Fermented food product and methods of producing

Abstract

The present invention relates to the field of fermented food products, more specifically, to non-animal dairy substitutes. The application discloses a novel method of growing fungal inoculants on a vegetable feedstock. The obtained fermented food product comprises fungal biomass and fermented vegetable feedstock.

Description

[0001] FERMENTED FOOD PRODUCT AND METHODS OF PRODUCING

[0002] FIELD OF THE INVENTION

[0003] The present invention relates to the field of fermented food products, more specifically, to non-animal dairy substitutes. The application discloses a novel method of growing fungal inoculants on a vegetable feedstock. The obtained fermented food product comprises fungal biomass and fermented vegetable feedstock.

[0004] BACKGROUND

[0005] In view of animal wellbeing and the impact of livestock on our environmental footprint, more and more people adapt their eating habits towards a non-animal based diet. Although the field of plant-based and microbial-based meat and dairy alternatives is booming, many products have suboptimal taste, aromas and texture. While over the years the focus of the food industry was mainly on vegetable based meat alternatives, there is an increasing interest in developing non-animal dairy products, such as yogurt or cheese.

[0006] WO2022219170A1 discloses methods of growing edible fibrous mycelium mass and homogenizing the mycelium mass with water to obtain a non-animal dairy substitute product.

[0007] WO2020176758A1 describes methods of producing dairy analog food products wherein particles of a filamentous fungus are the source for at least about 50% of protein in the cultured dairy analog food product.

[0008] On the other hand, WO2021174184A1 discloses a method of preparing vegan dairy products like cheese. The food products are almost exclusively made of vegetable sources that are inoculated with microbes to produce the rind and some additional flavors. Along the same line several online recipes can be found describing cheeses such as camembert that are made of cashew nuts and that are inoculated with for example Penicillium camemberti to produce the rind and the camembert characteristic flavors. However, since P. camemberti is aerobic, it grows only at the outside of the cheese and thus during the production of these cheese analogs no significant microbial biomass is produced. Fermentation of plant-based substrates is known in the art for enhancing the digestibility of legumes or other plant based material. Asensio-Grau et al., 2020 disclosed the use of fermentation for enhancing digestibility of lentil and Tripathi et al., 2017 fermented soybean to produce tempeh. Soybean fermentation is also disclosed in KR20230091313. Zwinkels Jasper et al. 2023 relates to the fermentation of barley and rice into tempeh and koji. WO2014110540, EP3603407 and https : / / web. archive. org / web / 20220319013041 / https: / / fullofpla nts. com / vegan- aged-camembert-cheese disclose the production of cheese analogs from nuts and seeds. US2022232854 discloses a method for producing a meat substitute using fungal fermentation. None of these documents disclose a cheese or dairy analog that combines both an elevated protein level after fermentation and a substantial amount of fungal biomass. Increased protein content and significant fungal biomass are essential for achieving a balanced nutritional profile as well as the aroma and texture of a credible cheese alternative.

[0009] There remains a need for tasty dairy alternatives that resemble the aromas and texture of the animal based original. The present invention addresses this need by providing a dairy analog that combines the texture, taste and smell of both fungal biomass and fermented vegetable feedstocks.

[0010] SUMMARY OF THE INVENTION

[0011] The present invention addresses the aforementioned challenges by developing a novel method of microbial biomass fermentation using an edible plant-based feedstock. The protein content, texture, taste and smell of the final fermented product can be attributed to both the produced fungal biomass and the fermented feedstock.

[0012] In a first aspect, the application provides a fermented product comprising a protein content of at least 25 w% of the dry matter of the fermented product and comprising a fungal biomass and a fermented vegetable feedstock, characterized in that the fungal biomass forms at least 40 w% and the fermented vegetable feedstock at least 10 w% of the dry matter of the fermented product. In one embodiment, the ratio of the fungal biomass to fermented vegetable feedstock is between about 9: 1 to about 1 : 1.5. In another embodiment, the fermented product is derived from an unfermented vegetable feedstock and a fungal inoculant. In yet another embodiment, the w% protein by dry matter of the fermented product is higher compared to the w% protein by dry matter of the unfermented vegetable feedstock.

[0013] In a particular embodiment, the unfermented vegetable feedstock comprises or consists of cauliflower.

[0014] In another embodiment, the fungal biomass comprises a fungal strain of the Penicillium genus, more particularly the fungal strain is selected from the list consisting of P. camemberti, P. candidum, P. roqueforti, P. glaucum, P. nalgiovense, P. caseifulvum, P. album, P. solitum, P. commune, P. carneum, P. carymbiferum, P. crustosum and P. nordicum. In another embodiment, the fungal biomass present in the fermented product has a protein content of at least 30 w% of the dry matter of the fungal biomass.

[0015] In a particular embodiment, the fermented product is a cheese-like product, more particularly a camembert-like product.

[0016] In a second aspect, the application provides a method of producing a fermented product comprising the steps of i) inoculating a vegetable feedstock with a fungal strain, wherein the vegetable feedstock is prepared by blending one or more vegetable or plant products in water; ii) fermenting the vegetable feedstock thereby producing fungal biomass; and iii) collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids. In one embodiment, the vegetable feedstock is sterilized before inoculating the feedstock with the fungal strain. In another embodiment, the method further comprises a step of blending the wet solid phase that is formed by the fungal biomass and the fermented feedstock solids to form a homogenous end product. In another embodiment, one or more plant-based, fungi-based or algae-based components are added during said blending step, more particularly one or more components selected from the group consisting of: flavors, spices, flavor enhancers, fats, fat replacers, oils, starch, proteins, preservatives, sweeteners, color additives, nutrients, emulsifiers, stabilizers, thickeners, gelling agents, pH control agents, acidulants, leavening agents, anti-caking agents, humectants, yeast nutrients, dough strengtheners, dough conditioners, firming agents, enzyme preparations, gasses, and combinations thereof.

[0017] In another embodiment, the method herein provided is a method of producing a cheese-like product. Said method further comprises a final step of transferring the homogenous end product to a cheese mold and optionally forming a rind on the fermented product by adding a fungal or microbial strain to the outside of the fermented product. In a more particular embodiment, said fungal or microbial strain is selected from the list consisting of Penicillium spp., Mucor spp., Pseudomonas spp., Candida spp., Phizomucor spp., Geotrichum candidum and Brevibacterium linens, more particular the fungal strain is selected from the list consisting of P. camemberti, P. candidum, P. roqueforti, P. glaucum, P. nalgiovense, P. caseifulvum, P. album, P. solitum, P. commune, P. carneum, P. carymbiferum, P. crustosum and P. nordicum.

[0018] BRIEF DESCRIPTION OF THE FIGURES

[0019] Figure 1 shows a 20 liter stirred tank fermentation vessel after the completion of the fermentation step. This fermentation was prepared with a cauliflower feedstock inoculated with Penicillium camemberti, as described in Example 2.

[0020] Figure 2 shows the fermented product which was collected following the fermentation step being blended with additional plant-, algae-, or fungi-based ingredients to prepare the homogenous end product, as described in Example 4.

[0021] Figure 3 shows two non-limiting examples (A, B) of the homogenous end product following blending with additional ingredients and transferring to a cheese mold to prepare a final product, as described in Example 5. Thes examples do not include a rind.

[0022] Figure 4 shows an image of a small slice of the homogenous end product following blending with additional ingredients and transferring to a cheese mold to prepare a final product, as described in Example 5. This example does not include a rind.

[0023] Figure 5 shows an example of a small prepared sample of a vegan Camembert-like cheese with a rind formed by applying a liquid formulation of P. camemberti on the outside of the cheese and aging for a minimum of 7 days, as described in Example 5.

[0024] DETAILED DESCRIPTION OF THE INVENTION

[0025] Unless otherwise defined, all terms used in disclosing the invention, including technical and scientific terms, have the meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. By means of further guidance, term definitions are included to better appreciate the teaching of the present invention.

