (ii) and the one or more functional ingredients may include a non-fungal protein, which may, but need not, be selected from the group consisting of bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof.

[0060] In embodiments, the inducing step may include (ii) and the one or more functional ingredients may include one or more enzymes, which may, but need not, be selected from the group consisting of catalase, chymosin, lactase, lipase, transglutaminase, and combinations thereof.

[0061] In embodiments, the inducing step may include (i), where the inducing step may involve lowering the pH of the liquid dispersion. The pH of the liquid dispersion may, but need not, be lowered by adding an acid to the liquid dispersion. The acid may, but need not, be selected from the group consisting of sorbic acid, benzoic acid, formic acid, acetic acid, dehydroacetic acid, lactic acid, propionic acid, boric acid, malic acid, fumaric acid, ascorbic acid, erythorbic acid, citric acid, tartaric acid, phosphoric acid, metatartaric acid, adipic acid, succinic acid, thiodipropionic acid, phytic acid, alginic acid, hydrochloric acid, sulfuric acid, gluconic acid, glutamic acid, guanylic acid, inosinic acid, cyclamic acid, cholic acid, and combinations thereof. The pH of the liquid dispersion may, but need not, be lowered by adding an acidifying microbial culture to the liquid dispersion. The inducing step may, but need not, further include heating the liquid dispersion. The liquid dispersion can, but need not, be heated to a temperature of about 150°F to about 180°F (about 65.5°C to about 83°C). The method can, but need not, include a step after the inducing step of further heating the liquid dispersion to a temperature of about 180°F to about 200°F (about 83°C to about 94°C). In the inducing step, the pH can, but need not, be adjusted to a pH of about 2 to about 4. In the inducing step, the pH can, but need not be adjusted to a pH of about 3.5.

[0062] In embodiments, the liquid dispersion may include at least one salt of calcium or magnesium, and / or the inducing step may include (iii), and the one or more salts may include at least one salt of calcium or magnesium. The at least one salt of calcium or magnesium may be selected from the group consisting of calcium carbonate, calcium sorbate, calcium benzoate, calcium sulfite, calcium bisulfite, calcium formate, calcium acetate, calcium propionate, calcium ascorbate, calcium lactate, monocalcium citrate, dicalcium citrate, tricalcium citrate, monocalcium phosphate, dicalcium phosphate, tricalcium phosphate, calcium malate, calcium hydrogen malate, calcium tartrate, calcium fumarate, calcium glyceryl phosphate, disodium calcium ethylenediaminetetraacetate, calcium lactobionate, calcium alginate, dicalcium diphosphate, dihydrogen calcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, The additive may, but need not, be selected from the group consisting of calcium ferrocyanide, dicalcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium silicate, calcium aluminosilicate, calcium stearate, calcium gluconate, synthetic calcium aluminate, calcium diglutamate, calcium guanylate, calcium inosinate, calcium 5'-ribonucleotide, calcium iodate, calcium bromate, calcium peroxide, calcium cyclamate, calcium saccharinate, magnesium lactate, monomagnesium phosphate, dimagnesium phosphate, magnesium citrate, magnesium salts of fatty acids, magnesium carbonate, magnesium bicarbonate, magnesium chloride, magnesium sulfate, magnesium oxide, magnesium silicate, magnesium trisilicate, magnesium stearate, magnesium gluconate, magnesium diglutamate, and combinations thereof.

[0063] In embodiments, the separating step may include pressing the fungal curd through a mesh filter. The mesh filter may, but need not, comprise a cloth. The cloth may, but need not, be cheesecloth. The mesh filter may, but need not, comprise a fine mesh sieve. The method may, but need not, further comprise forming the fungal curd into a block.

[0064] In embodiments, the liquid dispersion may further comprise at least one of a flavoring agent, a taste modifier, and a plant masking agent.

[0065] In an embodiment, at least a portion of the filamentous fungal particles may be produced by size reduction of a cohesive filamentous fungal mycelial biomass, which may be, but need not be, produced by liquid surface fermentation or solid-state fermentation.

[0066] In an embodiment, at least a portion of the filamentous fungal particles may be produced by submerged fermentation.

[0067] In embodiments, the filamentous fungal particles may be in the form of a powder having a particle size of about 30 μm to about 400 μm.

[0068] In embodiments, the filamentous fungal particles may consist essentially of fungal mycelium.

[0069] In embodiments, the filamentous fungal particles may comprise at least about 50% by weight of fungal mycelia, may comprise at least about 75% by weight of fungal mycelia, or may comprise at least about 95% by weight of fungal mycelia.

[0070] In embodiments, the solids content of the liquid dispersion may be from about 4% to about 7% by weight.

[0071] In another aspect of the present disclosure, a fungal tofu analog food product is made by the method of making a fungal tofu analog food product as disclosed herein.

[0072] While specific embodiments and applications have been illustrated and described, the disclosure is not limited to the precise configuration and components described herein. Various modifications, changes, and variations, which will be apparent to those skilled in the art, can be made in the arrangement, operation, and details of the methods and systems disclosed herein without departing from the spirit and scope of the overall disclosure.

[0073] As used herein, unless otherwise specified, the terms "about," "approximately," and the like, when used in connection with a numerical limit or range, mean that the recited limit or range may vary by up to 10%. As a non-limiting example, "about 750" may mean a minimum of 675 or a maximum of 825, or any value therebetween. When used in connection with a ratio or relationship between two or more numerical limits or ranges, the terms "about," "approximately," and the like mean that the limits or ranges, respectively, may vary by up to 10%. As a non-limiting example, a statement that two quantities are "approximately equal" may mean that the ratio between the two quantities is a minimum of 0.9:1.1 or a maximum of 1.1:0.9 (or any value therebetween); a statement that a four-way ratio is "about 5:3:1:1" may mean that the first number in the ratio can be any value at least 4.5 and no more than 5.5; the second number in the ratio can be any value at least 2.7 and no more than 3.3; etc.

[0074] The embodiments and configurations described herein are not complete or exhaustive, and it will be understood that other embodiments are possible that utilize, alone or in combination, one or more of the features set forth above or described in detail below. [Brief explanation of the drawings]

[0075] [Figure 1]1 is a flow chart illustrating a method for inducing aggregation of fungal proteins by acid addition, according to an embodiment of the present disclosure. [Figure 2] 1 is a flow chart illustrating a method for inducing aggregation of fungal proteins by addition of an acid-producing microbial culture according to an embodiment of the present disclosure. [Figure 3] 1 is a flow chart illustrating a method for inducing aggregation of fungal proteins by the addition of one or more functional ingredients, according to an embodiment of the present disclosure. [Figure 4] 1 is a flow chart illustrating a method for inducing aggregation of fungal proteins by the addition of one or more salts, according to an embodiment of the present disclosure. [Figure 5] 1 is a flow chart illustrating a method for inducing aggregation of fungal proteins by a combination of the addition of one or more functional ingredients and the addition of one or more acids, acid-producing microbial cultures, and / or salts, according to an embodiment of the present disclosure. [Figure 6] 1 is a flow chart illustrating a method for inducing aggregation of fungal proteins by acidification and / or addition of one or more salts according to an embodiment of the present disclosure. [Figure 7] 1 is a flow chart illustrating a method for making a fungal tofu analog food product from a fungal curd according to an embodiment of the present disclosure. [Figure 8] 1 is an image of an aqueous dispersion of Fusarium strain flavolapis according to an embodiment of the present disclosure. [Figure 9] 9 is an image of the aqueous dispersion shown in FIG. 8 shortly after the initiation of fungal curd formation, according to an embodiment of the present disclosure. [Figure 10] 9 is an image of the aqueous dispersion shown in FIG. 8 shortly after the initiation of fungal curd formation, according to an embodiment of the present disclosure. [Figure 11] 11A-11C are images of the aqueous dispersions illustrated in FIGS. 8-10 after further heating and fungal curd formation, according to embodiments of the present disclosure. [Figure 12]11A-11C are images of the aqueous dispersions illustrated in FIGS. 8-10 after further heating and fungal curd formation, according to embodiments of the present disclosure. [Figure 13] 13A-13C are images illustrating the separation of fungal curd from the liquid phase of the aqueous dispersions illustrated in FIGS. 8-12, according to embodiments of the present disclosure. [Figure 14] 1 is an image of a soft spreadable or ricotta-like cheese-like fungal curd composition according to an embodiment of the present disclosure. [Figure 15] 1 is a graph of oil-in-water emulsion stability of liquid dispersions of various mycelial biomass formats, according to embodiments of the present disclosure. [Figure 16] 1 is a graph of viscosity of liquid dispersions of various mycelial biomass formats as a function of shear rate, according to an embodiment of the present disclosure. [Figure 17] 1 is a graph of the viscosity of a liquid dispersion of biomat pieces at various pH values ​​as a function of shear rate, according to an embodiment of the present disclosure. [Figure 18] 1 is a graph of storage and loss moduli of liquid dispersions of biomat pieces at various pH values ​​as a function of shear frequency according to an embodiment of the present disclosure. [Figure 19] 1 is a graph of storage and loss moduli of a fungal gel formed from a liquid dispersion of biomat pieces as a function of time after aggregation of fungal proteins according to an embodiment of the present disclosure. [Figure 20-1] FIG. 20A: A graph of the moisture content of fungal tofu analog food products made using a liquid dispersion of soaked dough, according to an embodiment of the present disclosure. [Figure 20-2] FIG. 20B: A graph of moisture content of fungal tofu analog food products produced using a liquid dispersion of bio-mat pieces according to an embodiment of the present disclosure. [Figure 21-1] FIG. 21A: A graph of the protein content of fungal tofu analog food products made using a liquid dispersion of soaked dough, according to an embodiment of the present disclosure. [Figure 21-2]FIG. 21B: A graph of the protein content of fungal tofu analog food products produced using liquid dispersions of bio-mat pieces according to embodiments of the present disclosure. [Figure 22-1] FIG. 22A: A graph of the firmness of fungal tofu analog food products produced using a liquid dispersion of soaked dough, according to an embodiment of the present disclosure. [Figure 22-2] FIG. 22B: A graph of the firmness of a fungal tofu analog food product produced using a liquid dispersion of bio-mat pieces according to an embodiment of the present disclosure. [Figure 23-1] FIG. 23A: A graph of the cohesive strength of a fungal tofu analog food product made using a liquid dispersion of soaked dough, according to an embodiment of the present disclosure. [Figure 23-2] FIG. 23B: A graph of the cohesive strength of a fungal tofu analog food product produced using a liquid dispersion of bio-mat pieces according to an embodiment of the present disclosure. [Figure 24-1] FIG. 24A: A graph of the adhesive force of a fungal tofu analog food product made using a liquid dispersion of soaked dough, according to an embodiment of the present disclosure. [Figure 24-2] FIG. 24B: A graph of the adhesive force of a fungal tofu analog food product produced using a liquid dispersion of bio-mat pieces according to an embodiment of the present disclosure. [Figure 25] 1A-1C are scanning electron microscopy (SEM) images of fungal tofu analog food products produced using a liquid dispersion of soaked dough according to embodiments of the present disclosure. The image marked "A" is an image of the surface of the tofu analog food product, and the image marked "B" is an image of a cross section of the tofu analog food product. [Figure 26] (Same as the explanation for Figure 25.) [Figure 27] (Same as the explanation for Figure 25.) [Figure 28] (Same as the explanation for Figure 25.) [Figure 29] (Same as the explanation for Figure 25.) [Figure 30] (Same as the explanation for Figure 25.) [Figure 31] (Same as the explanation for Figure 25.) [Figure 32] (Same as the explanation for Figure 25.) [Figure 33] (Same as the explanation for Figure 25.) [Figure 34] (Same as the explanation for Figure 25.) [Figure 35] (Same as the explanation for Figure 25.) [Figure 36] (Same as the explanation for Figure 25.) [Figure 37] (Same as the explanation for Figure 25.) [Figure 38] (Same as the explanation for Figure 25.) [Figure 39] (Same as the explanation for Figure 25.) [Figure 40] (Same as the explanation for Figure 25.) [Figure 41] (Same as the explanation for Figure 25.) [Figure 42] (Same as the explanation for Figure 25.) [Figure 43] 1 shows SEM images of a fungal tofu analog food product produced using a liquid dispersion of bio-mat pieces according to an embodiment of the present disclosure. The image marked "A" is an image of the surface of the tofu analog food product, and the image marked "B" is an image of the cross section of the tofu analog food product. [Figure 44] (Same as in Figure 43.) [Figure 45] (Same as in Figure 43.) [Figure 46] (Same as in Figure 43.) [Figure 47] (Same as in Figure 43.) [Figure 48] (Same as in Figure 43.) [Figure 49] (Same as in Figure 43.) [Figure 50] (Same as in Figure 43.) [Figure 51] (Same as in Figure 43.) [Figure 52] (Same as in Figure 43.) [Figure 53] (Same as in Figure 43.) [Figure 54] (Same as in Figure 43.) [Figure 55] (Same as in Figure 43.) [Figure 56](Same as in Figure 43.) [Figure 57] (Same as in Figure 43.) [Figure 58] (Same as in Figure 43.) [Figure 59] (Same as in Figure 43.) [Figure 60] (Same as in Figure 43.) DETAILED DESCRIPTION OF THE INVENTION

[0076] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. All patents, applications, published applications, and other publications referenced herein are incorporated by reference in their entirety. If there are multiple definitions for terms herein, the definitions set forth in the Summary of the Invention shall prevail unless otherwise stated.

[0077] As used herein, unless otherwise specified, the term "analog" or "analogous food product" refers to a food product comprising an edible fungus that has aesthetic, culinary, nutritional, and / or sensory equivalence or similarity to an identified non-fungal food product. As non-limiting examples, an "ice cream analog food product," as the term is used herein, refers to a food product comprising an edible fungus that has aesthetic, culinary, nutritional, and / or sensory equivalence or similarity to conventional ice cream made from animal milk, and a "mayonnaise analog food product," as the term is used herein, refers to a food product comprising an edible fungus that has aesthetic, culinary, nutritional, and / or sensory equivalence or similarity to conventional mayonnaise made using animal products.

[0078] As used herein, unless otherwise specified, the term "coalesce" and its derivatives (e.g., "coalescing," "coalescence," etc.) refers to the phenomenon in which fungal proteins in a liquid dispersion of particles of filamentous fungal mycelial biomass are attracted to each other to form a fungal curd (i.e., a solid mass of aggregated filamentous fungal mycelial biomass proteins (and optionally other components) that is separable from the remaining liquid phase, similar to the manner in which proteins are coagulated from animal milk (to form curd) or soy milk (to form tofu)) or a gel material (i.e., a phase that retains its shape and resists flow, in which the liquid phase is dispersed throughout the network formed by the filamentous fungal proteins (and optionally other compounds)). Fungal protein "aggregation," as the term is used herein, can be induced by, as non-limiting examples, adjusting the pH of the liquid dispersion, adding one or more functional ingredients to the liquid dispersion, and / or adding one or more salts to the liquid dispersion.

[0079] As used herein, unless otherwise specified, the terms "cohesive mycelial biomass," "biomat," and "cohesive biomat" are used interchangeably and refer to mycelial biomass produced by a non-submerged fermentation process, such as liquid surface fermentation, membrane or mesh fermentation, or solid substrate fermentation, respectively. Non-limiting examples of cohesive mycelial biomass, as the term is used herein, include biomats composed substantially entirely of mycelium, and composite mycelium / raw biomats.

[0080] As used herein, unless otherwise specified, the term "flocculent mycelial biomass form" refers to a mycelial biomass form in which the mycelial biomass has been produced by a non-submerged fermentation process.

[0081] As used herein, unless otherwise specified, the term "colloid" refers to a mixture in which particles of one substance (the "dispersed phase") are dispersed throughout a volume of a different substance (the "dispersion medium"); for example, the dispersed phase may comprise or consist of fine bubbles, particles, etc. When the dispersed phase and dispersion medium of a colloid are specifically identified herein, they are separated by a hyphen and the dispersed phase is identified first; for example, reference herein to an "oil-water colloid" refers to a colloid in which oil is the dispersed phase and water is the dispersion medium.

[0082] As used herein, unless otherwise specified, the term "emulsion" refers to a colloid in which both the dispersed phase and the dispersion medium are liquid. Examples of emulsions, as this term is used herein, include, but are not limited to, butter (when melted), margarine (when melted), mayonnaise, and milk.

[0083] As used herein, unless otherwise specified, the term "foam" refers to a colloid in which the dispersed phase is a gas and the dispersion medium is a liquid. Examples of foams, as the term is used herein, include, but are not limited to, egg white foam (i.e., the product of whipping or otherwise incorporating air into egg whites) and whipped cream.

[0084] As used herein, unless otherwise specified, the term "foam stability" refers to the percentage of the foam's initial volume retained by the foam after a specified interval. Thus, as a non-limiting example, a foam having an initial volume of 5 liters and a volume of 4 liters after 14 days has 80% stability over 14 days. Unless otherwise specified, a "stable" foam, as that term is used herein, is one that has at least 50% stability after a specified interval.

[0085] As used herein, unless otherwise specified, the term "fungal curd" refers to any mass of filamentous fungal particles and optionally other components that (1) is formed by aggregation of fungal proteins in a liquid dispersion of filamentous fungal particles and (2) behaves as a substantially solid material such that it can be separated from the remaining liquid phase of the liquid dispersion by conventional means of liquid-solid separation (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.).

[0086] As used herein, unless otherwise specified, the term "gel" refers to a colloid in which the dispersed phase is a liquid and the dispersion medium is a solid. Examples of gels, as this term is used herein, include, but are not limited to, blancmange, butter (when cooled), custard (after it has been cooked), jam, jelly (after it has set), margarine (when cooled), and yogurt. Gels, as this term is used herein, can behave as solids or semi-solids and typically have an elastic modulus greater than their dynamic (or loss) modulus and therefore do not flow easily.

[0087] As used herein, unless otherwise specified, the term "functional ingredient" refers to any edible carbohydrate or protein.

[0088] As used herein, unless otherwise specified, the term "liquid aerosol" refers to a colloid in which the dispersed phase is a liquid and the dispersion medium is a gas.

[0089] As used herein, unless otherwise specified, the term "mycelial biomass" refers to filamentous fungal biomass that (i) contains at least 50% mycelium on a dry weight basis, and optionally may contain at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% mycelium on a dry weight basis, and (ii) has been produced by a submerged or non-submerged fermentation process. Any portion of the dry weight of the mycelial biomass that is not mycelium may consist of impurities and / or other filamentous fungal tissues (e.g., conidia, fruiting bodies, or parts thereof, etc.).

[0090] As used herein, unless otherwise specified, the term "mycelial biomass form" refers to mycelial biomass that has been produced by a particular type of fermentation process and then subjected to one or more particular post-production processing steps, such as dewatering, blending with liquid, drying, and grinding / milling, size reduction, spray drying, etc. Non-limiting examples of mycelial biomass formats, as the term is used herein, include: (1) biomass-containing fermentation broth or washed biomass resuspended in a liquid (referred to herein as "steep slurry"); (2) mycelial biomass in the form of a dough produced by dewatering a steep slurry (referred to herein as "steep dough"); (3) mycelial biomass in the form of a flour produced by drying the steep slurry or steep dough and grinding or milling it (referred to herein as "steep flour"); (4) a liquid dispersion of mycelial biomass produced by (i) blending the steep slurry and / or (ii) mixing the steep dough with a liquid and blending the mixture (referred to herein as "steep dispersion"); and (5) a liquid dispersion of mycelial biomass produced by spray drying a steep dispersion. (6) biomass in the form of a powder obtained by drying and grinding or milling a biomat or biomat pieces (referred to herein as "biomat powder"); (7) mycelial biomass in the form of a powder produced by drying and grinding or milling a biomat or biomat pieces (referred to herein as "biomat powder"); (8) a liquid dispersion of mycelial biomass produced by mixing a biomat, biomat pieces, or biomat powder with a liquid (referred to herein as "biomat dispersion"); and (9) mycelial biomass in the form of a powder produced by spray drying a biomat dispersion or other fluid containing biomat, biomat pieces, or biomat powder (referred to herein as "spray-dried biomat powder").

[0091] As used herein, unless otherwise specified, the term "mycelium" refers to the vegetative part of a filamentous fungus consisting of a mass of hyphae.

[0092] As used herein, unless otherwise specified, the term "non-submerged fermentation process" refers to any fungal fermentation process in which at least a portion of the mycelium formed by the process is not submerged in a liquid fermentation medium. Non-limiting examples of non-submerged fermentation processes, as this term is used herein, include liquid surface fermentation processes, solid surface (or solid substrate) fermentation processes, membrane surface fermentation processes, and mesh surface fermentation processes. Non-limiting examples of liquid surface fermentation processes include those described in PCT Application Publication Nos. 2017 / 151684, 2019 / 099474, 2020 / 176758, and 2023 / 021264.

[0093] As used herein, unless otherwise specified, the term "sol" refers to a colloid in which the dispersed phase is a solid and the dispersion medium is a liquid. Examples of sols, as the term is used herein, include, but are not limited to, custard (before it is cooked) and jelly (before it sets).

[0094] As used herein, unless otherwise specified, the term "solid aerosol" refers to a colloid in which the dispersed phase is a solid and the dispersion medium is a gas.

[0095] As used herein, unless otherwise specified, the term "solid foam" refers to a colloid in which the dispersed phase is a gas and the dispersion medium is a solid. Examples of solid foams, as the term is used herein, include, but are not limited to, bread, cake, ice cream, and meringue.

[0096] As used herein, unless otherwise specified, the term "solid sol" refers to a colloid in which both the dispersed phase and the dispersion medium are solids.

[0097] As used herein, unless otherwise specified, the term "submerged fermentation process" refers to any fungal fermentation process in which substantially all of the mycelium formed by the process is submerged in a liquid fermentation medium. Non-limiting examples of submerged fermentation processes include those described in British Patent No. 1,346,062, U.S. Patent No. 4,555,485, and PCT Publication Nos. 2022 / 157326 and 2022 / 236165.

[0098] As used herein, unless otherwise specified, the term "submerged mycelial biomass" refers to mycelial biomass produced by a submerged fermentation process.

[0099] As used herein, unless otherwise specified, the term "submerged mycelial biomass form" refers to a mycelial biomass form in which the mycelial biomass has been produced by a submerged fermentation process.

[0100] As used herein, unless otherwise specified, the term "vegan" refers to a food product that is substantially free of food components or ingredients, such as protein, derived from animals. Specific examples of non-vegan food ingredients or products include blood, eggs, isinglass, meat (and its components, e.g., animal fat), milk, rennet, and foods made using any one or more of these ingredients (e.g., ice cream, mayonnaise, etc.). As disclosed herein, some vegan food products may be analogs of non-vegan food products.

[0101] The present disclosure provides methods for inducing aggregation of filamentous fungal proteins in liquid dispersions of filamentous fungal proteins, as well as solid and / or colloidal fungal food products (e.g., cheese and cheese curd analog food products, tofu analog food products, edible gels, etc.) made by such methods. According to various embodiments of the present disclosure, aggregation of filamentous fungal proteins is induced by one or more of the following: (1) adjusting the pH of the liquid dispersion of filamentous fungal particles (e.g., by adding one or more acids or bases); (2) adding one or more functional ingredients (i.e., proteins and / or carbohydrates) to the liquid dispersion of filamentous fungal particles; and / or (3) adding one or more salts to the liquid dispersion of filamentous fungal particles. In some embodiments, this aggregation results in the formation of a fungal curd, i.e., a solid mass of filamentous fungal proteins (and possibly other components) that is separable from the remaining liquid phase, similar to the technique of coagulating proteins from animal milk (to form curd) or soy milk (to form tofu), while in other embodiments, aggregation results in the formation of a gel material, i.e., a phase that retains its shape and resists flow, in which the liquid phase is dispersed throughout the network formed by the filamentous fungal proteins (and possibly other compounds). The fungal curd or gel material can then be processed into any of a wide range of fungal food products, such as, by way of non-limiting examples, cheese and cheese curd-like food products, tofu-like food products, edible gels, etc.

[0102] edible filamentous fungi Edible filamentous fungi, particularly filamentous fungal mycelial biomass, can be used as a source of nutrients such as protein, either alone or incorporated into foodstuffs, such as the disclosed solid and / or colloidal fungal food products. Described herein are solid and / or colloidal fungal food products comprising aggregated proteins of edible filamentous fungi.

[0103] Suitable filamentous fungi for use in the disclosed methods are selected from the phyla or divisions Zygomycota, Glomermycota, Chytridiomycota, Basidiomycota, or Ascomycota. The Basidiomycota phylum includes, among other orders, Agaricales, Russulales, Polyporales, and Ustilaginales; the Ascomycota phylum includes, among other orders, Pezizales and Hypocreales; and the Zygomycota phylum includes, among other orders, Mucorales. In some embodiments, the edible filamentous fungus particles of the present invention belong to an order selected from Ustilaginales, Russulales, Polyporales, Agaricales, Pezizales, Hypocreales, and Mucorales. In some embodiments, the filamentous fungi of the order Ustilaginales are selected from the family Ustilaginaceae. In some embodiments, the filamentous fungi of the order Russulales are selected from the family Hericiaceae. In some embodiments, the filamentous fungi of the order Polyporales are selected from the families Polyporaceae or Grifolaceae.In some embodiments, the filamentous fungi of the order Agaricales are selected from the families Lyophyllaceae, Strophariaceae, Lycoperdaceae, Agaricomaceae, Pleurotaceae, Physalacriaceae, or Omphalotaceae. In some embodiments, the filamentous fungi of the order Pezizales are selected from the families Tuberaceae or Morchellaceae. In some embodiments, the filamentous fungi of the order Mucorales are selected from the family Mucoraceae. In some embodiments, the filamentous fungus is from the genera Fusarium, Aspergillus, Trichoderma, and / or Rhizopus.

[0104] Examples of species of filamentous fungi suitable for use in the methods provided by the present disclosure include Ustilago esculenta, Hericululm erinaceus, Polyporous squamosus, Grifola fondrosa, Hypsizygus marmoreus, Hypsizygus ulmariuos (Elm Oyster), Calocybe gambosa, Pholiota nameko, Calvatia gigantea, Agaricus bisporus, Stropharia rugosoannulata, Hypholoma lateritium, Pleurotus eryngii, and the like. eryngii, Pleurotus ostreatus (pearl), Pleurotus ostreatus var. columbinus (blue oyster), Tuber borchii, Morchella esculenta, Morchella conica, Morchella importuna, Sparassis crispa (cauliflower), Fusarium venenatum, Fusarium strain flavolapis (ATCC accession number PTA-10698), Disciotis venosa, Cordyceps militaris, Ganoderma lucidum (Reishi), Flammulina velutipes, Lentinula edodes, and Ophiocordyceps sinensis.Further examples include, but are not limited to, Trametes versicolor, Ceriporia lacerate, Pholiota gigantea, Leucoagaricus holosericeus, Pleurotus djamor, Calvatia fragilis, Handkea utriformis, and Rhizopus oligosporus.

[0105] In some embodiments, the filamentous fungus is a Fusarium species. In some embodiments, the filamentous fungus is Fusarium strain flavolapis (ATCC Accession No. PTA-10698), which may also be referred to herein as "F. flavolapis" or "Ff." In some embodiments, the filamentous fungus is Fusarium venenatum.

[0106] Because they are edible, filamentous fungi suitable for use in the methods and compositions provided by the present disclosure typically have a low mycotoxin content. In some embodiments, the total amount of mycotoxins in the filamentous fungi utilized in the disclosed methods and compositions is less than about 10 ppm.

