Protein dispersion

Abstract

The present invention relates to a method for preparing a plant-based protein hydrogel slurry, and a method for preparing a plant-based structured material (such as a film, a casting, a molded product, etc.) from the plant-based protein hydrogel slurry.

Description

Technical Field

[0001] The present invention relates to a method for preparing a plant-based protein hydrogel slurry, and a method for preparing a plant-based structured material (such as a film, a casting, a molded part, etc.) from the plant-based protein hydrogel slurry. The present invention also relates to the plant-based protein hydrogel slurry and the plant-based structured material itself, and uses thereof. Background Art

[0002] There is an increasing need to reduce the environmental impact of many everyday activities and to reduce the amount of non-renewable resources involved in these activities. An example of this is the increasing use of biodegradable or renewable packaging to replace traditional plastics such as polyethylene and polypropylene (e.g. edible films for food).

[0003] Efforts have therefore focused on using natural or naturally derived materials, such as cellulose, alginates, starches, collagen, and collagen-derived proteins, as film and packaging forming materials. However, many of these naturally derived films often have issues such as limited tensile strength or sensitivity to moisture or limited barrier properties, which limit the range of applications for which they are suitable. One option is to chemically modify the film-forming material, typically using cross-linking chemicals that can cross-link long-chain polymers, but this introduces complexity and additional chemistries that may not be suitable for the end use, such as in an edible film or product with high biodegradability. Similarly, composites that combine natural and synthetic materials to overcome any limitations of one material will also be more complex to produce and may not be edible or recyclable.

[0004] Among the different types of biopolymers that can serve as building blocks to generate new functional materials, such as membranes, proteins are interesting candidates due to their ability to self-assemble into functionalized structures.

[0005] Currently, the use of these materials in commercial applications is limited to highly soluble proteins of animal origin. Animal-based proteins commonly used in food, such as whey proteins, have good biocompatibility, biodegradability, amphiphilicity and functional properties, such as water solubility, emulsification and foaming ability. However, there is a growing demand to replace animal-based proteins with plant-based proteins, not only because of their lower environmental impact, but also because of their lower allergenicity and reduced cost.

[0006] The formation of self-assembled hydrogel materials, such as films of plant-based proteins, has been reported, where hydrogels can be obtained from soy and pea proteins under a range of experimental conditions. However, the mechanical properties obtained from current plant-based materials are generally lower compared to those obtained from animal-derived materials, as plant proteins are more difficult to process, at least in part due to their inherently low solubility in water. The same is true for the optical and barrier properties of current plant-based materials.

[0007] Therefore, a simple route to develop plant-based material products with improved physical properties is needed to increase the range of applications in which plant-based materials can be used to further replace synthetic materials. Summary of the invention

[0008] From a first aspect, the present invention provides a method for preparing a plant-based protein hydrogel slurry, the method comprising:

[0009] (a) forming a solution comprising one or more plant-based proteins in a solvent system, wherein the solvent system comprises miscible co-solvents; wherein a first co-solvent increases the solubility of the plant-based protein and a second co-solvent decreases the solubility of the plant-based protein;

[0010] (b) inducing a sol-gel transition of the protein in the solution to form a plant-based protein hydrogel; and

[0011] (c) subjecting the plant-based protein hydrogel to a shearing treatment to form a plant-based protein hydrogel slurry.

[0012] From another aspect, the present invention provides a plant-based protein hydrogel slurry prepared according to the method described above.

[0013] From another aspect, the present invention provides a method for preparing a plant-based structured material, the method comprising:

[0014] (a) preparing a plant-based protein hydrogel slurry according to the above method; and

[0015] (b) subjecting the plant-based protein hydrogel slurry to one or more solvent level reduction steps to reduce the level of the first co-solvent and / or the second co-solvent to obtain the plant-based structuring material.

[0016] From another aspect, the present invention provides a plant-based structured material prepared according to the method described above.

[0017] Viewed from another aspect, the present invention provides use of a plant-based protein hydrogel slurry as hereinbefore described to produce a plant-based structured material.

[0018] From another aspect, the present invention provides a plant-based protein hydrogel slurry, wherein the protein solid content is 5wt% to 25wt% based on the total weight of the plant-based protein hydrogel slurry, and the solid content of the protein is 5wt% to 25wt% based on the total weight of the plant-based protein hydrogel slurry. -1 and a viscosity at 20° C. in the range of 10 to 10,000 cps, wherein the plant-based protein hydrogel slurry comprises a d 50 Particle sizes range from 0.5 to 150 microns.

[0019] From another aspect, the present invention provides a plant-based protein hydrogel slurry, wherein the protein solid content is 5wt% to 25wt% based on the total weight of the plant-based protein hydrogel slurry, and the solid content of the protein is 5wt% to 25wt% based on the total weight of the plant-based protein hydrogel slurry. -1 and a viscosity at 20° C. in the range of 10 to 10,000 cps, wherein the plant-based protein hydrogel slurry comprises a d 50 Fragments with a particle size less than 500nm.

[0020] Viewed from another aspect the invention provides a film comprising a plant based protein hydrogel slurry as hereinbefore described.

[0021] definition

[0022] As used herein, the term "low shear step" refers to a process step in which a low level of mechanical energy is applied to a material, preferably by a cutting action, to cause it to be primarily broken or fragmented into large discrete pieces. "Low shear" generally does not include any grinding step that comminutes or breaks the material by high-speed impact, such as at speeds greater than 2 ms. -1 Differential impact. It also generally does not include cavitation-based grinding processes. In a particular embodiment, during the low shear step, the hydrogel is broken to produce fragments such that at least 80 wt % of the hydrogel fragments have a maximum size of 1 mm to 50 mm as determined by sieving. Sieving of the hydrogel slurry can be performed according to the methods described herein.

[0023] As used herein, the term "high shear step" refers to a process step in which energy is applied to reduce the hydrogel into small fragments, for example, to form, for example, a colloidal dispersion. During the high shear step, the hydrogel may be broken up to produce a hydrogel having a d measured by dynamic light scattering (DLS) or laser diffraction. 50 The hydrogel is fragmented during the high shear step to produce particles having a particle size of less than 500 nm as measured by dynamic light scattering (DLS). 50 In an alternative embodiment, during the high shear step, the hydrogel is fragmented to produce fragments having a particle size of d 50Particle size fragments range from 0.5 to 150 microns. DLS and laser diffraction may be performed according to the methods defined herein.

[0024] For the avoidance of doubt, the high shear step subjects the hydrogel to a higher level of shear than the low shear step. Where the process involves both a low shear step and a high shear step, the high shear step must occur after the low shear step (ie they are discrete steps occurring in this particular order).

[0025] Detailed description

[0026] The present application describes a method for making a pourable and pumpable hydrogel slurry that can be dried to form a strong film, coating, molding and other structured objects. Therefore, the present invention provides a method for preparing a plant-based protein hydrogel slurry, the method comprising:

[0027] (a) forming a solution comprising one or more plant-based proteins in a solvent system, wherein the solvent system comprises miscible co-solvents; wherein a first co-solvent increases the solubility of the plant-based protein and a second co-solvent decreases the solubility of the plant-based protein;

[0028] (b) inducing a sol-gel transition of the protein in the solution to form a plant-based protein hydrogel; and

[0029] (c) subjecting the plant-based protein hydrogel to a shearing treatment to form a plant-based protein hydrogel slurry.

[0030] In embodiments, prior to step (b), it may be preferred to remove solvent from the protein solution so as to form a more concentrated protein solution prior to step (b). This can be accomplished by applying heating and / or vacuum as well as other techniques. This initial reduction in solvent can provide advantages such as simplifying subsequent drying. Suitable equipment may include a scraped wall evaporator or a twin screw extruder to which vacuum is applied.

[0031] Any suitable plant-based protein can be used in the present invention. Different plant-based proteins can produce hydrogel slurries to produce structured objects with different properties. For example, soy protein may produce hydrogels (and materials formed by these hydrogels) that are stronger than pea protein, and may need to be treated differently to obtain optimal performance. However, the applicability of the present invention depends not only on the hydrogel properties, but also needs to be balanced with other factors (such as the availability of protein raw materials, lack of competition with food supply, protein allergy, etc.). In a preferred method of the present invention, the plant-based protein is selected from soy protein, pea protein, rice protein, potato protein, wheat protein, zein or sorghum protein. Preferably, the plant-based protein is selected from soy protein, pea protein, potato protein and / or rice protein. More preferably, the plant-based protein is selected from soy protein and / or pea protein.

[0032] In a preferred method of the present invention, the plant-based protein is selected from soy protein, pea protein, rice protein, potato protein, wheat protein, zein, rapeseed protein or sorghum protein. Preferably, the plant-based protein is selected from soy protein, pea protein, potato protein, rapeseed protein and / or rice protein. More preferably, the plant-based protein is selected from soy protein and / or pea protein.