[0026] As used herein, the following terms have the following meanings:

[0027] "A", "an", and "the" as used herein refers to both singular and plural referents unless the context clearly dictates otherwise. By way of example, "a compartment" refers to one or more than one compartment.

[0028] "About" as used herein referring to a measurable value such as a parameter, an amount, a temporal duration, and the like, is meant to encompass variations of + / - 20% or less, preferably + / -10% or less, more preferably + / -5% or less, even more preferably + / -1% or less, and still more preferably + / -0.1% or less of and from the specified value, in so far such variations are appropriate to perform in the disclosed invention. However, it is to be understood that the value to which the modifier "about" refers is itself also specifically disclosed.

[0029] "Comprise", "comprising", and "comprises" and "comprised of" as used herein are synonymous with "include", "including", "includes" or "contain", "containing", "contains" and are inclusive or open-ended terms that specifies the presence of what follows e.g. component and do not exclude or preclude the presence of additional, non-recited components, features, element, members, steps, known in the art or disclosed therein.

[0030] Furthermore, the terms first, second, third and the like in the description and in the claims, are used for distinguishing between similar elements and not necessarily for describing a sequential or chronological order, unless specified. It is to be understood that the terms so used are interchangeable under appropriate circumstances and that the embodiments of the invention described herein are capable of operation in other sequences than described or illustrated herein.

[0031] The recitation of numerical ranges by endpoints includes all numbers and fractions subsumed within that range, as well as the recited endpoints.

[0032] The expression "% by weight", "weight percent", "w%", "%w", "%wt" or "wt%", here and throughout the description unless otherwise defined, refers to the relative weight of the respective component (e.g. protein) based on the overall weight of the formulation (e.g. the fermented product or fungal biomass). The expression "% protein by dry weight", "w% protein by dry weight", "protein w% by dry weight", "wt% protein by dry weight", "protein wt% by dry weight" refers to the protein content of for example the fermented product or fungal biomass expressed in weight percentage of protein based on the dry weight of for example the fermented product or fungal biomass. In current application "dry weight" and "dry matter" are used interchangeably. The dry matter can for example be obtained by drying or dehydrating the wet solid phase.

[0033] Whereas the terms "one or more" or "at least one", such as one or more or at least one member(s) of a group of members, is clear per se, by means of further exemplification, the term encompasses inter alia a reference to any one of said members, or to any two or more of said members, such as, e.g., any >3, >4, >5, >6 or >7 etc. of said members, and up to all said members.

[0034] Reference throughout this specification to "one embodiment" or "an embodiment" means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrases "in one embodiment" or "in an embodiment" in various places throughout this specification are not necessarily all referring to the same embodiment but may. Furthermore, the particular features, structures or characteristics may be combined in any suitable manner, as would be apparent to a person skilled in the art from this disclosure, in one or more embodiments. Furthermore, while some embodiments described herein include some, but not other features included in other embodiments, combinations of features of different embodiments are meant to be within the scope of the invention, and form different embodiments, as would be understood by those in the art. For example, in the following claims, any of the claimed embodiments can be used in any combination.

[0035] "Fermentation" as used herein refers to the metabolic process during which microorganisms, such as bacteria, yeast, or fungi, convert a feedstock such as sugars or other organic compounds into various fermentation products, such as alcohol, acids, gasses, energy or convert the feedstock in a fermented feedstock with modified characteristics such as taste, texture, smell, stability, or nutritional profile. During "traditional fermentation" the aim is to produce or increase the production of said fermentation product or fermented feedstock. Although the microorganisms originated from the inoculant will reproduce and may form a significant microbial biomass, said biomass is not aimed for and will in some cases be removed from the final product for example by sedimentation or filtration. In "microbial biomass fermentation" or simply "biomass fermentation" on the other hand, the focus is on cultivating and propagating microorganisms to achieve a high yield of microbial biomass, which can for example be used as microbial protein in the production of food or animal feed.

[0036] "Fermented product" as used herein refers to a product directly or indirectly obtained by one or more fermentation processes. In the context of the current application, a non-limiting example of the fermented product is the solid fraction including fungal biomass and fermented feedstock solids that is obtained after the fermentation step. The liquid that is removed from the wet solids is referred to as the fermentation filtrate.

[0037] "Homogenous end product" as used herein refers to the fermented product that has been processed further, more particularly by a homogenizing step in the presence of fermentation filtrate, oil and / or other ingredients. A non-limiting example of a homogenous end product in the context of current application is the cheese-like product herein described.

[0038] "Feedstock" or "fermentation medium" as used herein refers to the raw materials, processed or semi-processed substances coming from said raw material or substances that serve as the input for a fermentation process. These materials are used as a starting point for microbial growth and metabolic activities. A "vegetable feedstock" is a feedstock that comprises one or more fresh, frozen, and / or cooked vegetables. The vegetable materials include but are not limited to unprocessed, processed, fermented, unfermented, isolated, concentrated and / or purified plant materials. Examples of unprocessed material are whole plant parts such as leaves, fruits, roots, tubers, stems, flowers or seeds. Processed plant materials are those that have been subjected to drying, cooking, slicing or freezing. Isolated or purified plant materials include plant flour, plant milk, plant extracts and plant residues. In a preferred embodiment, the vegetable materials comprise fresh or frozen whole plant parts, which are either unprocessed or else processed via cooking, slicing or freezing.

[0039] "Unfermented feedstock" as used herein refers to the original feedstock that serves as input for the fermentation process before the fermentation process is initiated. "Fermented feedstock" is a feedstock that has been modified during the fermentation process, for example partly or fully digested by the microorganisms so that the texture, aroma, nutritional profile, and appearance has changed. In the application "fermented feedstock" and "fermented vegetable feedstock" are used interchangeably. "Fermented feedstock solids" as used herein refers to the solid fraction of the fermented feedstock or fermentation medium at the end of the fermentation process. Since the feedstock can be largely digested during fermentation, some part of the feedstock can get dissolved in the medium, while other parts remain as small or larger particles that can form a solid phase.

[0040] "Fungal biomass" as used herein refers to the mass or total amount of fungal cells that has been produced as a result of a fermentation process. It represents the accumulated growth of fungi, more specifically of the fungal strain that was used during inoculation, under specific conditions that are conducive to their proliferation.

[0041] "Inoculation" as used herein refers to the deliberate introduction of a small amount of microorganisms, for example a fungal strain, often referred to as an "inoculum" or "inoculant" or "starter culture", into a larger volume of feedstock or fermentation medium. The purpose of inoculation is to initiate and promote the growth of the desired microorganisms, in order to carry out a fermentation process. The inoculum serves as the initial population of microorganisms that will multiply and transform the feedstock into the desired end products. Inoculation is a strategic step that kickstarts the fermentation process by introducing the right microorganisms into the feedstock, setting the stage for the microorganisms to multiply and carry out the desired biochemical transformations. The inoculum is often prepared under controlled conditions to ensure its viability and potency before being introduced to the larger fermentation vessel.

[0042] "Blending" or "mixing" or "homogenizing" refers to the process of combining two or more substances or materials thoroughly to create a homogeneous mixture. The goal of blending is to achieve a uniform distribution of the components throughout the mixture, ensuring that the final product has consistent properties and characteristics. Blending methods can vary depending on the materials being mixed and the desired outcome. Common blending techniques include using mechanical equipment such as mixers, agitators, blenders, and homogenizers.

[0043] "Homogenous" as used herein refers to a state in which a mixture or substance has a uniform composition and consistent properties throughout. In a homogeneous mixture, the different components are evenly distributed on a microscopic level, giving the impression that the mixture is a single, uniform substance. There is no clear boundary or separation between different regions of the mixture and can therefore be referred to as a single-phase system. Unlike in a heterogeneous mixture, where components might settle or separate over time, components in a homogeneous mixture remain evenly dispersed.