[0107] The amount of edible filamentous fungal biomass used in the disclosed methods can vary based on the desired protein content, texture, and / or flavor of the final solid and / or colloidal product. In various embodiments, the disclosed liquid dispersions can be prepared using an amount of fungal biomass ranging from about 0.25% to about 10% by weight, or any subrange therein. As used herein, weight percent is expressed as the weight of the component divided by the total weight of the composition; for example, if 1 g of filamentous fungal biomass is dispersed in 100 mL (i.e., 100 g) of water, the resulting dispersion would be expressed as a 1% by weight fungal dispersion. In some embodiments, the amount of edible filamentous fungal biomass used in the disclosed liquid dispersions is from about 1.0% to about 10.0% by weight, or any value within any subrange therein. In some embodiments, the amount of edible filamentous fungal biomass used in the disclosed liquid dispersions is from about 2.5% to about 4.0% by weight. In some embodiments, the amount of edible filamentous fungal biomass used in the disclosed liquid dispersions is from about 2.7% to about 4.0% by weight. In some embodiments, the amount of edible filamentous fungus used in the disclosed liquid dispersions is selected from 2.0 wt%, 2.1 wt%, 2.2 wt%, 2.3 wt%, 2.4 wt%, 2.5 wt%, 2.6 wt%, 2.7 wt%, 2.8 wt%, 2.9 wt%, 3.0 wt%, 3.1 wt%, 3.2 wt%, 3.3 wt%, 3.4 wt%, 3.5 wt%, 3.6 wt%, 3.7 wt%, 3.8 wt%, 3.9 wt%, 4.0 wt%, 4.1 wt%, 4.2 wt%, 4.3 wt%, 4.4 wt%, 4.5 wt%, 4.6 wt%, 4.7 wt%, 4.8 wt%, 4.9 wt%, and 5.0 wt%, and any subrange from about 1.0 wt% to about 10.0 wt%.

[0108] Liquid dispersion of filamentous fungal particles The methods provided by the present disclosure utilize liquid dispersions made from one or more filamentous fungi. The liquid dispersions include particles of filamentous fungi dispersed in a liquid medium, most typically an aqueous medium (i.e., a medium that is or includes water). In the case of cohesive fungal biomass, the fungal biomass is typically reduced in size for use in the disclosed dispersions, which can be accomplished by cutting, shredding, dicing, chopping, grinding, blending, sonication, or other means. Typically, some form of size reduction occurs prior to mixing with the liquid medium.

[0109] In various embodiments, liquid dispersions are prepared by combining and blending one or more mycelial biomass forms with a liquid phase, most commonly an aqueous (i.e., water or water-containing) phase. The liquid dispersions utilized in the disclosed methods are preferably stable so that the filamentous fungal particles do not readily separate from the liquid medium in which they are dispersed. For example, in various embodiments, upon forming the dispersion, the resulting fluid composition may appear homogeneous in appearance and / or may not visually separate into distinct phases, such that no visually noticeable or significant fungal sedimentation forms at the bottom of any container holding the dispersion.

[0110] Blending can be performed at various speeds and times. It will be appreciated that typical blending equipment combines two or more components through the use of moving or rotating blades. Therefore, longer blending times and / or higher blending speeds will often result in smaller fungal particles in the dispersion. In some embodiments, the fungal material is blended in the liquid medium until it is reduced to fine particles. In some embodiments, the particle size of the fungal material is the same as or similar to the particle size of conventional flours, such as wheat flour, typically from about 30 μm to about 400 μm, most typically from about 75 μm to about 120 μm. In some other embodiments, finer particles can be used, and the average particle size of the fungal material may be about 10 μm or less, and / or at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles may have a particle size of about 1 μm to about 10 μm, or alternatively, any subrange within this range. In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles are less than 10 μm, less than 9 μm, less than 8 μm, less than 7 μm, less than 6 μm, less than 5 μm, less than 4 μm, less than 3 μm, less than 2 μm, or less than 1 μm. In some embodiments, at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90% of the particles may have a particle size less than about 1 μm.

[0111] Typically, blending is carried out at a speed and for a period of time sufficient to produce filamentous fungal particles in the liquid having a particle size of about 500 μm or less. In some embodiments, blending is or includes high-shear blending, where the fungus and water are blended at a speed of at least about 10,000 rpm for at least 2 minutes. Additionally or alternatively, in some embodiments, blending is or includes high-pressure homogenization, where the fungus and water are blended at very high pressure (e.g., about 20,000 psi) to cause lysis of the fungal cells. In some embodiments, the blended mixture is gradually heated during blending, in some embodiments to the boiling point of water, to facilitate production of the dispersion. Once blending is complete, the heated mixture may, in some embodiments, be cooled before further use in the disclosed methods.

[0112] The mass ratio of filamentous fungal biomass to liquid phase (e.g., water) can be adjusted to produce a liquid dispersion with a desired consistency and density. The biomass to water ratio is generally about 1:50 to about 50:1, or any range therebetween. In some embodiments, the fungal biomass to water mass ratio is about 1:30, about 1:20, about 1:10, about 1:9, about 1:8, about 1:7, about 1:6, about 1:5, about 1:4, about 1:3, about 1:2, about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, or about 10:1.

[0113] In some embodiments, the filamentous fungal particles in the dispersion consist essentially of fungal mycelia. In some embodiments, the filamentous fungal particles in the dispersion comprise from about 50% to about 95% fungal mycelia by weight. In some embodiments, the filamentous fungal particles in the dispersion comprise at least about 50% fungal mycelia by weight, in some embodiments at least about 75% fungal mycelia by weight, and in some embodiments at least about 95% fungal mycelia by weight. In some embodiments, the dispersion comprises at least about 4% fungal solids. In other embodiments, the dispersion has a solids content of from about 4% to about 30%, typically from about 5% to about 15%, and most typically from about 6.25% to about 10%.

[0114] In some embodiments, the filamentous fungal particle includes both fungal mycelium and one or more fruiting bodies (or portions thereof). Some species of fungi produce multicellular fruiting bodies due to sexual reproduction and spore development. Fruiting bodies are the part of fungi that typically comes to mind when considering the term "mushroom." Fruiting bodies include mushroom caps, stems, gills, skirts, scales, vases, etc. While fruiting bodies grow above a growth substrate, mycelium typically grows within or beneath the substrate from which the mycelium obtains resources (e.g., soil, dead wood, etc.). The fungal threads that form mycelium are known as hyphae and are the nutritional structures of filamentous fungi, through which the fungus absorbs air and / or nutrients from its environment. Fungi typically must produce a certain mass of mycelium before they can produce fruiting bodies; i.e., the mycelium must collect the resources the fungus needs for fruiting body production. When fruiting body-forming filamentous fungi are used in liquid dispersions according to the present disclosure, the filamentous fungal particles of the liquid dispersion may, in some embodiments, be formed entirely or predominantly from fruiting bodies. Additionally or alternatively, the filamentous fungal particles in the liquid dispersions according to the present disclosure may be derived from fungal biomass that includes conidia. In some embodiments, the filamentous fungal particles of the liquid dispersion may comprise a mixture of mycelium, conidia, and fruiting body material in any proportion.

[0115] In some embodiments, the liquid dispersion is prepared under nitrogen, which results in a creamier liquid dispersion with less of a fungal odor. Preparation under nitrogen can be achieved by bubbling nitrogen gas into a sealed container to displace the available oxygen (and other non-nitrogen gases) in the container with nitrogen. This can be done during blending while the initial dispersion is being created.

[0116] The amount of liquid dispersion used in the methods disclosed herein, i.e., the total amount of edible filamentous fungal biomass and liquid dispersion medium, can vary based on the desired protein content, texture, and / or flavor of the final solid and / or colloidal food product. In various embodiments, an amount of dispersion ranging from about 80% to about 95% by weight can be used in the disclosed methods. In some embodiments, the amount of liquid dispersion used in the disclosed methods is from about 83% to about 91% by weight.

[0117] Mixed mycelial biomass composition Some embodiments of the present disclosure include mycelial biomass compositions comprising two or more different mycelial biomass forms. In some embodiments, particularly liquid dispersions of filamentous fungal biomass, the compositions may comprise two or more different mycelial biomass forms, the proteins of which can aggregate under a desired set of conditions (e.g., within a selected pH range) to form a fungal curd, gel, or other stable colloid. These compositions may comprise (i) at least one cohesive mycelial biomass form and at least one submerged mycelial biomass form, (ii) at least two different submerged mycelial biomass forms, and / or (iii) at least two different cohesive mycelial biomass forms; compositions characterized by one or more of these conditions (i) through (iii) are referred to herein as "mixed-form mycelial biomat compositions." A first non-limiting example of a mixed-type mycelial biomass composition according to these embodiments is a mycelial biomass composition comprising soaked dough as a first mycelial biomass type and biomat pieces (or a liquid dispersion thereof) as a second mycelial biomass type. A second non-limiting example of a mixed-type mycelial biomass composition according to these embodiments is a mycelial biomass composition comprising soaked dough as a first mycelial biomass type and soaked flour as a second mycelial biomass type. A third non-limiting example of a mixed-type mycelial biomass composition according to these embodiments is a mycelial biomass composition comprising biomat pieces (or a liquid dispersion thereof) as a first mycelial biomass type and soaked flour as a second mycelial biomass type.

[0118] The mixed-form mycelial biomass composition according to the present disclosure may have significant and important advantages and / or benefits over many conventional mycelial biomass compositions. In particular, the inventors have surprisingly and unexpectedly found that by combining at least one cohesive mycelial biomass form and at least one immersed mycelial biomass form, and / or at least two different immersed mycelial biomass forms, and / or at least two different cohesive mycelial biomass forms into a single mycelial biomass composition in the form of a liquid dispersion, the liquid dispersion can form a fungal curd or a stable gel by the method for aggregating fungal proteins disclosed herein and / or under conditions that can be controlled, optimized, selected, and / or adjusted for a desired application (e.g., a specific pH range, for example, particularly a pH value of about 4 or less, even more particularly a pH value of about 3.5), and that previous and / or conventional mycelial biomass compositions cannot be made to form a fungal curd or a gel or other stable colloid by the same method or under the same conditions. This is an important or necessary characteristic for making many fungal food products (e.g., mayonnaise-like food products, colloidal sauces, salad dressings, etc.). In certain embodiments of the present disclosure, mycelial biomass compositions of the present disclosure can be formed that behave more fluidly at a pH above about 4, begin to gel when the pH is reduced to below about 4, and form a viscous gel when the pH is further reduced to about 3.5, and the change in rheology under each of these conditions can be determined by a rheometer. Without wishing to be bound by theory, the inventors hypothesize that this advantageous and beneficial characteristic of the mycelial biomass compositions of the present disclosure is the result of molecular interactions between proteins of one mycelial biomass form and proteins of another mycelial biomass form, and / or between proteins of one mycelial biomass form and oligosaccharides and / or polysaccharides of another mycelial biomass form.

[0119] A further advantage and / or benefit of the mixed format mycelial biomass composition of the present disclosure compared to many conventional mycelial biomass compositions is that the formation of fungal curds or stable colloids may be reversible, and / or the stable colloid may be configured to transition from one type of colloid to another type of colloid by reversing the process step that caused the aggregation of fungal proteins in the mixed format mycelial biomass composition (e.g., by changing the pH to a pH outside the range in which the proteins of the composition aggregate). As a non-limiting example, a mixed format mycelial biomass composition according to the present disclosure may form a stable gel (i.e., a stable colloid in which the dispersed phase is a liquid and the dispersion medium is a solid) at a pH of about 4 or below (and in some embodiments, specifically a pH of about 3.5) due to the aggregation of fungal proteins, but raising the pH to at least about 5 may reverse the aggregation and cause the gel to collapse (i.e., the gel may separate into separate phases) and / or the gel may transition to a different type of colloid (e.g., an emulsion, sol, etc.).

[0120] Most typically, the mixed-format mycelial biomass compositions of the present disclosure comprise (i) at least one cohesive mycelial biomass format and at least one submerged mycelial biomass format, and / or (ii) at least two different submerged mycelial biomass formats. Without wishing to be bound by any particular theory, the inventors hypothesize that these combinations of multiple mycelial biomass formats allow for the production of mycelial biomass compositions having a combination of advantageous or beneficial characteristics of each of the two (or more) mycelial biomass formats, or even that in some embodiments, molecular interactions between the two or more mycelial biomass formats in the mycelial biomass composition allow for synergistic advantages or benefits (i.e., advantages or benefits that cannot be achieved by any single mycelial biomass format alone).

[0121] A first non-limiting example of a submerged mycelial biomass format suitable for use in a mixed-format mycelial biomass composition according to the present disclosure is a submerged slurry. Submerged slurries generally consist of a biomass-containing fluid (e.g., fermentation broth or water) and are produced by growing filamentous fungal biomass in a fermentation medium using a submerged fermentation process. In embodiments, the fermentation broth itself, together with accumulated biomass, constitutes the submerged slurry. In embodiments, the submerged slurry is produced by separating the fungal biomass from the liquid fermentation broth, such as by filtration or decanting, washing the separated biomass, such as with water, to remove the fermentation medium, and resuspending the washed biomass in water to obtain the submerged slurry. In embodiments, the submerged slurry has a water content level of approximately 85% to 99%.

[0122] A second non-limiting example of a suitable steeped mycelial biomass format for use in a mixed-format mycelial biomass composition according to the present disclosure is steeped dough. Steeped doughs are biomasses that generally have a dough-like consistency (i.e., they behave like a wet, solid mass) and are produced by dewatering steeped mycelial biomass, such as steeped slurry. Steeped doughs have a water content of approximately 60-85%.

[0123] A third non-limiting example of a steeped mycelial biomass format suitable for use in a mixed-format mycelial biomass composition according to the present disclosure is steeped flour. Steeped flour is a relatively fine, low-moisture fungal particle produced by drying and grinding or milling steeped slurry or dough. In embodiments, steeped flour has a water content of approximately 1-15%. In embodiments, steeped flour has a water content of 5-10%. Most typically, the production of steeped flour requires the removal of a greater proportion of water from the output of the steeped-fermentation process than the production of steeped dough (i.e., the steeped dough has a higher water content than the steeped flour).

[0124] A fourth non-limiting example of a suitable steeped mycelial biomass format for use in a mixed-format mycelial biomass composition according to the present disclosure is a steeped dispersion. A steeped dispersion is a flowable composition (and in some embodiments, a sol, i.e., a colloid in which the mycelial biomass is in particulate form and dispersed throughout a liquid dispersion medium) produced by (i) blending a steeped slurry and / or (ii) mixing a steeped dough with a liquid and blending the mixture. In embodiments, the steeped dispersion has a water content of 80-99%. In embodiments, the steeped dispersion has a water content of 85-95%.

[0125] A fifth non-limiting example of a suitable soaked mycelium biomass format for use in a mixed-format mycelium biomass composition according to the present disclosure is soaked spray-dried powder. Soaked spray-dried powders are relatively fine fungal particles produced by spray-drying a soaked dispersion. In embodiments, the soaked spray-dried powder has a water content of 1-10% or less. In embodiments, the soaked spray-dried powder has a water content of less than 6%.

[0126] A first non-limiting example of a suitable cohesive mycelial biomass format for use in a mixed-format mycelial biomass composition according to the present disclosure is biomat pieces. Biomat pieces are pieces of biomat (i.e., mycelial biomass formed by a suitable non-submerged fermentation process, such as a liquid surface fermentation process, a solid surface (or solid substrate) fermentation process, a membrane surface fermentation process, and a mesh surface fermentation process) that have been reduced in size or otherwise divided into pieces by cutting or other size-reducing techniques. In embodiments, the biomat pieces have 60-85% water. In embodiments, the biomat pieces have 70-80% water. In embodiments, the cohesive mycelial biomass and biomat pieces derived therefrom do not include the feedstock on which the filamentous fungal biomass was grown.

[0127] A second non-limiting example of a cohesive mycelial biomass format suitable for use in a mixed-format mycelial biomass composition according to the present disclosure is biomat flour. Biomat flour is a relatively fine, low-moisture particle in the form of a powder produced by drying and grinding biomat. In embodiments, the biomat flour has a moisture content of 4-14%. In embodiments, the biomat flour has a moisture content of 5-10%. In embodiments, the biomat flour has a moisture content of less than 6%.

[0128] A third non-limiting example of a suitable cohesive mycelial biomass format for use in a mixed-format mycelial biomass composition according to the present disclosure is a biomat dispersion. A biomat dispersion is a flowable composition (and in some embodiments, a sol, i.e., a colloid in which the mycelial biomass is in particulate form and dispersed throughout a liquid dispersion medium) produced by mixing biomat, biomat pieces, or biomat flour with a liquid. In embodiments, the biomat dispersion has a water content of 80-99.9%. In embodiments, the biomat dispersion has a water content of 85-95%.

[0129] A fourth non-limiting example of a suitable cohesive mycelial biomass format for use in the mixed-format mycelial biomass compositions of the present disclosure is spray-dried biomat powder. Spray-dried biomat powder is a relatively fine, multi-layer fungal particle produced by spray-drying a biomat dispersion or other fluid containing biomat, biomat pieces, or biomat powder. In embodiments, the spray-dried biomat powder has a water content of less than 1-10%. In embodiments, the spray-dried biomat powder has a water content of less than 6%, less than 5%, less than 4%, less than 3%, or less than 2%.

[0130] In the mixed-form mycelial biomass compositions of the present disclosure comprising at least a first mycelial biomass form and a second mycelial biomass form, the weight ratio of the first mycelial biomass form to the second mycelial biomass form can be any value in any range that enables the mycelial biomass composition to have any one or more of the advantageous and beneficial chemical and / or material properties described herein, such as the ability to form stable gels or other stable colloids under acidic conditions. More generally, the weight ratio of the first mycelial biomass form to the second mycelial biomass form can be in any range having a lower limit of A:B and an upper limit of C:D, where A and B are integers whose sum is 100, C and D are integers whose sum is 100, and A is less than C, and / or in certain embodiments, the weight ratio of the first mycelial biomass form to the second mycelial biomass form can be from about 1:10 to about 10:1 (or any value of any subrange thereof, e.g., about 1:9, about 1:8, about 1:7, about 1:6, about 1:5, about 1:4, about 1:3, about 1:2, about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, or about 9:1). As a first non-limiting example, if the first mycelial biomass form is a soaked dough and the second mycelial biomass form is a biomat piece (or a liquid dispersion thereof), the weight ratio of soaked dough to biomat piece can be about 75:25. As a second non-limiting example, if the first mycelial biomass form is a soaked dough and the second mycelial biomass is a soaked powder, the weight ratio of soaked dough to soaked powder can be about 40:60.

[0131] A first non-limiting example of a type of food material that can be made from a mixed mycelial biomass composition according to the present disclosure is flour, which can have a particle size of 30-400 μm, but need not be. The powder may be about 400 μm or less, about 390 μm or less, about 380 μm or less, about 370 μm or less, about 360 μm or less, about 350 μm or less, about 340 μm or less, about 330 μm or less, about 320 μm or less, about 310 μm or less, about 300 μm or less, about 290 μm or less, about 280 μm or less, about 270 μm or less, about 260 μm or less, about 250 μm or less, about 240 μm or less, about 230 μm or less, about 220 μm or less, about 210 μm or less, about 200 μm or less, about 190 μm or less, about 180 μm or less, about 170 μm or less, about 160 μm or less, about 150 μm or less, about 140 μm or less The particle size may, but need not, be less than about 130 μm, less than about 120 μm, less than about 110 μm, less than about 100 μm, less than about 90 μm, less than about 80 μm, less than about 70 μm, less than about 60 μm, less than about 50 μm, less than about 40 μm, less than about 30 μm, less than about 20 μm, less than about 10 μm, less than about 9 μm, less than about 8 μm, less than about 7 μm, less than about 6 μm, less than about 5 μm, less than about 4 μm, less than about 3 μm, less than about 2 μm, or less than about 1 micron, or alternatively less than about any integer number of μm between about 1 micron and about 400 μm. 10 Particle size, D 25 Particle size, D 50 Particle size, D 75 Particle size, D 90 The particle size may be any one or more of the particle size, weight average particle size, or weight average particle size. In some embodiments, substantially all of the particles may have a particle size of at least about 30 μm and no greater than about 400 μm.

[0132] In embodiments where the food material made from the mixed format mycelial biomass composition of the present disclosure is a flour, the particle size and particle size distribution can be the same as or similar to those conventional for powdery materials such as wheat or other flours. In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles are in the range of 0.03 mm to about 0.4 mm, or alternatively, any subrange within this range, e.g., about 0.03 mm to 0.07 mm, about 0.07 mm to about 0.12 mm, about 0.12 mm to about 0.15 mm, about 0.15 mm to about 2.0 mm, about 0.04 mm to about 0.2 mm, or 0.06 mm to about 0.120 mm, or 0.2 mm to about 0.4 mm. In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles are in the range of 0.075 mm to about 0.12 mm.

[0133] In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the mass of the particles is within the range of 0.03 mm to about 0.4 mm, or alternatively any subrange within this range, e.g., about 0.03 mm to 0.07 mm, about 0.07 mm to about 0.12 mm, about 0.12 mm to about 0.15 mm, about 0.15 mm to about 2.0 mm, about 0.04 mm to about 0.2 mm, or 0.06 mm to about 0.120 mm, or 0.2 mm to about 0.4 mm. In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the mass of the particles is within the range of 0.075 mm to about 0.12 mm.

[0134] In some embodiments, the particles of one or more mycelial biomass forms present in the flour can be size reduced. Size reduction can be performed using a mill, grinder, or other conventional equipment for size reduction.

[0135] In some embodiments, the moisture content of the particles of one or more mycelial biomass forms is less than about 15%, about 14%, about 13%, about 12%, about 11%, about 10%, about 9%, about 8%, about 7%, about 6%, about 5%, about 4%, about 3%, about 2%, or about 1%. Low moisture levels help prevent particle agglomeration.

[0136] Flours made from the mixed form mycelial biomass compositions according to the present disclosure are useful in preparing food ingredients such as baked goods, including but not limited to bread, rolls, muffins, cakes, cookies, pies, etc., or can be sprinkled on other food products.

[0137] A second non-limiting example of a type of food material that can be made from a mixed mycelial biomass composition according to the present disclosure is a plurality of solid particles other than flour. In such a plurality of solid particles other than flour, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles can have a particle length of about 0.05 mm to about 500 mm, a particle width of about 0.03 mm to about 7 mm, and a particle height of about 0.03 mm to about 1.0 mm, or alternatively, any subrange within these ranges. For example, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles may have a particle length of about 0.08 mm to about 100 mm, or 10 mm to about 70 mm, or 130 mm to about 200 mm, a particle width of about 0.05 mm to about 2 mm, or about 1 mm to about 3 mm, or about 4 mm to about 6 mm, and a particle height of about 0.03 mm to about 0.06 mm, or about 0.04 mm to about 0.07 mm, or about 0.08 mm to about 1.0 mm.

[0138] In some embodiments, at least one mycelial biomass form in a mixed form mycelial biomass composition according to the present disclosure is in the form of particles, wherein at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the mass of the particles have a particle length of about 0.05 mm to about 500 mm, a particle width of about 0.03 mm to about 7 mm, and a particle height of about 0.03 mm to about 1.0 mm, or alternatively any subrange within these ranges. For example, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% by mass of the particles may have a particle length of about 0.08 mm to about 100 mm, or 10 mm to about 70 mm, or 130 mm to about 200 mm, a particle width of about 0.05 mm to about 2 mm, or about 1 mm to about 3 mm, or about 4 mm to about 6 mm, and a particle height of about 0.03 mm to about 0.06 mm, or about 0.04 mm to about 0.07 mm, or about 0.08 mm to about 1.0 mm. For example, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles may have a particle length of about 0.08 mm to about 100 mm, or 10 mm to about 70 mm, or 130 mm to about 200 mm, a particle width of about 0.05 mm to about 2 mm, or about 1 mm to about 3 mm, or about 4 mm to about 6 mm, and a particle height of about 0.03 mm to about 0.06 mm, or about 0.04 mm to about 0.07 mm, or about 0.08 mm to about 1.0 mm.

[0139] In some embodiments of food materials made from a mixed format mycelial biomass composition according to the present disclosure in the form of a plurality of solid particles other than flour, at least one mycelial biomass format can mimic the texture and chewiness of meat products such as chicken nuggets or hamburgers, and thus the food materials may be useful in the production of meat analogs and / or meat fillers and / or extenders. In use, meat product fillers or extenders according to the present disclosure may be adapted to be mixed with animal meat in a ratio of 10:90 to 90:10, or any subrange therebetween.

[0140] In some embodiments of food materials made from a mixed-format mycelial biomass composition according to the present disclosure in the form of multiple solid particles other than flour, at least one mycelial biomass format may comprise particles, wherein at least 90% of the particles have a length of less than about 1.5 mm, with the majority of the length being 1 mm or less, a width of less than about 1 mm, and a height of less than about 0.75 mm. Mycelial biomass formats comprising such particles may be characterized by a higher perceived density in the mouth, may be easier to chew, may present a creamier mouthfeel and a more refined eating experience, and / or may be used to prepare food materials similar to hamburgers found in fine dining establishments.

[0141] In some embodiments of food materials made from mixed-format mycelial biomass compositions according to the present disclosure in the form of multiple solid particles other than flour, at least one mycelial biomass format may comprise particles, wherein at least about 90% of the particles have a length of about 4 mm to about 10 mm, a width of about 1.0 mm to about 3 mm, and a height of less than 0.75 mm. Mycelial biomass formats comprising such particles may be found to provide a more substantial eating experience, similar to the type of burger commonly found at specialty burger restaurants or barbecues.

[0142] A third non-limiting example of the type of food material that can be made from mixed-form mycelial biomass compositions according to the present disclosure is a liquid dispersion, particularly a dispersion of particles of two or more mycelial biomass forms in an aqueous liquid. In some embodiments, the liquid dispersion can be a substitute for milk or milk analogs.

[0143] In some embodiments of food materials made from mixed-type mycelial biomass compositions according to the present disclosure in the form of a liquid dispersion, at least one mycelial biomass form may comprise particles less than about 10 μm. In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles of the at least one mycelial biomass form may be within the particle size range of about 1 μm to about 10 μm, or alternatively, any subrange within this range. In some embodiments, at least 50%, or at least 60%, or at least 70%, or at least 80%, or at least 90% of the particles of the at least one mycelial biomass form may have a particle size less than 10 μm, less than 9 μm, less than 8 μm, less than 7 μm, less than 6 μm, less than 5 μm, less than 4 μm, less than 3 μm, less than 2 μm, or less than 1 micron. In some embodiments, at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90% of the particles of the at least one mycelial biomass form may have a particle size of less than about 1 micron.

[0144] In some embodiments of food materials made from mixed-type mycelial biomass compositions according to the present disclosure in the form of a liquid dispersion, the weight ratio of one or more mycelial biomass forms to water can be adjusted to produce a liquid dispersion of appropriate consistency and density. The ratio of one or more mycelial biomass forms to water can range from about 1:10 to about 10:1, or any range therebetween. In some embodiments, the ratio of one or more mycelial biomass forms to water can be about 1:10, about 1:9, about 1:8, about 1:7, about 1:6, about 1:5, about 1:4, about 1:3, about 1:2, about 1:1, about 2:1, about 3:1, about 4:1, about 5:1, about 6:1, about 7:1, about 8:1, about 9:1, or about 10:1.