[0033] Suitable plant-based proteins also include:

[0034] - Brassicas: including Brassica balearica: Mallorca cabbage, Brassica carinata: Abyssinian mustard or Abyssinian cabbage, Brassica elongata: elongated mustard, Brassica fruticulosa: Mediterranean cabbage, Brassica hilarionis: St Hilarion cabbage, Brassica juncea: Indian mustard, brown and leaf mustards, Sarepta mustard mustard, Brassica anapus: rapeseed, canola, rutabaga, Brassica anarinosa: broadbeaked mustard, Brassica nigra: black mustard, Brassica oleracea: cabbage, cabbage, kale, broccoli, cauliflower, kale, Brussels sprouts, kohlrabi, Brassica perviridis: tender greens, mustard spinach, Brassica rapa (synonym B. campestris): Chinese cabbage, turnip, rapini, komatsuna, Brassica rupestris: brown mustard, Brassica tournefortii): Asian mustard

[0035] - Solanaceae: includes tomatoes, potatoes, eggplants, bell peppers and hot peppers;

[0036] - Cereals: including corn, rice, wheat, barley, sorghum, millet, oats, rye, triticale, fonio

[0037] - Pseudocereals: including amaranth (millet, red amaranth, prince-of-Wales-feather), bread nuts, buckwheat, chia, cockscomb (also called quail grass or soko), pitseed goosefoot, Quinoa and wattle seeds (also called acacia seeds);

[0038] -Fabaceae: includes Acacia alata (winged acacia), Acacia decipiens, Acacia saligna (commonly known by various names including coojong, golden wreath acacia, orange acacia, blue-leaved acacia), Arachis hypogaea (peanut), Astragalus galegiformis, Cytisus laburnum (common laburnum, golden chain or golden rain), Cytisus supinus, Dolichios lablab (common names include lablab, lablab-bean bonavist bean / pea, dolichos bean, seim bean, lablabbean, Egyptian kidney bean, Indian bean, bataw and Australian bean), Lentil, Genista tinctorial (common names include dyer'swhin, waxen woad, and waxen wood), Glycine max (soybean), Lathyrus clymenum (pea or Lathyrus), Lathyrus odoratus (pea or Lathyrus), Lathyrus staivus (pea or Lathyrus), Lathyrus silvetris (pea or Lathyrus), Lotus tetragonolobus (four-winged bean or winged pea), Lupinus albus (lupin), Lupinus angustifolius (lupin), Lupinus truncatula (lupin), luteus)(Lupinus), Lupinus polyphyllus)(Lupinus), Medicagosativa(Alfalfa), Phaseolus aureus(Mung bean), Phaseolus coccineus(Runner bean), Phaseolus nanus(Green bean / French bean),Phaseolus vulgaris (Green bean / French bean), Pisum sativum (pea), Trifolium hybridum (clover), Trifolium pretense (red clover), Vicia faba (broad bean), Vicia sativa (wild pea), Vigna unguiculate (cowpea),

[0039] -Non-legumes: Includes: Acanshosicyos horrida (Acanshosicyos horrida), Aesculus hippocastanum (Horse Chestnut), Anacardium occidentale (Cashew Tree), Balanitesaegyptica, Bertholletia excels (Brazil Nut), Beta vulgaris (Beet), Brassica napus (Rapeseed), Brassica juncea (Brown Mustard), Brassicanigra (Black Mustard), Brassica hirta (Eurasian Mustard), Cannabis sativa (Cannabis), Citrus aurantiaca (Citrus), Cucurbita maxima (Pumpkin), Buckwheat (Polygonum), Gossypium barbadensis (Extra-Long Staple Cotton), Heianthus annuus (Sunflower) annuus (sunflower), Nicotiana sp. (tobacco plant), Prunus avium (cherry), Prunus cerasus (sour cherry), Prunus domestica (plum), Prunus amygdalus (almond tree), Ricinus communis (castor bean / castor oil plant), Sasamum indicum (sesame), Sinapis alba (white mustard), Terlfalrea pedata (pomace fruit).

[0040] For the avoidance of doubt, the plant-based hydrogels and structured materials according to the present invention do not include plants in their natural state, for example naturally occurring plant cells, organelles or vesicles are not the plant-based hydrogels or structured materials of the present invention.

[0041] In the method according to the present invention, the plant-based protein hydrogel is formed by adding the plant-based protein to a solvent system, wherein the solvent system comprises two or more miscible co-solvents as defined herein. By selecting a solvent system comprising miscible co-solvents, the properties of the hydrogel and the associated sol-gel conditions can be controlled, wherein the first co-solvent increases the solubility of the plant-based protein and the second co-solvent reduces the solubility of the plant-based protein.

[0042] The first co-solvent increases the solubility of the plant-based protein. The first co-solvent can be considered a solubilizing co-solvent. There can be one or more solubilizing co-solvents and the solubilizing co-solvent can fully or partially solubilize the plant-based protein.

[0043] Examples of solubilizing co-solvents are organic acids. Organic acids are organic compounds with acidity. Suitable organic acids include acetic acid, formic acid, propionic acid or alpha-hydroxy acids. Suitable organic acids include acetic acid, formic acid, propionic acid, alpha-hydroxy acids or beta-hydroxy acids. Suitable alpha-hydroxy acids include glycolic acid, lactic acid, malic acid, citric acid and tartaric acid. Suitable beta-hydroxy acids include beta-hydroxypropionic acid, beta-hydroxybutyric acid, beta-hydroxy beta-methylbutyric acid, 2-hydroxybenzoic acid and carnitine. Particularly preferred organic acids are volatile organic acids, i.e., those with a boiling point below 130°C. This is because volatile organic acids can be easily removed from the plant-based protein hydrogel (hydrogen) slurry in the subsequent drying step, so that the final plant-based structured material contains almost no (if any) residual organic acid. Examples of volatile organic acids include acetic acid and formic acid. Preferred organic acids are acetic acid and lactic acid. The use of organic acids can dissolve plant proteins and can also gently hydrolyze proteins. For example, without wishing to be bound by theory, the solubility of plant-based proteins in organic acids may be due to: i) protonation of the protein and ii) the presence of an anionic solvation layer that helps reduce hydrophobic interactions.

[0044] In a preferred process of the invention, the first co-solvent is an organic acid.

[0045] In a preferred process of the present invention, the boiling point of the first co-solvent is less than 130°C, more preferably less than 120°C.

[0046] The second co-solvent reduces the solubility of the plant-based protein compared to the first co-solvent. The second co-solvent may be considered a desolvating co-solvent. One or more desolvating co-solvents may be present.

[0047] An example of a second co-solvent for dissolving is a buffered aqueous solution. Preferably, the second co-solvent can be water, ethanol, methanol, acetone, acetonitrile, dimethyl sulfoxide, dimethylformamide, formamide, 2-propanol, 1-butanol, 1-propanol, hexanol, tert-butanol, ethyl acetate or hexafluoroisopropanol. Particularly preferably, the second co-solvent is water and / or ethanol. Most preferably, the second co-solvent is water.

[0048] In a preferred process of the invention, the second co-solvent has a boiling point of less than 130°C, more preferably less than 120°C.

[0049] In a preferred method of the present invention, the concentration of the plant-based protein in the solvent system in step (a) is 50-250 mg / ml, preferably 50-150 mg / ml. The proportion of the solubilizing co-solvent (usually an organic acid) can be varied according to the protein concentration, for example, a higher organic acid ratio is used as the protein concentration increases.

[0050] In preferred methods of the invention, the solvent system has a cosolvent ratio of the first cosolvent to the second cosolvent of about 20-80% v / v, about 20-60% v / v, about 25-55% v / v, about 30-50% v / v, about 20%, about 30%, about 40%, about 50% or about 60% v / v, most preferably about 30-50% v / v. Such ratios result in functionally useful materials.

[0051] In a preferred method of the present invention, the protein solution is heated to a first temperature above the sol-gel temperature of the one or more plant-based protein solutions and then reduced to a second temperature below the sol-gel temperature of the one or more plant-based protein solutions to form a hydrogel.

[0052] In a preferred method of the invention, the extent of protein hydrolysis (i.e., the percentage of peptide bonds cleaved in the protein hydrolysate) is controlled to alter the properties of the resulting hydrogel. For example, increasing the concentration of organic acid present during formation increases the extent of protein hydrolysis. A higher degree of protein hydrolysis results in the formation of a less rigid hydrogel.

[0053] In a preferred method of the present invention, the extent of protein hydrolysis is from 0.1 to 10%, preferably from 0.1 to 5%, more preferably from 0.1 to 2.5%.

[0054] In order to form a solution comprising one or more plant-based proteins, it may be necessary to apply a physical stimulus to the protein / solvent system mixture to dissolve the protein. Suitable physical stimuli include heating, ultrasonic treatment, stirring, high shear mixing or other physical techniques. The preferred technique is heating, optionally simultaneously or subsequently ultrasonic treatment.