[0044] "Plant-based" refers to something that is derived from or primarily composed of plants, i.e. living organisms belonging to the kingdom Plantae.

[0045] "Algae-based" refers to something that is derived from or primarily composed of algae. Algae are diverse photosynthetic organisms that can range from microscopic single-celled forms to larger multicellular types. They are found in various aquatic environments, including freshwater and marine habitats.

[0046] "Fungi-based" refers to something that is derived from or primarily composed of fungi. Fungi are a diverse group of organisms that include mushrooms, molds, yeasts, and other related species. In a "fungi-based" context, the emphasis is on using fungi as a foundational material, ingredient, or source for various applications and products. Non-limiting examples include powders or extracts of mushrooms or fungi such as yeast extract or nutritional yeast.

[0047] "Dairy" as used herein refers to products that are derived from the milk of mammals, particularly cows, goats, and sheep. These products include various types of milk and milk-based foods that are commonly consumed by humans. Non-limiting examples of dairy are milk, cheese, butter, yogurt, ice cream, cream.

[0048] "Non-animal dairy" are dairy analogs or dairy substitutes that are produced without any animal derived product. The milk which is that basis (starting) ingredient of dairy is replaced for example by plant seed milk, or the solid dairy such as cheeses are substituted by cheese-like products comprising purees of for example nuts.

[0049] "Cheese mold" or "mold" as used herein refers to a device or container used in cheese making to shape and press fresh curds into specific forms. Cheese molds are used to give cheeses their characteristic shapes and sizes, as well as to facilitate the drainage of whey from the curds during the cheese-making process. Cheese molds come in various shapes, sizes, and materials, depending on the type of cheese being made and the desired outcome. They can be made from materials such as plastic, stainless steel, wood, or even natural materials like bamboo leaves. "Rind" refers to the outer layer or surface of a cheese. It is the part of the cheese that develops as it interacts with the environment during the aging and maturation process. The rind can greatly influence the flavor, aroma, and texture of the cheese and is an important aspect of the cheese's overall character. In the case of for example Camembert and Brie, the cheeses are inoculated with specific molds (such as Penicillium camembert!) on the surface. These molds grow and form a fuzzy, white layer on the exterior of the cheese. The molds on the surface of the cheese contribute to the ripening process. They break down the proteins and fats in the cheese, leading to the development of characteristic flavors and aromas.

[0050] In a first aspect, the invention relates to a fermented product, more particularly to a fermented food product or an edible fermented product. Said product can also be referred to as a formulation, a composition, ingredient, or a food material. In one embodiment, said fermented product is a dairy analog or a non-animal dairy substitute. Non-limiting examples of a dairy analog comprise milk analogs, yogurt analogs, butter analogs, cream analogs, cheese analogs, or cheese-like products such as dips and sauces.

[0051] Non-limiting examples of cheese analog food products are hard cheese analogs (e.g. Parmesan), a semi-hard cheese analog (e.g. Gouda), a semi-soft cheese analog (e.g. Havarti), a soft or soft ripened cheese analog (e.g. Brie), a cream cheese analog, a sour milk cheese analog, a blue cheese analog, a mascarpone cheese analog, a pasta filata (e.g. Mozzarella) cheese analog, a brined cheese analog (e.g. Feta), a whey cheese analog (e.g. Ricotta or Brunost), or a fresh cheese analog (e.g. cottage cheese).

[0052] The dairy analog food products according to the present invention may, in some embodiments, be vegan food products, i.e. food products that contain no animal products, and thus allow followers of vegan diets to incorporate such dairy analogs into their diet.

[0053] In one embodiment, the fermented product is a product of biomass fermentation and comprises a fungal biomass and a fermented vegetable feedstock. The fungal biomass is derived from an inoculation of a vegetable feedstock, more particularly an unfermented vegetable feedstock, with a fungal inoculant. In one embodiment, said fungal inoculant is a fungal species of a genus selected from the list consisting of Penicillium, Geotrichum, Candida, Rhizomucor, Mucor, Scopulariopsis, Fusarium and Aspergillus. In a particular embodiment, the fungal species is selected from the list consisting of Penicillium spp., Mucor spp., Candida spp., Rhizomucor spp., Geotrichum candidum, Agaricus spp., Aspergillus spp., Lentinula spp., Pleurotus spp., Cantharellus spp., Ganoderma spp., Rhizopus spp., Fusarium spp., Trichoderma spp., Marasmius spp., Lepista spp., or Boletus spp.. In a more particular embodiment, the fungal species is a Penicillium species, even more particularly a fungal species selected from the list consisting of P. camemberti, P. candidum, P. roqueforti, P. glaucum, P. nalgiovense, P. caseifulvum, P. album, P. solitum, P. commune, P. carneum, P. carymbiferum, P. crustosum and P. nordicum. In a most particular embodiment, the fungal species is selected from the list consisting of P. camemberti, P. candidum, P. commune, P. carneum, and P. roqueforti. In another or further embodiment, the fungal species is a fungal strain or a combination of one or more fungal strains. The fungal strains may be selected from the list consisting of Penicillium camemberti LMG 9126, LMG 34569, LMG 297, LMG 39345, LMG 8446, LMG 31569, LMG 21928, LMG 11510, LMG 29036, LMG 29157, LMG 14158, NRRL 885, LMG 18985, LMG 28971, LMG 15913, and LMG 29790; Penicillium roqueforti NRRL 849, LMG 29083, and LMG 2915; and Penicillium carneum NRRL 25170; and Penicillium commune MUCL 39355.

[0054] In another embodiment, the fungal biomass is derived from an inoculation of a vegetable feedstock with multiple fungal species or multiple different fungal strains.

[0055] In another or further embodiment, the fungal biomass that is obtained at the end of the fermentation process or the fungal biomass comprised in the fermented product has a protein content of at least 30 w% by dry matter or at least 31 w%,

[0056] 32 w%, 33 w%, 34 w%, 35 w%, 36 w%, 37 w%, 38 w%, 39 w%, 40 w%, 41 w%,

[0057] 42 w%, 43 w%, 44 w%, 45 w%, 46 w%, 47 w%, 48 w%, 49 w%, 50 w%, 51 w%,

[0058] 52 w%, 53 w%, 54 w%, 55 w%, 56 w%, 57 w%; 58 w%, 59 w% or 60 w% by dry matter, wherein said dry matter is the dry matter of the fungal biomass. In another embodiment, the fungal biomass has a protein content of between 30 and 60 w% by dry matter, or between 32 and 55 w% by dry matter, or between 35 and 50 w% by dry matter, or between 38 and 48 w% by dry matter, or between 40 and 45 w% by dry matter, wherein said dry matter is the dry matter of the fungal biomass.

[0059] As mentioned earlier, the fermented product is a product of biomass fermentation. Traditional fermentation is the process of changing a feedstock through microbial aerobic or anaerobic digestion. This is how must is processed into wine, and milk in yogurt or cheese. Traditional fermentation results in products with unique flavor and nutritional profiles and modified texture, but the fermented product almost exclusively consists of the fermented feedstock, or more particularly the protein content of the fermented product is almost exclusively derived from the feedstock and not from the microbes. In some cases (e.g. in beer and wine production) the majority of the produced microbial mass is removed from the final product.

[0060] Biomass fermentation on the other hand uses a feedstock or fermentation medium for the rapid growth of microorganisms to efficiently de novo make large amounts of microbial biomass and proteins. The microorganisms that reproduce through this process are thus themselves ingredients for alternative proteins.

[0061] The protein content in w% of dry matter of many vegetables is between 8 w% and 30 w%, while the protein content of the fungal biomass is between 30 and 60 w%. As a result of the biomass fermentation of the vegetable feedstock, de novo proteins are made during the process. Hence, if a vegetable feedstock is used with a protein content of between 8 and 30 w%, the protein content of the fermented product comprising fungal biomass will be higher than that of the vegetable feedstock because of the production and presence of fungal proteins.