[0145] In some embodiments, the liquid dispersion food material according to the present disclosure is stable such that at least one mycelial biomass form does not readily separate from the liquid medium, and preferably such that none of the mycelial biomass forms readily separate. For example, upon forming the dispersion, the dispersion may appear homogeneous in appearance and may not separate into visually distinct phases, and / or no visually discernible or significant sedimentation may form at the bottom of the container holding the dispersion. In some embodiments, the liquid dispersion can be stored at room temperature and / or refrigerated temperatures, e.g., about 35°F (1.6°C), for at least about 1 hour, at least about 2 hours, at least about 3 hours, at least about 4 hours, at least about 5 hours, at least about 6 hours, at least about 7 hours, at least about 8 hours, at least about 9 hours, at least about 10 hours, at least about 11 hours, at least about 12 hours, at least about 13 hours, at least about 14 hours, at least about 15 hours, at least about 16 hours, at least about 17 hours, at least about 18 hours, at least about 19 hours, at least about 20 hours, at least about 21 hours, at least about 22 hours, at least about 23 hours, at least about 24 hours, at least about 25 hours, at least about 26 hours, at least about 27 hours, at least about 28 hours, at least about 30 hours, at least about 31 hours, at least about 32 hours, at least about 33 hours, at least about 34 hours, at least about 35 hours, at least about 36 hours, at least about 37 hours, at least about 38 hours, at least about 39 hours, at least about 40 hours, at least about 41 hours, at least about 42 hours, at least about 43 hours, at least about 44 hours, at least about 45 hours, at least about 46 hours, at least about 47 hours, at least about 48 hours, at least about 49 hours, at least about 50 hours, at least about 51 hours, at least about 52 hours, at least about 53 hours, at least about 54 hours, at least about 55 hours, at least about 56 hours, at least about 57 hours, at least about 58 hours, at least about 59 hours, at least about 60 hours, at least about 61 or may remain stable for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 1 week, at least about 2 weeks, at least about 3 weeks, at least about 4 weeks, at least about 1 month, at least about 2 months, at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 12 months, at least about 13 months, at least about 14 months, at least about 15 months, at least about 16 months, at least about 17 months, or at least about 18 months.

[0146] In some embodiments, a liquid dispersion food material according to the present disclosure may comprise at least about 4%, at least about 5%, at least about 6%, at least about 7%, at least about 8%, at least about 9%, at least about 10%, at least about 11%, at least about 12%, at least about 13%, at least about 14%, at least about 15%, at least about 16%, at least about 17%, at least about 18%, at least about 19%, or at least about 20% solids. In other embodiments, a liquid dispersion food material according to the present disclosure may have a solids content of from about 4% to about 30%, or any subrange between 4% and 30%, and particularly, the solids content may be from about 5% to about 15%, or from about 6.25% to about 10%, in some embodiments.

[0147] The liquid dispersion food ingredients according to the present disclosure can be used as drinks or beverages, including as a substitute for any dairy product such as cow's milk, almond milk, rice milk, soy milk, etc. These food ingredients can also be used in numerous recipes, including recipes for soups, ice cream, yogurt, smoothies, fudge, and candies such as caramels and truffles.

[0148] A fourth non-limiting example of a type of food material that can be made from a mixed-form mycelial biomass composition according to the present disclosure is an emulsion. In an emulsion food material according to the present disclosure, either or both of the dispersed liquid phase and the liquid dispersion medium can be part of one or more mycelial biomass forms or can be separate components from the mycelial biomass forms.

[0149] In some embodiments, the food material may be a particle-stabilized emulsion, otherwise known as a Pickering emulsion. In these embodiments, one or more mycelial biomass forms may stabilize the colloid by adsorbing to the interface between the dispersed phase and the dispersion medium, for example, the interface between the air bubbles and the solid phase in an ice cream-like food product, or the interface between the oil droplets and water in a mayonnaise-like food product.

[0150] The one or more mycelial biomass forms in the emulsion food material according to the present disclosure may have a desired hydrophilic-lipophilic balance (HLB) of from about 3 to about 16, in some embodiments from about 3 to about 6 (e.g., for stabilizing water-in-oil emulsions) or from about 8 to about 16 (e.g., for stabilizing oil-in-water emulsions such as mayonnaise-like food products).

[0151] Another important parameter related to the stability of emulsion food materials according to the present disclosure is the contact angle, i.e., the angle formed by two phase interfaces (typically between a liquid-gas interface, such as at the surface of a liquid droplet, and a liquid-solid interface, such as when the droplet rests on a solid substrate). A low contact angle (e.g., near 0°) indicates high surface energy, as the droplet will tend to spread across and adhere to the solid surface, while a high contact angle (e.g., near 90°) indicates the tendency of the solid surface to repel the droplet. In emulsion food materials according to the present disclosure, the contact angle of a colloidal food composition on a solid surface, such as a silicon wafer, at ambient conditions (e.g., about 25°C and about 1 atm pressure) can generally be between about 45° and about 75°; the surface energy, and therefore the contact angle, of the emulsion food material can, in some embodiments, be controlled, selected, and / or adjusted by the selection of one or more particularly suitable mycelial biomass forms. Without wishing to be bound by any particular theory, it is believed that the selection of one or more particularly suitable mycelial biomass forms allows for the formulation of emulsion food materials with excellent stability, for example, comprising filamentous fungal particles with high water wettability (for stable oil-in-water emulsions), high oil wettability (for stable water-in-oil emulsions), and / or a balance of these two properties.

[0152] A fifth non-limiting example of the type of food material that can be made from the mixed format mycelial biomass composition according to the present disclosure is a foam, particularly a foam that is stable in that it does not spontaneously collapse immediately upon cessation of the foaming process, which may include whipping with a whisk, incorporating compressed gas, or other conventional foaming processes. In some embodiments, a foam food material according to the present disclosure can be made by subjecting a liquid dispersion food material, emulsion food material, and / or sol food material according to the present disclosure to such a foaming process.

[0153] Foam food materials according to the present disclosure are smooth and creamy in appearance and exhibit the presence of bubbles with a distribution of sizes, with larger bubbles tending to pop after standing or pouring, while smaller bubbles persist for a longer period of time to form a stable foam product. Foam food materials according to the present disclosure may stably incorporate air or other gases into the food material to have compositional properties similar to those of liquid dispersion food materials according to the present disclosure. For example, foam food materials according to the present disclosure may have an increase in volume (i.e., overrun) of at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 75%, at least about 100%, at least about 200%, at least about 300%, at least about 400%, or at least about 500% compared to the starting volume of the liquid material before foaming due to the incorporation of air. In various embodiments, the foam food material according to the present disclosure may be refrigerated for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 31 days, at least about 32 days, at least about 33 days, at least about 34 days, at least about 35 days, at least about 36 days, at least about 37 days, at least about 38 days, at least about 39 days, at least about 40 days, at least about 41 days, at least about 42 days, at least about 43 days, at least about 44 days, at least about 45 days, at least about 46 days, at least about 47 days, at least about 48 days, at least about 49 days, at least about 50 days, at least about 51 days, at least about 52 days, at least about 53 days, at least about 54 days, at least about 55 days, at least about 56 days, at least about 57 days, at least about 58 days, at least about 59 days, at least about 60 days, at least about 61 days, at least about 62 days, at least about The foam stability may be at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, or at least about 95% for at least about 1 day, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 12 months, at least about 13 months, at least about 14 months, at least about 15 months, at least about 16 months, at least about 17 months, or at least about 18 months.

[0154] In foam food materials according to the present disclosure, an average overrun of about 12% may be suitable for preparing ice cream (with more fat and emulsifiers), frozen yogurt, cheesecake batter, whipped toppings, etc. In some embodiments, the foam food materials may incorporate nitrogen to provide different overrun characteristics.

[0155] A sixth non-limiting example of a type of food material that can be made from a mixed-format mycelial biomass composition according to the present disclosure is a gel. In particular, as disclosed more fully elsewhere throughout this disclosure, a food material according to the present disclosure can be a gel at certain pH ranges (e.g., a pH of about 4 or less and / or a pH of about 3.5), but can take on different physical forms (e.g., a liquid dispersion, emulsion, foam, sol, etc.) at different pH ranges (e.g., a pH of at least about 5).

[0156] A seventh non-limiting example of a type of food material that can be made from the mixed-form mycelium biomass composition according to the present disclosure is a solid foam. In a solid foam food material according to the present disclosure, one important stability parameter is foam stability, i.e., the percentage of the initial volume of the solid foam that is retained by the solid foam after a certain interval, which allows for the creation of a solid foam that does not rapidly collapse spontaneously. The foaming process may involve whipping with a whisk, incorporating compressed gas, or other conventional foaming processes, which will usually result in the formation of gas bubbles of various sizes. Larger bubbles tend to pop after standing or pouring, while smaller bubbles can remain suspended for a long time to form a stable solid foam. A solid foamed food material according to the present disclosure may have an increase in volume (i.e., overrun) due to the incorporation of air, compared to the starting volume of the solid dispersion medium prior to foaming, of at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 75%, at least about 100%, at least about 200%, at least about 300%, at least about 400%, or at least about 500%.In various embodiments, the solid foam food material according to the present disclosure may be refrigerated for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 31 days, at least about 32 days, at least about 33 days, at least about 34 days, at least about 35 days, at least about 36 days, at least about 37 days, at least about 38 days, at least about 39 days, at least about 40 days, at least about 41 days, at least about 42 days, at least about 43 days, at least about 44 days, at least about 45 days, at least about 46 days, at least about 47 days, at least about 48 days, at least about 49 days, at least about 50 days, at least about 51 days, at least about 52 days, at least about 53 days, at least about 54 days, at least about 55 days, at least about 56 days, at least about 57 days, at least about 58 days, at least about 59 days, at least about 60 days, at least about 61 days, at least about 62 days, at least The foam may have at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, or at least about 95% foam stability for 8 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months, at least about 4 months, at least about 5 months, at least about 6 months, at least about 7 months, at least about 8 months, at least about 9 months, at least about 10 months, at least about 11 months, at least about 12 months, at least about 13 months, at least about 14 months, at least about 15 months, at least about 16 months, at least about 17 months, or at least about 18 months.

[0157] Further ingredients As described in the following sections of this disclosure, other components that impart beneficial qualities to the aggregated protein composition can be added to the liquid dispersion before, during, or after adjusting the pH, adding functional ingredients, and / or adding salt. For example, flavoring agents can be added to help the aggregated protein composition more similar in taste to a selected non-fungal food product (e.g., cheese, tofu, etc.), dairy fortifiers can be added to impart a creamier texture, and / or taste modifiers can be added to impart a desired mouthfeel, mask undesirable flavor notes, and / or improve the overall taste of the disclosed food products.

[0158] Suitable flavoring agents can include, for example, half and half flavoring, cottage cheese flavoring, milk flavoring, cheese culture flavoring, cream flavoring, butter flavoring, and the like. In the context of vegan foods, such flavoring agents are commercially available preparations containing flavorings that mimic half and half, cottage cheese, milk, butter, and the like. Most commercially available products are water-soluble flavoring agents suitable for addition to aqueous dispersions before, during, or after pH adjustment, addition of functional ingredients, and / or addition of salt. Flavoring agents can be mixed and matched in various ways to achieve the desired flavor profile in the final aggregated protein-containing food product. The amount of any one flavoring agent used in the disclosed methods can vary based on the desired flavor of the final aggregated protein product. In various embodiments, a single flavoring agent can be used in an amount of 0.05% to 3.0% by weight. In some embodiments, the amount of a single flavoring agent used in the disclosed methods is selected from 0.05%, 0.1%, 0.15%, 0.2%, 0.25%, and 0.3% by weight.

[0159] Dairy fortifiers can also be added to the disclosed liquid dispersions before, during, or after pH adjustment, addition of functional ingredients, and / or salt addition. Dairy fortifiers are typically made from natural or artificial flavoring agents and are used to add dairy-like flavor to non-dairy products, to mask and hide flavors, to reduce or increase fat content, to reduce sodium content, and / or to reduce sugar content. Dairy fortifiers are commercially available preparations that are water-soluble and suitable for addition to aqueous dispersions before, during, or after pH adjustment, addition of functional ingredients, and / or addition of salt. Dairy fortifiers can be mixed and matched in various ways to achieve the desired flavor profile in the final aggregated protein-containing food product, although in some particular embodiments, only a single dairy fortifier can be used.

[0160] The amount of any one dairy fortifier used in the disclosed methods can vary based on the desired flavor of the final aggregated protein-containing food product. In various embodiments, a single dairy fortifier can be used in an amount of 0.05% to 3.0% by weight. In some embodiments, the amount of a single dairy fortifier used in the disclosed methods is selected from 0.05%, 0.1%, 0.15%, 0.2%, 0.25%, and 0.3% by weight.

[0161] Taste modifiers can also be added to the disclosed dispersions before, during, or after pH adjustment, addition of functional ingredients, and / or addition of salt. Taste modifiers, such as Modumax® (Royal DSM, NL), are natural compositions that help create an improved taste profile in food products that may contain high-intensity sweeteners, are low in fat, or contain a higher protein content that results in undesirable flavor notes. These modifiers mask certain undesirable flavors and enhance other desirable flavors. For example, botanical masking agents are preparations that mask the green and / or beany flavors of certain botanical ingredients. These are commercially available preparations, typically water-soluble, suitable for addition to liquid dispersions before, during, or after pH adjustment, addition of functional ingredients, and / or addition of salt. Taste modifiers are typically used one at a time in a single formulation, but they can also be mixed and matched in various ways to achieve the desired flavor profile in the final aggregated protein-containing food product.

[0162] The amount of any one taste modulating agent used in the disclosed methods can vary based on the desired flavor of the final aggregated protein-containing food product. In various embodiments, a single taste modulating agent can be used in an amount of 0.01% to 1.0% by weight. In some embodiments, the amount of a single taste modulating agent used in the disclosed methods is selected from 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, and 0.1% by weight.

[0163] Sugars and salts (other than the sugars and / or salts used to induce aggregation of fungal proteins, as further described below) can also be added to the disclosed liquid dispersions to enhance the flavor of the final aggregated protein-containing food product. Any suitable form of food-grade sugar can be used (e.g., dextrose, sucrose, etc.). In some embodiments, the amount of sugar used in the disclosed methods is 1.0% to 3.0% by weight. Any suitable form of food-grade salt, typically sodium salt and / or chloride salt, most typically sodium chloride, can also be used (e.g., sea salt, kosher salt, etc.). In some embodiments, the amount of salt used in the disclosed methods is 0.5% to 0.8% by weight.

[0164] In some embodiments of the methods disclosed herein, an oil and / or solid fat, or one or more oils and / or solid fats, are incorporated into the liquid dispersion of filamentous fungal particles. In a typical embodiment, an oil and / or solid fat can be included in the disclosed liquid dispersion to impart a cheese-like texture to the fungal curd and / or gel formed upon aggregation of the fungal proteins. Any suitable food-grade oil and / or solid fat, or a blend of food-grade oils and / or solid fats, can be used in the disclosed methods. The oils and / or solid fats can be selected for their taste, texture, melting temperature, perceived moisture content, or any combination thereof. For example, some oils, such as olive oil, have a distinctive flavor that can be imparted to the aggregated protein composition. Alternatively, a neutral-flavored oil, such as coconut oil or soybean oil, may be desirable to avoid imparting a strong flavor to the aggregated protein composition.

[0165] Suitable oils for use in the disclosed methods, either alone or in combination with one or more other oils and / or solid fats, include acai oil, almond oil, amaranth oil, apricot oil, argan oil, artichoke oil, avocado oil, bean oil (Moringa oleifera), and the like. oleifera) seeds), blackcurrant seed oil, borage seed oil, Borneo tallow nut oil, buffalo gourd oil, canola oil, carob pod oil, cashew oil, coconut oil, coriander seed oil, corn oil, cottonseed oil, evening primrose oil, false flax oil, grape seed oil, hazelnut oil, hemp oil, kapok seed oil, macadamia nut oil, meadowfoam seed oil, mustard oil, okra seed oil, olive oil, palm oil, peanut oil, pecan oil, perilla oil, peki oil, pine nut oil, pine seed oil, pistachio oil, poppy seed oil, prune kernel oil, pumpkin seed oil, quinoa oil, ramekins oil, rice bran oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, thistle oil, walnut oil, and wheat germ oil.

[0166] Suitable solid fats for use in the disclosed methods, either alone or in combination with one or more other oils and / or solid fats, include marine tallow, butter, chicken fat, clarified butter, cocoa butter, dripping, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, and vegetable shortening.

[0167] The amount of oil and / or solid fat, or combination of oils and / or solid fats, used in the disclosed methods can vary based on the desired texture and / or flavor of the final aggregated protein composition. In various embodiments, amounts of oil and / or solid fat ranging from 4% to 10% by weight can be used in the disclosed methods. In some embodiments, the amount of oil and / or solid fat used in the disclosed methods is selected from 4.0%, 4.5%, 5.0%, 5.5%, 6.0%, 6.5%, 7.0%, 7.5%, 8.0%, 8.5%, 9.0%, 9.5%, and 10% by weight.

[0168] Incorporating oils and / or solid fats into liquid dispersions of filamentous fungal particles can be difficult because oils and / or solid fats are typically immiscible with water, which in most embodiments constitutes all or most of the liquid phase of the liquid dispersion. Therefore, in various embodiments, it is useful to include an emulsifier to promote dispersion stability. Suitable emulsifiers for use in the disclosed methods are typically FDA-approved food additives. Suitable emulsifiers may be synthetic or naturally occurring. For example, many hydrocolloids function as thickeners, aiding in the structure, texture, flavor, and shelf life of various food products. Hydrocolloids are often simply referred to as gums because of the food texture and consistency they produce. Hydrocolloids include emulsifiers made from plant, animal, and aquatic sources. Plant-based hydrocolloids include locust bean gum, carrageenan, pectin, and starch, while animal-sourced variants include chitosan made from crustacean shells. Hydrocolloids such as xanthan gum may also be from microbial sources.

[0169] In some embodiments, emulsifiers suitable for use in the disclosed methods include carboxymethylcellulose, carrageenan, cellulose, guar gum, lecithin, mono- and diglycerides of fatty acids, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, polysorbates, stearoyl lactylate, sorbitan esters, sucrose esters, sucroglycerides, xanthan gum, and combinations thereof.

[0170] The amount of emulsifier used in the disclosed methods can vary. In various embodiments, an amount of emulsifier between 0.1% and 1.0% by weight is used. In some embodiments, the amount of emulsifier used in the disclosed methods is between 0.2% and 0.5% by weight. In some embodiments, the amount of emulsifier used in the disclosed methods is selected from 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, and 1.0% by weight.

[0171] Aggregation of fungal proteins by pH adjustment In some embodiments of the present disclosure, aggregation of the filamentous fungal proteins is induced by adjusting the pH of a liquid dispersion of filamentous fungal particles (e.g., by adding one or more acids or bases). In some embodiments, this aggregation results in the formation of a fungal curd, i.e., a solid mass of filamentous fungal proteins (and optionally other components) that is separable from the remaining liquid phase, similar to the method of coagulating proteins from animal milk (to form curd) or soy milk (to form tofu), while in other embodiments, aggregation results in the formation of a gel material, i.e., a phase that retains its shape and resists flow, in which the liquid phase is dispersed throughout a network formed by the filamentous fungal proteins (and optionally other compounds).

[0172] The filamentous fungal particles in liquid dispersions according to the present disclosure are characterized by an isoelectric point (pI), i.e., a specific pH at which the surface of the particles has no net charge or is electrically neutral; therefore, the relationship between the pH of the liquid dispersion and the pI of the filamentous fungal particles strongly influences the relative affinity with which the filamentous fungal particles interact electrostatically with each other, with molecules of the liquid phase (e.g., water molecules), and / or with molecules of other compounds within the liquid dispersion (e.g., functional components, salts, etc.). Accordingly, without wishing to be bound by any particular theory, the inventors hypothesize that adjusting the pH of a liquid dispersion of filamentous fungal particles may cause aggregation of the filamentous fungal particles by any one or more of several mechanisms.

[0173] As a first non-limiting example, when the liquid dispersion is a mixed-type mycelial biomass composition containing at least two mycelial biomass types with different isoelectric points, there may be a pH range in which particles of the two mycelial biomass types are electrostatically attracted to each other, forming a fungal protein network. In particular, as further described elsewhere throughout this disclosure, the inventors have discovered that mixed-type mycelial biomass compositions containing, for example, biomat pieces and soaked dough, or soaked flour and soaked dough in various ratios, can form a non-flowable gel due to aggregation of fungal proteins within a certain pH range (e.g., around pH 3.5), but that adjusting the pH outside this range (e.g., below about 3 or above about 4) reverses the aggregation, thereby "collapses" the gel, and returns it to a flowable liquid state.

[0174] As a second non-limiting example, when the liquid phase of the liquid dispersion includes a significant amount of polar liquid (e.g., water), there may be a pH range within which the filamentous fungal particles in the liquid dispersion are repelled by, or at least have little or no electrostatic attraction to, the molecules of the polar liquid and are therefore more likely to aggregate with each other or other solid components within the liquid dispersion, whereas outside this pH range, the filamentous fungal particles are attracted to the polar molecules and are therefore more likely to remain stably dispersed in the liquid dispersion. In particular, as further described elsewhere throughout this disclosure, the inventors have found that liquid dispersions of biomat pieces can be prepared that form a non-flowing gel due to aggregation of fungal proteins within a particular pH range (e.g., around about pH 3.5), but that adjusting the pH outside this range (e.g., below about 3 or above about 4) reverses the aggregation, thereby causing the gel to collapse and return to a flowing liquid state.

[0175] As a third non-limiting example, when a liquid dispersion includes a non-fungal protein and / or oligosaccharides and / or polysaccharides, the fungal protein in the liquid dispersion may be subject to different electrostatic interactions with the non-fungal protein and / or oligosaccharides and / or polysaccharides at different pH ranges, depending on the pI of the non-fungal protein and / or oligosaccharides and / or polysaccharides. In particular, as further described elsewhere throughout this disclosure, the inventors have determined that in certain liquid dispersions of filamentous fungal particles containing non-fungal protein (e.g., potato protein, chickpea protein, etc.) and / or oligosaccharides and / or polysaccharides (e.g., maltodextrin), the pH at which biopolymer complexes form (hereinafter referred to as pH Φ If the pI of the fungal particles is different from the pH, Φ When the pH is adjusted toward pH 6.0, fungal proteins and non-fungal components can begin to form soluble complexes ("Stage I" aggregation), and then when ... ΦWe found that when the pH was further adjusted to a range between the pH and the pI of the fungal particles, larger interpolymer complexes could begin to form ("Stage II" aggregation), and finally, when the pH was adjusted even further above the pI of the fungal particles, highly networked gels could begin to form ("Stage III" aggregation).

[0176] As a fourth non-limiting example, when the liquid dispersion includes salt, there may be a pH range in which ionic salt "bridges" form between fungal protein molecules. In particular, as further described elsewhere throughout this disclosure, the inventors have found that fungal proteins in certain liquid dispersions of filamentous fungal particles may aggregate into "fine" networks (i.e., networks with a relatively low concentration of ionic salt "bridges") when the pH is relatively far from the pI of the fungal material and the salt is present at a relatively low ionic strength, and / or may aggregate into "particulate" or "coarse" networks (i.e., networks with a relatively high concentration of ionic salt "bridges") when the pH is relatively near the pI of the fungal material and the salt is present at a relatively high ionic strength.

[0177] Aggregation of fungal proteins by pH adjustment occurs when one or more food-safe acids or bases are introduced into the liquid dispersion to cause aggregation. These acids or bases can be added directly and / or they can be produced by live microbial cultures. In embodiments in which an acid or base is added directly, suitable acids for lowering the pH of the liquid dispersion of filamentous fungal particles include sorbic acid, benzoic acid, formic acid, acetic acid, dehydroacetic acid, lactic acid, propionic acid, boric acid, malic acid, fumaric acid, ascorbic acid, erythorbic acid, citric acid, tartaric acid, phosphoric acid, metatartaric acid, adipic acid, succinic acid, thiodipropionic acid, phytic acid, alginic acid, hydrochloric acid, sulfuric acid, gluconic acid, glutamic acid, guanylic acid, inosinic acid, cyclamic acid, cholic acid, and combinations thereof, and suitable bases for raising the pH of the liquid dispersion of filamentous fungal particles include iron hydroxide, sodium hydroxide, potassium hydroxide, calcium hydroxide, ammonium hydroxide, magnesium hydroxide, and combinations thereof. In embodiments in which the acid or base is produced by a live microbial culture, the microbial culture may include one or more bacteria, molds, or yeasts suitable for fermentation of food products; fermentation by these microorganisms typically results in the conversion of sugars (e.g., lactose) to acids (e.g., lactic acid), thus acidifying the liquid dispersion. Non-limiting examples of microorganisms suitable for use in these embodiments include starter lactic acid bacteria (e.g., Lactococcus lactis ssp. lactis, Lactococcus lactis ssp. cremoris, Streptococcus salivarius ssp. thermophilus, Lactobacillus helveticus, etc.), and non-starter lactic acid bacteria (e.g., Lactobacillus casei ssp. casei, etc.). ssp. casei, Lactobacillus plantarum, etc.In some embodiments, pH adjustment-induced aggregation of fungal proteins can be enhanced by heating and / or cooling the liquid dispersion before, during, and / or after addition of acid or base.

[0178] 1, one non-limiting embodiment of a method 100 for making a composition comprising aggregated fungal proteins (e.g., fungal curd, fungal gel, etc.) by pH adjustment is illustrated. The embodiment of method 100 illustrated in FIG. 1 includes a liquid dispersion preparation step 110, an optional component addition step 120, an optional lipid addition step 130, an acid addition step 140, and an optional heating step 150.

[0179] In liquid dispersion preparation step 110, a liquid dispersion of edible filamentous fungi is prepared by combining water and one or more mycelial biomass forms in a ratio described herein, e.g., about 30:1, 27:1, or 20:1, or alternatively, any ratio that results in a dispersion having a solids content of about 0.1% to about 15% by weight (e.g., in some embodiments where the liquid phase is water, a water content of about 85% to about 99.9% by weight). The water and mycelial biomass forms are placed in a device with rotating blades, such as a blender or impeller, and sheared until smooth. In this embodiment, shearing is carried out by high-speed blending or mixing (e.g., greater than about 10,000 rpm) continuously for a period of at least 2 to 10 minutes, or until a stable, homogenous dispersion is achieved; shear mixing is desirable to achieve sufficient homogeneity of the fungal material in the water and completely break down any aggregates possible. This step can be performed without the addition of heat, but if desired, heat (e.g., 90°F to 120°F, 32.2°C to 48.9°C) can be applied to facilitate the production of a homogeneous mixture.

[0180] In the optional ingredient addition step 120, ingredients such as additional (e.g., non-fungal) protein, flavorings, salt, sugar, etc., can be added to the liquid dispersion. In some embodiments, a mixture of non-fungal proteins is added to the dispersion in a total amount of about 0.25-10% by weight, in some embodiments in a total amount of about 3.0-4.0% by weight, and in some embodiments, about 3.5% by weight. For example, soy protein powder (e.g., soy flour) and an 80% concentration hemp protein powder preparation can be added in equal amounts or in various concentrations. Calcium or magnesium salts can also be added in amounts of about 0.01-0.5% by weight, typically about 0.1-0.3% by weight, and most typically about 0.2% by weight. Similarly, any desired flavoring agents (e.g., half-and-half flavoring, cottage cheese flavoring, milk flavoring, etc.) can be added in a total amount of about 0.4-0.8% by weight, and in some embodiments, about 0.6% by weight. Dairy fortifiers can be added in amounts of about 0.1-0.3% by weight, and in some embodiments, about 0.2% by weight. Any desired flavor modifiers can be added in amounts of about 0.01-0.05% by weight, and in some embodiments, about 0.03% by weight. Salt and sugar can also be added as desired (e.g., in amounts of about 0.5% by weight and about 1.0% by weight, respectively). All of these components are then mixed into the dispersion until completely dissolved. In some embodiments, this optional component addition step 120 may include heating the liquid dispersion, for example, to about 90°F to 120°F (32.2°C to 48.9°C), in some embodiments, 110°F (43.3°C), and allowing the liquid dispersion to stand at this temperature for about 5 to 20 minutes, in some embodiments, about 10 minutes; this heating may be done to ensure that any powdered components added to the mixture are fully hydrated, thereby minimizing agglomerates or pockets of dry, inactive components.

[0181] In the optional lipid addition step 130, an oil and / or solid fat, or a blend of two or more oils and / or solid fats, can be added to the liquid dispersion in an amount of about 4% to 10% by weight, and in one particular embodiment, coconut oil is added in an amount of about 10% by weight. The lipid-dispersion combination is then mixed in any manner suitable to ensure homogeneity, i.e., complete incorporation of the oil and / or solid fat into the liquid dispersion. As a first non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be mixed with a blender or impeller until it is visually homogeneous. As a second non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be subjected to continuous high-shear mixing (e.g., greater than about 10,000 rpm) for a period of at least 2 to 10 minutes, or until a stable, homogeneous lipid-dispersion combination is achieved. As a third non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be homogenized under pressure (about 160 bar) using a high-pressure homogenizer.