[0055] Preferably, the protein / solvent system mixture is subjected to a physical stimulus of heating, wherein the solution is heated to about or above 70° C. More preferably, the protein / solvent system mixture is heated to about or above 75° C., about or above 80° C., about or above 85° C., or about 90° C. Even more preferably, the protein / solvent system mixture is heated to 85° C.

[0056] Preferably, the protein / solvent system mixture is subjected to physical stimulation, including heating for a period of about 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes or more than 30 minutes. The preferred heating period is about 30 minutes. The heated protein / solvent system mixture is optionally simultaneously or subsequently subjected to ultrasonic treatment.

[0057] The resulting protein solution is heated so that the protein solution is then maintained above the sol-gel transition of the protein solution. The sol-gel transition temperature of the protein can be changed by changing the solvent system (e.g., by selecting the choice of organic acid, the ratio of organic acid to other solvents, or by other means). By appropriate selection of conditions, the sol-gel transition of the protein can be carefully controlled, thereby controlling the formation of the hydrogel.

[0058] Preferably, the protein solution is heated to about or above 70° C. More preferably, the protein solution is heated to about or above 75° C., about or above 80° C., about or above 85° C., or about 90° C. Even more preferably, the protein solution is heated to about 85° C.

[0059] The protein solution can be maintained at an elevated temperature for a period of about 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes, 45 minutes or 1 hour. The preferred period is at least 30 minutes so that the protein can be dissolved fully. The protein solution can be maintained at an elevated temperature for a longer period of time. This can be used for commercial batch processing or for fluid handling steps, where it is necessary to keep the protein solution in liquid form for a longer period of time.

[0060] After the protein solution is heated to a temperature higher than the sol-gel transition temperature, the temperature of the protein solution can be reduced to a second temperature lower than the sol-gel transition temperature to promote the formation of a hydrogel. The second temperature can be room temperature. The second temperature can be in the range of 5 to 25°C, preferably 10 to 20°C. The protein solution can be kept at a reduced temperature for a long period of time, such as several days, weeks. The protein solution can be kept at a reduced temperature for a period of about 5 minutes, 10 minutes, 15 minutes, 20 minutes, 25 minutes or about 30 minutes. The specific reduced time period is about 5 minutes. However, the method of the present invention allows the protein to remain in the solution for a long period of time. Therefore, as needed, the protein solution can be kept above the sol-gel transition temperature as long as it is necessary to keep the protein in liquid form. This may be several hours, several days or longer, but preferably on the order of several minutes or hours. In addition, since the process is reversible, for example, the solution can be kept at a lower temperature (e.g., room temperature) at which a hydrogel will be formed, but then heated to a temperature higher than the sol-gel transition temperature to restore the solution to a liquid state for further processing. In this way, protein hydrogels can be preserved for hours, days, weeks, months or years because the hydrogels remain stable for long periods of time.

[0061] The specific temperature will depend on the properties of the protein source, the solvent conditions used and therefore the sol-gel transition temperature. Alternatively, the elevated and lowered temperatures can be relatively fixed (e.g., about 85°C, then about room temperature), and the co-solvent mixture conditions adjusted to ensure the appropriate sol-gel transition temperature for the selected plant-based protein.

[0062] Therefore, a preferred method of the present invention comprises:

[0063] (ai) forming a protein solution comprising one or more plant-based proteins and a solvent system, wherein the solvent system comprises miscible co-solvents; wherein a first co-solvent increases the solubility of the plant-based protein and a second co-solvent decreases the solubility of the plant-based protein;

[0064] (aii) subjecting the protein solution to a physical stimulus, such as heating and / or sonication, for a period of time;

[0065] (bi) raising the temperature of the protein solution to a first elevated temperature above the sol-gel transition temperature for a period of time;

[0066] (bii) reducing the temperature of the protein solution to below the sol-gel transition temperature, so that the plant-based protein self-aggregates into a plant-based protein hydrogel; and

[0067] (c) subjecting the plant-based protein hydrogel to a shearing treatment to form a plant-based protein hydrogel slurry.

[0068] The protein solution may be maintained at an elevated temperature in step (bi) while it is formed in a suitable mould, wherein the protein is allowed to form a hydrogel after the temperature is reduced in step (bii).

[0069] Without wishing to be bound by theory, it is believed that when plant proteins are added to a solvent system and subjected to physical stimuli such as heating and / or ultrasonic treatment, plant proteins partially unfold, resulting in exposure of hydrophobic amino acids that were originally buried in the native structure of the protein. Once partially unfolded, cosolvents are able to interact with the unfolded protein molecules. For example, organic acids are more likely to acquire protonated amino acid residues and are able to form anionic salt bridges that stabilize hydrophobic interactions. In addition, upon heating at elevated temperatures, non-covalent intermolecular contacts of protein-protein are destroyed.

[0070] Furthermore, it is believed that upon cooling the protein solution below the sol-gel temperature, protein-protein noncovalent intermolecular contacts are achieved, thereby promoting the self-assembly of plant protein molecules into a hydrogel of interconnected protein aggregates.

[0071] In addition, it is believed that the application of mechanical agitation, such as sonication, breaks up large colloidal protein aggregates into smaller ones, as well as disrupting the interactions between protein molecules. Using this method, the size of protein aggregates can be significantly reduced to a particle size of less than 100 nm.

[0072] Preferably, the process of the present invention produces a plant-based protein solution comprising protein aggregates having an average size of less than 200 nm, preferably less than 150 nm, less than 125 nm, less than 100 nm, less than 90 nm, less than 80 nm, less than 70 nm, less than 60 nm, less than 50 nm, less than 40 nm, or less than 30 nm.

[0073] It is believed that the method of the present invention allows plant proteins to aggregate into supramolecular structures held by intermolecular hydrogen bonding interactions, particularly between beta-strands.The method of the present invention enables the formation of materials in which high levels of beta-sheet intermolecular interactions exist.

[0074] A feature of the method of the present invention is that no cross-linking agent is required, as the plant protein will form a hydrogel on its own. Thus, in a preferred method of the present invention, the plant-based protein hydrogel contains no or substantially no cross-linking agent.

[0075] However, in an alternative preferred method of the invention, the plant-based protein hydrogel may comprise a cross-linking agent. Suitable cross-linking agents include microbial transglutaminase, glutaraldehyde, formaldehyde, glyoxal, phenolic compounds, epoxy compounds, genipin or dialdehyde starch.

[0076] In the method of the present invention, the plant-based protein hydrogel is subjected to a shearing treatment to form a plant-based protein hydrogel slurry. As will be appreciated by those skilled in the art, the shearing treatment does not include centrifugation. Without wishing to be bound by theory, it is believed that the shearing treatment changes the composition of the hydrogel so that the resulting slurry is composed of small fragments of controlled size. This means that the plant-based protein hydrogel slurry can be poured, pumped and processed. In addition, the small fragments can then be well combined together in subsequent processing, such as forming a film. The inventors of the present invention were surprised to find that, in general, the smaller the fragments of the hydrogel slurry, the better the tensile strength and optical properties of the resulting film.

[0077] The inventors have also discovered that it is desirable to control the rheological properties of the hydrogel within certain limits to aid in the processing of the hydrogel and the properties of the final film.

[0078] In a preferred method of the present invention, the shear treatment comprises a high shear step. Preferably, the high shear step involves breaking the plant-based protein hydrogel into fragments.

[0079] In a preferred method of the invention, the fragments produced in the high shear step have a d value of less than 500 nm, preferably less than 300 nm, more preferably less than 200 nm, even more preferably less than 50 nm as determined by dynamic light scattering (DLS). 50 .

[0080] In an alternative preferred process of the present invention, the fragments produced in the high shear step have a d of 0.5 to 150 microns, preferably 0.6 to 100 microns, more preferably 0.7 to 70 microns, even more preferably 0.8 to 50 microns, more preferably 0.9 to 25 microns, more preferably 1 to 20 microns, more preferably 1 to 10 microns, even more preferably 1 to 5 microns as determined by laser diffraction. 50 The inventors have surprisingly found that if the fragments are within these size ranges, the hydrogel slurry can be used to form films with improved properties such as tensile strength or clarity.

[0081] In a preferred process of the invention, the fragments produced in the high shear step have a d value as determined by laser diffraction of less than 10 microns, more preferably less than 8 microns, more preferably less than 6 microns, more preferably less than 4 microns, more preferably less than 2 microns, even more preferably less than 1 micron. 10 In a preferred process of the invention, the fragments produced in the high shear step have a d θ measured by laser diffraction of 0.05 to 10 microns, more preferably 0.1 to 8 microns, more preferably 0.2 to 6 microns. 10Without wishing to be bound by theory, it is believed that the presence of a proportion of relatively smaller sized fragments within the hydrogel slurry can help bind together relatively larger sized fragments also present therein, which means that any final structured material made using the plant-based protein hydrogel slurry (such as a film) has better mechanical properties.