[0062] In another or further embodiment, the fermented product as described herein is provided wherein the w% protein by dry weight of the fermented product is statistically significantly increased compared to the w% protein by dry weight of the unfermented vegetable feedstock. More particularly, the fermented product as described herein comprises a protein content in w% protein by dry weight that is at least 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 12%, 15%, 17%, 20%, 25%, 30%, 35%, 40%, 45% or 50% higher than that of the unfermented vegetable feedstock. In a more particular embodiment, the fermented product as described herein comprises a protein content in w% protein by dry weight that is at least between 15% and 40% or between 10% and 20% higher than that of the unfermented vegetable feedstock. In a most particular embodiment, the fermented product as described herein comprises a protein content in w% protein by dry weight that is at least between 12% and 16% higher than that of the unfermented vegetable feedstock.

[0063] In some preferred embodiments, the vegetable feedstock for the final fermented product is free of added protein isolates.

[0064] Non-limiting examples of the vegetable feedstock comprise cauliflower, lentils, peas, green peas, chickpeas, beans, soybean, potatoes, sweet potatoes, carrots, parsnip, leek, pumpkin, squash, broccoli, rice, peppers, red peppers, onions, spinach, kale, zucchini, mushrooms, artichokes, tomatoes, eggplant, beets, cabbage, brussels sprouts, cucumber, asparagus, corn, oats, wheat, barley, celeriac, turnip or sugar beet.

[0065] The vegetable of the vegetable feedstock is in some embodiments a starchy and / or sugary vegetable, such as but not limited to sugar beet, corn, turnip, or parsnip. Alternatively, the vegetable is not rich in proteins, such as but not limited to oats or barley.

[0066] In further embodiments, the vegetable is a neutral or mild colour selected from white, beige and / or yellow and any variation therebetween.

[0067] In a preferred embodiment, the vegetable contains sulfur compounds. Non-limiting examples of sulfur-containing vegetables are the members of Brassica and Allium families.

[0068] In alternative embodiments, the vegetable is starchy and / or sugary, has a neutral- mild colour and is sulfur-containing. In yet another embodiment the vegetable is starchy and / or sugary and has a neutral-mild colour. In yet another embodiment the vegetable is neutral-mild colour and is sulfur-containing. In yet another embodiment the vegetable is starchy and / or sugary and has a neutral-mild colour.

[0069] In an embodiment, the vegetable feedstock is free of nuts and seeds.

[0070] In a lesser preferred embodiment, the vegetable feedstock included added nuts or seeds. Non-limiting examples of nuts and seeds are cashew nuts, peanuts, almonds, macadamias, walnuts, pecans, brazil nuts, hazelnuts, pistachios, pine nuts, sunflower seeds, pumpkin seeds, sesame seeds.

[0071] In a particular embodiment, the vegetable feedstock comprises or consists of two or more vegetables or plant-based material. In another particular embodiment, the vegetable feedstock comprises or consists of a mixture of vegetable or plant-based material.

[0072] In a more particular embodiment, the vegetable feedstock comprises or consists of cauliflower. In a most particular embodiment, the vegetable feedstock comprises at least 5 w%, at least 10 w%, at least 15 w% or at least 20 w% cauliflower by wet weight or comprises between 5 and 50 w%, between 10 and 40%, between 12 and 35 w%, between 15 and 25 w% or around 20 w% cauliflower by wet weight. In another or further embodiment, the fermented product comprising fungal biomass and fermented vegetable feedstock, has a protein content of at least 25 w%, 26 w%, 27 w%, 28 w%, 29 w%, 30 w%, 31 w%, 32 w%, 33 w%, 34 w%, 35 w%, 36 w%, 37 w%, 38 w%, 39 w% or 40 w% of the dry weight of the fermented product. In a particular embodiment, said protein content is exclusively derived from the combination of fermented vegetable feedstock and fungal biomass.

[0073] In an embodiment of the fermented product disclosed herein, the fermented vegetable feedstock forms at least 1 w%, 2 w% , 3 w%, 4 w%, 5 w%, 6 w%, 7 w%, 8 w%, 9 w%, 10 w%, 15 w%, 20 w%, 25 w%, 30 w%, 35 w%, 40 w%, 45 w%, 50 w%, 55 w%, 60 w%, or 65 w% of the fermented product. Preferably, the fermented vegetable feedstock forms at least 10 w% of the fermented product.

[0074] In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 30 w% of the fermented product and the fermented vegetable feedstock between 40 and 70 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 35 w% of the fermented product and the fermented vegetable feedstock between 35 and 65 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 40 w% of the fermented product and the fermented vegetable feedstock between 30 and 60 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 45 w% of the fermented product and the fermented feedstock between 25 and 55 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 50 w% of the fermented product and the fermented feedstock between 20 and 50 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 55 w% of the fermented product and the fermented feedstock between 15 and 45 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 55 w% of the fermented product and the fermented feedstock between 35 and 45 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 60 w% of the fermented product and the fermented feedstock between 10 and 40 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 65 w% of the fermented product and the fermented feedstock between 5 and 35 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms at least 70 w% of the fermented product and the fermented feedstock between 1 and 30 w%. In a further particular embodiment, said w% is the w% per dry weight of the fermented product.

[0075] In another embodiment, the fermented product is characterized in that the fungal biomass forms between 25 and 40 w% of the fermented product and the fermented feedstock between 60 and 75 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms between 30 and 50 w% of the fermented product and the fermented feedstock between 50 and 70 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms between 35 and 60 w% of the fermented product and the fermented feedstock between 40 and 65 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms between 40 and 65 w% of the fermented product and the fermented feedstock between 35 and 60 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms between 45 and 70 w% of the fermented product and the fermented feedstock between 30 and 55 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms between 50 and 75 w% of the fermented product and the fermented feedstock between 25 and 50 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms between 55 and 65 w% of the fermented product and the fermented feedstock between 35 and 45 w%. In another embodiment, the fermented product is characterized in that the fungal biomass forms about 60 w% of the fermented product and the fermented feedstock about 40 w%. In a further particular embodiment, said w% is the w% per dry matter of the fermented product.

[0076] In another embodiment, the fermented product is characterized in that the ratio of the fungal biomass to fermented vegetable feedstock is about 10: 1 to about 1 :5, more particularly about 8: 1 to about 1:3, even more particularly about 5: 1 to about 2: 1, even more particularly about 2: 1 to about 1 :2, about 2: 1 to about 1: 1.5 or most particularly about 1.5: 1.

[0077] In an embodiment of the fermented product as disclosed herein, the ratio of the fungal biomass to fermented vegetable feedstock is between 9: 1 and 1 : 1.5, between 8: 1 and 1 : 1.5, between 7: 1 and 1: 1.5, between 6: 1 and 1 : 1.5, between 5: 1 and 1 : 1.5, between 4: 1 and 1 : 1.5, between 3: 1 and 1 : 1.5, between 2: 1 and 1 : 1.5, between or 1: 1 and 1: 1.5. In an alternative embodiment, the ratio of the fungal biomass to fermented vegetable feedstock is between 9: 1 and 1 : 1, between 9: 1 and 2: 1, between 9: 1 and 3: 1, between 9: 1 and 4: 1, between 9: 1 and 5: 1, between 9: 1 and 6: 1, between 9: 1 and 7: 1, or between 9: 1 and 8: 1.

[0078] Also provided herein is a fermented product comprising cauliflower, more particularly blended cauliflower, even more particularly fermented cauliflower and a filamentous fungus, more particularly fungal biomass, wherein the fermented product does not comprise a butter, more particularly coconut butter or cacao butter or wherein the fermented product does not comprise cauliflower milk and / or a yeast. In a more particular embodiment, said fungal biomass is the fungal biomass according to any of the embodiments herein described.