[0182] In the acid addition step 140, one or more acids are added to the liquid dispersion to lower the pH of the liquid dispersion to a pH at which a desired degree or type of aggregation of the fungal proteins occurs. As a first non-limiting example, if the liquid dispersion includes oil and / or solid fat and it is desired to form a soft fungal curd, useful, for example, in making soft cheese analog food products, the pH of the liquid dispersion can be lowered to about 5.5-6.0. As a second non-limiting example, if the liquid dispersion is a mixed-form mycelial biomass composition and it is desired to aggregate two or more mycelial biomass forms to convert the liquid dispersion into a non-flowing gel, the pH of the liquid dispersion can be lowered to about 3.5. Thus, the amount of acid added in acid addition step 140 depends on both the "target" pH and the particular acid used, which can be selected to achieve the target pH without exceeding regulatory or safety limits for acids in food ingredients / products; by way of non-limiting example, if it is desired to lower the pH of the liquid dispersion from about 7.0-7.5 to about 5.5-6.0, a weaker acid (e.g., lactic acid, citric acid, or a combination thereof) can be used in an amount of about 2-3 g / L, while if it is desired to lower the pH to a lower value of about 3.5, a smaller amount of a stronger acid (e.g., hydrochloric acid) can be used. The dispersion-acid combination is then mixed for a time sufficient to ensure dispersion of the acid, and optionally, the liquid dispersion may be allowed to stand without mixing, for example, for a period of about 5-15 minutes. Optionally, the liquid dispersion may be heated and / or mixed during the acid addition step 140 (e.g., to about 160°F to 175°F (71.1°C to 79.4°C)), which may help ensure that the liquid dispersion does not separate before aggregation of the fungal proteins is complete.

[0183] In optional heating step 150, the liquid dispersion is heated, with or without mixing, to about 185°F-200°F (85°C-93.3°C); in some embodiments, mixing and / or heating may help ensure complete aggregation of the fungal proteins, for example, by increasing physical contact between the acid and the fungal particles. The temperature is typically maintained for a period of at least about 30 seconds, and in some embodiments, about 1-10 minutes.

[0184] Optionally, although not shown in Figure 1, the aggregated fungal protein composition can then be further processed. As a first non-limiting example, if the aggregation results in the formation of a substantially solid fungal curd that is separable from the liquid phase, further processing can include any one or more conventional liquid-solid separation steps (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.). As a second non-limiting example, if the aggregation results in the formation of a gel (i.e., a colloid in which a liquid phase is dispersed throughout a solid dispersion medium formed by the aggregation of the fungal proteins), the gel can be further processed by any suitable technique for processing edible gels, as known to those of skill in the art. Most typically, the aggregated fungal protein composition is edible (i.e., safe for human consumption) without any further processing, although with further processing, the aggregated fungal protein composition may be capable of being made into a suitable food product (e.g., cheese or cheese curd-like food product, tofu-like food product, etc.).

[0185] Referring now to Figure 2, another non-limiting embodiment of a method 200 for making a composition comprising aggregated fungal proteins (e.g., fungal curd, fungal gel, etc.) by pH adjustment is illustrated. This embodiment is generally similar to the embodiment of method 100 illustrated in Figure 1, with the most notable difference being that the pH adjustment is achieved by adding a microbial culture suitable for fermentation of the food product rather than simply by directly adding an acid. In particular, the embodiment of method 200 illustrated in Figure 2 includes a liquid dispersion preparation step 210, an optional ingredient addition step 220, an optional lipid addition step 230, an optional homogenization step 240, and a microbial culture addition step 250.

[0186] In liquid dispersion preparation step 210, a liquid dispersion of edible filamentous fungi is prepared by combining water and one or more mycelial biomass forms in a ratio described herein, e.g., about 30:1, 27:1, or 20:1, or alternatively, any ratio that results in a dispersion having a solids content of about 0.1% to about 15% by weight (e.g., in some embodiments where the liquid phase is water, a water content of about 85% to about 99.9% by weight). The water and mycelial biomass forms are placed in a device with rotating blades, such as a blender or impeller, and sheared until smooth. In this embodiment, shearing is carried out by high-speed blending or mixing (e.g., greater than about 10,000 rpm) continuously for a period of at least 2 to 10 minutes, or until a stable, homogenous dispersion is achieved; shear mixing is desirable to achieve sufficient homogeneity of the fungal material in the water and completely break down any possible aggregates. This step can be performed without the addition of heat, but if desired, heat (e.g., 90°F to 120°F, 32.2°C to 48.9°C) can be applied to facilitate the production of a homogeneous mixture.

[0187] In the optional ingredient addition step 220, ingredients such as additional (e.g., non-fungal) protein, flavorings, salt, sugar, etc., can be added to the liquid dispersion. In some embodiments, a mixture of non-fungal proteins is added to the dispersion in a total amount of about 0.25-10% by weight, in some embodiments in a total amount of about 3.0-4.0% by weight, and in some embodiments, about 3.5% by weight. For example, soy protein powder (e.g., soy flour) and an 80% concentration hemp protein powder preparation can be added in equal amounts or in various concentrations. Calcium or magnesium salts can also be added in amounts of about 0.01-0.5% by weight, typically about 0.1-0.3% by weight, and most typically about 0.2% by weight. Similarly, any desired flavoring agents (e.g., half-and-half flavoring, cottage cheese flavoring, milk flavoring, etc.) can be added in a total amount of about 0.4-0.8% by weight, and in some embodiments, about 0.6% by weight. Dairy fortifiers can be added in amounts of about 0.1-0.3% by weight, and in some embodiments, about 0.2% by weight. Any desired flavor modifiers can be added in amounts of about 0.01-0.05% by weight, and in some embodiments, about 0.03% by weight. Salt and sugar can also be added as desired (e.g., in amounts of about 0.5% by weight and about 1.0% by weight, respectively). All of these components are then mixed into the dispersion until completely dissolved. In some embodiments, this optional component addition step 120 may include heating the liquid dispersion, for example, to about 90°F to 120°F (32.2°C to 48.9°C), in some embodiments, 110°F (43.3°C), and allowing the liquid dispersion to stand at this temperature for about 5 to 20 minutes, in some embodiments, about 10 minutes; this heating may be done to ensure that any powdered components added to the mixture are fully hydrated, thereby minimizing agglomerates or pockets of dry, inactive components.

[0188] In the optional lipid addition step 230, an oil and / or solid fat, or a blend of two or more oils and / or solid fats, can be added to the liquid dispersion in an amount of about 4% to 10% by weight, with coconut oil being added in an amount of about 10% by weight in one particular embodiment. The lipid-dispersion combination is then mixed using a blender or impeller until it appears visually smooth. Optionally, the lipid-dispersion combination may be heated to about 180°F to 190°F (82.2°C to 87.8°C), in some embodiments, about 185°F (85°C), and allowed to stand at this temperature for 10 to 20 minutes, in some embodiments, about 15 minutes.

[0189] In the optional homogenization step 240, the (optionally heated) lipid-dispersion combination can be homogenized under pressure (about 200 bar) using a high-pressure homogenizer to ensure that the oil and / or solid fat is fully incorporated into the liquid dispersion. In some embodiments, the high-pressure homogenizer forces the flow of the lipid-dispersion combination through a system that subjects the lipid-dispersion combination to one or more forces that ensure complete mixing of the oil and / or solid fat into the liquid dispersion, thus homogenizing the oil and / or solid fat into the liquid dispersion.

[0190] In the microbial culture addition step 250, the microbial culture is added to the (optionally homogenized) liquid dispersion, typically in an amount of about 0.01-0.05% by weight, along with any media or growth nutrients required by the microbial culture but not otherwise present in the liquid dispersion. The inoculated liquid dispersion is then maintained at a temperature conducive to the survival and growth of the microbial culture (e.g., about 90°F-120°F (32.2°C-48.9°C), in some embodiments about 110°F (43.3°C)) for a sufficient time for the nutrients in the liquid dispersion to be fermented by the microbial culture, typically about 4-12 hours, in certain embodiments about 6 hours, or overnight, depending on the microorganisms present in the culture. If the liquid dispersion is at an elevated temperature (e.g., after the optional heating of the optional lipid addition step 230), the microbial culture is typically added while or after the liquid dispersion is cooling to the maintenance temperature. Fermentation by the microbial culture produces acids or bases that lower or raise the pH of the liquid dispersion, thus causing aggregation of the fungal proteins in the liquid dispersion.

[0191] Optionally, although not shown in Figure 2, the aggregated fungal protein composition can then be further processed. As a first non-limiting example, if the aggregation results in the formation of a substantially solid fungal curd that is separable from the liquid phase, further processing can include any one or more conventional liquid-solid separation steps (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.). As a second non-limiting example, if the aggregation results in the formation of a gel (i.e., a colloid in which a liquid phase is dispersed throughout a solid dispersion medium formed by the aggregation of the fungal proteins), the gel can be further processed by any suitable technique for processing edible gels, as known to those of skill in the art. Most typically, the aggregated fungal protein composition is edible (i.e., safe for human consumption) without any further processing, although with further processing, the aggregated fungal protein composition may also be capable of being made into a suitable food product (e.g., cheese or cheese curd-like food product, tofu-like food product, etc.).

[0192] While much of the teaching of this disclosure relates to compositions (e.g., in some embodiments, mixed-format mycelial biomass compositions) that can form fungal curds or gels or other stable colloids under acidic conditions, particularly at a pH of about 4 or below, and even more particularly at a pH of about 3.5, by aggregation of fungal proteins, it should be clearly understood that mycelial biomass compositions that form fungal curds or stable gels can be produced at any of several selected pH conditions. As a first non-limiting example, mycelial biomass compositions according to the present disclosure may form a fungal curd or stable gel at a pH of about 14 or less, about 13.5 or less, about 13 or less, about 12.5 or less, about 12 or less, about 11.5 or less, about 11 or less, about 10.5 or less, about 10 or less, about 9.5 or less, about 9 or less, about 8.5 or less, about 8 or less, about 7.5 or less, about 7 or less, about 6.5 or less, about 6 or less, about 5.5 or less, about 5 or less, about 4.5 or less, about 4 or less, about 3.5 or less, about 3 or less, about 2.5 or less, about 2 or less, about 1.5 or less, about 1 or less, about 0.5 or less, or about 0 or less. and / or methods of making a fungal curd or stable gel according to the present disclosure may include adjusting the pH to about 14 or less, about 13.5 or less, about 13 or less, about 12.5 or less, about 12 or less, about 11.5 or less, about 11 or less, about 10.5 or less, about 10 or less, about 9.5 or less, about 9 or less, about 8.5 or less, about 8 or less, about 7.5 or less, about 7 or less, about 6.5 or less, about 6 or less, about 5.5 or less, about 5 or less, about 4.5 or less, about 4 or less, about 3.5 or less, about 3 or less, about 2.5 or less, about 2 or less, about 1.5 or less, about 1 or less, about 0.5 or less, or about 0 or less.As a second non-limiting example, mycelial biomass compositions according to the present disclosure may form a fungal curd or stable gel at a pH of at least about 0, at least about 0.5, at least about 1, at least about 1.5, at least about 2, at least about 2.5, at least about 3, at least about 3.5, at least about 4, at least about 4.5, at least about 5, at least about 5.5, at least about 6, at least about 6.5, at least about 7, at least about 7.5, at least about 8, at least about 8.5, at least about 9, at least about 9.5, at least about 10, at least about 10.5, at least about 11, at least about 11.5, at least about 12, at least about 12.5, at least about 13, at least about 13.5, or at least about 14. and / or methods of making a fungal curd or stable gel according to the present disclosure may include adjusting the pH to at least about 0, at least about 0.5, at least about 1, at least about 1.5, at least about 2, at least about 2.5, at least about 3, at least about 3.5, at least about 4, at least about 4.5, at least about 5, at least about 5.5, at least about 6, at least about 6.5, at least about 7, at least about 7.5, at least about 8, at least about 8.5, at least about 9, at least about 9.5, at least about 10, at least about 10.5, at least about 11, at least about 11.5, at least about 12, at least about 12.5, at least about 13, at least about 13.5, or at least about 14. As a third non-limiting example, a mycelial biomass composition according to the present disclosure may form a fungal curd or stable gel at any pH value in any range having a lower pH value limit of 0.0 to 14.0 to any tenth place and an upper pH value limit of 0.0 to 14.0 to any other tenth place, and / or a method of making a fungal curd or stable gel according to the present disclosure may include adjusting the pH to any pH value in any range having a lower pH value limit of 0.0 to 14.0 to any tenth place and an upper pH value limit of 0.0 to 14.0 to any other tenth place.

[0193] Aggregation of fungal proteins by adding functional ingredients In some embodiments of the present disclosure, aggregation of filamentous fungal proteins is induced by adding one or more functional ingredients (i.e., proteins and / or carbohydrates) to a liquid dispersion of filamentous fungal particles. In some embodiments, this aggregation results in the formation of a fungal curd, i.e., a solid mass of filamentous fungal proteins (and optionally other components) that is separable from the remaining liquid phase, similar to the method of coagulating proteins from animal milk (to form curd) or soy milk (to form tofu), while in other embodiments, aggregation results in the formation of a gel material, i.e., a phase that retains its shape and resists flow, in which the liquid phase is dispersed throughout the network formed by the filamentous fungal proteins (and optionally other compounds).

[0194] The one or more functional ingredients can assist or induce aggregation of fungal proteins by any of several mechanisms. In particular, as further described elsewhere throughout this disclosure, the inventors have found that, depending on the pI of the fungal proteins and the pH of the liquid dispersion, the addition of selected non-fungal proteins and / or oligosaccharides and / or polysaccharides having a pI different from the pI of the fungal proteins can subject the fungal proteins to a desired type or degree of electrostatic interaction with each other, the non-fungal proteins and / or oligosaccharides and / or polysaccharides, or other components in the liquid dispersion. As a first non-limiting example, non-fungal proteins (e.g., potato proteins, chickpea proteins, etc.) and / or oligosaccharides and / or polysaccharides (e.g., maltodextrin) can be used at a pH of the liquid dispersion below the desired pH of the liquid dispersion. ΦThe fungal proteins can be selected based on their pI to result in the formation of soluble complexes between the fungal protein and the non-fungal protein and / or oligosaccharides and / or polysaccharides at the desired pH ("Stage I" aggregation). As a second non-limiting example, the non-fungal proteins (e.g., potato proteins, chickpea proteins, etc.) and / or oligosaccharides and / or polysaccharides (e.g., maltodextrin) can be selected based on their pI to result in the formation of soluble complexes between the fungal protein and the non-fungal protein and / or oligosaccharides and / or polysaccharides at the desired pH ("Stage I" aggregation). Φ and the pI of the fungal protein, which can result in the formation of larger interpolymer complexes between the fungal protein and the non-fungal protein and / or oligosaccharides and / or polysaccharides at the desired pH ("Stage II" aggregation). As a third non-limiting example, non-fungal proteins (e.g., potato proteins, chickpea proteins, etc.) and / or oligosaccharides and / or polysaccharides (e.g., maltodextrin) can be selected based on their pI so that the desired pH of the liquid dispersion is in the range between the pH of the liquid dispersion and the pI of the fungal protein, thereby resulting in the formation of larger interpolymer complexes between the fungal protein and the non-fungal protein and / or oligosaccharides and / or polysaccharides at the desired pH ("Stage II" aggregation). Φ and the pI of the fungal protein, thereby resulting in the formation of a highly networked gel of fungal proteins with non-fungal proteins and / or oligosaccharides and / or polysaccharides at the desired pH ("Stage III" aggregation).

[0195] In some embodiments, non-fungal proteins and / or oligosaccharides and / or polysaccharides can be selected that effectively "balance" the surface charge on the fungal protein at the desired pH; as a non-limiting example, if the desired pH (e.g., 4.2) is above the pI of the fungal protein (e.g., 2.2 for soaked dough in a liquid dispersion) and below the pI of the non-fungal protein and / or oligosaccharides and / or polysaccharides (e.g., 5.1 for commercially available potato protein), the negative surface charge of the fungal protein can be offset by the approximately equal but opposite positive charge of the non-fungal protein and / or oligosaccharides and / or polysaccharides at the desired pH. In particular, if the non-fungal protein and / or oligosaccharides and / or polysaccharides are selected such that the pI of the non-fungal protein and / or oligosaccharides and / or polysaccharides is sufficiently higher or sufficiently lower than the pI of the fungal protein (e.g., if the difference between the pI of the fungal protein and the pI of the non-fungal protein and / or oligosaccharides and / or polysaccharides is at least about 2.2, at least about 2.3, at least about 2.4, at least about 2.5, at least about 2.6, at least about 2.7, at least about 2.8, or at least about 2.9), a range of intermediate pH values ​​may exist between the pI of the fungal protein and the pI of the non-fungal protein and / or oligosaccharides and / or polysaccharides, in which case the attractive electrostatic interactions between the fungal protein and the non-fungal protein and / or oligosaccharides and / or polysaccharides allow the latter to essentially "reinforce" the former to form a gel structure.

[0196] The addition of non-fungal proteins and / or oligosaccharides and / or polysaccharides can also provide additional structural properties to the resulting aggregated protein composition and / or enhance the nutritional value of food materials and products made from the aggregated protein composition. As a non-limiting example, hemp protein is considered a "complete" protein because it is known to contain all the essential amino acids humans need to ingest through food, and hemp seeds contain more than 30 g of protein per 100 g of seeds. Additionally, soy protein, like hemp, is considered a "complete" protein because it is known to contain all the essential amino acids humans need to ingest through food, and soybeans contain approximately 18 g of protein per 100 g. Other non-fungal proteins contain similar benefits and can be mixed and matched in the disclosed manner to achieve a desired nutritional profile. Suitable proteins for use in the disclosed methods and compositions include bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof. Suitable oligosaccharides and / or polysaccharides for use in the disclosed methods and compositions include cellobiose, isomaltose, isomaltulose, lactose, lactulose, maltose, sucrose, trehalose, turanose, maltotriose, melezitose, raffinose, stachyose, acarbose, fructooligosaccharides, galactooligosaccharides, isomaltooligosaccharides, maltodextrin, beta-glucan, chitosan, dextrin, dextran, fructose, fructan, galactose, galactan, glucose, glucan, hemicellulose, levan, lignin, mannan, pectin, starch or fractions thereof (e.g., amylopectin, amylose), xanthan gum, and combinations thereof.

[0197] In some embodiments, one or more of the non-fungal proteins added to the liquid dispersion can be an enzyme, and aggregation of the fungal proteins can be induced, at least in part, by enzymatic action. Suitable enzymes for use in the disclosed methods and compositions include catalase, chymosin, lactase, lipase, transglutaminase, and combinations thereof.

[0198] The amount of non-fungal protein and / or oligosaccharides and / or polysaccharides used in the disclosed methods can vary based on the desired aggregation behavior of the liquid dispersion, as described above, and / or the desired texture and / or flavor of the resulting aggregated protein composition. In various embodiments, amounts of non-fungal protein and / or oligosaccharides and / or polysaccharides can be used in amounts of 1.0% to 4.0% by weight. This amount can reflect the use of a single source of non-fungal protein and / or oligosaccharides and / or polysaccharides, or a combination of sources of non-fungal protein and / or oligosaccharides and / or polysaccharides, again depending on the desired aggregation behavior, texture, flavor, etc. In some embodiments, the amount of non-fungal protein and / or oligosaccharides and / or polysaccharides used in the disclosed methods is selected from 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, and 4.0% by weight, or any range between 1.0% and 4.0% by weight.

[0199] Referring now to Figure 3, one non-limiting embodiment of a method 300 for making a composition comprising aggregated fungal proteins (e.g., fungal curd, fungal gel, etc.) by adding a functional ingredient is illustrated. The embodiment of method 300 illustrated in Figure 3 includes a liquid dispersion preparation step 310, a functional ingredient addition step 320, an optional lipid addition step 330, an optional acidification step 340, and an optional heating step 350.

[0200] In liquid dispersion preparation step 310, a liquid dispersion of edible filamentous fungi is prepared by combining water and one or more mycelial biomass forms in a ratio described herein, e.g., about 30:1, 27:1, or 20:1, or alternatively, any ratio that results in a dispersion having a solids content of about 0.1% to about 15% by weight (e.g., in some embodiments where the liquid phase is water, a water content of about 85% to about 99.9% by weight). The water and mycelial biomass forms are placed in a device with rotating blades, such as a blender or impeller, and sheared until smooth. In this embodiment, shearing is carried out by high-speed blending or mixing (e.g., greater than about 10,000 rpm) continuously for a period of at least 2 to 10 minutes, or until a stable, homogenous dispersion is achieved; shear mixing is desirable to achieve sufficient homogeneity of the fungal material in the water and completely break down any aggregates possible. This step can be performed without the addition of heat, but if desired, heat (e.g., 90°F to 120°F, 32.2°C to 48.9°C) can be applied to facilitate the production of a homogeneous mixture.

[0201] In the functional ingredient addition step 320, one or more functional ingredients, i.e., non-fungal proteins and / or oligosaccharides and / or polysaccharides, are added to the liquid dispersion. In some embodiments, a mixture of non-fungal proteins is added to the dispersion in a total amount of about 0.25-10% by weight, in some embodiments, in a total amount of about 3.0-4.0% by weight, and in some embodiments, about 3.5% by weight. As a first non-limiting example, soy protein powder (e.g., soy flour) and an 80% concentration hemp protein powder preparation can be added in equal amounts or in various concentrations. As a second non-limiting example, a protein having a selected isoelectric point (e.g., commercially available potato protein having an isoelectric point of about pH 5.1 and / or commercially available chickpea protein having an isoelectric point of about pH 4.5) can be added, typically in a total amount of about 1.0% by weight. As a third non-limiting example, oligosaccharides and / or polysaccharides (e.g., maltodextrin) selected to exert an electrostatic effect on the fungal proteins in the liquid dispersion (thus inducing aggregation) can be added.

[0202] The functional ingredient addition step 320 may optionally further include adding components such as flavoring agents, salts, and sugars to the liquid dispersion. Calcium or magnesium salts may also be added in amounts of about 0.01-0.5% by weight, typically about 0.1-0.3% by weight, and most typically about 0.2% by weight. Similarly, any desired flavoring agents (e.g., half and half flavoring, cottage cheese flavoring, milk flavoring, etc.) may be added in a total amount of about 0.4-0.8% by weight, and in some embodiments, about 0.6% by weight; dairy fortifiers may be added in amounts of about 0.1-0.3% by weight, and in some embodiments, about 0.2% by weight; and any desired flavor modifiers may be added in amounts of about 0.01-0.05% by weight, and in some embodiments, about 0.03% by weight. Salt and sugar may also be added to taste (e.g., in amounts of about 0.5% by weight and about 1.0% by weight, respectively).

[0203] All of these components are then mixed into the dispersion until completely dissolved. In some embodiments, this functional ingredient addition step 320 may optionally include heating the liquid dispersion, for example, to about 90°F to 120°F (32.2°C to 48.9°C), in some embodiments, 110°F (43.3°C), and allowing the liquid dispersion to stand at this temperature for about 5 to 20 minutes, in some embodiments, about 10 minutes; this heating may be done to ensure that any powdered components added to the mixture are fully hydrated, thereby minimizing agglomerates or pockets of dry, inactive components.

[0204] In the optional lipid addition step 330, an oil and / or solid fat, or a blend of two or more oils and / or solid fats, can be added to the liquid dispersion in an amount of about 4% to 10% by weight; in one particular embodiment, coconut oil is added in an amount of about 10% by weight. The lipid-dispersion combination is then mixed in any manner suitable to ensure homogeneity, i.e., complete incorporation of the oil and / or solid fat into the liquid dispersion. As a first non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be mixed with a blender or impeller until it is visually homogeneous. As a second non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be subjected to continuous high-shear mixing (e.g., greater than about 10,000 rpm) for a period of at least 2 to 10 minutes, or until a stable, homogeneous lipid-dispersion combination is achieved. As a third non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be homogenized under pressure (about 160 bar) using a high-pressure homogenizer.

[0205] In the optional acidification step 340, one or more acids and / or acidifying microbial cultures can be added to the liquid dispersion to lower the pH of the liquid dispersion to a pH at which a desired degree or type of aggregation of the fungal proteins occurs. As a first non-limiting example, if the liquid dispersion contains oil and / or solid fat and it is desired to form a soft fungal curd, useful, for example, in making soft cheese analog food products, the pH of the liquid dispersion can be lowered to about 5.5-6.0. As a second non-limiting example, if the liquid dispersion is a mixed-form mycelial biomass composition and it is desired to aggregate two or more mycelial biomass forms to convert the liquid dispersion into a non-flowing gel, the pH of the liquid dispersion can be lowered to about 3.5. Thus, the amount of acid / acidifying microbial culture added in the optional acidification step 340 can depend on both the "target" pH and the particular acid / acidifying microbial culture used, which can be selected to achieve the target pH without exceeding regulatory or safety limits for acids in food ingredients / products; by way of non-limiting example, if it is desired to lower the pH of the liquid dispersion from about 7.0-7.5 to about 5.5-6.0, a weaker acid (e.g., lactic acid, citric acid, or a combination thereof) can be used in an amount of about 2-3 g / L, while if it is desired to lower the pH to a lower value of about 3.5, a smaller amount of a stronger acid (e.g., hydrochloric acid) can be used. The dispersion-acid combination can then be mixed for a time sufficient to ensure dispersion of the acid; optionally, the liquid dispersion can be allowed to stand without mixing, for example, for a period of about 5-15 minutes. Optionally, the liquid dispersion may be heated and / or mixed during the acidification step 340 (e.g., to about 160°F to 175°F (71.1°C to 79.4°C)), which may help ensure that the liquid dispersion does not separate before aggregation of the fungal proteins is complete.

[0206] It is emphasized that the acidification step 340 is an optional step. Specifically, in some embodiments of the method 300 illustrated in Figure 3, adjustment of the pH of the liquid dispersion after the functional ingredient addition step 320 may not be necessary; or, in other words, the "natural" pH of the liquid dispersion after the functional ingredient addition step 320 may be an appropriate target pH to achieve the desired type and degree of aggregation of the fungal proteins.

[0207] In optional heating step 350, the liquid dispersion is heated, with or without mixing, to about 185°F-200°F (85°C-93.3°C); in some embodiments, mixing and / or heating may help ensure complete aggregation of the fungal proteins, for example, by increasing physical contact between the functional ingredient and the fungal particles. The temperature is typically maintained for a period of at least about 30 seconds, and in some embodiments, about 1-10 minutes.

[0208] Optionally, although not shown in Figure 3, the aggregated fungal protein composition can then be further processed. As a first non-limiting example, if the aggregation results in the formation of a substantially solid fungal curd that is separable from the liquid phase, further processing can include any one or more conventional liquid-solid separation steps (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.). As a second non-limiting example, if the aggregation results in the formation of a gel (i.e., a colloid in which a liquid phase is dispersed throughout a solid dispersion medium formed by the aggregation of the fungal proteins), the gel can be further processed by any suitable technique for processing edible gels, as known to those of skill in the art. Most typically, the aggregated fungal protein composition is edible (i.e., safe for human consumption) without any further processing, although with further processing, the aggregated fungal protein composition may be capable of being made into a suitable food product (e.g., cheese or cheese curd-like food product, tofu-like food product, etc.).

[0209] Aggregation of fungal proteins by the addition of salt In some embodiments of the present disclosure, aggregation of filamentous fungal proteins is induced by adding one or more salts to a liquid dispersion of filamentous fungal particles. In some embodiments, this aggregation results in the formation of a fungal curd, i.e., a solid mass of filamentous fungal proteins (and optionally other components) that is separable from the remaining liquid phase, similar to the method of coagulating proteins from animal milk (to form curd) or soy milk (to form tofu), while in other embodiments, aggregation results in the formation of a gel material, i.e., a phase that retains its shape and resists flow, in which the liquid phase is dispersed throughout a network formed by the filamentous fungal proteins (and optionally other compounds).