[0082] Thus, in a preferred process of the invention, the fragments produced in the high shear step have a d value of from 0.5 to 150 microns as determined by laser diffraction. 50 and d of 0.05 to 10 μm as determined by laser diffraction 10 , preferably 0.6 to 100 μm d 50 and 0.1 to 8 μm d 10 , more preferably 0.7 to 70 microns d 50 and 0.2 to 6 μm d 10 .

[0083] In a preferred process of the invention, said fragments produced in said high shear step have a d5 as determined by laser diffraction of less than 3 microns, more preferably less than 1 micron, even more preferably less than 0.15 microns.

[0084] In a preferred method of the invention, the high shear step involves ultrasonic treatment (for example using equipment such as Bandelin HD4200, TS 113 probe or Hielscher UIP1000hdT), high shear mechanical stirring (for example using equipment such as Silverson rotor-stator high shear mixer) or cavitation, preferably ultrasonic treatment.

[0085] In a preferred method of the invention, the high shear step involves ultrasonic treatment (for example using equipment such as Bandelin HD4200, TS 113 probe or Hielscher UIP1000hdT), high shear mechanical stirring (for example using equipment such as Silverson rotor-stator high shear mixer), high pressure homogenization or cavitation, preferably ultrasonic treatment.

[0086] In a preferred process of the invention, the high shear step is carried out at a temperature below the sol-gel temperature of the plant-based protein solution. In a preferred process of the invention, the high shear step is carried out for a duration of at least 5 minutes, more preferably at least 1 minute.

[0087] In a preferred method of the invention, the shear treatment comprises two steps. Preferably, the shear treatment comprises a low shear step followed by a high shear step.

[0088] In a preferred method of the present invention, the low shear step involves breaking the plant-based protein hydrogel into fragments. Preferably, at least 50 wt% of the fragments produced in the low shear step have a particle size of 1 mm to 50 mm, preferably 5 mm to 30 mm, more preferably 10 mm to 30 mm. More preferably, at least 80 wt% of the fragments produced in the low shear step have a particle size of 1 mm to 50 mm, preferably 5 mm to 30 mm, more preferably 10 mm to 30 mm. This can be measured by collecting the fragments on a series of sieves with decreasing mesh sizes and weighing the amount on the different sieves.

[0089] The low shear step is carried out at a temperature below the sol-gel temperature of the plant-based protein solution.

[0090] In a preferred method of the present invention, the low shear step comprises mechanical cutting. Mechanical cutting refers to cutting using a blade (eg, a knife, an extruder blade, etc.).

[0091] In an alternative preferred method of the present invention, the low shear step comprises extrusion. For example, the plant-based protein solution formed in step (a) can be extruded into a non-solubilizing solvent (e.g., water) to form a plant-based protein hydrogel of large discrete fragments, for example, large discrete fragments can take the form of an extrudate having a linear or ribbon-like form. In this way, the fragments can be directly subjected to a solvent reduction step, as described in more detail below. A low shear step of this nature is more suitable for large-scale processing. In this case, the low shear step can reduce at least one dimension of the large fragments to between 1 mm and 50 mm, such as the diameter of the extrudate. Preferably, at least 50 wt% of the fragments produced in the low shear step have at least one internal dimension in the range of 1 mm to 50 mm, preferably 5 mm to 30 mm, more preferably 10 mm to 30 mm. More preferably, at least 80 wt% of the fragments produced in the low shear step have at least one internal dimension in the range of 1 mm to 50 mm, preferably 5 mm to 30 mm, more preferably 10 mm to 30 mm.

[0092] In a preferred method of the present invention, the high shear step comprises further fragmenting the plant-based protein hydrogel. Preferably, the fragments produced in the high shear step have a d value of less than 500 nm, preferably less than 300 nm, more preferably less than 200 nm, even more preferably less than 50 nm as measured by dynamic light scattering. 50 Alternatively, the fragments produced in the high shear step have a d as determined by laser diffraction of 0.5 to 150 microns, preferably 0.6 to 100 microns, more preferably 0.7 to 70 microns, even more preferably 0.8 to 50 microns, more preferably 0.9 to 25 microns, more preferably 1 to 20 microns, more preferably 1 to 10 microns, even more preferably 1 to 5 microns.50 The inventors have surprisingly found that if the fragments are within these particle size ranges, the hydrogel slurry can be used to form films with improved properties such as tensile strength.

[0093] In a preferred process of the invention, the fragments produced in the high shear step have a d value as determined by laser diffraction of less than 10 microns, more preferably less than 8 microns, more preferably less than 6 microns, more preferably less than 4 microns, more preferably less than 2 microns, even more preferably less than 1 micron. 10 In a preferred process of the invention, the fragments produced in the high shear step have a d θ measured by laser diffraction of 0.05 to 10 microns, more preferably 0.1 to 8 microns, more preferably 0.2 to 6 microns. 10 Without wishing to be bound by theory, it is believed that the presence of a proportion of relatively smaller sized fragments within the hydrogel slurry can help bind together relatively larger sized fragments also present therein, which means that any final structured material made using the plant-based protein hydrogel slurry (such as a film) has better mechanical properties.

[0094] Thus, in a preferred process of the invention, the fragments produced in the high shear step have a d value of from 0.5 to 150 microns as determined by laser diffraction. 50 and d of 0.05 to 10 μm as determined by laser diffraction 10 , preferably 0.6 to 100 μm d 50 and 0.1 to 8 μm d 10 , more preferably 0.7 to 70 microns d 50 and 0.2 to 6 μm d 10 .

[0095] In a preferred process of the invention, said fragments produced in said high shear step have a d5 as determined by laser diffraction of less than 3 microns, more preferably less than 1 micron, even more preferably less than 0.15 microns.

[0096] In a preferred method of the present invention, the particle size distribution of the hydrogel fragments in the plant-based protein hydrogel slurry can be adjusted by changing the nature and intensity of the high shear step. In another preferred method, the particle size distribution of the hydrogel fragments in the plant-based protein hydrogel slurry can be adjusted by mixing or combining two or more different hydrogel slurries that have undergone different high shear steps and have different particle size distributions.

[0097] In a preferred process of the present invention, the high shear step is carried out at a temperature below the sol-gel temperature of the vegetable based protein solution.

[0098] In a preferred method of the present invention, the high shear step is carried out at a temperature below the protein denaturation temperature of the plant-based protein solution.

[0099] The high shear step is carried out for a duration of at least 5 minutes, more preferably at least 1 minute.

[0100] In a preferred method of the invention, the high shear step involves ultrasonic treatment (for example using equipment such as Bandelin HD4200, TS 113 probe or Hielscher UIP1000hdT), high shear mechanical stirring (for example using equipment such as Silverson rotor-stator high shear mixer) or cavitation, preferably ultrasonic treatment.

[0101] In a preferred method of the invention, the high shear step involves ultrasonic treatment (for example using equipment such as Bandelin HD4200, TS 113 probe or Hielscher UIP1000hdT), high shear mechanical stirring (for example using equipment such as Silverson rotor-stator high shear mixer), high pressure homogenization or cavitation, preferably ultrasonic treatment.

[0102] In a preferred method of the present invention, step (c) further comprises subjecting the plant-based protein hydrogel slurry to a solvent reduction step, most preferably a solubilizing solvent reduction step, between said low shear step and said high shear step.

[0103] The solubilizing solvent refers to a solvent or a mixture of solvents in which the plant-based protein hydrogel slurry can be dissolved. Examples include organic acids such as acetic acid, formic acid, propionic acid and / or α-hydroxy acids; Examples include organic acids such as acetic acid, formic acid, propionic acid, α-hydroxy acids and / or β-hydroxy acids. The α-hydroxy acid may preferably be selected from glycolic acid, lactic acid, malic acid, citric acid and / or tartaric acid. The β-hydroxy acid may preferably be selected from β-hydroxypropionic acid, β-hydroxybutyric acid, β-hydroxy β-methylbutyric acid, 2-hydroxybenzoic acid and carnitine.

[0104] In a preferred method of the present invention, the solvent reduction step comprises the following steps:

[0105] (i) contacting the fragments of the plant-based hydrogel slurry with a non-solubilizing solvent;

[0106] (ii) separating the fragments of the plant-based hydrogel slurry from the non-solubilizing solvent to obtain a washed plant-based protein hydrogel; and

[0107] (iii) optionally repeating steps (i) and (ii).

[0108] Step (i) comprises contacting the fragments of the plant-based protein hydrogel slurry with a non-solubilizing solvent. A non-solubilizing solvent refers to a solvent or a mixture of solvents in which the plant-based protein hydrogel slurry is not soluble. Examples include water or a mixture of water and ethanol.

[0109] In a preferred method of the invention, step (ii) involves screen filtration. More preferably, screen filtration involves the use of a plurality of screens of decreasing size.