[0079] "Cauliflower milk" as used herein refers to WO2021174184A1, wherein cauliflower milk is defined by a puree comprising cauliflower, coconut butter and pink salt. According to WO2021174184A1, cauliflower milk may be created by mixing approximately 800.0 grams of finely sliced cauliflower with approximately 75.0 grams of coconut butter and approximately 4.0 grams of a pink salt to form a mixture, stirring and heating the mixture until the mixture changes in color, and blending the mixture with approximately 1230.0 grams of water to form a puree.

[0080] Also provided herein is a fermented product comprising cauliflower, more particularly blended cauliflower, even more particularly fermented cauliflower and a filamentous fungus, more particularly fungal biomass, wherein the dry weight of the fermented product comprises between 30 w% and 99 w% or between 30 w% and 90 w%, or between 35 w% and 85 w%, or between 40 w% and 75 w%, or between 45 w% and 70 w% or between 50 w% and 68 w% or between 55% and 65 w% or around 60 w% of dry weight of the filamentous fungus. In one embodiment, said weight percentages are measured after RIMA deactivation. In a further particular embodiment, said dry weight of the fermented product comprises around 40 w%, around 41 w%, around 42 w%, around 43 w%, around 44 w% or around 45 w% of dry weight of the filamentous fungus, wherein said dry weight was measured after RNA deactivation.

[0081] Also provided herein is a fermented product comprising cauliflower, more particularly blended cauliflower, even more particularly fermented cauliflower and a filamentous fungus, wherein the filamentous fungus is the source for between 30 w% and 90 w%, or between 35 w% and 80w%, or between 40 and 70 w%, or between 42 w% and 65 w% or between 45 and 60 w% or between 48 and 55 w% or around 50 w% of the protein content of the fermented product or wherein the filamentous fungus comprises between 30 and 90 w%, or between 35 w% and 80 w%, or between 40 and 70 w%, or between 42 w% and 65 w% or between 45 and 60 w% or between 48 and 55 w% or around 50 w% of protein content of the fermented product. In a more particular embodiment, said fungal biomass is the fungal biomass according to any of the embodiments herein described.

[0082] Also provided herein is a fermented product comprising at least 30 w%, 35 w%, 40 w%, 45 w% or 50 w% of a fungal biomass per dry weight of the fermented product, wherein the fermented product is prepared or derived from a plant or vegetable source and a fungal inoculant. In a particular embodiment, said plant or vegetable source is cauliflower. In another or further embodiment, said fungal inoculant can be any fungal inoculant, fungal species or fungal strain herein described.

[0083] In embodiments of the current disclosure, the fermented product comprises aroma compounds that give specific cheese flavours such as compounds with butter, creamy, sulfurous, sweet, fruity, floral, sour, pungent, vegetal, mushroomy, earthy, or malty aroma. Non-limiting examples of such aroma compounds include but are not limited to dimethyl sulfide, dimethyl disulfide, dimethyl trisulfide, 2-butanone, 2-pentanone, 2-hexanone, 2-heptanone, 2-nonanone, diacetyl, acetonin, acetic acid, propanoic acid, butanoic acid, 3-methyl butanal, methanethiol, 2-nonenal, butyric acid, caproic acid, octanoic acid, propionic acid, isovaleric acid, phenylacetic acid, lactic acid, 1- or 2-butanol, 2-pentanol, isoamylacetate, ethanethioic acid s- methyl ester, 3-(methylthio)butanal, 2,4-dithiapentane, 2- or 3-methyl butanal, 2- butanone,

[0084] Diacetyl and acetoin are compounds typically found in cheese. Diacetyl is commonly found in dairy products and cheeses, especially in those with a creamy or buttery flavour profile, such as certain types of soft cheeses (e.g., Brie and Camembert).

[0085] Acetoin is found in various kinds of cheese, particularly those that undergo prolonged fermentation or aging. It is less intense than diacetyl but still contributes to the dairy-like aroma.

[0086] Dimethyl disulfide and dimethyl trisulfide can be found in certain cheeses, particularly those that have a strong, pungent aroma, such as blue cheeses (e.g., Roquefort) and some aged cheeses. 2-Heptanone has a fruity or banana-like aroma and can be found in various cheeses, contributing to their overall aroma.

[0087] 2-Nonanone has a fruity or floral aroma and is found in some cheeses, particularly those with complex flavours, though not as characteristic as diacetyl.

[0088] Alcohol and ester compounds tend to contribute fermented, wine-like, sweet, and fruity aromas to specific cheese types.

[0089] In a second aspect, a method is provided of producing a fermented product comprising the steps of:

[0090] - preparing a vegetable feedstock by blending one or more vegetables or plant products in water;

[0091] - inoculating the vegetable feedstock with one or more fungal strains;

[0092] - fermenting the vegetable feedstock thereby producing fungal biomass; and

[0093] - collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids.

[0094] Also provided is a method of producing a fermented product comprising the steps of: inoculating a vegetable feedstock with one or more fungal strains, wherein the vegetable feedstock is obtained by blending one or more vegetable or plant products in water;

[0095] - fermenting the vegetable feedstock thereby producing fungal biomass; collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids.

[0096] In a particular embodiment, said wet solid phase comprises the fungal biomass and the fermented feedstock solids in a ratio between about 10: 1 to about 1 :5, more particularly about 8: 1 to about 1 :3, even more particularly about 5: 1 to about 2: 1, even more particularly about 2: 1 to about 1 :2, most particularly about 1.5: 1.

[0097] In an embodiment of the method as disclosed herein, the ratio of the fungal biomass to fermented vegetable feedstock in the wet solid phase is between 9: 1 and 1 : 1.5, between 8: 1 and 1 : 1.5, between 7: 1 and 1: 1.5, between 6: 1 and 1 : 1.5, between 5: 1 and 1 : 1.5, between 4: 1 and 1 : 1.5, between 3: 1 and 1 : 1.5, between 2: 1 and 1 : 1.5, between or 1: 1 and 1: 1.5. In an alternative embodiment, the ratio of the fungal biomass to fermented vegetable feedstock in the wet solid phase is between 9: 1 and 1 : 1, between 9: 1 and 2: 1, between 9: 1 and 3: 1, between 9: 1 and 4: 1, between 9: 1 and 5: 1, between 9: 1 and 6: 1, between 9: 1 and 7: 1, or between 9: 1 and 8: 1.

[0098] In another particular embodiment, the fungal biomass forms at least 30 w%, at least 35 w%, at least 40 w%, at least 50 w%, at least 55 w% or at least 60 w% of the wet solid phase per dry weight of said solid phase.

[0099] In another particular embodiment, the fermented product that is produced by said method is any of the fermented products disclosed in current application.

[0100] In another particular embodiment, the fermenting step is performed in a stirred tank with controlled agitation, aeration and / or pH control.

[0101] In another or further embodiment, said fermented product is a dairy analog or a non-animal dairy substitute. In a particular further embodiment, said dairy analog is milk analog, yogurt analog, butter analog, cream analog, cheese analog, cream cheese analog, mozzarella analog, gouda analog, Cheddar analog, brie analog, Camembert analog, goats cheese analog, Roquefort analog, parmesan analog, Emmental analog, gruyere analog, feta analog or cheese-like product such as a dip or a creamy sauce.

[0102] In one embodiment, the one or more vegetable or plant products are fresh, frozen, cooked or sliced and / or are blended in a concentration of at least 5%, 10%, 15%, 20%, 25%, 35%, 40%, 45% or 50% by wet weight in water, more particularly distilled water. In another or further embodiment, one or more of a variety of soluble additives are added to the vegetable feedstock in the blending step. Non-limiting examples of additives include salts, citric acid, buffering agents, glucose, amino acids, trace elements such as zinc, iron and copper.

[0103] In another or further embodiment, the one or more vegetable or plant products comprise at least cauliflower, more particularly cauliflower florets.