[0210] One or more salts can assist or induce aggregation of fungal proteins through any of several mechanisms. In particular, as further described elsewhere throughout this disclosure, the inventors have discovered that, depending on the pI of the fungal protein and the pH of the liquid dispersion, the addition of selected salts can result in the formation of ionic salt "bridges" between fungal protein molecules. As a first, non-limiting example, if the pH of the liquid dispersion is relatively far from the pI of the fungal material, salts (e.g., calcium salts, magnesium salts) can be added to the liquid dispersion at a relatively low ionic strength to result in the formation of a relatively low concentration of salt bridges, which can cause the fungal protein to aggregate into a "fine" network. As a second, non-limiting example, if the pH of the liquid dispersion is relatively close to the pI of the fungal material, salts (e.g., calcium salts, magnesium salts) can be added to the liquid dispersion at a relatively high ionic strength to result in the formation of a relatively high concentration of salt bridges, which can cause the fungal protein to aggregate into a "particulate" or "coarse" network. It should therefore be clearly understood that in some embodiments, it is desirable to add calcium and / or magnesium salts to liquid dispersions when the pH of the liquid dispersion is approximately equal to the pI of the fungal protein (e.g., within about 1 pH unit, about 0.95 pH units, about 0.9 pH units, about 0.85 pH units, about 0.8 pH units, about 0.75 pH units, about 0.7 pH units, about 0.65 pH units, about 0.6 pH units, about 0.55 pH units, about 0.5 pH units, about 0.45 pH units, about 0.4 pH units, about 0.35 pH units, about 0.3 pH units, about 0.25 pH units, about 0.2 pH units, about 0.15 pH units, about 0.1 pH units, or about 0.05 pH units), while in other embodiments it is desirable to add calcium and / or magnesium salts to liquid dispersions when the pH of the liquid dispersion is not approximately equal to the pI of the fungal protein.

[0211] The inventors have found that, generally, food-grade salts containing divalent cations, i.e., edible calcium and / or magnesium salts, can be effective in inducing aggregation of fungal proteins in liquid dispersions, and in particular, as further described elsewhere throughout this disclosure, salts containing monovalent cations (e.g., sodium chloride) have been found to generally be ineffective, or significantly less effective than salts containing divalent cations, in inducing aggregation. Furthermore, the inventors have found that different divalent cations result in different rheologies and / or textures for the resulting aggregated protein compositions, and specifically, the use of calcium salts generally results in the formation of "softer" curd compositions, having a texture similar to that of, for example, ricotta cheese (or curds suitable for use in making thereof), while the use of magnesium salts generally results in the formation of "firmer" curd compositions, having a texture similar to that of, for example, tofu (or curds suitable for use in making thereof).

[0212] The addition of calcium and / or magnesium salts can also enhance the nutritional value of food ingredients and products made from the aggregated protein composition. As a non-limiting example, edible and / or food-grade calcium and / or magnesium salts can be added such that the aggregated protein composition, when consumed by a human in a selected amount (e.g., a single recommended serving), provides at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% of the Recommended Dietary Allowance (RDA) of calcium and / or magnesium.Suitable calcium salts for use in the disclosed methods and compositions include calcium carbonate, calcium sorbate, calcium benzoate, calcium sulfite, calcium bisulfite, calcium formate, calcium acetate, calcium propionate, calcium ascorbate, calcium lactate, monocalcium citrate, dicalcium citrate, tricalcium citrate, monocalcium phosphate, dicalcium phosphate, tricalcium phosphate, calcium malate, calcium hydrogen malate, calcium tartrate, calcium fumarate, calcium glyceryl phosphate, disodium calcium ethylenediaminetetraacetate, calcium lactobionate, calcium alginate, dicalcium diphosphate, dihydrogen calcium diphosphate Suitable magnesium salts for use in the disclosed methods and compositions include sodium calcium polyphosphate, calcium polyphosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, calcium ferrocyanide, dicalcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium silicate, calcium aluminosilicate, calcium stearate, calcium gluconate, synthetic calcium aluminate, calcium diglutamate, calcium guanylate, calcium inosinate, calcium 5'-ribonucleotides, calcium iodate, calcium bromate, calcium peroxide, calcium cyclamate, calcium saccharinate, and combinations thereof. Suitable magnesium salts for use in the disclosed methods and compositions include magnesium lactate, monomagnesium phosphate, dimagnesium phosphate, magnesium citrate, magnesium salts of fatty acids, magnesium carbonate, magnesium bicarbonate, magnesium chloride, magnesium sulfate, magnesium oxide, magnesium silicate, magnesium trisilicate, magnesium stearate, magnesium gluconate, magnesium diglutamate, and combinations thereof.

[0213] The amount of one or more salts used in the disclosed methods can vary based on the desired aggregation behavior of the liquid dispersion, as described above, and / or the desired texture and / or flavor of the resulting aggregated protein composition. In various embodiments, calcium and / or magnesium salts can be used in an amount of about 0.01% to about 0.5% by weight. This amount can reflect the use of a single calcium salt, a single magnesium salt, a combination of two or more calcium salts, a combination of two or more magnesium salts, or a combination of at least one calcium salt and at least one magnesium salt, again depending on the desired aggregation behavior, texture, flavor, etc. In some embodiments, the amount of one or more salts used in the disclosed methods is selected from 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, and 0.5% by weight.

[0214] 4, one non-limiting embodiment of a method 400 for making a composition comprising aggregated fungal proteins (e.g., fungal curd, fungal gel, etc.) by the addition of salt is illustrated. The embodiment of method 400 illustrated in FIG. 4 includes a liquid dispersion preparation step 410, a salt addition step 420, an optional lipid addition step 430, an optional acidification step 440, and an optional heating step 450.

[0215] In liquid dispersion preparation step 410, a liquid dispersion of edible filamentous fungi is prepared by combining water and one or more mycelial biomass forms in a ratio described herein, e.g., about 30:1, 27:1, or 20:1, or alternatively, any ratio that results in a dispersion having a solids content of about 0.1% to about 15% by weight (e.g., in some embodiments where the liquid phase is water, a water content of about 85% to about 99.9% by weight). The water and mycelial biomass forms are placed in a device with rotating blades, such as a blender or impeller, and sheared until smooth. In this embodiment, shearing is carried out by high-speed blending or mixing (e.g., greater than about 10,000 rpm) continuously for a period of at least 2 to 10 minutes, or until a stable, homogenous dispersion is achieved; shear mixing is desirable to achieve sufficient homogeneity of the fungal material in the water and completely break down any possible aggregates. This step can be performed without the addition of heat, but if desired, heat (e.g., 90°F to 120°F, 32.2°C to 48.9°C) can be applied to facilitate the production of a homogeneous mixture.

[0216] In the salt addition step 420, one or more calcium or magnesium salts are added to the liquid dispersion. The one or more calcium or magnesium salts are generally added in an amount of about 0.01-0.5% by weight, typically about 0.1-0.3% by weight, and most typically about 0.2% by weight. As a first non-limiting example, in embodiments where it is desired to form a "softer" curd composition, e.g., with a texture similar to that of ricotta cheese (or a curd suitable for use in its preparation), one or more calcium salts (e.g., tricalcium citrate, tricalcium phosphate, calcium lactate, or a combination thereof) can be added. As a second non-limiting example, in embodiments where it is desired to form a "firmer" curd composition, e.g., with a texture similar to that of tofu (or a curd suitable for use in its preparation), one or more magnesium salts (e.g., magnesium chloride) can be added. As a first non-limiting example, soy protein powder (e.g., soy flour) and an 80% concentration hemp protein powder preparation can be added in equal amounts or at various concentrations. As a second non-limiting example, proteins having selected isoelectric points (e.g., commercially available potato proteins having an isoelectric point of about pH 5.1 and / or commercially available chickpea proteins having an isoelectric point of about pH 4.5) can be added, typically in a total amount of about 1.0 wt. As a third non-limiting example, oligosaccharides and / or polysaccharides (e.g., maltodextrin) selected to exert an electrostatic effect on the fungal proteins in the liquid dispersion (thus inducing aggregation) can be added.

[0217] The salt addition step 420 may optionally further include adding components to the liquid dispersion, such as flavoring agents, non-fungal proteins and / or oligosaccharides and / or polysaccharides, sugars, etc. In some embodiments, a mixture of non-fungal proteins may be added to the dispersion in a total amount of about 0.25-10% by weight, in some embodiments in a total amount of about 3.0-4.0% by weight, and in some embodiments, about 3.5% by weight; similarly, any desired flavoring agents (e.g., half and half flavoring, cottage cheese flavoring, milk flavoring, etc.) may be added to the dispersion in a total amount of about 0.4-0.8% by weight, and in some embodiments, about 0.6% by weight; dairy fortifiers may be added to the dispersion in an amount of about 0.1-0.3% by weight, and in some embodiments, about 0.2% by weight; and any desired flavor modifiers may be added to the dispersion in an amount of about 0.01-0.05% by weight, and in some embodiments, about 0.03% by weight. Salts other than those used to induce aggregation (e.g., sodium salts such as sodium chloride) and sugars may be added as desired (e.g., in amounts of about 0.5% by weight and about 1.0% by weight, respectively).

[0218] All of these components are then mixed into the dispersion until completely dissolved. In some embodiments, this salt addition step 420 may optionally include heating the liquid dispersion to, for example, about 90°F to 120°F (32.2°C to 48.9°C) or about 160°F to 175°F (71.1°C to 79.4°C), in some embodiments 110°F (43.3°C) or 175°F (79.4°C), and allowing the liquid dispersion to stand at this temperature for about 5 to 30 minutes, in some embodiments about 10 minutes or about 30 minutes, this heating may be done to ensure that any powdered components added to the mixture are fully hydrated, thereby minimizing agglomerates or pockets of dry, inactive components.

[0219] In the optional lipid addition step 430, an oil and / or solid fat, or a blend of two or more oils and / or solid fats, can be added to the liquid dispersion in an amount of about 4% to 10% by weight, and in one particular embodiment, coconut oil is added in an amount of about 10% by weight. The lipid-dispersion combination is then mixed in any manner suitable to ensure homogeneity, i.e., complete incorporation of the oil and / or solid fat into the liquid dispersion. As a first non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be mixed with a blender or impeller until it is visually homogeneous. As a second non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be subjected to continuous high-shear mixing (e.g., greater than about 10,000 rpm) for a period of at least 2 to 10 minutes, or until a stable, homogeneous lipid-dispersion combination is achieved. As a third non-limiting example of a suitable mixing technique, the lipid-dispersion combination can be homogenized under pressure (about 160 bar) using a high-pressure homogenizer.

[0220] In the optional acidification step 440, one or more acids and / or acidifying microbial cultures can be added to the liquid dispersion to lower the pH of the liquid dispersion to a pH at which a desired degree or type of aggregation of the fungal proteins occurs. As a first non-limiting example, if the liquid dispersion contains oil and / or solid fat and it is desired to form a soft fungal curd, useful, for example, in making soft cheese analog food products, the pH of the liquid dispersion can be lowered to about 5.5-6.0. As a second non-limiting example, if the liquid dispersion is a mixed-form mycelial biomass composition and it is desired to aggregate two or more mycelial biomass forms to convert the liquid dispersion into a non-flowing gel, the pH of the liquid dispersion can be lowered to about 3.5. Thus, the amount of acid / acidifying microbial culture added in optional acidification step 440 can depend on both the "target" pH and the particular acid / acidifying microbial culture used, which can be selected to achieve the target pH without exceeding regulatory or safety limits for acids in food ingredients / products; by way of non-limiting example, if it is desired to lower the pH of the liquid dispersion from about 7.0-7.5 to about 5.5-6.0, a weaker acid (e.g., lactic acid, citric acid, or a combination thereof) can be used in an amount of about 2-3 g / L, while if it is desired to lower the pH to a lower value of about 3.5, a smaller amount of a stronger acid (e.g., hydrochloric acid) can be used. The dispersion-acid combination may then be mixed for a time sufficient to ensure dispersion of the acid; optionally, the liquid dispersion may be allowed to stand without mixing, for example, for a period of about 5-15 minutes. Optionally, the liquid dispersion may be heated and / or mixed during the acidification step 440 (e.g., to about 160°F to 175°F (71.1°C to 79.4°C)), which may help ensure that the liquid dispersion does not separate before aggregation of the fungal proteins is complete.

[0221] It is emphasized that the acidification step 440 is an optional step. Specifically, in some embodiments of the method 400 illustrated in Figure 4, adjustment of the pH of the liquid dispersion after the salt addition step 420 may not be necessary; or, in other words, the "natural" pH of the liquid dispersion after the salt addition step 420 may be an appropriate target pH to achieve the desired type and degree of aggregation of the fungal proteins.

[0222] In optional heating step 450, the liquid dispersion is heated, with or without mixing, to about 185°F-200°F (85°C-93.3°C); in some embodiments, mixing and / or heating may help ensure complete aggregation of the fungal proteins, for example, by increasing physical contact between the salt and the fungal particles. The temperature is typically maintained for a period of at least about 30 seconds, and in some embodiments, about 1-10 minutes.

[0223] Optionally, although not shown in Figure 4, the aggregated fungal protein composition can then be further processed. As a first non-limiting example, if the aggregation results in the formation of a substantially solid fungal curd that is separable from the liquid phase, further processing can include any one or more conventional liquid-solid separation steps (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.). As a second non-limiting example, if the aggregation results in the formation of a gel (i.e., a colloid in which a liquid phase is dispersed throughout a solid dispersion medium formed by the aggregation of the fungal proteins), the gel can be further processed by any suitable technique for processing edible gels, as known to those of skill in the art. Most typically, the aggregated fungal protein composition is edible (i.e., safe for human consumption) without any further processing, although with further processing, the aggregated fungal protein composition may also be capable of being made into a suitable food product (e.g., cheese or cheese curd-like food product, tofu-like food product, etc.).

[0224] Aggregation of fungal proteins by a combination of the above techniques It should be clearly understood that in some embodiments, fungal protein aggregation can be induced by a combination of any two or all three of the methods described in the preceding sections of this disclosure. As a first non-limiting example, fungal protein aggregation can be induced by a combination of pH adjustment and the addition of one or more functional ingredients. As a second non-limiting example, fungal protein aggregation can be induced by a combination of pH adjustment and the addition of one or more salts. As a third non-limiting example, fungal protein aggregation can be induced by a combination of one or more functional ingredients and the addition of one or more salts. As a fourth non-limiting example, fungal protein aggregation can be induced by a combination of pH adjustment, the addition of one or more functional ingredients, and the addition of one or more salts.

[0225] Referring now to Figure 5, one non-limiting embodiment of a method 500 for making a fungal curd is illustrated. The embodiment of method 500 illustrated in Figure 5 includes a liquid dispersion preparation step 510, a component addition step 520, a lipid addition step 530, an acidification and / or salt addition step 540, and a heating step 550, and can be particularly effective for producing a relatively soft fungal curd suitable for processing into, for example, a cheese curd like food product or a soft cheese (such as ricotta) like food product.

[0226] A liquid dispersion of edible filamentous fungi is prepared in liquid dispersion preparation step 510. Liquid dispersion preparation step 510 may be substantially similar to the analogous steps 110, 210, 310, 410 described above and illustrated in Figures 1-4.

[0227] In the component addition step 520, components such as functional ingredients (e.g., non-fungal proteins and / or oligosaccharides and / or polysaccharides), flavorings, salts, sugars, emulsifiers, etc. are added to the liquid dispersion. The component addition step 520 can be generally similar to the analogous steps 120, 220, 320 described above and illustrated in Figures 1-3.

[0228] In the lipid addition step 530, an oil and / or solid fat, or a blend of two or more oils and / or solid fats, is added to the liquid dispersion. The lipid addition step 530 may be generally similar to the analogous steps 130, 230, 330, 430 described above and illustrated in Figures 1-4.

[0229] In the acidification and / or salt addition step 540, the liquid dispersion is acidified (i.e., its pH is reduced), or one or more calcium or magnesium salts are added to the liquid dispersion, or both. In some embodiments, the acidification and / or salt addition step 540 comprises adding an acid and / or an acid-producing microbial culture to the liquid dispersion and thus may be generally similar to (or may include substantially similar substeps of) the analogous steps 140, 250, 340, and 440 illustrated in FIGS. 1-4 and described above. Additionally or alternatively, in some embodiments, the acidification and / or salt addition step 540 comprises adding one or more calcium or magnesium salts to the liquid dispersion and thus may be generally similar to (or may include substantially similar substeps of) the analogous step 420 illustrated in FIG. 4 and described above. In those embodiments in which the acidification and / or salt addition step 540 comprises both acidification and salt addition, it should be clearly understood that these two substeps may occur simultaneously or sequentially in any order.

[0230] In heating step 550, the liquid dispersion is heated with or without agitation to ensure complete aggregation of the fungal proteins. Heating step 550 can be generally similar to analogous steps 150, 350, 450 described above and illustrated in Figures 1, 3, and 4.

[0231] Optionally, although not shown in Figure 5, the fungal curd produced by method 500 can then be further processed. By way of non-limiting example, further processing can include any one or more conventional liquid-solid separation steps (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.). Most typically, the fungal curd composition is edible (i.e., safe for human consumption) without any further processing, although further processing may enable the aggregated fungal protein composition to be made into a suitable food product (e.g., cheese or a cheese curd-like food product).

[0232] Referring now to Figure 6, one non-limiting embodiment of a method 600 for making a fungal curd is illustrated. The embodiment of method 600 illustrated in Figure 6 includes a liquid dispersion preparation step 610, an acidification and / or salt addition step 620, and a heating step 630.

[0233] A liquid dispersion of edible filamentous fungi is prepared in liquid dispersion preparation step 610. Liquid dispersion preparation step 610 may be substantially similar to analogous steps 110, 210, 310, 410, 510 described above and illustrated in Figures 1-5.

[0234] In the acidification and / or salt addition step 620, the liquid dispersion is acidified (i.e., its pH is reduced), one or more calcium or magnesium salts are added to the liquid dispersion, or both. In some embodiments, the acidification and / or salt addition step 620 comprises adding an acid and / or an acid-producing microbial culture to the liquid dispersion and thus may be substantially similar to (or may include substantially similar substeps of) the analogous steps 140, 250, 340, and 440 illustrated in Figures 1-4 and described above. Additionally or alternatively, in some embodiments, the acidification and / or salt addition step 620 comprises adding one or more calcium or magnesium salts to the liquid dispersion and thus may be substantially similar to (or may include substantially similar substeps of) the analogous step 420 illustrated in Figure 4 and described above. Thus, the acidification and / or salt addition step 620 may, in its entirety, be substantially similar to the analogous step 540 illustrated in Figure 5 and described above. In those embodiments in which the acidification and / or salt addition step 620 includes both acidification and salt addition, it should be clearly understood that these two sub-steps may be performed simultaneously or sequentially in any order.

[0235] In heating step 630, the liquid dispersion is heated with or without agitation to ensure complete aggregation of the fungal proteins. Heating step 630 can be generally similar to analogous steps 150, 350, 450, 550 described above and illustrated in Figures 1 and 3-5.

[0236] Optionally, although not shown in Figure 6, the fungal curd produced by method 600 can then be further processed. By way of non-limiting example, further processing can include any one or more conventional liquid-solid separation steps (e.g., decanting, pressing, filtration, gravity separation, screw separation, centrifugation, etc.). Most typically, the fungal curd composition is edible (i.e., safe for human consumption) without any further processing, although further processing may enable the aggregated fungal protein composition to be made into a suitable food product (e.g., a tofu analog food product).

[0237] Fungal curd composition containing aggregated fungal proteins The present disclosure provides fungal curd compositions comprising aggregated fungal proteins, optionally produced by the methods disclosed herein. In many embodiments, these compositions may be cheese curd analog food products, such as those resembling ricotta cheese curds, mozzarella cheese curds, or other cheese curds. In other embodiments, these compositions may be tofu (or tofu curd) analog food products.

[0238] In many, but not all, embodiments, the curd compositions of the present disclosure comprise an oil. Oils that may be present in the curd compositions are described elsewhere throughout this disclosure, but in certain embodiments, the oil may be selected from acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof.

[0239] In many, but not all, embodiments, the curd compositions of the present disclosure comprise a solid fat. Solid fats that may be present in the curd compositions are described elsewhere throughout this disclosure, but in particular embodiments the solid fat may be selected from marine tallow, butter, chicken fat, clarified butter, cocoa butter, drippings, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof.

[0240] In some embodiments, the fungal curd composition comprises a non-fungal protein, such as hemp protein, soy protein, pea protein, chickpea protein, rice protein, or a combination thereof. As mentioned above, such non-fungal proteins may induce or assist in the aggregation of the fungal proteins.

[0241] In some embodiments, the filamentous fungal biomass used to make the curd can comprise particles produced by size reduction of cohesive filamentous fungal mycelial biomass. This biomass can be produced by any of several methods, including liquid surface fermentation, solid-state fermentation, or submerged fermentation. Additionally or alternatively, the curd can be or can be made from a mixed-type mycelial biomass composition as disclosed herein; as a non-limiting example, a fungal curd composition can be made by a fungal protein agglomeration method as disclosed herein, in which case the liquid dispersion used in the method is a liquid dispersion comprising particles of at least two different mycelial biomass types.

[0242] In one embodiment, the filamentous fungal particles consist essentially of fungal mycelia, hi other embodiments, the filamentous fungal particles comprise at least about 50% by weight fungal mycelia, at least about 75% by weight fungal mycelia, or at least about 95% by weight fungal mycelia.

[0243] Typically, fungal curd compositions are edible per se. Depending on the combination of flavoring agents added to the dispersion before, during, or after aggregation of the fungal proteins, they can achieve a variety of cheese-like, tofu-like, etc. flavors. They are also suitable for the production of other cheese-like food products, as described below.

[0244] In some embodiments, the fungal curd composition is a basic food ingredient, similar to, for example, dairy curd or tofu, and therefore may contain no components or ingredients other than the fungal material, a residual liquid phase (e.g., water), and any acids / bases, functional ingredients, and / or salts used to induce flocculation. In these embodiments, the fungal biomass and / or proteins, alone or in combination with acids / bases, functional ingredients, and / or salts used to induce flocculation, comprise at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, or at least about 80% by weight of the curd composition on a dry weight basis. or may account for about 80% by weight, at least about 85% by weight, at least about 90% by weight, at least about 95% by weight, at least about 96% by weight, at least about 97% by weight, at least about 98% by weight, at least about 99% by weight, at least about 99.1% by weight, at least about 99.2% by weight, at least about 99.3% by weight, at least about 99.4% by weight, at least about 99.5% by weight, at least about 99.6% by weight, at least about 99.7% by weight, at least about 99.8% by weight, at least about 99.9% by weight, or substantially all of theIn particular, the fungal curd compositions according to these embodiments may have a very low content or may be completely free of non-fungal gelling agents, emulsifiers, thickeners, and / or hydrocolloids, such as algae-derived or plant-derived polysaccharides, e.g., alginates and carrageenans, and the total content of non-fungal gelling agents, emulsifiers, thickeners, and / or hydrocolloids in the fungal curd compositions according to these embodiments may be less than about 2% by weight, less than about 1.9% by weight, less than about 1.8% by weight, less than about 1.7% by weight, less than about 1.6% by weight, less than about 1.5% by weight, less than about 1.4% by weight, less than about 1.3% by weight, less than about 1.2% by weight, less than about 1.1% by weight, less than about 0.9 ...5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about 1.5% by weight, less than about %, less than about 0.8%, less than about 0.7%, less than about 0.6%, less than about 0.5%, less than about 0.4%, less than about 0.3%, less than about 0.2%, less than about 0.19%, less than about 0.18%, less than about 0.17%, less than about 0.16%, less than about 0.15%, less than about 0.14%, less than about 0.13%, less than about 0.12%, less than about 0.11%, less than about 0.1%, less than about 0.09%, less than about 0.08%, less than about 0.07%, less than about 0.06%, less than about 0.05%, less than about 0.04%, less than about 0.03%, less than about 0.02%, less than about 0.01%, or 0% by weight.

[0245] The fungal curd compositions according to the present disclosure may have any desired moisture content or may be further processed as described elsewhere herein to have this. In some embodiments, the moisture content of the fungal curd composition may be similar to the moisture content of a conventional food product that the fungal curd composition is intended to resemble, such as tofu. As a first non-limiting example, if it is desired to make a "soft" (relatively moist) tofu analog food product, the fungal curd composition may have a moisture content of at least about 65% by weight, at least about 66% by weight, at least about 67% by weight, at least about 68% by weight, at least about 69% by weight, at least about 70% by weight, at least about 71% by weight, at least about 72% by weight, at least about 73% by weight, at least about 74% by weight, at least about 75% by weight, at least about 76% by weight, at least about 77% by weight, at least about 78% by weight, at least about 79% by weight, at least about 80% by weight, at least about 81% by weight, at least about 82% by weight, at least about 83% by weight, at least about 84% by weight, at least about 85% by weight, at least about 86% by weight, at least about 87% by weight, at least about 88% by weight, at least about 89% by weight, or at least about 90% by weight. As a second non-limiting example, if it is desired to make a "firm" (relatively low moisture) tofu analog food product, the fungal curd composition may have a moisture content of about 90% by weight or less, about 89% by weight or less, about 88% by weight or less, about 87% by weight or less, about 86% by weight or less, about 85% by weight or less, about 84% by weight or less, about 83% by weight or less, about 82% by weight or less, about 81% by weight or less, about 80% by weight or less, about 79% by weight or less, about 78% by weight or less, about 77% by weight or less, about 76% by weight or less, about 75% by weight or less, about 74% by weight or less, about 73% by weight or less, about 72% by weight or less, about 71% by weight or less, about 70% by weight or less, about 69% by weight or less, about 68% by weight or less, about 67% by weight or less, about 66% by weight or less, or about 65% by weight or less.

[0246] Fungal curd compositions according to the present disclosure may have any desired protein content. In some embodiments, the protein content of the fungal curd composition may be comparable to, or desirably higher than, the protein content of a conventional food product that the fungal curd composition is intended to resemble, such as tofu. As a non-limiting example, the protein content of the fungal curd composition may be at least about 40% by weight (on a dry weight basis), more particularly about 40% to about 60% by weight (or any value within any subrange thereof, e.g., any value within any range having a lower integer dry weight percentage limit of 40% to 60% by weight and an upper integer dry weight percentage limit of 40% to 60% by weight).

[0247] The fungal curd compositions according to the present disclosure may have any desired hardness value as measured by an appropriate texture profile analysis method (e.g., the method described in Example 14 below), or may be further processed as described elsewhere herein to have such a value. Hardness is defined as the maximum force during the first compression cycle. In some embodiments, the hardness of the fungal curd composition may be comparable to, or desirably higher or lower than, the hardness of a conventional food product that the fungal curd composition is intended to resemble, such as tofu. As a non-limiting example, the hardness of the fungal curd composition (as measured by the method described in Example 14 below) may be from about 1 N to about 50 N, or alternatively, any value within any range having a lower limit of any integer between 1 N and 50 N Newtons and an upper limit of any other integer between 1 N and 50 N Newtons.

[0248] A fungal curd composition according to the present disclosure may have, or may be further processed as described elsewhere herein to have, any desired cohesive strength value as measured by an appropriate texture profile analysis method (e.g., the method described in Example 14 below). In some embodiments, the cohesive strength of the fungal curd composition may be comparable to, or desirably higher or lower than, the cohesive strength of a conventional food product that the fungal curd composition is intended to resemble, such as tofu. As a non-limiting example, the cohesive strength of the fungal curd composition (as measured by the method described in Example 14 below) may be from about 0.001 N·mm to about 60 N·mm, or alternatively, any value within any range having a lower limit of any integer in Newton-millimeters to three decimal places from 0.001 N·mm to 60 N·mm, and an upper limit of any other integer in Newton-millimeters to three decimal places from 0.001 N·mm to 60 N·mm.