[0110] As will be appreciated by those skilled in the art, if the fragments produced in the low shear step are too small, the solvent reduction step may prove difficult as the fragments will eventually clog the mesh. However, if the fragments produced in the low shear step are too large, the solvent reduction step may take an excessive amount of time as the mass transfer of solvent from the core of the fragments is slow.

[0111] Without wishing to be bound by theory, it is believed that due to the porous nature of the hydrogel, the solvent reduction step may remove some or all of the solvent (eg, organic acid) from the hydrogel by solvent exchange.

[0112] It is important that the hydrogel fragments are fragile and deformable enough, and small enough to bond together well during subsequent processing to form a final film or other structured material. If the hydrogel fragments cannot deform sufficiently, the strength and integrity of any film or structured material will be reduced. In addition, strong hydrogels are more difficult to disperse to form a slurry.

[0113] It is also important that the hydrogel fragments are not too soft and deformable. Too soft can make any intermediate processing steps (such as washing and solvent exchange) difficult. Overly soft hydrogel fragments are often caused by insufficient levels of preformed macrostructure in the hydrogel, which usually results in weaker films or structured materials.

[0114] The strength of protein hydrogels can be altered by changing the concentrations of protein and organic acid, as well as other variables.

[0115] Therefore, it is important that the strength of the hydrogel used to form the hydrogel slurry is within certain limits. This can be measured by an oscillatory rheometer. A suitable measure of hydrogel strength is the storage modulus G' of the hydrogel. Suitable test conditions are 1% strain applied at an oscillation frequency of 1 Hz at 20°C. A suitable apparatus is an Anton Paar MCR 92 rheometer with a 50 mm diameter, 1 degree cone and plate measurement geometry.

[0116] Therefore, in a preferred method of the present invention, before washing, the plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of greater than 1000 Pa, preferably greater than 2000 Pa, more preferably greater than 5000 Pa, even more preferably greater than 6000 Pa, and most preferably greater than 8000 Pa. As will be appreciated by those skilled in the art, 2πrad / s is equal to 1 Hz.

[0117] In a preferred method of the invention, the plant-based protein hydrogel has a storage modulus (G') of less than 20,000 Pa, preferably less than 15,000 Pa, more preferably less than 10,000 Pa at 10 rad / s before washing.

[0118] Furthermore, in a preferred method of the present invention, the washed plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of greater than 500 Pa, preferably greater than 1000 Pa, more preferably greater than 2500 Pa, even more preferably greater than 3000 Pa, and most preferably greater than 4000 Pa.

[0119] In a preferred method of the present invention, the washed plant-based protein hydrogel has a storage modulus (G') of less than 20,000 Pa, preferably less than 15,000 Pa, more preferably less than 10,000 Pa at 10 rad / s.

[0120] The preferred method of the present invention further comprises the following steps:

[0121] (d) Changing the pH of the plant-based protein hydrogel slurry.

[0122] In a preferred method of the present invention, step (d) is performed after step (c). In an alternative preferred method of the present invention, step (d) is performed sequentially with step (c).

[0123] When adjusting the pH of the plant-based protein hydrogel slurry, there is a possibility that the slurry passes through the isoelectric point of the protein. Due to the lack of charge repulsion at the isoelectric point, the dispersed protein fragments in the plant-based protein hydrogel slurry can quickly agglomerate. To avoid this, pH adjusting materials can be used to quickly change the pH, thereby minimizing the time that the slurry is at the isoelectric point.

[0124] Therefore, in a preferred method of the present invention, step (d) involves adding a pH adjusting material to the plant-based protein hydrogel slurry. Preferably, the pH adjusting material is an aqueous solution containing monovalent metal ions, divalent metal ions such as calcium or ammonium ions, preferably an alkaline aqueous solution containing monovalent metal ions, divalent metal ions or ammonium ions. More preferably, the pH adjusting material is an aqueous hydroxide solution, such as sodium hydroxide, potassium hydroxide or ammonium hydroxide.

[0125] In a preferred method of the present invention, the pH of the plant-based protein hydrogel slurry after step (d) is at least 1 pH unit below the isoelectric point of the plant-based protein.

[0126] In a preferred method of the present invention, the pH of the plant-based protein hydrogel slurry after step (d) is at least 1 pH unit higher than the isoelectric point of the plant-based protein. The preferred method of the present invention further comprises adding additional ingredients to the plant-based protein hydrogel slurry. Preferably, the additional ingredients are selected from plasticizers, sunscreens, preservatives, pigments and other inorganic nanoparticles (e.g., clay) or mixtures thereof.

[0127] Particularly preferably, the additional ingredient is a plasticizer. In a preferred method of the invention, the plasticizer is selected from ethylene glycol, diethylene glycol, triethylene glycol, tetraethylene glycol, polyethylene glycol, propylene glycol, sorbitol, mannitol, xylitol, lactic acid, glycolic acid, triethyl citrate, fatty acids, glucose, mannose, fructose, sucrose, ethanolamine, urea, triethanolamine, vegetable oils, lecithin, waxes and amino acids.

[0128] Particularly preferably, the additional ingredient is a plasticizer. In a preferred method of the invention, the plasticizer is selected from glycerol, ethylene glycol, diethylene glycol, triethylene glycol, tetraethylene glycol, polyethylene glycol, propylene glycol, sorbitol, mannitol, xylitol, lactic acid, citric acid, glycolic acid, triethyl citrate, fatty acids, glucose, mannose, fructose, sucrose, ethanolamine, urea, triethanolamine, vegetable oils, lecithin, waxes and amino acids.

[0129] The amount of plasticizer to be incorporated will depend on the use of the final material, such as a film. Preferably, the plant-based protein hydrogel slurry may include about 1% wt of plasticizer, based on the total weight of the protein present in the plant-based protein hydrogel slurry, about 2% wt, about 5% wt, about 10% wt, about 20% wt, about 30% wt, about 40% wt, about 50% wt, about 60% wt or more. More preferably, based on the total weight of the protein present in the plant-based protein hydrogel slurry, the plant-based protein hydrogel slurry may include about 5-50% wt plasticizer, about 10-50% wt, about 20-40% wt, about 15-35% wt or about 20% wt plasticizer.

[0130] The addition of plasticizers affects the mechanical properties of a material. Typically, adding plasticizers increases the elasticity of the material, but this often reduces the strength of the resulting material, such as tensile strength.

[0131] Composite films with improved physical and / or barrier properties to particles can be formed by adding inorganic particles such as clay flakes to the protein hydrogel slurry.

[0132] In the preferred method of the present invention, the plant-based protein hydrogel slurry is heated for 50 seconds. -1 The viscosity is in the range of 10 to 10000 cps, preferably in the range of 10 to 8000 cps, preferably in the range of 12 to 6000 cps, preferably in the range of 15 to 5000 cps, preferably 20 to 2000 cps, such as in 50s -1 The viscosity is measured by an Anton Paar MCR 92 rheometer using a plate and cone measuring geometry with a 50 mm plate and a 1° angle. Control of the viscosity facilitates the subsequent processing of the plant-based protein hydrogel slurry. For example, if the viscosity is too high, it becomes more difficult to form a film or coating using the slurry, such as when spreading the slurry to form a film. If the viscosity is too low, it may also be difficult to form structured objects, such as films, because the slurry spreads too easily. Unless otherwise stated, the viscosity is measured at 20°C.

[0133] In a preferred method of the present invention, the plant-based protein hydrogel slurry has a protein solids content of 5 wt% to 25 wt% based on the total weight of the plant-based protein hydrogel slurry, preferably 6 wt% to 20 wt% based on the total weight of the plant-based protein hydrogel slurry, more preferably 7 wt% to 15 wt%, more preferably 7.5 wt% to 12.5 wt%. The protein level affects the rheology of the hydrogel. Hydrogels containing less than 5 wt% protein solids content are generally not strong enough to provide processable fragments. Although hydrogels containing greater than 25 wt% protein solids content are stronger, they are more difficult to disperse and have been found to form weaker structured materials (e.g., films).

[0134] The plant-based hydrogel slurries described herein allow for the formation of a range of useful plant-based biomaterials. The use of plant-based materials has many advantages over previously used animal or petrochemical sources. First, plant resources are renewable and can be efficiently obtained in an environmentally friendly manner. Second, plant resources are biodegradable and are therefore an environmentally friendly alternative to other plastics. Third, plant-based proteins have a significant advantage over animal-derived proteins in that they do not introduce animal-derived proteins into the human body. This has positive implications from both a pharmacological and pharmaceutical perspective, as materials of animal origin must undergo rigorous inspection and processing to ensure that no adverse factors are present (e.g., removal of prions, etc.); and also because these products are suitable for vegetarians / strict vegetarians.

[0135] Since plant-based proteins are naturally present in the human (or other animal) diet, the biomaterials prepared according to the invention exhibit a higher degree of digestibility than other biopolymers such as polysaccharides (e.g., alginate or chitosan). This makes them particularly suitable for pharmaceutical, food and / or cosmetic applications.