[0104] In one embodiment, the method further comprises a step of sterilizing the vegetable feedstock before the feedstock is inoculated with one or more fungal strains. "Sterilizing" or "sterilization" as used herein refers to the complete destruction or elimination of all viable organisms, more particularly microorganisms such as bacteria, yeast, fungi, viruses, in or on an object being sterilized. Sterilization procedures are well known by the person skilled in the art and involve the use of heat, radiation, chemicals or physical removal of cells. In a particular embodiment, said sterilizing step uses heat, more particularly steam. In another particular embodiment, said sterilizing step is an in situ sterilization step. In situ sterilization or in situ batch sterilization or sterilization-in-place of a liquid medium refers to a process where the medium is sterilized without being moved. In this process, the fermenter vessel containing the feedstock is heated using steam and held at the sterilization temperature for a period of time, for example at 121°C for 20 min. Cooling water is then used to bring the temperature back to normal operating conditions. In another particular embodiment, said sterilizing is autoclaving. Typically and known by the skilled person, an autoclaving process comprises a heating step at high pressure, for example using a high-pressure cooking device that is operated at 5-15 psi and 112°C-127°C for 10-50 min.

[0105] In another or further embodiment, the collecting step of the wet solid phase is a step of separating the wet solid phase comprising fungal biomass and fermented feedstock solids from a liquid filtrate and collecting said wet solid phase. Said separation step or dewatering step can be performed by filtration or centrifugation techniques known by the skilled person in the art.

[0106] In another or further embodiment, the method further comprises a step of RIMA deactivation and removal by washing. In one embodiment, said step is performed after the fermentation step but before the wet solid phase is collected. The fermentation mix is then heated to for example 60-65°C for 10 minutes and subsequently the wet solid phase is collected by filtration or centrifugation. In another embodiment, said RNA deactivation and removal step is performed after the first collection of the wet solid phase. Said solid phase is then diluted in water again, heated to for example 60-65°C for 10 minutes whereafter a second collection step is performed by filtration or centrifugation to recover the wet solid phase comprising the fungal biomass and the fermented feedstock solids.

[0107] In another or further embodiment, the method further comprises a step of homogenizing the collected wet solid phase into a homogenous end product. In a particular embodiment, the wet solid phase is homogenized by a further blending and / or stirring step in the presence of or prior to adding one or more ingredients. Non-limiting examples of said ingredients are the fermentation filtrate (i.e. the liquid from which the wet solid phase was separated), an oil, or one or more plant-based or algae-based components. In one embodiment, said one or more plant-based or algae-based components are selected from the list consisting of flavors, spices, flavor enhancers, fats, fat replacers, oils, starch, proteins, preservatives, sweeteners, color additives, nutrients, emulsifiers, stabilizers, thickeners, gelling agents, pH control agents, acidulants, leavening agents, anti-caking agents, humectants, yeast nutrients, dough strengtheners, dough conditioners, firming agents, enzyme preparations, gasses, and combinations thereof. The further homogenizing step has the purpose of making the fermented product more smooth and homogenous.

[0108] In another or further embodiment, the method further comprises a step of pasteurizing the homogenous fermented product. Pasteurization is well known by the skilled person. A non-limiting example of a typical pasteurization step comprises heating the product to for example 60°C for 30 minutes up to 90°C for 8 minutes. Alternative and similarly effective pasteurization methods are known by the skilled person.

[0109] In another or further embodiment, the method as described herein is provided wherein the fermented product is a cheese-like product and wherein the method further comprises a step of transferring the homogenous end product in one or more cheese molds. In a further embodiment, the method further comprises a step of removing the cheese-like product from the cheese molds.

[0110] In another or further embodiment, the method further comprises a step of providing a rind to the cheese-like product. The rind can be formed by either applying (e.g. spraying) a liquid formulation of a microbial strain or combination of microbial strains on the outside of the cheese, or blending one or more active microbial strains into the final mixture following the pasteurization step and before molding.

[0111] In a more particular embodiment, the method comprises a step of adding one or more microbial strains to the outside of the cheese-like product for making the rind. In another particular embodiment, the method comprises a step of blending one or more microbial strains into the final mixture for making the rind, preferably following the pasteurization step and prior to the molding step.

[0112] In a most particular embodiment, said one or more microbial strains are selected from the list consisting of Penicillium spp., Mucor spp., Pseudomonas spp., Candida spp., Rhizomucor spp., Geotrichum candidum and Brevibacterium linens. In a preferred embodiment the microbial strain for rind formation is a fungal strain. In an even more particular embodiment, said fungal strain is a fungal species from the Penicillium genus. In a most particular embodiment, said one or more fungal strains is selected from the list consisting of P. camemberti, P. candidum, P. roqueforti, P. glaucum, P. nalgiovense, P. caseifulvum, P. album, P. solitum, P. commune, P. carymbiferum, P. crustosum and P. nordicum.

[0113] In another embodiment, a fermented product is provided, wherein said fermented product is obtained by any of the methods herein described.

[0114] In a particular embodiment, a fermented product is provided obtained or produced by the method comprising the steps of: inoculating a vegetable feedstock with one or more fungal strains, wherein the vegetable feedstock is obtained by blending one or more vegetable or plant products in water;

[0115] - fermenting the vegetable feedstock thereby producing fungal biomass; collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids.

[0116] In another embodiment, a homogenous end product is provided, wherein said homogenous end product is obtained by any of the methods herein described.

[0117] In a particular embodiment, a homogenous end product is provided obtained or produced by the method comprising the steps of: inoculating a vegetable feedstock with one or more fungal strains, wherein the vegetable feedstock is obtained by blending one or more vegetable or plant products in water;

[0118] - fermenting the vegetable feedstock thereby producing fungal biomass; collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids; blending the wet solid phase to form a homogenous end product.

[0119] In a further embodiment, during said blending step one or more components are added. In a particular embodiment, said one or more components are fermentation filtrate. In another or further particular embodiment, said one or more components are one or more plant-based, fungi-based or algae-based components, even more particularly components selected from the group consisting of: flavors, spices, flavor enhancers, fats, fat replacers, oils, starch, proteins, preservatives, sweeteners, color additives, nutrients, emulsifiers, stabilizers, thickeners, gelling agents, pH control agents, acidulants, leavening agents, anti-caking agents, humectants, yeast nutrients, dough strengtheners, dough conditioners, firming agents, enzyme preparations, gasses, and combinations thereof.

[0120] In another or further embodiment, said homogenous end product is a dairy analog or a non-animal dairy substitute. Non-limiting examples of a dairy analog comprise milk analogs, yogurt analogs, butter analogs, cream analogs, cheese analogs, cream cheese analog, mozzarella analog, gouda analog, Cheddar analog, brie analog, Camembert analog, goats cheese analog, Roquefort analog, parmesan analog, Emmental analog, gruyere analog, feta analog or cheese-like products such as dips or creamy sauces.

[0121] In another or further embodiment, the homogenous end product is a cheese-like product.

[0122] In another particular embodiment, a cheese-like product is provided obtained or produced by the method comprising the steps of: inoculating a vegetable feedstock with one or more fungal strains, wherein the vegetable feedstock is obtained by blending one or more vegetable or plant products in water;

[0123] - fermenting the vegetable feedstock thereby producing fungal biomass; collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids; blending the wet solid phase to form a cheese-like product; optionally transferring the cheese-like product to a cheese mold and forming a rind on the cheese-like product.

[0124] EXAMPLES

[0125] Example 1. Preparing the vegetable feedstock

[0126] A vegetable feedstock was prepared with cauliflower as the main ingredient. Cauliflower florets were added in a 20 w% concentration by wet weight to distilled water. A variety of additives such as salts, citric acid, glucose can be added. Next, the obtained mix was blended. The type of blender is not crucial as long as the cauliflower florets are significantly decreased in particle size. A non-limiting example of a blender is an immersion blender. After blending the mix, the feedstock was autoclaved (21 minutes at 121 °C at 15 psi) to sterilize the feedstock. If some additives would undergo Maillard reactions or other degradation reactions due to the autoclaving step, these additives are added after the autoclaving step, optionally these additives are sterilized prior to adding to the autoclaved feedstock.