[0249] The fungal curd compositions according to the present disclosure may have any desired cohesive strength value as measured by an appropriate texture profile analysis method (e.g., the method described in Example 14 below), or may be further processed as described elsewhere herein to have such a value. Cohesive strength refers to how well a material can withstand a second deformation compared to a first deformation, which in the context of the materials of the present disclosure can be interpreted as the "tightness" of the aggregation of fungal proteins in the aggregated fungal protein composition to resist deformation; generally, a product with strong cohesion will be more able to withstand the stresses of fabrication, packaging, and shipping, and therefore more likely to be presented to the consumer / user in its expected condition. In some embodiments, the cohesive strength of the fungal curd composition may be comparable to, or desirably higher or lower than, the cohesive strength of a conventional food product that the fungal curd composition is intended to resemble, such as tofu. As a non-limiting example, the cohesive strength of the fungal curd composition (as measured by the method described in Example 14 below) can be from about 0.001 to about 4, or alternatively, any value within any range having a lower limit of any integer from 0.001 to 4 to three decimal places and an upper limit of any other integer from 0.001 to 4 to three decimal places.

[0250] Fungal curd products can be made to be very tasty and can provide a wide variety of taste and nutritional experiences depending on the additives, flavors, etc. with which the fungal curd product is combined to form the food product. In some embodiments, the curds produced by the disclosed methods using the disclosed liquid dispersions of filamentous fungal particles are characterized by one or more nutritional profiles shown in Tables 1-5:

[0251] [Table 1]

[0252] [Table 2]

[0253] [Table 3]

[0254] [Table 4]

[0255] [Table 5]

[0256] Fungal gel compositions containing aggregated fungal proteins In many embodiments, particularly those utilizing a mixed format mycelial biomass composition as disclosed herein, the aggregated fungal protein composition is in the form of a non-flowing gel. Such a gel is, in many embodiments, a food ingredient that can be consumed as a food product by itself or converted into a food product by mixing or otherwise combining with one or more other food components, such as flavorings, herbs, spices, flavor enhancers, fats, fat substitutes, preservatives, sweeteners, color additives, nutrients, emulsifiers, stabilizers, thickeners, pH adjusters, acidulants, leavening agents, anti-caking agents, water retention agents, yeast nutrients, dough enhancers, dough conditioners, firming agents, enzymes, gases, vegetables, fruits, non-animal-derived proteins such as plant proteins (e.g., pea protein, soy protein, and textured plant proteins), meat products, and the like. In some embodiments, the gel may be a food product or may be suitable for combining with one or more other food components to form a food product similar to conventional or known food products that contain dairy or otherwise animal-derived ingredients (milk, eggs, etc.), and the mycelial biomass may be provided in addition to or in place of the animal-derived ingredients. In some embodiments, the gel may be a non-dairy composition, or more specifically a vegan composition (i.e., a composition that does not contain animal-derived components). Examples of food products that can be made using the flocculated fungal protein-containing gels of the present disclosure include, but are not limited to, blancmange, bread, butter, cake, custard, egg white foam, ice cream, jam, jelly, margarine, mayonnaise, meringue, milk, whipped cream, yogurt, cream cheese, emulsified meat products (e.g., hot dogs), vegetarian and vegan analogs of emulsified meat products, and many sauces and spreads (e.g., béchamel sauce, espagnole sauce, hollandaise sauce, hummus, Russian dressing, tartar sauce, thousand island dressing, velouté, etc.), and / or the like. In some embodiments, the gels and / or food products made therefrom may be gluten-free.

[0257] In some embodiments, the fungal gel composition is a basic food ingredient that may contain no components or ingredients other than the fungal material, a residual liquid phase (e.g., water), and any acids / bases, functional ingredients, and / or salts used to induce flocculation. In these embodiments, the fungal biomass and / or protein may comprise at least about 25% by weight, at least about 30% by weight, at least about 35% by weight, at least about 40% by weight, at least about 45% by weight, at least about 50% by weight, at least about 55% by weight, at least about 60% by weight, at least about 65% by weight, at least about 70% by weight, at least about 75% by weight, at least about 80% by weight, at least about 85% by weight, at least about 90% by weight, at least about 95% by weight, at least about 96% by weight, at least about 97% by weight, at least about 98% by weight, at least about 99% by weight, at least about 99.1% by weight, at least about 99.2% by weight, at least about 99.3% by weight, at least about 99.4% by weight, at least about 99.5% by weight, at least about 99.6% by weight, at least about 99.7% by weight, at least about 99.8% by weight, at least about 99.9% by weight, or substantially all of the gel composition on a dry weight basis.In particular, the fungal gel compositions according to these embodiments may have very low or no non-fungal gelling agents, emulsifiers, thickeners, and / or hydrocolloids, such as algae-derived or plant-derived polysaccharides, e.g., alginates and carrageenans, and the total content of non-fungal gelling agents, emulsifiers, thickeners, and / or hydrocolloids in the fungal gel compositions according to these embodiments may be less than about 2% by weight, less than about 1.9% by weight, less than about 1.8% by weight, less than about 1.7% by weight, less than about 1.6% by weight, less than about 1.5% by weight, less than about 1.4% by weight, less than about 1.3% by weight, less than about 1.2% by weight, less than about 1.1% by weight, less than about 1% by weight, less than about 0.9 ... %, less than about 0.8%, less than about 0.7%, less than about 0.6%, less than about 0.5%, less than about 0.4%, less than about 0.3%, less than about 0.2%, less than about 0.19%, less than about 0.18%, less than about 0.17%, less than about 0.16%, less than about 0.15%, less than about 0.14%, less than about 0.13%, less than about 0.12%, less than about 0.11%, less than about 0.1%, less than about 0.09%, less than about 0.08%, less than about 0.07%, less than about 0.06%, less than about 0.05%, less than about 0.04%, less than about 0.03%, less than about 0.02%, less than about 0.01%, or about 0% by weight.

[0258] Post-processing of compositions of aggregated fungal proteins Once the agglomerated fungal protein composition is produced, it can be further processed to produce food products such as cheese analogue food products, tofu analogue food products, etc. In particular, the fungal curd compositions produced by the methods disclosed herein are suitable for preparing non-dairy, often vegan, cheese analogue and tofu analogue food products.

[0259] In some embodiments, the fungal curd compositions according to the present disclosure can be further processed to produce cultured cheese analog food products, i.e., products in which microbial food cultures (i.e., live bacteria, yeast, or molds) have been introduced into the fungal curd composition. As a non-limiting example, the fungal food material according to the present invention can be cultured with Lactobacillus spp. or other lactic acid bacteria (e.g., to make a cheese analog food product or other dairy analog food product). In some embodiments, the fungal curd composition can be cultured with two or more microbial food cultures, either simultaneously or sequentially, to produce an analog of a cheese food product made by fermentation of the two or more microbial cultures; by way of non-limiting example, cultured food products according to the present disclosure can include, among others, a semi-soft ripened cheese analog food product (made by subjecting the fungal curd to a first culture of Lactobacillus spp. or other lactic acid bacteria and a second culture of cheese-ripening yeast), a blue cheese analog food product (made by subjecting the fungal curd to a first culture of Lactobacillus spp. or other lactic acid bacteria and a second culture of a mold such as Penicillium roqueforti), or a soft-ripened cheese (e.g., Brie or Camembert) analog food product (made by subjecting the fungal curd to a first culture of Lactobacillus spp. or other lactic acid bacteria and a second culture of a mold such as Penicillium roqueforti). spp. or other lactic acid bacteria and a second culture with Penicillium camemberti).

[0260] In some embodiments, the cheese analog food product comprises a thickening or gelling agent. Such agents are known in the art and include agar, gelatin, starches (e.g., arrowroot, tapioca, corn, potato), higher fat liquids (e.g., coconut milk), fats (e.g., coconut flakes, deodorized or otherwise), chickpea water, flaxseed, xanthan gum, guar gum, psyllium husk, ground chia seeds, nut and / or seed butters, pumpkin puree, cooked mashed yams ... Included are mashed sweet potatoes, applesauce, mashed overripe bananas or plantains, pureed dates or prunes, soaked and stewed figs, shredded fruits / vegetables, shredded coconut, gluten-free flours (e.g., teff flour, buckwheat flour, amaranth flour, chickpea flour, sorghum flour, almond flour), cooked pureed beans, cocoa powder, vegetable gums, polysaccharides, vegetable mucilages, seaweed derivatives, pectin, gluten, soy, and egg analogs. The thickener can be a fat, which can be a liquid such as coconut milk, or a solid such as deodorized coconut flakes.

[0261] In some embodiments, cheese analog food products that can be produced from the fungal curd compositions disclosed herein contain lactic acid bacteria (LAB). These bacteria produce lactic acid as a major metabolic end product of carbohydrate fermentation. Examples of LAB include Lactobacillus, Leuconostoc, Pediococcus, Lactococcus, and Streptococcus. In some embodiments, the cheese analog food product contains the bacteria Lactobacillus bulgaricus and / or Streptococcus thermophilus.

[0262] In some embodiments, the cheese analog food product further comprises rennet. The rennet can be derived from animal, plant, or microbial sources. In vegetarian or vegan food products, the rennet is derived from vegetarian and / or microbial sources.

[0263] In some embodiments, the cheese analog food product further comprises enzyme water, which is produced as follows: 100 g of whole rye or durum wheat seeds (or other suitable whole grain seeds) are combined with 1 liter of water and allowed to germinate for 2-4 hours. Once the seeds begin to sprout and the first roots appear, the seeds are transferred to a clean jar containing 1 liter of water. The jar is covered with a permeable cloth (e.g., linen or cotton) and incubated at room temperature for 24 hours, at the end of which the water in the jar changes color and odor. This water, called enzyme water, can be used to produce cheese analog food products.

[0264] In some embodiments, the cheese analog food product further comprises probiotics, which are mixtures of live microorganisms such as bacteria and yeast that confer health benefits, including improved digestion.

[0265] In some embodiments, the cheese analog food product comprises milk solids derived from animal milk. In some embodiments, the cheese analog food product does not comprise milk solids derived from animal milk.

[0266] As non-limiting examples, cheese analog food products produced from the curds provided by the present disclosure may, in some embodiments, be a hard cheese (e.g., Parmesan) analog food product, a semi-hard cheese (e.g., Gouda) analog food product, a semi-soft cheese (e.g., Havarti) analog food product, a soft or soft ripened cheese (e.g., Brie) analog food product, a cream cheese analog food product, a sour milk cheese analog food product, a blue cheese analog food product, a mascarpone cheese analog food product, a pasta filata (mozzarella) cheese analog food product, a blind cheese (feta) analog food product, a whey cheese (ricotta or brunost) analog food product, or a fresh cheese (cottage cheese) analog food product. In other embodiments, the fungal curd provided by the present disclosure can be used to make a cream analog food product such as a creme fraiche analog food product, a smetana analog food product, a sour cream analog food product, a half and half analog food product, a table cream analog food product, a whipped cream analog food product, a double cream analog food product, a clotted cream analog food product, a sour cream analog food product, a pasteurized cream analog food product, or a condensed cream analog food product.

[0267] Referring now to Figure 7, one non-limiting embodiment of a method 700 for making a tofu analog food product is illustrated. The embodiment of method 700 illustrated in Figure 7 includes a fungal curd formation step 710, a curd separation step 720, a first pressing step 730, an optional breaking step 740, an optional pasteurization step 750, and an optional second pressing step 760.

[0268] In fungal curd formation step 710, a fungal curd is formed by any of the methods for forming a fungal curd disclosed herein. As a first non-limiting example, the fungal curd can be formed by inducing aggregation of fungal proteins by pH adjustment, e.g., by adjusting the pH of a liquid dispersion of filamentous fungal particles to a pH of about 3.5 at a temperature of about 175°F (79.4°C) with slow mixing and / or stirring. As a second non-limiting example, the fungal curd can be formed by agglomeration of fungal proteins by adding salt, e.g., by adding calcium chloride and / or magnesium chloride to a liquid dispersion of filamentous fungal particles at a temperature of about 175°F (79.4°C) with slow mixing and / or stirring without pH adjustment.

[0269] In curd separation step 720, the fungal curd is separated from the liquid phase of the liquid dispersion by any suitable liquid-solid separation technique, non-limiting examples of which include decanting, pressing, filtration, gravity separation, screw separation, and centrifugation.

[0270] In a first pressing step 730, the fungal curd is mechanically pressed to squeeze out at least a portion of the trapped water (thus reducing the moisture content and increasing the solids content of the fungal curd) and compress the fungal curd into a shape suitable for packaging and / or further processing. Most typically, the fungal curd is pressed into a rectangular block in the first pressing step 730 by any suitable type of mechanical press or other similar machine or device commonly used in making tofu (e.g., a conventional screw-type tofu mould).

[0271] In an optional breaking step 740, the pressurized tofu analog food product may be broken down into smaller particles for subsequent processing. In some embodiments, the optional breaking step 740 may be performed to improve the effectiveness of subsequent processing steps, such as the optional pasteurization step 750. Additionally or alternatively, the optional breaking step 740 may be performed to incorporate additional ingredients (e.g., chopped vegetables, ground meat, etc.) into the tofu analog food product to create a cooking ingredient.

[0272] The tofu analog food product may be pasteurized to remove potentially pathogenic microorganisms in the tofu analog food product in optional pasteurization step 750. The pasteurization step may be carried out by any suitable means and under any suitable conditions known to those skilled in the art, and as a non-limiting example, optional pasteurization step 750 may be achieved by rapidly heating the tofu analog food product to a temperature of about 90-100°C and maintaining the tofu analog food product at this temperature for about 5-30 minutes.

[0273] In an optional second pressing step 760, the tofu analog food product may again be mechanically pressed to further squeeze out trapped water (thus further reducing the moisture content and increasing the solids content of the fungal curd) and again compress the fungal curd into a shape suitable for packaging and / or further processing. The optional second pressing step 760 may be performed to produce tofu analog food products with particularly low moisture contents; by way of non-limiting example, while a single pressing step 730 may be effective to produce soft / "silky" tofu or firm tofu analog food products (e.g., tofu analog food products with a surface firmness similar to fresh meat and an internal texture similar to firm custard), a second pressing step 760 may be required to produce extremely firm tofu analog food products (e.g., tofu analog food products with a firmness similar to cooked meat and a chewy or crumbly texture similar to that of paneer).

[0274] A first pressurizing step 730 and an optional second pressurizing step 760 can be performed to pressurize residual water out of the tofu analog product to the desired extent. In certain embodiments, the pressurizing step can reduce the water content of the fungal curd and / or tofu analog product to an amount that is equivalent to the water content of a conventional tofu product, for example, about 65% to about 90% by weight (or any value within any subrange thereof). As a first non-limiting example, if it is desired to make a "soft" (relatively moist) tofu analog food product, the first pressing step 730 and optionally the second pressing step 760 can reduce the water content of the fungal curd and / or tofu analog to at least about 65% by weight, at least about 66% by weight, at least about 67% by weight, at least about 68% by weight, at least about 69% by weight, at least about 70% by weight, at least about 71% by weight, at least about 72% by weight, at least about 73% by weight, at least about 74% by weight, at least about 75% by weight, at least about 76% by weight, at least about 77% by weight, at least about 78% by weight, at least about 79% by weight, at least about 80% by weight, at least about 81% by weight, at least about 82% by weight, at least about 83% by weight, at least about 84% by weight, at least about 85% by weight, at least about 86% by weight, at least about 87% by weight, at least about 88% by weight, at least about 89% by weight, or at least about 90% by weight. As a second non-limiting example, if it is desired to make a "firm" (relatively low moisture) tofu analog food product, the first pressing step 730 and optionally the second pressing step 760 can reduce the water content of the fungal curd and / or tofu analog to about 90% by weight or less, about 89% by weight or less, about 88% by weight or less, about 87% by weight or less, about 86% by weight or less, about 85% by weight or less, about 84% by weight or less, about 83% by weight or less, about 82% by weight or less, about 81% by weight or less, about 80% by weight or less, about 79% by weight or less, about 78% by weight or less, about 77% by weight or less, about 76% by weight or less, about 75% by weight or less, about 74% by weight or less, about 73% by weight or less, about 72% by weight or less, about 71% by weight or less, about 70% by weight or less, about 69% by weight or less, about 68% by weight or less, about 67% by weight or less, about 66% by weight or less, or about 65% by weight or less.

[0275] Method 700, illustrated in Figure 7, can provide a resultant fungal tofu analog product having any desired firmness value as measured by an appropriate texture profile analysis method (e.g., the method described in Example 14 below). In certain embodiments, method 700 can provide a resultant fungal tofu analog product having a firmness that is comparable to, or desirably higher or lower than, the firmness of a conventional tofu product. As a non-limiting example, the firmness of the tofu analog product (as measured by the method described in Example 14 below) can be from about 1 N to about 50 N, or alternatively, any value within any range having a lower integer limit between 1 N and 50 N Newtons and an upper integer limit between 1 N and 50 N Newtons.

[0276] The method 700 illustrated in Figure 7 can provide a resultant fungal tofu analog product having any desired stickiness value as measured by an appropriate texture profile analysis method (e.g., the method described in Example 14 below). In certain embodiments, the method 700 can provide a resultant fungal tofu analog product having a stickiness that is comparable to, or desirably higher or lower than, the stickiness of a conventional tofu product. As a non-limiting example, the stickiness of the tofu analog product (as measured by the method described in Example 14 below) can be any value between about 0.001 N·mm and about 60 N·mm, or alternatively, any value within any range having a lower limit of any integer in Newton-millimeters to three decimal places from 0.001 N·mm to 60 N·mm, and an upper limit of any other integer in Newton-millimeters to three decimal places from 0.001 N·mm to 60 N·mm.

[0277] Method 700, illustrated in Figure 7, can provide a resultant fungal tofu analog product having any desired cohesive strength value as measured by an appropriate texture profile analysis method (e.g., the method described in Example 14 below). In certain embodiments, method 700 can provide a resultant fungal tofu analog product having a cohesive strength that is comparable to, or desirably higher or lower than, that of a conventional tofu product. As a non-limiting example, the cohesive strength of the tofu analog product (as measured by the method described in Example 14 below) can be any value from about 0.001 to about 4, or alternatively, any value within any range having a lower limit of any integer from 0.001 to 4 to three decimal places and an upper limit of any other integer from 0.001 to 4 to three decimal places.

[0278] The concepts disclosed herein are further illustrated by the following non-limiting examples, which are presented solely for the purpose of illustrating particular embodiments of the disclosure and in no way limit the scope or breadth of the disclosure.

[0279] [Example 1] Fungal curd formation by bacterial culture or acid addition Two batches of cheese curd analogs were prepared from an edible filamentous fungus, specifically the filamentous acidophilic Fusarium strain flavolapis (also known as "F. flavolapis" or "Ff"). This experiment was designed to test the difference in curd production between the addition of bacterial culture (Batch 1, as shown in Table 6 below) and the addition of acid (Batch 2, as shown in Table 7 below) to the fungal preparation. Biomat growth and preparation of F. flavolapis was performed as described in PCT / US2020 / 020152 (WO 2020 / 176758).

[0280] [Table 6]

[0281] Water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. All powdered ingredients (soy flour, dextrose, hemp protein, salt, tricalcium citrate, dairy fortifier, flavoring, taste modifier) ​​were added to the aqueous dispersion and mixed until completely dissolved. The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0282] Coconut oil was then added and the entire combination mixed until smooth. The mixture was then heated to 185°F for 15 minutes and homogenized while hot at 200 bar pressure. The homogenate was then cooled to 110°F, at which point 0.02% bacterial culture preparation (VEGA™ Vibe, Chr. Hansen Holding A / S, DK) was added along with 0.5% triglyceride mixture. The batch was held at 110°F for 6 hours, at which point curd formation was observed.

[0283] [Table 7]

[0284] Water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. All powdered ingredients (soy flour, dextrose, hemp protein, salt, tricalcium citrate, dairy fortifier, flavoring, taste modifier) ​​were added to the aqueous dispersion and mixed until completely dissolved. The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0285] Coconut oil was then added and the entire combination mixed at high shear (e.g., 10,000 rpm for 2 minutes) until smooth. The mixture was then heated to 160°F with slow mixing (speed 0.5), 2g of 50% citric acid was added, and the resulting mixture was slowly heated to 190°F over a period of 15 minutes. At this stage, mixing was stopped and the batch was allowed to rest at 190°F for an additional 10 minutes, at which stage curd formation was observed.

[0286] Although curd formation was observed in both batches, these data suggest that the acid formation of the curd occurs more quickly than the curd formed by the bacterial culture.

[0287] [Example 2] Fungal curd formation with the addition of citric acid A third batch of cheese curd analog was prepared from an edible filamentous fungus, specifically the filamentous acidophilic Fusarium strain F. flavolapis, as shown in Table 8 below. This experiment was conducted to confirm the reproducibility of curd formation using the addition of acid to the fungal preparation. Biomat growth and preparation of F. flavolapis was performed as described in PCT / US2020 / 020152 (WO 2020 / 176758).

[0288] [Table 8]

[0289] Water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. All powdered ingredients (dextrose, hemp protein, salt, tricalcium citrate, flavorings, taste modifiers) were added to the aqueous dispersion and mixed until completely dissolved. The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0290] Coconut oil was then added, and the entire combination was mixed at high shear (10,000 rpm for 2 minutes) until smooth. The mixture was then heated to 160°F with slow mixing (speed 0.5), 2g of 50% citric acid was added, and the resulting mixture was slowly heated to 190°F over a period of 15 minutes. Mixing was then stopped and the batch was allowed to rest at 190°F for an additional 10 minutes, as in Batch 2 of Example 1, at which stage curd formation was observed, confirming the reproducibility of curd production using acid.

[0291] [Example 3] Fungal curd formation with higher fungal protein content A fourth batch of cheese curd analog was prepared from an edible filamentous fungus, specifically the filamentous acidophilic Fusarium strain F. flavolapis. This experiment was conducted to test higher concentrations of fungus, increasing from 2.70%-3.00% to 4.00% of the total composition (as shown in Table 9 below), along with homogenization and their overall effect on curd formation. F. flavolapis biomat growth and preparation were performed as described in PCT / US2020 / 020152 (WO 2020 / 176758).

[0292] [Table 9]

[0293] Water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. All powdered ingredients (dextrose, hemp protein, salt, tricalcium citrate, flavorings, taste modifiers) were added to the aqueous dispersion and mixed until completely dissolved. The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0294] Coconut oil was then added, and the entire combination was mixed until smooth and then homogenized at 160 bar pressure. The mixture was then heated to 165°F with slow mixing (speed 0.5), 2g of 50% citric acid was added, and the resulting mixture was further heated to 190°F over a period of 15 minutes. Mixing was then stopped, and the batch was allowed to rest at 190°F for an additional 5 minutes, similar to Batch 2 in Example 1. Curd formation was not observed in this batch, which may be due to an excessively high concentration of F. flavolapis or the homogenization process.

[0295] [Example 4] Effect of hemp protein and texturizing agents on fungal curd formation Two additional batches 5 and 6 of cheese curd analogs were prepared from edible filamentous fungi, specifically the filamentous acidophilic Fusarium strain F. flavolapis. The experiments were conducted first to determine the effect of hemp protein on curd production (Batch 5, as shown in Table 10 below) and to determine whether the addition of texturizing agents could improve the resulting properties of the curd (Batch 6, as shown in Table 11 below), such as cut and yield. F. flavolapis biomat growth and preparation were performed as described in PCT / US2020 / 020152 (WO 2020 / 176758).

[0296] Batch 5 - This batch was prepared twice: once with 2.7% F. flavolapis and 2% hemp protein, and once with 2.7% F. flavolapis and 3% hemp protein.

[0297] [Table 10]

[0298] For each batch made, water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. All powdered ingredients (dextrose, hemp protein, salt, tricalcium citrate, flavorings, taste modifiers) were added to the aqueous dispersion and mixed until completely dissolved. The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0299] Coconut oil was then added, and the entire combination was mixed until smooth, resulting in a pH of 7.0 for the 2% hemp protein batch and 7.2 for the 3% hemp protein batch. The mixture was then heated to 170°F with slow mixing (speed 0.5), and 2g of 50% citric acid was added. At this stage, the approximate pH of the 2% hemp protein batch was 5.8, and the approximate pH of the 3% hemp protein batch was 6.0. The resulting mixture was slowly heated to 190°F over a period of 15 minutes. Mixing was then stopped, and each batch was allowed to rest at 190°F for an additional 5 minutes, at which point curd formation was observed. The curd yield for both the 2% hemp protein and 3% hemp protein batches was approximately 50%.

[0300] Batch 6 - This batch was also prepared twice: both contained 3% F. flavolapis and 2% hemp protein, and included sunflower lecithin as an emulsifier.

[0301] [Table 11]

[0302] For each batch made, water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. All powdered ingredients (sunflower lecithin, dextrose, hemp protein, salt, tricalcium phosphate, flavorings, taste modifiers) were added to the aqueous dispersion and sheared until completely dissolved (e.g., 10,000 rpm for 2 minutes). The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0303] Coconut oil was then added, and the entire combination was sheared (e.g., at 10,000 rpm for 2 minutes) until emulsified, resulting in a pH range of 7.5 to 7.6 in the batches. The mixture was then heated to 170°F with slow mixing (speed 0.5), and 3 g of 50% citric acid was added. At this stage, the approximate pH range for both batches was 5.9 to 6.0. Mixing was stopped, and both batches were allowed to rest at 170°F without mixing. Slow mixing (speed 0.5) was then resumed, and the batches were slowly heated to 190°F over a 15-minute period. Again, mixing was stopped, and each batch was allowed to rest for an additional 5 minutes at 190°F, at which point curd formation was observed. The curd yield for both batches was approximately 45%. Despite the lower yield, Batch 6 had a clearer curd that exhibited cleaner cuts than Batch 5.

[0304] [Example 5] Effect of subsequent addition of calcium on fungal curd formation. An additional batch 7 of cheese curd analog was prepared from an edible filamentous fungus, specifically the filamentous acidophilic Fusarium strain F. flavolapis. This experiment was conducted to determine the effect of adding calcium much later in the curd formation process. Additionally, as shown in Table 12 below, while all previous batches used citric acid, this batch used a different acid (here, lactic acid). F. flavolapis biomat growth and preparation was performed as described in PCT / US2020 / 020152 (WO 2020 / 176758).

[0305] [Table 12]

[0306] As in previous examples, water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion. However, in this batch, all powdered ingredients (sunflower lecithin, dextrose, hemp protein, salt, flavorings, taste modifiers) except for calcium lactate were added to the aqueous dispersion and sheared until completely dissolved (e.g., 10,000 rpm for 2 minutes). The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0307] Coconut oil was then added, and the entire combination was sheared (e.g., at 10,000 rpm) for 10 minutes until emulsified, resulting in a pH of 7.5-7.6. The mixture was then heated to 170°F with slow mixing (speed 0.5), and both calcium lactate and 2 g of 90% lactic acid were added. At this stage, the pH dropped to approximately 5.9-6.0. Mixing was then stopped, and the batch was allowed to rest at 170°F for 10 minutes. The temperature was then increased to 190°F, and the batch was held at this temperature for 5 minutes, at which stage curd formation was observed. The curd yield was approximately 45%. This batch 7 had a clearer curd that exhibited a cleaner cut than batch 6.

[0308] [Example 6] Formation of fungal cheese curd-like material An additional batch 8 of cheese curd analog was prepared from an edible filamentous fungus, specifically the filamentous acidophilic Fusarium strain F. flavolapis. Biomat growth and preparation of F. flavolapis was performed as described in PCT / US2020 / 020152 (WO 2020 / 176758), as shown in Table 13 below.