[0136] Preferably, the plant-based hydrogel slurry of the present invention can be used to form a film, such as a thin film. Plant protein-derived films have many applications, including forming biodegradable flexible films for food packaging applications.

[0137] The advantage of the plant-based materials of the present invention over animal-based materials or starch-based / cellulose materials is their inherent water insolubility. Most biopolymer films are easily soluble in water and therefore cannot be used alone for food packaging applications and require additional synthetic polymer coatings. These problems can be overcome with the present invention. The films of the present invention are also soluble in alkaline conditions, or in the presence of proteolytic enzymes, or in the presence of chaotropic agents.

[0138] The present invention also provides a plant-based protein hydrogel slurry prepared according to the method.

[0139] The present invention also provides a method for preparing a plant-based structured material, the method comprising:

[0140] (a) preparing a plant-based protein hydrogel slurry according to the above method; and

[0141] (b) subjecting the plant-based protein hydrogel slurry to one or more solvent level reduction steps to reduce the level of the first co-solvent and / or the second co-solvent to obtain the plant-based structuring material.

[0142] In preferred methods of the invention, one or more solvent level reduction steps reduce the level of a first co-solvent (eg, an organic acid).

[0143] In preferred methods of the invention, one or more solvent level reduction steps reduce the level of a second co-solvent (eg an alcohol such as ethanol).

[0144] In a preferred method of the invention, step (b) involves placing the plant-based protein hydrogel slurry on a surface prior to performing one or more solvent level reducing steps.

[0145] In a preferred method of the invention, the solvent level reducing step comprises heating. Preferably, the solvent level reducing step comprises heating at a temperature in the range of 50°C to 100°C, more preferably at a temperature in the range of 55°C to 95°C. In such a solvent level reducing step, the solvent is thus removed by evaporation.

[0146] In a preferred method of the invention, the solvent level reduction step involves forced convection of dry air.

[0147] In a preferred method of the invention, the plant-based structuring material is a film.

[0148] In an alternative preferred method of the invention, the plant-based structural material is a casting (ie a plant-based structural material formed by moulding, preferably non-thermo-reversible moulding, more preferably injection moulding).

[0149] In an alternative preferred method of the invention, the plant-based structuring material is a coating. Preferably, the coating is a food coating, a seed coating, a pharmaceutical coating or a surface coating (eg a paper coating).

[0150] The coatings of the present invention are fully biodegradable, thus providing an environmentally friendly alternative to conventional coatings made from synthetic materials (e.g., chemically modified natural polymers or fossil fuel-derived polymers). For example, the food coating of the present invention provides a fully biodegradable coating that meets food standards and can extend the shelf life of the coated food. The drug coating of the present invention provides a fully biodegradable coating that can be used to replace traditional enteric coatings and can mask any unpleasant taste associated with the coated drug ingredient.

[0151] In a preferred method of the present invention, the plant-based structuring material comprises a plant-based protein having a secondary structure, wherein the secondary structure has at least 40% intermolecular β-sheets, at least 50% intermolecular β-sheets, at least 60% intermolecular β-sheets, at least 70% intermolecular β-sheets, at least 80% intermolecular β-sheets, or at least 90% intermolecular β-sheets. Preferably, the % intermolecular β-sheet content is measured by FTIR.

[0152] In a preferred method of the invention, the plant-based structural material has a Young's modulus greater than 20 MPa; preferably greater than 50 MPa, greater than 80 MPa, greater than 100 MPa, greater than 200 MPa, greater than 300 MPa, greater than 400 MPa, greater than 500 MPa or greater than 600 MPa. Young's modulus is a measure of the strength of a structured article.

[0153] In a preferred method of the invention, the plant-based structuring material is a film.

[0154] Preferably, the thickness of the film is in the range of 1 to 1000 μm, preferably 10 to 150 μm, more preferably 20 to 100 μm, even more preferably 30 to 70 μm, most preferably 35 to 60 μm. This can be measured with a micrometer.

[0155] Preferably, the film has a percent elongation at break of 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 100% or more, or more.

[0156] The films produced according to the method of the invention can be micropatterned with features ranging from 100 nm to 1000 μm, thereby achieving new functional properties such as: superhydrophobicity (lotus effect) or structural color (due to Mie scattering).

[0157] Functional composite membranes can be produced by embedding inorganic nanoparticles (e.g., gold or silver nanoparticles) into a protein matrix. Applications may include materials suitable for flexible electronics or membranes with antimicrobial properties. Composite membranes with improved physical and / or barrier particles can be formed by embedding particles such as clay flakes into a protein matrix.

[0158] The present invention also provides a plant-based structured material prepared according to the above method.

[0159] The preferred plant-based structured material according to the present invention is a film, a casting or a coating. Preferably, the coating is a food coating, a seed coating, a drug coating or a surface coating (e.g., a paper coating). The preferred properties of the plant-based structured material of the present invention are as described above. The plant-based structured material of the present invention can be used in a variety of applications, including food, beverages, cosmetics, formulations (e.g., coatings), and packaging.

[0160] The clarity and high strength of the films of the present invention make them particularly suitable for packaging applications.

[0161] The present invention also provides the use of the plant-based protein hydrogel slurry as defined above to produce a plant-based structured material. Preferably, the plant-based structured material is a film, a casting, a molding or a coating. Preferably, the coating is a food coating, a seed coating, a drug coating or a surface coating (e.g. a paper coating).

[0162] The present invention also provides a plant-based protein hydrogel slurry, wherein the protein solid content is 5wt% to 25wt% based on the total weight of the plant-based protein hydrogel slurry, and the solid content of the protein is 5wt% to 25wt% based on the total weight of the plant-based protein hydrogel slurry. -1 and a viscosity at 20° C. in the range of 10 to 10,000 cps, wherein the plant-based protein hydrogel slurry comprises a d 50 Particle sizes range from 0.5 to 150 microns.

[0163] The preferred plant-based protein hydrogel slurry of the present invention comprises a d 50 Particle sizes of fragments are from 0.6 to 100 microns, more preferably from 0.7 to 70 microns, even more preferably from 0.8 to 50 microns, more preferably from 0.9 to 25 microns, more preferably from 1 to 20 microns, more preferably from 1 to 10 microns, even more preferably from 1 to 5 microns.

[0164] The preferred plant-based protein hydrogel slurry of the present invention has a protein solid content of 6 wt % to 20 wt %, more preferably 7 wt % to 15 wt %, even more preferably 7.5 wt % to 12.5 wt %, based on the total weight of the plant-based protein hydrogel slurry.

[0165] Preferably, the viscosity of the plant-based protein hydrogel slurry is measured by an Anton Paar MCR 92 rheometer using a plate and cone measurement geometry with a 50 mm plate and a 1° angle at 50 s. -1 The preferred plant-based protein hydrogel slurry of the present invention is measured at 50s -1 and a viscosity range of 10 to 8000 cps at 20°C, more preferably within 50s -1 and 12 to 6000 cps at 20°C, more preferably at 50s -1 and 15 to 5,000cps at 20°C.

[0166] The present invention also provides a plant-based protein hydrogel slurry, wherein the protein solid content of the plant-based protein hydrogel slurry is 5wt% to 25wt% of the total weight of the plant-based protein hydrogel slurry, and the solid content of the protein of the plant-based protein hydrogel slurry is 5wt% to 25wt% of the total weight of the plant-based protein hydrogel slurry. -1 and a viscosity at 20° C. in the range of 10 to 10,000 cps, wherein the plant-based protein hydrogel slurry comprises a d 50 Particle size less than 500nm.

[0167] The preferred plant-based protein hydrogel slurry of the present invention comprises a d 50 Fragments having a particle size of less than 300 nm, more preferably less than 200 nm, even more preferably less than 50 nm.

[0168] The preferred plant-based protein hydrogel slurry of the present invention has a protein solid content of 6 wt % to 20 wt %, more preferably 7 wt % to 15 wt %, even more preferably 7.5 wt % to 12.5 wt %, based on the total weight of the plant-based protein hydrogel slurry.

[0169] Preferably, the viscosity of the plant-based protein hydrogel slurry is measured by an Anton Paar MCR 92 rheometer using a plate and cone measurement geometry with a 50 mm plate and a 1° angle at 50 s. -1 The preferred plant-based protein hydrogel slurry of the present invention is measured at 50s -1 and a viscosity range of 10 to 8000 cps at 20°C, more preferably within 50s -1 and 12 to 6000 cps at 20°C, more preferably at 50s -1 and 15 to 5,000cps at 20°C.

[0170] The present invention also provides the use of the plant-based protein hydrogel slurry as defined above to produce a plant-based structured material. Preferably, the plant-based structured material is a film, a casting, a molding or a coating. Preferably, the coating is a food coating, a seed coating, a drug coating or a surface coating (e.g. a paper coating).