[0127] Example 2. Fermenting the vegetable feedstock

[0128] Different types of microorganisms can be used to ferment the vegetable feedstock. As a first example, the feedstock is fermented by the filamentous fungus Penicillium camemberti. The P. camemberti strain was initially grown on potato dextrose agar plates from which a 10% by volume inoculant was taken to start a liquid culture in 10 mL tubes in potato dextrose medium. Next, the fungal strain was grown at 4% by volume inoculation in 250-500 mL aerated flasks in the vegetable feedstock as prepared in Example 1. Finally a full-scale fermentor with the vegetable feedstock was inoculated with a 10% by volume liquid sample of the previous step.

[0129] The vegetable feedstock was fermented for 24 to 48 hours at 26.9°C in a stirred tank bioreactor with controlled agitation, aeration and pH control (Figure 1).

[0130] Example 3. Collecting the fermented product

[0131] After the fermentation step, the content of the bioreactor was divided into phases: on one hand the wet solids including fungal biomass and fermented feedstock and on the other hand the liquid. This dewatering step can be performed in different ways as long as water is being removed from the solid phase. One possibility is to filter the fermentation product for example over a Miracloth filter to separate the liquid filtrate from the wet solids and to squeeze it to remove additional water. Another possibility is to use centrifugation or other standard filtration techniques.

[0132] Next, the wet solid biomass was diluted in water and heated to 65°C for 10 minutes to deactivate and remove RNA. A subsequent filter step was performed to recover the solid biomass and wash away the RNA contained in the liquid phase. This RNA deactivation step could also be performed in the fermentation vessel immediately after the fermentation was completed and before the first filtration step.

[0133] The obtained fermented product that consists of filamentous fungal biomass and any residual vegetable feedstock solids is kept wet and should be used immediately fresh or should be stored frozen and / or vacuum packed.

[0134] A sample of the fermented product was collected following the dewatering step, and analyzed to determine the ratio of fungal biomass to fermented vegetable feedstock. This ratio was determined by performing one of a variety of possible analytical techniques (e.g. protein analyses such as Kjeldahl, Dumas or other type of tests; mass spectrometry; qPCR; chitosan assay; chitin assay) on the fermented product. The values were compared to samples of the vegetable feedstock before fermentation, and samples of pure fungal biomass that were grown on a standard sugar-based liquid solution, so that all of the collected solid mass was derived solely from the fungus and not the feedstock.

[0135] One example of a straightforward analytical technique to estimate the ratio of fungal biomass to fermented vegetable feedstock in the fermented product, expressed as percentages, is to measure the protein content as a percentage of dry matter using a Kjeldahl or Dumas test for the fermented product (F), vegetable feedstock (VQ, and pure fungal biomass (8). This approach works as long as the protein contents of the vegetable feedstock and pure fungal biomass are different, and the protein content of the pure fungal biomass is higher than that of the fermented product.

[0136] The percentage of fungal biomass, X, can then be expressed by the following equation: X = 100*(F-V) / (B-V), and the remaining percentage for the fermented vegetable feedstock = 100-X.

[0137] In a representative example, the protein content as a percentage of dry matter for the vegetable feedstock (VQ, in this case autoclaved cauliflower, was found to be 20.6%. The protein content of the fermented product (F) grown with Penicillium camemberti and following RIMA deactivation was 30.6±5.7% determined as the mean of 4 independent measurements. The w% protein by dry matter of the fermented product was thus 4.3% to 15.7% higher compared to the w% protein by dry matter of the unfermented vegetable feedstock.

[0138] The protein content as a percentage of dry matter for the pure fungal biomass (B) of Penicillium camemberti grown on a sugar solution was 41.6% and following RNA deactivation 43%. Using the equation provided in the previous paragraph, the percentage of fungal biomass in the fermented product was estimated as 61%, and the percentage of fermented vegetable feedstock was estimated as 39%.

[0139] A variety of other techniques could be employed to estimate the ratio of fungal biomass to fermented vegetable feedstock in the fermented product, such as qPCR, quantitative mass spectrometry, quantitative LCMS (liquid chromatography-mass spectrometry) and quantitative assays for chitosan and chitin content, where the latter two components should only be found within the fungal biomass. Example 4. Final product preparation

[0140] Depending on the additional flavours and tastes one wants to achieve, several additional ingredients can be added to make a cheese-like product. Preferably these are plant-based, fungi-based or algae-based ingredients. Non-limiting examples include an oil, for example sunflower oil or olive oil; starch, for example tapioca starch or potato starch; protein, for example chickpea protein or fava bean protein; gelling agents, for example carrageenan; salt, for example sea salt; acidity regulators, for example citric acid or lactic acid; yeast extracts or any other flavoring components.

[0141] The wet solid phase comprising the fungal biomass and the fermented feedstock solids was homogenized with some of the fermentation filtrate of the previous steps to make a more smooth preparation (Figure 2). This can be achieved by any small- scale or industrial blender. The homogenization step may also include adding oils or other ingredients. Finally the mixture was pasteurized. A typical example is an 8 minute heating step at 90°C.

[0142] Example 5. A vegan Camembert-like cheese

[0143] The fermented product as obtained in Example 4 can now be used for several food applications. One example is for the production of a Camembert-like cheese.

[0144] The mixture as obtained in Example 4 was transferred to cheese molds and allowed to set. Figures 3-5 illustrate the obtained cheese-like product after removing the molds. Then a rind was created by adding Penicillium camemberti to the outside of the molded cheese analog. The rind can be formed by either applying (e.g. spraying) a liquid formulation of P. camemberti on the outside of the cheese, or blending some active P. camemberti into the final mixture following the pasteurization step and before molding.

[0145] After the P. camemberti inoculation step, the Camembert-like cheese was aged for 1 to 2 weeks (Figure 6).

[0146] Example 6. A vegan Roquefort-like cheese

[0147] Another example of the use of the obtained mixture is for the production of a Roquefort-like cheese.

[0148] The mixture as obtained in Example 4 was transferred to cheese molds and allowed to set before removing the molds. Then a rind was created by adding Penicillium roqueforti to the outside of the molded cheese analog. The rind can be formed by either applying (e.g. spraying) a liquid formulation of P. roqueforti on the outside of the cheese, or blending some active P. roqueforti into the final mixture following the pasteurization step and before molding.

[0149] After the P. roqueforti inoculation step, the Roquefort-like cheese was aged for 1 to 2 weeks.

[0150] Example 7. A vegan cream cheese analog

[0151] Another example of the use of the obtained mixture is for the production of a cream cheese analog.

[0152] The mixture as obtained in Example 4 is homogenized once more to obtain a soft, creamy and spreadable texture. A variety of spices and herbs such as garlic, basil, lemon zest, parsley, coriander, dill, ginger, lavender, oregano, rosemary, saffron, thyme, among others are added. The cream cheese like product is placed in the fridge overnight and can be served with crackers or crostini the day after.

[0153] Example 8. A vegan goat cheese analog

[0154] Another example of the use of the obtained mixture is for the production of a goat cheese analog.

[0155] The mixture as obtained in Example 4 is homogenized once more to obtain a soft, creamy and spreadable texture, then transferred to cheese molds and allowed to set before removing the molds. These cheeses can either be used as-is after removing the molds, or they can contain a rind formed by aging the cheese following addition of yeast or mold cultures.

[0156] Example 9. Aroma compounds in cheese analog fermented product

[0157] A vegetable feedstock was prepared from cauliflower or green peas, as described in Example 1. The vegetable feedstock was further fermented using Penicillium camemberti or Penicillium roqueforti as described in Example 2. A fermented product was collected as described in Example 3.