[0309] [Table 13]

[0310] Water and F. flavolapis were placed in a blender (Vitamix®, Vita-Mix Corporation, USA) and sheared until smooth (e.g., 10,000 rpm for 2 minutes) to create an aqueous dispersion as shown in Figure 8. The aqueous dispersion was then transferred to a Thermomix multicooker. For this batch, all powdered ingredients (sunflower lecithin, dextrose, hemp protein, salt, flavorings) except for tricalcium phosphate were added to the aqueous dispersion and sheared until completely dissolved (e.g., 10,000 rpm for 2 minutes). The resulting mixture was heated to 110°F for 10 minutes to hydrate the powders.

[0311] Coconut oil was then added, and the entire combination was sheared (e.g., at 10,000 rpm) for 10 minutes until emulsified, resulting in a pH of 7.0-7.6. The mixture was then heated to 170°F with slow mixing (speed 0.5). Tricalcium phosphate was then added while whisking. Immediately thereafter, 2 g of 90% lactic acid was added while whisking. As shown in Figures 9 and 10, a loose curd immediately began to form. At this stage, the approximate pH dropped to 5.5-6.0. Mixing was then stopped, and the batch was allowed to rest at 170°F for 10 minutes. The temperature was then increased to 190°F, and the batch was held at this temperature for 5 minutes, at which stage curd formation was observed, as shown in Figures 11 and 12. The curd was then poured through cheesecloth to separate it from the liquid, as shown in Figure 13. The resulting curd is cooled (e.g., to 34°F-40°F, 1°C-4°C) over a period of approximately 24 hours, after which the curd resembles a soft spreadable or ricotta-like cheese, as shown in Figure 14.

[0312] [Example 7] Gelation of soaked fabric / biomat dispersion mycelium biomass blends Steeped Dough: Fungal biomass from an edible filamentous fungus, specifically the filamentous acidophilic Fusarium strain F. flavolapis, was prepared by steeped fermentation in a stirred-tank reactor. After growth, steam was injected into the fermenter until the temperature reached approximately 80°C to inactivate the biomass. After inactivation, the fermentation broth was dehydrated to produce a biomass with a dough-like consistency, with an isoelectric point of 2.20 and an average particle size of 61.71 μm. The steeped dough biomass was pureed using a grinder, and the moisture content of the pureed material was measured and determined to be 83-86% by weight. Water was added to the pureed biomass at a water:biomass ratio of 3:1 (w / w). The water / biomass mixture was placed in a Vitamix blender and blended at the maximum setting for 1 minute, then homogenized for 2 minutes using a high-speed (10,000 rpm) homogenizer. The homogenized mixture was transferred to a Thermomix multi-cooker and cooked at 175°F and speed 3.0 for 30 minutes, and the resulting liquid dispersion (hereafter referred to in this example as "dough milk") was then cooled to room temperature. The pH of this milk ranged from 5.8 to 6.2.

[0313] 1M hydrochloric acid was added dropwise to the batter milk. A pH meter was used to record the pH throughout the acidification process. The acidified batter milk became more viscous but did not form a gel when the pH was lowered to 3.5. Two samples of the acidified batter milk were stored for 10 minutes, one at room temperature and one refrigerated. The container holding the acidified batter milk was then inverted, and the milk poured out of the container.

[0314] Biomat pieces: Biomat of Fusarium strain F. flavolapis was produced by a surface fermentation process. Biomat pieces with an isoelectric point of 2.25 and an average particle size of 66.21 μm were pureed using a grinder. The moisture content of the pureed material was measured and determined to be 78-80% by weight. Water was added to the pureed biomass at a water:biomass ratio of 3:1 (w / w). The water / biomass mixture was placed in a Vitamix blender and blended at the maximum setting for 1 minute, then homogenized for 2 minutes using a high-speed (10,000 rpm) homogenizer. The homogenized composition was transferred to a Thermomix multicooker and heat-treated at 175°F and speed 3.0 for 30 minutes. The resulting liquid dispersion (hereafter referred to as "biomat milk") was then cooled to room temperature. The pH of this milk ranged from 6.4 to 6.5.

[0315] 1M hydrochloric acid was added dropwise to the BioMat milk. A pH meter was used to record the pH throughout the acidification process. The pH was reduced to 3.5, and two samples of the acidified BioMat milk were stored for 10 minutes, one at room temperature and one refrigerated. The container holding the acidified BioMat milk was then inverted, and the gel system that formed did not flow out of the container.

[0316] Blend #1: Biomat milk and dough milk (non-acidified) were prepared as described above and combined in a 1:3 ratio of Biomat milk:dough milk. The blend was mixed on a stir plate at 7,000 rpm for 5 minutes at room temperature. The pH was then adjusted to at least 3.5, but not more than 4, by adding 1 M hydrochloric acid to produce an acidified blend. After the pH adjustment, two samples of the acidified blend were allowed to rest for at least 10 minutes, one at room temperature and one in the refrigerator. After this resting period, the containers of both samples were inverted and no flow was observed, indicating that the adjustment of the pH to between 3.5 and 4 resulted in gelation, i.e., the formation of a gel system in the previously fluid mycelial biomass blend, as shown in Table 14 below.

[0317] [Table 14]

[0318] [Example 8] Gelation of soaked dough / soaked flour mycelial biomass compositions Steeped dough: The steeped dough was processed into dough milk as described in Example 7. The dry solids content of the dough milk was calculated to be 15%. As described in Example 7, the steeped dough became more viscous but did not form a gel, even when the pH was lowered to 3.5.

[0319] Immersion spray-dried powder: A fungal biomass of Fusarium strain F. flavolapis was prepared by stirred-tank fermentation. After growth, steam was injected into the fermenter until the temperature reached approximately 80°C to inactivate the biomass. After inactivation, the biomass was washed with deionized water and collected as an approximately 25% solids mixture. The wet mixture was then spray-dried to produce an immersion spray-dried powder with a final solids content of approximately 98%, an isoelectric point of 2.55, and an average particle size of 20.48 μm. A solution of the immersion spray-dried powder was prepared using tap water at a concentration of 6.5% (w / v). The water / biomass mixture was placed in a Vitamix blender and blended at the maximum setting for 1 minute, then homogenized for 2 minutes using a high-speed (10,000 rpm) homogenizer. The homogenized composition was transferred to a Thermomix multicooker and heated at 175°F and speed 3.0 for 30 minutes. The resulting liquid dispersion (hereafter referred to as "milk powder") was then cooled to room temperature. 1 M hydrochloric acid was added dropwise to the resulting milk powder. The pH was recorded throughout the acidification process using a pH meter. The pH was reduced from an initial pH of 5.5-6.3 to 3.5, and two samples of the acidified milk powder were stored for 10 minutes, one at room temperature and one refrigerated. The container holding the acidified milk powder was then inverted, allowing the milk to flow out of the container.

[0320] Blend #2: Dough milk (non-acidified) and soaked spray-dried powder were prepared as described above. The dry solids content of the dough milk was calculated to be approximately 6.5%. Based on the calculated dry solids content, the soaked spray-dried powder and dough milk were combined in a 2:3 powder:dough milk ratio. The powder / dough milk composition was mixed on a stir plate at 7,000 rpm at room temperature for at least 30 minutes to ensure all of the flour molecules were hydrated. The pH was then adjusted to at least 3.5 but not more than 4.0 by adding 1 M hydrochloric acid to produce an acidified blend. After the pH adjustment, two samples of the acidified blend were allowed to rest for at least 10 minutes, one at room temperature and one in the refrigerator. After this resting period, the containers of both samples were inverted and no flow was observed, indicating that adjusting the pH to between 3.5 and 4 resulted in gelation, i.e., the formation of a gel system in the previously fluid mycelium biomass blend, as shown in Table 15 below.

[0321] [Table 15]

[0322] [Example 9] Emulsion stability of pH-adjusted fungal liquid dispersions containing oils For each of the dough milks described in Example 7, powdered milks described in Example 8, and BioMat milks described in Example 7, 60 g of room temperature milk and 120 g of water were added to a 350 mL beaker and stirred for 30 seconds to ensure thorough mixing. 20 g of vegetable oil was added to each beaker, and the resulting fungal dispersion-oil mixture was homogenized at 10,000 rpm for 2 minutes to form an oil-in-water emulsion. Samples of each emulsion were taken from the bottom of each beaker and transferred to glass vials or 100 mL graduated cylinders using a 10 mL automatic pipettor. The pH of each sample was adjusted to either pH 4 or pH 7.

[0323] The emulsion stability index (ESI) of each emulsion was calculated 1, 2, and 24 hours after the samples were transferred to the vials or graduated cylinders; the ESI is defined as the height of the emulsified phase in the vial or cylinder, expressed as a percentage of the total height of the liquid in the vial or cylinder (i.e., the height of the emulsified phase plus the height of the aqueous phase that separated from the emulsion and sank to the bottom of the vial / cylinder). These results are shown in Figure 15 (for each milk type / pH combination, the left-most bar in the bar graph in Figure 15 represents t=1 hour, the middle bar represents t=2 hours, and the right-most bar represents t=24 hours). As shown in Figure 15, liquid dispersions made from all three mycelial biomass formats had oil-in-water emulsion stability greater than 50% for more than 24 hours in both pH ranges. In particular, both dough milk and biomat milk formed very stable emulsions at pH 4, and biomat pieces formed very stable emulsions at pH 7.

[0324] [Example 10] Rheological properties of fungal liquid dispersions and gels made from them The apparent viscosity of each of batter milk as described in Example 7, powdered milk as described in Example 8, and BioMat milk as described in Example 7 was measured over a range of shear rates at their unadjusted pH values. As illustrated in Figure 16, all three milks exhibited shear thinning behavior, with the viscosity of the powdered milk (diamond data points) being significantly lower than that of the BioMat milk (square data points) and batter milk (circle data points) at all shear rates.

[0325] Additionally, as illustrated in Figure 17, the biomat milk of Example 7 exhibits shear thinning behavior, i.e., viscosity decreases with increasing shear rate, at all pH levels: starting pH (pH 6.4-6.5, bottom curve), pH 4.5 (middle curve), and pH 3.5 (upper curve). Furthermore, regardless of shear rate, the biomat milk was observed to thicken (i.e., exhibit increasing viscosity) as the pH decreased toward pH 3.5; we hypothesize that this phenomenon may be due to gelation due to aggregation of fungal proteins, as supported by the frequency sweep data presented in Figure 18. Specifically, as illustrated in Figure 18, when BioMat Milk is subjected to a frequency sweep test at 0.1% strain (in the linear viscoelastic region), it exhibits higher storage modulus (G') values ​​as the pH is decreased from the starting pH (filled triangle data points) through pH 4.5 (upper set of open circle data points) to pH 3.5 (filled circle data points), suggesting that the "stiffness" of the gel is enhanced by decreasing the pH to approximately 3.5.

[0326] Another sample of BioMat Milk was loaded onto the rheometer immediately after adjusting the pH to 3.5 to determine the time required for the liquid dispersion to transition to a true gel system. As illustrated in Figure 19, the transition from a liquid dispersion to a true gel system occurs almost instantly upon lowering the pH to 3.5; the storage modulus (G', upper curve) of the sample was over 1000 Pa at t = 0 seconds (equivalent to the gel composition at pH 3.5 illustrated in Figure 18) and remained essentially constant over the 3 hour test.

[0327] [Example 11] Gelation by adding non-fungal proteins and adjusting pH Commercially available potato protein with an isoelectric point (pI) of 5.1 was added in an amount of 1% by weight to the dough milk as described in Example 7, and the pH of the milk was then adjusted to 4.2 by dropwise addition of 1 M hydrochloric acid. At this pH, due to their respective isoelectric points, the potato proteins have a significant positive surface charge and the proteins of the soaked dough have a significant negative surface charge; therefore, these two types of proteins are prone to attractive electrostatic interactions. The potato proteins therefore induced aggregation of the fungal proteins to form a non-fluid gel structure.

[0328] In contrast, when the same procedure was repeated using a commercially available chickpea protein with a pI of 4.5, no gelation was observed and the chickpea protein-enriched dough milk remained in a substantially fluid liquid form. The inventors hypothesize that this result is due to the small difference between the pI of the chickpea protein (4.5) and the pH of the liquid dispersion (4.2); as a result of this small difference, the surface charge on the chickpea protein was much smaller than that on the potato protein, and therefore the chickpea protein had a much lower affinity for electrostatic interactions with the fungal proteins and could not therefore induce them to aggregate to form a gel structure.

[0329] [Example 12] Production of tofu-like products by aggregation of fungal proteins in liquid dispersions Water was added to the soaked dough and biomat pieces as described in Example 7 to achieve mixtures with the fungal biomass concentrations shown in Table 16 below; the milks were prepared by placing the water and biomass in a Vitamix blender, blending at the maximum setting for 1 minute, and then homogenizing for 2 minutes using a high-speed (10,000 rpm) homogenizer. Each homogenized mixture was transferred to a Thermomix multicooker and heat-processed at 175°F and speed 3.0 for 30 minutes. The pH of each milk was either left unadjusted at its respective starting pH (5.1-5.8 for BioMat milks, 5.5-6.2 for dough milks) or adjusted to pH 3.5 by dropwise addition of 1 M hydrochloric acid, after which a divalent cation salt (either calcium chloride, CaCl2, or a mixture of about 90% by weight magnesium chloride, MgCl2, and about 10% by weight calcium chloride; the latter of these compositions will be referred to simply as magnesium chloride salt in this and the following examples) was added to some of the milks, as shown in Table 16. After pH adjustment (if necessary) and salt addition (if necessary), each milk was heat treated in a Thermomix multicooker at 175°F and speed 1.0 for 30 seconds to induce aggregation of the fungal proteins and production of fungal curd.

[0330] Each of the resulting curd compositions was poured into a 16 cm x 16 cm x 10.5 cm (6.3 in x 6.3 in x 4.13 in) tofu mold lined with cotton cloth. The cloth was folded once over the curd, and a 5 kg weight was placed on the mold cover for 30 minutes to pressurize each curd composition into a tofu-like product. Each tofu-like product was removed from the mold, crushed by hand, transferred to a glass beaker, and pasteurized at 98°C for 5 minutes (sufficient for the internal temperature of each sample to reach 72°C or higher for at least 15 seconds, in accordance with U.S. Food and Drug Administration pasteurization guidelines). The pasteurized tofu-like products were then transferred back into the mold and pressed with the 5 kg weight for an additional 15 minutes, after which each product was sealed in a vacuum sealer bag and refrigerated for 48 hours.

[0331] It was observed that when BioMat Milk was used, a curd could not be formed without the addition of salt at the unadjusted (starting) pH, regardless of the milk's solids content, but when the pH was lowered to 3.5, a curd could be formed without the addition of salt. When steep dough milk was used, a curd could not be formed at either pH range at 6.25% solids without the addition of salt, but when the pH was adjusted to 3.5, a curd could be formed at 10% solids without the addition of salt. It was also noted that steep dough milk with 10% solids content, without added salt or pH adjustment (Sample ID9 in Table 16 below), formed a cohesive mass when heated; however, as explained in the Examples below, the inventors conclude that this is likely simply a result of dehydrating the "sticky" steep dough milk and not due to true aggregation of fungal proteins.

[0332] [Table 16]

[0333] [Example 13] Protein and moisture content of tofu-like products Two grams of each tofu-like product produced in Example 12 was dried on an aluminum pan in an oven at 110°C for 24 hours. The difference between the mass of the original sample (2 grams) and the mass of the dried sample was determined for each product; this difference is assumed to be the total moisture content of the original sample and is expressed as a percentage of the original sample mass in Figures 20A (soaked dough, Sample IDs 1-18) and 20B (biomat pieces, Sample IDs 19-36); in each of these figures, the results for each sample are presented in numerical order of Sample ID from left to right, except that the right-most bar in each figure represents the moisture content of a commercially available, extra-firm tofu product obtained at a grocery store, which was measured for comparison.

[0334] Separately, samples of each tofu-like product produced in Example 7 were freeze-dried and ground using a coffee grinder, and the protein content (on a dry weight basis) of each ground freeze-dried sample was determined by nitrogen combustion on a LECO analyzer according to American Association of Cereal Chemists (AACC) Method 46-30.01, using an assumed nitrogen-to-protein conversion factor of 6.25. These results are illustrated in Figures 21A (soaked dough, Sample IDs 1-18) and 21B (biomat pieces, Sample IDs 19-36); in each of these figures, the results for each sample are presented in numerical order of Sample ID from left to right, except that the right-most bar in each figure represents the protein content of a commercially available, extra-firm tofu product obtained at a grocery store, which was measured for comparison.

[0335] [Example 14] Texture profile analysis of tofu-like products A texture profile analysis (TPA) was performed on samples of each tofu analog product produced in Example 12 using a TA.XTPlus texture analyzer. Specifically, a 30 mm cylinder of each product was cut using a circular stainless steel cookie cutter and compressed to 75% of the sample thickness using a cylindrical (50 mm diameter) stainless steel probe attached to a 2 kg load cell; a pre-test speed of 1 mm / s, a test speed of 2 mm / s, and a post-test speed of 2 mm / s were used, with a trigger force of 5.0 g and a rest period of 5 seconds between cycles. All samples were removed from refrigeration and allowed to rest at room temperature for 10 minutes before TPA testing.

[0336] Hardness, cohesion, and adhesive strength data were collected for each TPA test. Hardness data are shown in Figures 22A (soaked dough, Sample IDs 1–18) and 22B (biomat pieces, Sample IDs 19–36); in each of these figures, the results for each sample are presented in numerical order by sample ID from left to right, except that the rightmost bar in each figure represents the hardness of a commercially available, extra-firm tofu product obtained at a grocery store, measured for comparison. Adhesion data are shown in Figures 23A (soaked dough, Sample IDs 1–18) and 23B (biomat pieces, Sample IDs 19–36); in each of these figures, the results for each sample are presented in numerical order by sample ID from left to right, except that the rightmost bar in each figure represents the cohesion of a commercially available, extra-firm tofu product obtained at a grocery store, measured for comparison. The cohesive strength data are shown graphically in Figures 24A (soaked dough, Sample IDs 1-18) and 24B (biomat pieces, Sample IDs 19-36); in each of these figures, the results for each sample are presented in numerical order of the Sample ID from left to right, except that the right-most bar in each figure represents the cohesive strength of a commercially available, very hard tofu product obtained at a grocery store, which was measured for comparison.

[0337] Figures 22A-24B illustrate some trends regarding the effects of pH and salt concentration. At higher (unadjusted) pH values, increasing salt concentration resulted in increased hardness, regardless of biomass type, solids content, or salt type. Without wishing to be bound by any particular theory, the inventors hypothesize that a combination of magnesium or calcium cations, which act to weaken the repulsive forces between negatively charged fungal protein molecules, and the formation of salt bridges and / or water ionic bonds played a role in enhancing ionic bonds in the fungal curd compositions. At pH 3.5, the opposite phenomenon was observed, where sample hardness decreased with increasing salt concentration. Without wishing to be bound by any particular theory, the inventors hypothesize that because salt was added after the pH adjustment, when aggregation may have already occurred, the salt molecules exerted a shielding effect on the protein surface charge, reducing the rate of fungal protein aggregation.

[0338] Figures 22A-24B further illustrate some trends regarding the effects of salt type and dry solids content. For samples prepared using soaked dough, magnesium chloride produced harder fungal curds than calcium chloride at lower solids contents (6.25 wt%), regardless of pH, while the opposite trend was observed at higher solids contents (10 wt%). Samples prepared using biomat pieces exhibited a different pattern: magnesium chloride produced harder fungal curds than calcium chloride at pH 3.5, but calcium chloride produced harder fungal curds than magnesium chloride at higher (unadjusted) pHs. Without wishing to be bound by any particular theory, the inventors hypothesize that these differences are due to differences in surface chemistry and affinity for different salts between the two mycelial biomass forms.

[0339] Additionally, Figures 22A-24B illustrate that, in general, the harder fungal cards were also less cohesive, indicating greater stiffness and less structural flexibility upon applied deformation. Samples prepared using soaked dough generally had lower hardness, but higher cohesive strength and much higher adhesive strength than samples prepared using biomat pieces; without wishing to be bound by any particular theory, the inventors hypothesize that these differences are due to differences in the water-binding properties of fungal proteins between the two mycelial biomass forms.

[0340] Additionally, our conclusion that the cohesive mass formed upon heating steep dough milk with 10% solids content without salt addition or pH adjustment (Sample ID 9) was simply a dehydrated mass of "sticky" steep dough material and not a true aggregated protein composition is supported by the observation that Sample 9 had the lowest firmness (Figure 22A, "as is" pH, 10% DS, 5th bar) of all samples. Without being bound by any particular theory, we hypothesize that the lack of firmness in this sample indicates that while a cohesive mass was able to form, aggregation of fungal proteins did not occur.

[0341] [Example 15] Microstructure of tofu-like products Freeze-dried samples of some of the tofu-like products produced in Example 12 were platinum-coated and imaged by scanning electron microscopy (JEOL Ltd., Tokyo, Japan) using an accelerating voltage of 5 kV. Surface and cross-sectional images of each of Samples ID1-36 are shown in Figures 25A-60B; the images are presented in numerical order of the sample ID (i.e., images of Sample ID1 are shown in Figures 25A and 25B, images of Sample ID2 are shown in Figures 26A and 26B, etc.), while in each pair of figures, "A" is an image of the surface of the sample and "B" is an image of the cross-section of the sample.

[0342] Figures 25A-60B reveal several trends in the microstructure of the tofu-like products produced. For samples prepared using liquid dispersions containing 6.25 wt.% soaked dough solids, increasing the salt concentration resulted in the formation of a layered, rod-like structure with higher surface porosity (regardless of pH or salt type), and magnesium chloride provided a more uniform, less coarse microstructure than calcium chloride. For samples prepared using liquid dispersions containing 10 wt.% soaked dough solids, increasing the salt concentration resulted in the formation of a denser, more uniform microstructure (regardless of pH or salt type); the microstructure also tended to be denser and more uniform with calcium chloride, while magnesium chloride produced irregular aggregate particles with a rougher surface. For samples prepared using liquid dispersions containing 6.25 wt% biomat piece solids, increasing the salt concentration (regardless of pH or salt type) resulted in the formation of layered, rod-like structures with smoother surfaces, and magnesium chloride provided a more uniform, less rough microstructure than calcium chloride. For samples prepared using liquid dispersions containing 8.33 wt% biomat piece solids, increasing the salt concentration (regardless of pH or salt type) resulted in the formation of a denser, more uniform microstructure with lower surface porosity; magnesium chloride tended to produce a denser, more uniform layered microstructure, while calcium chloride tended to produce irregular, aggregate particles with a more porous surface.

[0343] Additionally, our conclusion that the cohesive mass formed by heating steeped dough milk with 10% solids content without salt addition or pH adjustment (sample ID 9) is simply a dehydrated mass of "sticky" steeped dough material and not a true cohesive protein composition is supported by comparing the SEM cross-section of this material (FIG. 33B) with SEM cross-sections of curds formed by pH adjustment and / or salt addition of the same milk composition (samples IDs 5-8 and 14-18; FIGS. 29B, 30B, 31B, 32B, 38B, 39B, 40B, 41B, and 42B). Notably, FIG. 33B shows a granular structure, while the other figures each show a more layered structure. Without being bound by any particular theory, the inventors hypothesize that the layered structures shown in Figures 29B, 30B, 31B, 32B, 38B, 39B, 40B, 41B, and 42B result from aggregation of the filamentous fungal biomass, which, due to its fine filamentous structure, tends to form layered structures upon aggregation, at least for the particle sizes of the materials used in Examples 12-15.

[0344] The concepts illustratively disclosed herein may be suitably practiced in the absence of any element not specifically disclosed herein. However, it will be apparent to those skilled in the art that many changes, variations, modifications, other uses, and applications of the present disclosure are possible, and that changes, variations, modifications, other uses, and applications that do not depart from the spirit and scope of the present disclosure are deemed to be encompassed by the present disclosure.

[0345] The foregoing discussion has been presented for purposes of illustration and description. The foregoing is not intended to limit the disclosure to the form or forms disclosed herein. In the foregoing Detailed Description, for example, various features are grouped together in one or more embodiments for the purpose of streamlining the disclosure. Features of the embodiments may be combined in alternative embodiments other than the embodiments discussed above. This method of disclosure is not to be interpreted as reflecting an intention that the claims require more features than are expressly recited in each claim. Rather, as reflected in the following claims, inventive aspects lie in less than all features of a single foregoing disclosed embodiment. Thus, the following claims are hereby incorporated into this Detailed Description, with each claim standing on its own as a separate embodiment.

[0346] Furthermore, while the present disclosure includes a description of one or more embodiments and certain variations and modifications, other variations, combinations, and modifications are within the scope of the present disclosure, as would be within the skill and knowledge of one of ordinary skill in the art after understanding the present disclosure. The right to include alternative embodiments to the extent permitted, including alternative, interchangeable, and / or equivalent structures, functions, ranges, or steps to those claimed, regardless of whether such alternative, interchangeable, and / or equivalent structures, functions, ranges, or steps are disclosed herein, and without the intention of offering any patentable subject matter to the public, is intended. [Explanation of symbols]

[0347] 100 ways 110 Liquid Dispersion Preparation 120 Component addition 130 Lipid addition 140 Acid addition 150 heating 200 ways 210 Liquid Dispersion Preparation 220 Component addition 230 Lipid addition 240 Homogenization 250 Microbial culture addition 300 ways 310 Liquid Dispersion Preparation 320 Addition of functional ingredients 330 Lipid addition 340 Acidification 350 heating 400 ways 410 Liquid Dispersion Preparation 420 Salt added 430 Lipid addition 440 Acidification 450 heating 500 ways 510 Liquid Dispersion Preparation 520 Component addition 530 Lipid addition 540 Acidification and / or Salting 550 heating 600 ways 610 Liquid Dispersion Preparation 620 Acidification and / or Salting 630 Heating 700 methods 710 Fungal curd formation 720 Fungal Card Separation 730 First Pressurization 740 Crush 750 Pasteurization 760 Second Pressurization

Claims

1. 1. A method for making a solid and / or colloidal fungal food material, comprising: Inducing aggregation of fungal proteins in a liquid dispersion of filamentous fungal particles A method comprising:

2. the inducing step comprising: (i) adjusting the pH of the liquid dispersion; (ii) adding one or more functional ingredients to the liquid dispersion; and (iii) adding one or more salts to the liquid dispersion; The method of claim 1 , comprising at least one of:

3. The method of claim 2 , wherein the inducing step comprises: (i).

4. The method of claim 2 , wherein the inducing step comprises (ii).

5. The method of claim 2 , wherein the inducing step comprises (iii).

6. The method of claim 2 , wherein the inducing step comprises (i) and (ii).

7. 3. The method of claim 2, wherein the inducing step comprises (i) and (iii).

8. 3. The method of claim 2, wherein the inducing step comprises (ii) and (iii).

9. The method of claim 2 , wherein the inducing step includes (i), (ii), and (iii).

10. 10. The method of claim 1, wherein the liquid dispersion of filamentous fungal particles comprises an oil and / or a solid fat.

11. 11. The method of claim 10, comprising, prior to the inducing step, combining a liquid phase, the filamentous fungal particles, and the oil and / or solid fat to form the liquid dispersion.

12. 12. The method of claim 11, wherein the combining step comprises blending the liquid phase and the filamentous fungus particles with the oil and / or solid fat.

13. The method of claim 12 wherein the blending comprises high shear.

14. 14. The method of claim 13, wherein the high speed shearing comprises shearing the liquid phase, the filamentous fungus particles, and the oil and / or solid fat at a rotational speed of at least about 10,000 rpm for at least about 2 minutes.

15. 15. The method of any one of claims 11 to 14, wherein the combining step comprises adding an emulsifier.

16. 16. The method of claim 15, wherein the emulsifier is selected from the group consisting of carboxymethylcellulose, carrageenan, cellulose, guar gum, lecithin, mono- and diglycerides of fatty acids, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, polysorbates, stearoyl lactylate, sorbitan esters, sucrose esters, sucroglycerides, xanthan gum, and combinations thereof.

17. 17. The method of any one of claims 10 to 16, wherein the oil and / or solid fat comprises an oil selected from the group consisting of acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof.