[0171] The present invention provides a film comprising the plant-based protein hydrogel slurry as described above.

[0172] Preferred films of the present invention:

[0173] (a) comprises a plant-based protein having a secondary structure having at least 40% intermolecular beta-sheets, at least 50% intermolecular beta-sheets, at least 60% intermolecular beta-sheets, at least 70% intermolecular beta-sheets, at least 80% intermolecular beta-sheets, or at least 90% intermolecular beta-sheets, preferably wherein the % intermolecular beta-sheet content is measured by FTIR; and / or

[0174] (b) having a tensile strength of 4 to 25 MPa, preferably 6 to 15 MPa, more preferably 8 to 12 MPa; and / or

[0175] (c) having an elongation at break percentage of 10% or more, preferably 20% or more, preferably 30% or more, preferably 40% or more, preferably 50% or more, preferably 60% or more, preferably 70% or more, preferably 80% or more, preferably 90% or more, preferably 100% or more or more.

[0176] The present invention also provides a film obtained from the plant-based protein hydrogel slurry described above. For example, the film can be obtained by subjecting the plant-based protein hydrogel slurry to one or more solvent level reduction steps. Preferred features of the solvent level reduction step are as described above. Industrial methods for producing structured objects such as films from hydrogel slurries include casting, in which the hydrogel slurry is poured or extruded in a carefully controlled manner onto a moving surface such as a belt or drum and subjected to controlled drying conditions. BRIEF DESCRIPTION OF THE DRAWINGS

[0177] Figure 1 A transmission electron microscope (TEM) image of the diluted pea protein-based slurry prepared in Example 1 is shown.

[0178] Figure 2a is a photograph showing the transparency of the film prepared in Example 2, Figure 2b is a photograph showing the transparency of the film prepared in Example 3.

[0179] Figure 3Shown is a scanning electron microscope (SEM) image of a coating on a paper substrate prepared from a pea protein based slurry as described in Example 5. Also shown is a portion of an uncoated paper.

[0180] Figure 4 a shows a scanning electron microscope (SEM) image of a coating on strawberries prepared from a pea protein-based slurry as described in Example 6, Figure 4 b shows uncoated strawberries.

[0181] Figure 5 a shows a scanning electron microscope (SEM) image of the coating layer on the acetaminophen tablet prepared from the pea protein-based slurry as described in Example 7, Figure 5 b shows the uncoated tablet.

[0182] Figure 6 a shows a scanning electron microscope (SEM) image of a coating on wheat seeds prepared from a pea protein based slurry as described in Example 8, Figure 6 b shows uncoated seeds. Example

[0183] Material

[0184] Pea protein isolate (PPI) (80% protein) was purchased from Cambridge Commodities Ltd.

[0185] Soy protein isolate was purchased from Cambridge Commodities Ltd.

[0186] Lactic acid (food grade, ≥80%) was purchased from Cambridge Commodities Ltd.

[0187] Acetic acid (glacial) was purchased from Fisher Scientific.

[0188] Measurement method

[0189] Viscosity measurements were performed using an Anton Paar MCR 92 rheometer using a plate and cone measurement geometry with a 50 mm plate and 1 degree angle, at a constant shear of 50 s at 20 °C. -1 .

[0190] Storage modulus (G') can be measured using an Anton Paar MCR 92 rheometer using a plate and cone measurement geometry with a 50 mm plate and a 1 degree angle, with a strain of 1% at an oscillation frequency of 1 Hz.

[0191] Particle size measurements were performed using either dynamic light scattering (DLS) techniques or laser diffraction techniques. DLS measurements were performed using a Zeta Sizer Nano S from Malvern Panalytical and were operated according to the manufacturer's instructions. It is important that the slurry is sufficiently diluted to avoid misleading results due to agglomeration of particles during testing. The hydrogel slurry was diluted 100 times with deionized water. It is also important that the pH of the sample be away from the isoelectric point of the sample to avoid misleading results due to agglomeration. The isoelectric point of the pea protein isolate material tested here was 4.5 and the pH of the slurry was adjusted to 3 with acetic or lactic acid before measurement. Typically, multiple dilutions and / or buffers should be tested to ensure proper dispersion. 200 μL of the diluted slurry was placed in a cuvette and placed in the equipment. The test was then performed according to the standard equipment procedure. The measured d 50 For weight / volume distribution. DLS can generally be used to measure particle sizes up to 500 nm. The upper limit is mainly determined by the onset of sedimentation. For particles larger than this, particle size measurements are performed using laser diffraction with an Anton Paar PSA 1190. The measurement is performed by diluting the protein slurry in an aqueous solution of acetic or lactic acid adjusted to the same pH. The slurry is diluted to the desired concentration to obtain the optical density required for the measurement (usually 5-15% obscuration). The referenced d 50 For volume distribution. 10 and d5 values ​​can also be obtained in this way using laser diffraction.

[0192] Other devices can be used, such as the NANO-flex from Colloid Metrix system.

[0193] The particle size of the hydrogel fragments from the low shear step can be determined by sieving. A suitable technique is to take 200 grams of the hydrogel mixture from the low shear step and quickly disperse it in 1000 ml of deionized water. The dispersed mixture is then quickly poured into a series of stacked sieves with decreasing mesh sizes between 50 mm and 1 mm. The sieves from Endecotts are suitable. The percentage of slurry in a specific size range can be calculated by combining the weights of the fragments on the different sieves and calculating them as a percentage of the total amount of slurry. Due to the short test cycle, the error caused by any additional solvent changes is minimal.

[0194] Transmission electron microscopy (TEM) measurements were performed using a Thermo Scientific FEI Talos F200X G2 TEM. A suitable technique is to prepare the test sample by diluting the fine plant-based hydrogel slurry to a 1:100 dilution with a 3% acetic acid solution and depositing the sample on a TEM grid (C400Cu, EM resolution) and then staining with uranyl acetate. The maximum fragment size is then determined by optical inspection of at least 30 fragments randomly selected from the field of view of the test sample. The maximum length is the maximum length of a line drawn between any two opposite boundaries of a fragment that does not cross any external boundary.

[0195] Scanning electron microscopy (SEM) images were taken using a MIRA 3FEG-SEM, TESCAN with 10 nm platinum coating.

[0196] Structural analysis was performed using an FTIR-Equinox 55 spectrometer (Bruker). The samples were used without further pretreatment and loaded into the FTIR holder. The atmospheric compensation spectrum was subtracted from the original FTIR spectrum, and the second-order derivative was applied for further analysis. Each FTIR measurement was repeated 3 times. The instrument detection sensitivity was 5%. In order to resolve the transition of the native structure of pea protein isolate to supramolecular aggregates, the vibration changes of amide I, which is closely related to the secondary structure of the protein, were tracked. The structural analysis was performed in solution (e.g. directly using a plant-based protein hydrogel slurry) or on the resulting dry film. In the latter case, the membrane sample for structural analysis was prepared by drying 200 μL of the relevant plant-based protein hydrogel slurry at 37 ° C for 6 hours.

[0197] Young's modulus and tensile strength of structured objects such as membranes can be measured using Tinius Olsen's 5ST electromechanical universal tester. A 10 cm by 1 cm strip is placed between the clamps and extended at a rate of 12.5 mm / min, and the force and extension are recorded. The thickness of the membrane before testing can be measured with a micrometer.

[0198] The light transmittance of structured objects (e.g., films) can be measured using a Cary 500 UV-Vis spectrometer. The measurements were performed in the wavelength range of 300 to 800 nm at a scan speed of 600 nm / min with an averaging time of 0.1 sec.

[0199] Example 1: Preparation of pea protein gel structured objects

[0200] (a) Protein hydrogel formation

[0201] 500 ml of a mixture consisting of 12.5% ​​(w / v) pea protein isolate in 40% (v / v) lactic acid solution was prepared.

[0202] The mixture was then heated in a water bath at 80° C. for 30 minutes and then subjected to a brief ultrasound step to disrupt large colloidal aggregates (Hielscher UIP1000hdT (1000 W, 20 kHz)), after which a clear solution was obtained. The energy applied was 16 Wh over 7 minutes.

[0203] The solution was then poured into a 220 mm Petri dish and cooled at 10°C for 72 hours to obtain a self-standing protein hydrogel.

[0204] (b) Application of shear in protein hydrogels

[0205] Shear was then applied to the hydrogel formed in step (a) as follows. The protein hydrogel was cut into approximately 1 cm cubes by a low shear cutting step. The cubes were placed in a 75 μm filter bag, which was then immersed in a bucket containing 5 L of deionized water. This formed a crude protein hydrogel slurry within the filter bag. The hydrogel cubes were soaked for 1 hour with occasional gentle stirring. This step was performed to reduce the concentration of lactic acid in the hydrogel by diffusion into the continuous aqueous phase, and was repeated 5 more times until the final pH of the aqueous solution was 3.28.