[0158] The aroma compounds of the fermented products were detected by GC-MS (gas chromatography mass-spectrometry) which detects and identifies volatile chemical compounds released by a sample. GC-MS results for 3 combinations of vegetable feedstock and fungal strain are shown in Table 1, where only the compounds detected with the highest intensity are listed, which were also detected in a dairybased Camembert cheese sample as reference.

[0159] Table 1. Combination of vegetable feedstocks and fungal strains used to produce fermented products and the aroma compounds detected in the fermented products.

[0160] The aroma compounds profile identified for the three vegetable feedstock and fungal strain combinations indicates that the resulting fermented products have a cheeselike flavour and aroma profile and are thus suitable as cheese alternatives.

[0161] Example 10. Nutritional value of a cheese analog fermented product

[0162] A vegetable feedstock was prepared from cauliflower, as described in Example 1. The vegetable feedstock was further fermented using Penicillium camemberti as described in Example 2. A fermented product was formed as described in Example 3.

[0163] The nutrient profile of the cheese analog product was determined using HPLC (high performance liquid chromatography) for the amino acid profile, Dumas and Kjeldahl tests for protein content, enzymatic-gravimetric method for dietary fibre content, gravimetric method for fats, gas chromatography for sugars, and direct calculation for carbohydrates (subtracting moisture, protein, lipids, and minerals from the total). The nutrient profile of the fermented cheese analog product was compared to the nutrient profile of standardized whole cow's milk (Tables 2 and 4). Table 2. Comparative values of macronutrients of the fermented product and whole cow's milk Table 3. Comparative values of essential amino acids of the fermented product and cow's milk

[0164] Table 4. Comparative values of non-essential amino acids of the fermented product and cow's milk

[0165] The fermented product has fewer calories and a lower fat content compared to whole cow's milk (Table 2), making it advantageous for those managing their calorie intake. Additionally, the product contains 3 g of fibre, which promotes digestive health, whereas cow's milk contains no fibre (Table 2). It also offers a well-balanced profile of essential and non-essential amino acids comparable to that found in cow's milk, supporting overall health, muscle repair, and immune function (Table 3 and Table 4).

[0166] Example 11. Dairy-analog specialty products

[0167] Various dairy analogs are created by combining different vegetable feedstocks with various fungal strains as described in examples 1-2. The resulting fermented product is collected as described in example 3, which can then be blended with additional ingredients as described in example 4 and optionally add a rind according to examples 5-6.

[0168] Penicillium camemberti is used to ferment corn, oats, chickpeas, or sugarbeet vegetable feedstock as described in examples 1-3 to produce a fermented product. The fermented product is further blended with additional ingredients to create a cheese analog as described in example 4, and a rind is formed as described in example 5. This process results in a dairy analog that mimics the characteristics of Camembert cheese. Penicillium camemberti is the mold responsible for Camembert's distinctive white rind and creamy, buttery texture. When fermented on these vegetable feedstock sources to create the fermented product, the resulting analog is expected to develop a creamy texture, and an earthy, mushroom-like flavor with slightly tangy notes.

[0169] Penicillium roqueforti is used to ferment white beans and onions, producing a dairy analog similar to Roquefort cheese, as described in examples 1-4 and 6. This analog is expected to have a firm and crumbly texture with blue-green veins, much like traditional blue cheese. The flavor profile would be strong, tangy, and slightly pungent, complemented by the earthy, savory undertone from the onions.

[0170] Pleurotus ostreatus is used to ferment corn or oats, resulting in a dairy analog with characteristics akin to semi-soft cheeses. Pleurotus ostreatus, commonly known as the oyster mushroom, imparts a mild, slightly sweet, and earthy flavor, which translates into a creamy and mildly flavored cheese analog.

[0171] Agaricus bisporus is used to ferment cabbage and white beans, creating a dairy analog with a unique blend of umami, earthy, and slightly nutty flavors. Agaricus bisporus, commonly known as the button mushroom, combined with cabbage and white beans, yields a firm yet smooth texture and a complex flavor profile.

[0172] The vegetable feedstocks significantly contribute to the flavor and texture of the resulting fermented product and dairy analog products formed after blending with additional ingredients and optionally forming a rind.

[0173] It is clear that the methods and products according to the invention, and its applications, are not limited to the presented examples.

Claims

CLAIMS1. A fermented product comprising a protein content of at least 25 w% of the dry matter of the fermented product and comprising a fungal biomass and a fermented vegetable feedstock, characterized in that the fungal biomass forms at least 40 w% of the dry matter of the fermented product and the fermented vegetable feedstock at least 10 w% of the dry matter of the fermented product.

2. The fermented product according to claim 1, wherein the ratio of the fungal biomass to fermented vegetable feedstock is between about 9: 1 to about 1: 1.5.

3. The fermented product according to any of claims 1-2, wherein the fermented product is derived from an unfermented vegetable feedstock and a fungal inoculant.

4. The fermented product according to claim 3, wherein the w% protein by dry matter of the fermented product is higher compared to the w% protein by dry matter of the unfermented vegetable feedstock.

5. The fermented product according to any of claims 3-4, wherein the unfermented vegetable feedstock comprises cauliflower.

6. The fermented product according to any of the preceding claims, wherein the fungal biomass comprises a fungal strain of the Penicillium genus.

7. The fermented product according to claim 6, wherein the fungal strain is selected from the list consisting of P. camemberti, P. candidum, P. roqueforti, P. glaucum, P. nalgiovense, P. caseifulvum, P. album, P. solitum, P. commune, P. carymbiferum, P. crustosum and P. nordicum.

8. The fermented product according to any of the preceding claims, wherein the fungal biomass has a protein content of at least 30 w% of the dry matter of the fungal biomass.

9. A method of producing a fermented product comprising the steps of: a. inoculating a vegetable feedstock with a fungal strain, wherein the vegetable feedstock is prepared by blending one or more vegetable or plant products in water; b. fermenting the vegetable feedstock thereby producing fungal biomass; and c. collecting the wet solid phase comprising the fungal biomass and the fermented feedstock solids.

10. The method according to claim 9, wherein the wet solid phase comprises the fungal biomass and the fermented feedstock solids in a ratio between about 9: 1 to about 1 : 1.5.

11. The method according to any of claims 9-10, wherein the vegetable feedstock is sterilized before inoculating the feedstock with the fungal strain.

12. The method according to any of claims 9-11, further comprising a step of blending the wet solid phase from step c to form a homogenous end product.

13. The method according to claim 12, wherein one or more plant-based, fungi- based or algae-based components are added during the blending step, wherein the one or more components are selected from the group consisting of: flavors, spices, flavor enhancers, fats, fat replacers, oils, starch, proteins, preservatives, sweeteners, color additives, nutrients, emulsifiers, stabilizers, thickeners, gelling agents, pH control agents, acidulants, leavening agents, anti-caking agents, humectants, yeast nutrients, dough strengtheners, dough conditioners, firming agents, enzyme preparations, gasses, and combinations thereof.

14. The method according to any of claims 12-13, further comprising a step of transferring the homogenous end product to a cheese mold and forming a rind on the homogenous end product, optionally by adding a microbial strain selected from the list consisting of Penicillium spp., Mucor spp., Pseudomonas spp., Candida spp., Rhizomucor spp., Geotrichum candidum and Brevibacterium linens.

15. The method according to claim 14, wherein the microbial strain is selected from the list consisting of P. camemberti, P. candidum, P. roqueforti, P. glaucum, P. nalgiovense, P. caseifulvum, P. album, P. solitum, P. commune, P. carymbiferum, P. crustosum and P. nordicum.

16. A fermented product obtained by the method according to any of claims 9-15.

17. The fermented product according to any of claims 1-8 or the method according to any of claims 9-15, wherein the fermented product does not comprise cashew nuts, coconut butter or cacao butter.

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