18. 18. The method of any one of claims 10 to 17, wherein the oil and / or solid fat comprises a solid fat selected from the group consisting of marine tallow, butter, chicken fat, clarified butter, cocoa butter, dripping, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof.

19. 19. The method of any one of claims 10 to 18, wherein the oil content of the liquid dispersion is from about 1% to about 5% by weight.

20. 10. The method of any one of claims 2, 4, 6, 8, and 9, wherein the one or more functional ingredients comprise a non-fungal protein.

21. 21. The method of claim 20, wherein the non-fungal protein is selected from the group consisting of bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof.

22. 10. The method of any one of claims 2, 4, 6, 8, and 9, wherein the one or more functional ingredients comprise one or more enzymes.

23. 23. The method of claim 22, wherein the one or more enzymes are selected from the group consisting of catalase, chymosin, lactase, lipase, transglutaminase, and combinations thereof.

24. 10. The method of any one of claims 2, 3, 6, 7, and 9, wherein the inducing step lowers the pH of the liquid dispersion.

25. 25. The method of claim 24, wherein the pH of the liquid dispersion is lowered by adding an acid to the liquid dispersion.

26. 26. The method of claim 25, wherein the acid is selected from the group consisting of sorbic acid, benzoic acid, formic acid, acetic acid, dehydroacetic acid, lactic acid, propionic acid, boric acid, malic acid, fumaric acid, ascorbic acid, erythorbic acid, citric acid, tartaric acid, phosphoric acid, metatartaric acid, adipic acid, succinic acid, thiodipropionic acid, phytic acid, alginic acid, hydrochloric acid, sulfuric acid, gluconic acid, glutamic acid, guanylic acid, inosinic acid, cyclamic acid, cholic acid, and combinations thereof.

27. 25. The method of claim 24, wherein the pH of the liquid dispersion is lowered by adding an acidifying microbial culture to the liquid dispersion.

28. 28. The method of any one of claims 24 to 27, wherein the inducing step further comprises heating the liquid dispersion.

29. 30. The method of claim 28, wherein the liquid dispersion is heated to a temperature of about 150°F to about 180°F (about 65.5°C to about 83°C).

30. 30. The method of claim 29, further comprising further heating the liquid dispersion to a temperature of about 180°F to about 200°F (about 83°C to about 94°C) after the inducing step.

31. 31. The method of any one of claims 1 to 30, wherein the liquid dispersion comprises at least one salt of calcium or magnesium.

32. 10. The method of any one of claims 2, 5, and 7-9, wherein the one or more salts comprise at least one salt of calcium or magnesium.

33. The at least one salt of calcium or magnesium is selected from the group consisting of calcium carbonate, calcium sorbate, calcium benzoate, calcium sulfite, calcium bisulfite, calcium formate, calcium acetate, calcium propionate, calcium ascorbate, calcium lactate, monocalcium citrate, dicalcium citrate, tricalcium citrate, monocalcium phosphate, dicalcium phosphate, tricalcium phosphate, calcium malate, calcium hydrogen malate, calcium tartrate, calcium fumarate, calcium glyceryl phosphate, disodium calcium ethylenediaminetetraacetate, calcium lactobionate, calcium alginate, dicalcium diphosphate, dihydrogen calcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, fumarate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, calcium phosphate ... phosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate, calcium phosphate 33. The method of claim 31 or 32, wherein the anti-inflammatory agent is selected from the group consisting of calcium ferrocyanide, dicalcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium silicate, calcium aluminosilicate, calcium stearate, calcium gluconate, synthetic calcium aluminate, calcium diglutamate, calcium guanylate, calcium inosinate, calcium 5'-ribonucleotide, calcium iodate, calcium bromate, calcium peroxide, calcium cyclamate, calcium saccharinate, magnesium lactate, monomagnesium phosphate, dimagnesium phosphate, magnesium citrate, magnesium salts of fatty acids, magnesium carbonate, magnesium bicarbonate, magnesium chloride, magnesium sulfate, magnesium oxide, magnesium silicate, magnesium trisilicate, magnesium stearate, magnesium gluconate, magnesium diglutamate, and combinations thereof.

34. 34. The method of any one of claims 1 to 33, wherein the liquid dispersion further comprises at least one of a flavoring agent, a taste modifier, and a plant masking agent.

35. 35. The method of any one of claims 1 to 34, wherein at least a portion of the filamentous fungal particles are produced by size reduction of a cohesive filamentous fungal mycelium biomass.

36. 36. The method of claim 35, wherein the cohesive filamentous fungal mycelial biomass is produced by liquid surface fermentation or solid state fermentation.

37. 37. The method of any one of claims 1 to 36, wherein at least a portion of the filamentous fungal particles are produced by submerged fermentation.

38. 38. The method of any one of claims 1 to 37, wherein the filamentous fungus particles are in the form of a powder having a particle size of about 30 μm to about 400 μm.

39. 39. The method of any one of claims 1 to 38, wherein the filamentous fungal particles consist essentially of fungal mycelium.

40. 40. The method of any one of claims 1 to 39, wherein the filamentous fungal particles comprise at least about 50% by weight fungal mycelium.

41. 41. The method of claim 40, wherein the filamentous fungal particles comprise at least about 75% by weight fungal mycelium.

42. 42. The method of claim 41, wherein the filamentous fungal particles comprise at least about 95% by weight fungal mycelium.

43. 43. The method of any one of claims 1 to 42, wherein the solids content of the liquid dispersion is from about 4% to about 7% by weight.

44. 44. The method of any one of claims 1 to 43, wherein the weight ratio of filamentous fungal particles to liquid in the liquid dispersion is from about 1:10 to about 10:

1.

45. 45. The method of any one of claims 1 to 44, wherein the liquid dispersion is stable at room temperature for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

46. 46. ​​The method of any one of claims 1 to 45, wherein the liquid dispersion is stable at refrigerated temperatures for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

47. 47. The method of any one of claims 1 to 46, wherein the liquid dispersion is a mixed-type mycelial biomass composition comprising a first mycelial biomass type and a second mycelial biomass type, wherein the first mycelial biomass type and the second mycelial biomass type are different from each other.

48. 48. The method of claim 47, wherein the first mycelial biomass form is a clumped mycelial biomass form and the second mycelial biomass form is a submerged mycelial biomass form.

49. 49. The method of claim 48, wherein the first mycelial biomass form is selected from the group consisting of biomat pieces, biomat flour, biomat dispersion, and spray-dried biomat flour.

50. 50. The method of claim 48 or 49, wherein the second mycelial biomass format is selected from the group consisting of a steeped slurry, a steeped dough, a steeped powder, a steeped dispersion, and a steeped spray-dried powder.

51. 48. The method of claim 47, wherein the first and second mycelial biomass forms are each a submerged mycelial biomass form.

52. 52. The method of claim 51, wherein the first and second mycelial biomass forms are each selected from the group consisting of a soaked liquid biomass, a soaked dough, a soaked powder, a soaked dispersion, and a soaked spray-dried powder.

53. the liquid dispersion is a combined liquid dispersion, and prior to the inducing step: blending the mixture of the first mycelial biomass form and a first liquid to form a first liquid dispersion; blending the mixture of the second mycelial biomass form and a second liquid to form a second liquid dispersion; combining the first liquid dispersion and the second liquid dispersion to form the combined liquid dispersion; 53. The method of any one of claims 47 to 52, further comprising:

54. 54. The method of any one of claims 1 to 53, wherein a gel is formed in the inducing step.

55. 55. The method of claim 54, wherein the inducing step comprises adjusting the pH of the liquid dispersion to a gelling pH of about 4 or less.

56. 56. The method of claim 55, wherein the gelling pH is about 3.

5.

57. a fungal curd is formed in the inducing step; Separating at least a portion of the liquid phase of the liquid dispersion from the fungal curd.

57. The method of any one of claims 1 to 56, further comprising:

58. 58. The method of claim 57, wherein in the separating step, at least a portion of the liquid phase is at least about 90% by weight of the liquid phase.

59. 59. The method of claim 57 or 58, wherein the inducing step comprises adjusting the pH of the liquid dispersion to a pH of from about 2 to about 4.

60. 60. The method of claim 59, wherein in the inducing step, the pH is adjusted to a pH of about 3.

5.

61. 61. The method of any one of claims 57 to 60, wherein the separating step comprises pressing the fungal curd through a mesh filter.

62. 62. The method of claim 61, wherein the mesh filter comprises a fabric.

63. 63. The method of claim 62, wherein the fabric is cheesecloth.

64. 63. The method of claim 61 or 62, wherein the mesh filter comprises a fine mesh sieve.

65. 65. The method of any one of claims 61 to 64, further comprising forming the fungal card into a block.

66. A food material comprising aggregated filamentous fungal mycelial biomass.

67. 67. The food material of claim 66, further comprising an oil and / or a solid fat.

68. 68. The food material of claim 67, wherein the oil and / or solid fat comprises an oil selected from the group consisting of acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof.

69. 69. The food material of claim 67 or 68, wherein the oil and / or solid fat comprises a solid fat selected from the group consisting of marine tallow, butter, chicken fat, clarified butter, cocoa butter, dripping, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof.

70. 70. The food material of any one of claims 66 to 69, further comprising a non-fungal protein.

71. 71. The food material of claim 70, wherein the non-fungal protein is selected from the group consisting of bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof.

72. 72. The food material of any one of claims 66 to 71, wherein at least a portion of the filamentous fungal particles are produced by size reduction of a cohesive filamentous fungal mycelium biomass.

73. 73. The food material of claim 72, wherein the cohesive filamentous fungal mycelial biomass is produced by liquid surface fermentation or solid-state fermentation.

74. 74. The food material of any one of claims 66 to 73, wherein at least a portion of the filamentous fungal particles are produced by submerged fermentation.

75. 75. The food material of any one of claims 66 to 74, wherein the filamentous fungal particles consist essentially of fungal mycelium.

76. 76. The food material of any one of claims 66 to 75, wherein the filamentous fungal particles comprise at least about 50% by weight fungal mycelium.

77. 77. The food material of claim 76, wherein the filamentous fungal particles comprise at least about 75% by weight fungal mycelium.

78. 78. The food material of claim 77, wherein the filamentous fungal particles comprise at least about 95% by weight fungal mycelium.

79. 79. The food material of any one of claims 66 to 78, wherein the fungal curd is in the form of a block.

80. 80. The food material of any one of claims 66 to 79, which does not contain a non-fungal gelling agent.

81. 81. The food material of any one of claims 66 to 80, consisting essentially of said aggregated filamentous fungal mycelial biomass.

82. 82. The food material of any one of claims 66 to 81, consisting of said filamentous fungal mycelium biomass and at least one acid or base.

83. 83. The food material of any one of claims 66 to 82, consisting of said filamentous fungal mycelium biomass and at least one functional ingredient.

84. 84. The food material of any one of claims 66 to 83, consisting of said filamentous fungal mycelium biomass and at least one salt.

85. 81. The food material of any one of claims 66 to 80, further comprising a microbial food culture.

86. 86. The food material according to any one of claims 66 to 85, having a hardness of from about 1 N to about 50 N.

87. 87. The food material of any one of claims 66 to 86, having an adhesive strength of from about 0.001 N mm to about 60 N mm.

88. 88. The food material of any one of claims 66 to 87, having a cohesive strength of from about 0.001 to about 4.

89. 89. The food material of any one of claims 66 to 88, wherein the food material is a fungal curd made by the method of any one of claims 57 to 65.

90. a first mycelial biomass form; and Second mycelial biomass form A mixed-type mycelial biomass composition comprising: A mixed-type mycelial biomass composition, wherein the first mycelial biomass type and the second mycelial biomass type are different mycelial biomass types.

91. 91. The mixed-form mycelial biomass composition of claim 90, which is a food material.

92. 92. The mixed-format mycelial biomass composition of claim 91, wherein the food material is selected from the group consisting of a flour, a plurality of solid particles other than flour, a liquid dispersion, an emulsion, a foam, a gel, a sol, and a solid foam.

93. 93. The mixed format mycelial biomass composition of claim 92, wherein the food material is a flour, the flour comprising filamentous fungal particles having a particle size of about 30 μm to about 400 μm.

94. 93. The mixed-format mycelial biomass composition of claim 92, wherein the food material is a plurality of solid particles other than flour, the plurality of solid particles comprising filamentous fungal particles having a particle length of about 0.05 mm to about 500 mm, a particle width of about 0.03 mm to about 7 mm, and a particle height of about 0.03 mm to about 1.0 mm.

95. 93. The mixed-type mycelial biomass composition of claim 92, wherein the food material is a liquid dispersion or sol, and the weight ratio of filamentous fungal particles to liquid in the liquid dispersion or sol is from about 1:10 to about 10:

1.

96. 96. The mixed format mycelial biomass composition of claim 92 or 95, wherein the food material is a liquid dispersion or sol, and the liquid dispersion or sol is stable for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

97. The food material has a foam stability of at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, or at least about 95% for at least about 1 day, at least about 2 days, at least about 3 days, at least about 4 days, at least about 5 days, at least about 6 days, at least about 7 days, at least about 8 days, at least about 9 days, at least about 10 days, at least about 11 days, at least about 12 days, at least about 13 days, at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 31 days, at least about 32 days, at least about 33 days, at least about 34 days, at least about 35 days, at least about 36 days, at least about 37 days, at least about 38 days, at least about 39 days, at least about 40 days, at least about 41 days, at least about 42 days, at least about 43 days, at least about 44 days, at least about 45 days, at least about 46 days, at least about 47 days, at least about 48 days, at least about 49 days, at least about 50 days, at least about 51 days, at least about 52 days, at least about 53 days, at least about 54 days, at least about 55 days, 93. The mixed-form mycelial biomass composition of claim 92, wherein the foam has a duration of at least about 14 days, at least about 15 days, at least about 16 days, at least about 17 days, at least about 18 days, at least about 19 days, at least about 20 days, at least about 21 days, at least about 22 days, at least about 23 days, at least about 24 days, at least about 25 days, at least about 26 days, at least about 27 days, at least about 28 days, at least about 29 days, at least about 30 days, at least about 1 month, at least about 2 months, or at least about 3 months.

98. 98. A food product comprising the food ingredient of any one of claims 91 to 97.

99. 98. The mixed-type mycelial biomass composition of any one of claims 91 to 97, wherein the first mycelial biomass type is an aggregated mycelial biomass type and the second mycelial biomass type is an imbibed mycelial biomass type.

100. 100. The mixed format mycelial biomass composition of claim 99, wherein said first mycelial biomass format is selected from the group consisting of biomat pieces, biomat flour, biomat dispersion, and spray-dried biomat flour.

101. 101. The mixed format mycelial biomass composition of claim 99 or 100, wherein the second mycelial biomass format is selected from the group consisting of a soaked slurry, a soaked dough, a soaked powder, a soaked dispersion, and a soaked spray-dried powder.

102. 102. The mixed-type mycelial biomass composition of any one of claims 99 to 101, which is a food material.

103. 103. The mixed-type mycelial biomass composition of claim 102, which is a gel.

104. 104. The mixed-form mycelial biomass composition of claim 103, which is a food product selected from the group consisting of a blancmange-like food product, a butter-like food product, a custard-like food product, a jam-like food product, a jelly-like food product, a margarine-like food product, and a yogurt-like food product.

105. 105. The mixed-type mycelial biomass composition of any one of claims 99 to 104, wherein the mass ratio of the first mycelial biomass type to the second mycelial biomass type is from about 1:10 to about 10:

1.

106. 109. The mixed-type mycelial biomass composition of any one of claims 90 to 97 or claims 99 to 108, wherein the first mycelial biomass type and the second mycelial biomass type are each a submerged mycelial biomass type.

107. 107. The mixed-form mycelial biomass composition of claim 106, wherein the first mycelial biomass form and the second mycelial biomass form are each selected from the group consisting of a soaked slurry, a soaked dough, a soaked powder, a soaked dispersion, and a soaked spray-dried powder.

108. 108. The mixed-type mycelial biomass composition of claim 106 or 107, which is a food material.

109. 109. The mixed-type mycelial biomass composition of claim 108, which is a gel.

110. 110. The mixed-form mycelial biomass composition of claim 109, which is a food product selected from the group consisting of a blancmange-like food product, a butter-like food product, a custard-like food product, a jam-like food product, a jelly-like food product, a margarine-like food product, and a yogurt-like food product.

111. 111. The mixed-type mycelial biomass composition of any one of claims 106 to 110, wherein the mass ratio of the first mycelial biomass type to the second mycelial biomass type is from about 1:10 to about 10:

1.

112. (i) adjusting the pH of the liquid dispersion; (ii) adding one or more functional ingredients to the liquid dispersion; and (iii) adding one or more salts to the liquid dispersion 1. A method for producing a fungal gel, comprising at least one of: The method, wherein the liquid dispersion is a mixed-type mycelial biomass composition comprising a first mycelial biomass type and a second mycelial biomass type, and the first mycelial biomass type and the second mycelial biomass type are different mycelial biomass types.

113. 113. The method of claim 112, wherein the fungal gel is a food product.

114. 114. The method of claim 113, wherein the food product is selected from the group consisting of a blancmange like food product, a butter like food product, a custard like food product, a jam like food product, a jelly like food product, a margarine like food product, and a yogurt like food product.

115. 115. The method of any one of claims 112 to 114, wherein the mass ratio of the first mycelial biomass form to the second mycelial biomass form is from about 1:10 to about 10:

1.

116. 116. The method of any one of claims 112 to 115, wherein the first mycelial biomass format is a cohesive mycelial biomass format selected from the group consisting of biomat pieces, biomat powder, biomat dispersion, and spray-dried biomat powder, and the second mycelial biomass format is a soaked mycelial biomass format selected from the group consisting of soaked slurry, soaked dough, soaked powder, soaked dispersion, and soaked spray-dried powder.

117. 117. The method of claim 116, wherein the first mycelial biomass form is selected from the group consisting of biomat pieces, biomat flour, and spray-dried biomat flour, and the second mycelial biomass form is selected from the group consisting of soaked dough and soaked flour.

118. 118. The method of any one of claims 112 to 117, wherein the first and second mycelial biomass forms are each a soaked mycelial biomass form selected from the group consisting of a soaked paste, a soaked powder, a soaked liquid dispersion, and a soaked spray-dried powder.

119. 119. The method of claim 118, wherein the first and second mycelial biomass forms are each selected from the group consisting of soaked dough, soaked flour, and soaked spray-dried flour.

120. The mixed type mycelial biomass composition comprises: blending the mixture of the first mycelial biomass form and a first liquid to form a first liquid dispersion; blending the mixture of the second mycelial biomass form and a second liquid to form a second liquid dispersion; combining the first liquid dispersion and the second liquid dispersion to form the mixed mycelial biomass composition; 120. The method of claim 117 or 119, wherein the compound is produced by a process comprising:

121. 121. The method of any one of claims 112 to 120, wherein the inducing step comprises (i), and wherein in the inducing step, the pH of the liquid dispersion is adjusted to a gelling pH of about 4 or less.

122. 122. The method of claim 121, wherein the gelling pH is about 3.

5.

123. 1. A method for making a fungal tofu analog food product, comprising: Inducing aggregation of fungal proteins in a liquid dispersion of filamentous fungal particles to form a fungal curd, the inducing step comprising: (i) adjusting the pH of the liquid dispersion; (ii) adding one or more functional ingredients to the liquid dispersion; and (iii) adding one or more salts to the liquid dispersion; and compressing the fungal curd to form a fungal tofu analogue food product; A method comprising:

124. 124. The method of claim 123, further comprising separating the fungal curd from the liquid phase of the liquid dispersion after the inducing step.

125. 125. The method of claim 123 or 124, wherein the liquid dispersion comprises an oil and / or a solid fat.

126. 126. The method of claim 125, further comprising the step of combining a liquid phase, the filamentous fungus particles, and the oil and / or solid fat to form the liquid dispersion prior to the inducing step.

127. 127. The method of claim 126, wherein the combining step comprises blending the liquid phase and the filamentous fungus particles with the oil and / or solid fat.

128. 128. The method of claim 127, wherein the blending comprises high shear.

129. 129. The method of claim 128, wherein the high speed shearing comprises shearing the liquid phase, the filamentous fungus particles, and the oil and / or solid fat at a rotational speed of at least about 10,000 rpm for at least about 2 minutes.

130. 130. The method of any one of claims 126 to 129, wherein the combining step comprises adding an emulsifier.

131. 131. The method of claim 130, wherein the emulsifier is selected from the group consisting of carboxymethylcellulose, carrageenan, cellulose, guar gum, lecithin, mono- and diglycerides of fatty acids, polyglycerol esters of fatty acids, polyglycerol polyricinoleate, polysorbates, stearoyl lactylate, sorbitan esters, sucrose esters, sucroglycerides, xanthan gum, and combinations thereof.

132. 132. The method of any one of claims 125 to 131, wherein the oil and / or solid fat comprises an oil selected from the group consisting of acai oil, almond oil, avocado oil, blackcurrant seed oil, borage seed oil, canola oil, cashew oil, coconut oil, corn oil, cottonseed oil, evening primrose oil, grapeseed oil, hazelnut oil, hemp oil, macadamia oil, olive oil, palm oil, peanut oil, pecan oil, pine seed oil, pistachio oil, pumpkin seed oil, safflower oil, sesame oil, soybean oil, sunflower oil, tea oil, walnut oil, and combinations thereof.

133. 133. The method of any one of claims 125 to 132, wherein the oil and / or solid fat comprises a solid fat selected from the group consisting of marine tallow, butter, chicken fat, clarified butter, cocoa butter, dripping, duck fat, back fat, lard, mango butter, margarine, schmaltz, shea butter, speck, suet, tail fat, tallow, vegetable shortening, and combinations thereof.

134. 134. The method of any one of claims 125 to 133, wherein the oil content of the liquid dispersion is from about 1% to about 5% by weight.

135. 135. The method of any one of claims 123 to 134, wherein the inducing step comprises (ii) and the one or more functional ingredients comprise a non-fungal protein.

136. 136. The method of claim 135, wherein the non-fungal protein is selected from the group consisting of bean protein, broccoli protein, chickpea protein, hemp protein, lentil protein, nut protein, pea protein, potato protein, quinoa protein, rice protein, seaweed protein, seed protein, soy protein, spinach protein, and combinations thereof.

137. 137. The method of any one of claims 123 to 136, wherein the inducing step comprises (ii) and the one or more functional ingredients comprise one or more enzymes.

138. 138. The method of claim 137, wherein the one or more enzymes are selected from the group consisting of catalase, chymosin, lactase, lipase, transglutaminase, and combinations thereof.

139. 139. The method of any one of claims 123 to 138, wherein the inducing step comprises (i), and wherein the inducing step lowers the pH of the liquid dispersion.

140. 140. The method of claim 139, wherein the pH of the liquid dispersion is lowered by adding an acid to the liquid dispersion.

141. 141. The method of claim 140, wherein the acid is selected from the group consisting of sorbic acid, benzoic acid, formic acid, acetic acid, dehydroacetic acid, lactic acid, propionic acid, boric acid, malic acid, fumaric acid, ascorbic acid, erythorbic acid, citric acid, tartaric acid, phosphoric acid, metatartaric acid, adipic acid, succinic acid, thiodipropionic acid, phytic acid, alginic acid, hydrochloric acid, sulfuric acid, gluconic acid, glutamic acid, guanylic acid, inosinic acid, cyclamic acid, cholic acid, and combinations thereof.

142. 140. The method of claim 139, wherein the pH of the liquid dispersion is lowered by adding an acidifying microbial culture to the liquid dispersion.

143. 143. The method of any one of claims 139 to 142, wherein the inducing step further comprises heating the liquid dispersion.

144. 144. The method of claim 143, wherein the liquid dispersion is heated to a temperature of about 150°F to about 180°F (about 65.5°C to about 83°C).

145. 145. The method of claim 144, further comprising, after the inducing step, further heating the liquid dispersion to a temperature of about 180°F to about 200°F (about 83°C to about 94°C).

146. 146. The method of any one of claims 139 to 145, wherein in the inducing step, the pH is adjusted to a pH of from about 2 to about 4.

147. 147. The method of claim 146, wherein in the inducing step, the pH is adjusted to a pH of about 3.

5.

148. 148. The method of any one of claims 123 to 147, wherein the liquid dispersion comprises at least one salt of calcium or magnesium.

149. 149. The method of any one of claims 123 to 148, wherein the inducing step comprises (iii) and the one or more salts comprise at least one salt of calcium or magnesium.

150. The at least one salt of calcium or magnesium is selected from the group consisting of calcium carbonate, calcium sorbate, calcium benzoate, calcium sulfite, calcium bisulfite, calcium formate, calcium acetate, calcium propionate, calcium ascorbate, calcium lactate, monocalcium citrate, dicalcium citrate, tricalcium citrate, monocalcium phosphate, dicalcium phosphate, tricalcium phosphate, calcium malate, calcium hydrogen malate, calcium tartrate, calcium fumarate, calcium glyceryl phosphate, disodium calcium ethylenediaminetetraacetate, calcium lactobionate, calcium alginate, dicalcium diphosphate, dihydrogen calcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium salts of fatty acids, calcium stearoyl-2-lactylate, calcium stearoyl fumarate, calcium chloride, calcium sulfate, calcium oxide, phenanthate ...

150. The method of claim 148 or 149, wherein the anti-inflammatory agent is selected from the group consisting of calcium cyanide, dicalcium diphosphate, sodium calcium polyphosphate, calcium polyphosphate, calcium silicate, calcium aluminosilicate, calcium stearate, calcium gluconate, synthetic calcium aluminate, calcium diglutamate, calcium guanylate, calcium inosinate, calcium 5'-ribonucleotide, calcium iodate, calcium bromate, calcium peroxide, calcium cyclamate, calcium saccharinate, magnesium lactate, monomagnesium phosphate, dimagnesium phosphate, magnesium citrate, magnesium salts of fatty acids, magnesium carbonate, magnesium bicarbonate, magnesium chloride, magnesium sulfate, magnesium oxide, magnesium silicate, magnesium trisilicate, magnesium stearate, magnesium gluconate, magnesium diglutamate, and combinations thereof.

151. 151. The method of any one of claims 123 to 150, wherein the separating step comprises pressing the fungal curd through a mesh filter.

152. 152. The method of claim 151, wherein the mesh filter comprises a fabric.

153. 153. The method of claim 152, wherein the cloth is cheesecloth.

154. 153. The method of claim 151 or 152, wherein the mesh filter comprises a fine mesh sieve.

155. 155. The method of any one of claims 151 to 154, further comprising forming the fungal card into a block.

156. 156. The method of any one of claims 123 to 155, wherein the liquid dispersion further comprises at least one of a flavoring agent, a taste modifier, and a plant masking agent.

157. 157. The method of any one of claims 123 to 156, wherein at least a portion of the filamentous fungal particles are produced by size reduction of a cohesive filamentous fungal mycelium biomass.

158. 158. The method of claim 157, wherein the cohesive filamentous fungal mycelium biomass is produced by liquid surface fermentation or solid state fermentation.

159. 159. The method of any one of claims 123 to 158, wherein at least a portion of the filamentous fungal particles are produced by submerged fermentation.

160. 160. The method of any one of claims 123 to 159, wherein the filamentous fungus particles are in the form of a powder having a particle size of from about 30 μm to about 400 μm.

161. 161. The method of any one of claims 123 to 160, wherein the filamentous fungal particles consist essentially of fungal mycelium.

162. 162. The method of any one of claims 123 to 161, wherein the filamentous fungal particles comprise at least about 50% by weight fungal mycelium.

163. 163. The method of claim 162, wherein the filamentous fungal particles comprise at least about 75% by weight fungal mycelium.

164. 164. The method of claim 163, wherein the filamentous fungal particles comprise at least about 95% by weight fungal mycelium.

165. 165. The method of any one of claims 123 to 164, wherein the solids content of the liquid dispersion is from about 4% to about 7% by weight.

166. 166. A fungal tofu analog food product made by the method of any one of claims 123 to 165.

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