[0206] The strained gel cubes were transferred to 500 ml bottles and probe sonicated for 10 min in a high shear step (~0.2 kJ / ml) to form a homogenous low viscosity protein dispersion. The viscosity of the slurry was 12 cps. The d 50 The size is 90nm.

[0207] (c) Preparation of pea protein film

[0208] The fine hydrogel slurry prepared in step (b) was adjusted to pH 2.6 by adding a small amount of lactic acid, then tilted onto a heated surface (maintained at 80° C.) and dried for 1 hour to form a structured object as a film with an average thickness of 18.1 μm. The resulting film was transparent and had a Young's modulus of 361 MPa and a tensile strength of 14 MPa.

[0209] Example 2: Preparation of pea protein gel structured objects

[0210] (a) Protein hydrogel formation

[0211] 800 ml of a mixture consisting of 10% (w / v) pea protein isolate in 40% (v / v) acetic acid solution was prepared.

[0212] The mixture was then heated in a water bath at 85° C. for 20 minutes and then subjected to a brief ultrasound step to disrupt large colloidal aggregates (Hielscher UIP500hdT (500 W, 20 kHz)), after which a clear solution was obtained. The energy applied was 200 KJ over 30 minutes.

[0213] The solution was poured into two 220 mm Petri dishes. The Petri dishes were sealed and cooled by storing in a refrigerator at 4°C for 20 hours to obtain a self-supporting protein hydrogel. The storage modulus of this hydrogel was 2640 Pa.

[0214] (b) Application of shear in protein hydrogels

[0215] Shear was then applied to the hydrogel formed in step (a) as follows. The protein hydrogel was cut into approximately 1 cm cubes by a low shear cutting step. The cubes were placed in a 75 μm filter bag, which was then immersed in a bucket containing 7 L of deionized water. This formed a crude protein hydrogel slurry within the filter bag. The hydrogel cubes were soaked for 1.5 hours with occasional gentle stirring. This step was performed to reduce the concentration of acetic acid in the hydrogel by diffusion into the continuous aqueous phase, and was repeated 1 more time until the final pH of the aqueous solution was 3.1.

[0216] The strained gel cubes were transferred to a 1 L bottle and subjected to high shear using a rotor stator (15,000 rpm for 5 min) and probe sonication on ice for 30 min (~0.5 kJ / ml) to form a homogenous low-viscosity protein dispersion. The viscosity of the slurry was 23 cps. The d 50 The size is 103nm.

[0217] (c) Preparation of pea protein film

[0218] The fine hydrogel slurry prepared in step (b) was mixed with 20 w / w% glycerol, poured onto a plastic petri dish, and dried at room temperature for 24 hours to form a structured object in the form of a film with an average thickness of 78.5 μm. The PSD and optical properties of the pea protein film of Example 2 were studied. The results are shown in Table 1. The resulting film was transparent, as shown in FIG. Figure 2a As shown, its transmittance at 600 nm is 81.1%.

[0219]

[0220] Table 1

[0221] The results show that functional films with high transparency can be formed using the plant-based protein hydrogel slurry of the present invention. Without wishing to be bound by theory, it is believed that achieving a controlled particle size distribution allows for a high level of transparency to be observed.

[0222] Example 3: Preparation of soy protein gel structured objects

[0223] (a) Protein hydrogel formation

[0224] 430 grams of a mixture consisting of 7.0% (w / w) soy protein isolate in 30% (v / v) acetic acid solution was prepared.

[0225] The mixture was then heated in a water bath at 85°C for 30 minutes, followed by a brief sonication step to break up large colloidal aggregates (Bandelin HD4200, TS 113 probe), after which a clear solution was obtained. The energy applied was 200 KJ during 30 minutes.

[0226] The solution was poured into two 220 mm Petri dishes. The Petri dishes were sealed and cooled by storing in a refrigerator at 4°C for 20 hours to obtain a self-supporting protein hydrogel.

[0227] (b) Application of shear in soybean protein hydrogel

[0228] Shear was then applied to the hydrogel formed in step (a) as follows. The protein hydrogel was cut into approximately 1 cm cubes by a low shear cutting step. The cubes were placed in a 75 μm filter bag, which was then immersed in a bucket containing 5 L of deionized water. This formed a crude protein hydrogel slurry within the filter bag. The hydrogel cubes were soaked for 1.5 hours with occasional gentle stirring. This step was performed to reduce the concentration of acetic acid in the hydrogel by diffusion into the continuous aqueous phase, and was repeated 1 more time until the final pH of the aqueous solution was 3.07.

[0229] The strained gel cubes (469 g) were transferred to a 0.5 L bottle and mixed with 40 g of deionized water. A colloidal suspension of large gel particles was obtained and divided into 50 g aliquots, which were then subjected to different levels of high shear to obtain samples with different particle size distributions, as detailed in Table 2:

[0230]

[0231]

[0232] Table 2

[0233] (c) Preparation of soybean protein film

[0234] The fine hydrogel slurry of sample B prepared in step (b) was mixed with 20 w / w% glycerol and poured onto a plastic petri dish and dried at room temperature for 24 h to form a structured object, which was a film with an average thickness of 61 μm. The PSD and optical properties of the soy protein film of Example 3 were studied. The results are shown in Table 3. The obtained film was transparent, as shown in FIG. Figure 2b As shown, its transmittance at 600 nm is 89.8%.

[0235]

[0236] Table 3

[0237] The results show that functional films with high transparency can be formed using the plant-based protein hydrogel slurry of the present invention. Without wishing to be bound by theory, it is believed that achieving a controlled particle size distribution allows for a high level of transparency to be observed.

[0238] Example 4: Preparation of pea protein gel structured objects

[0239] (a) Protein hydrogel formation

[0240] 450 grams of a mixture consisting of 11.11% (w / v) pea protein isolate in a 30% (v / v) acetic acid solution was prepared.

[0241] The mixture was then heated in a water bath at 85°C for 30 minutes, followed by a brief sonication step to break up large colloidal aggregates (Bandelin HD4200, TS 113 probe), after which a clear solution was obtained. The energy applied was 200 KJ during 30 minutes.

[0242] The solution was poured into two 220 mm Petri dishes. The Petri dishes were sealed and cooled by storing in a refrigerator at 4°C for 20 hours to obtain a self-supporting protein hydrogel.

[0243] (b) Application of shear in protein hydrogels

[0244] Shear was then applied to the hydrogel formed in step (a) as follows. The protein hydrogel was cut into approximately 1 cm cubes by a low shear cutting step. The cubes were placed in a 75 μm filter bag, which was then immersed in a bucket containing 5 L of deionized water. This formed a crude protein hydrogel slurry within the filter bag. The hydrogel cubes were soaked for 1.5 hours with occasional gentle stirring. This step was performed to reduce the concentration of acetic acid in the hydrogel by diffusion into the continuous aqueous phase, and was repeated 1 more time until the final pH of the aqueous solution was 3.02.

[0245] The strained gel cubes (400 g) were transferred to a 0.5 L bottle and mixed with 100 g of deionized water. A colloidal suspension of large gel particles was obtained and divided into 50 g aliquots, which were then subjected to different levels of high shear to obtain samples with different particle size distributions, as detailed in Table 4:

[0246]

[0247] Table 4

[0248] (c) Preparation of pea protein film

[0249] The fine hydrogel slurry prepared in step (b) was mixed with 20 w / w% glycerol and poured onto a PTFE evaporating dish and dried at room temperature for 24 h to form a structured object as a film. Table 5 describes the properties of the resulting film:

[0250] sample Film thickness (μm) Tensile strength(MPa) Elongation at break (%) A 140.8±6.5 4.9±0.1 17.7±6.6 B 111.0±49.9 12.0±0.5 35.9±20.4 C 97.7±4.0 8.0±0.4 7.8±5.5

[0251] Table 5

[0252] The results show that the method of the present invention allows the preparation of plant-based protein hydrogel slurries with controlled particle size distribution. The results also show that the plant-based protein hydrogel slurries of the present invention can be used to prepare structured objects, such as films, which have excellent tensile properties that can be controlled by controlling the slurry particle size. The produced films have a secondary structure with a high level of intermolecular β-sheets (e.g., at least 50% intermolecular β-sheets). This high degree of intermolecular interaction is believed to contribute to the enhanced mechanical properties observed.

[0253] Example 5: Using pea protein dispersion as a coating for paper

[0254] The colloidal pea protein dispersion obtained in Example 2 was sprayed onto an uncoated paperboard substrate using a spray gun. One layer was first applied and then dried in an oven at 80°C for 1 minute to evaporate the remaining solvent. This procedure was repeated 15 times until a uniform, transparent coating was applied. The results are shown in Figure 2. Figure 3 shown.

[0255] Example 6: Using pea protein dispersion as a coating for food

[0256] The colloidal pea protein dispersion obtained from Example 2 was applied as a coating to fresh strawberries by a dip coating step. The strawberries were first dipped into 50 ml of the pea protein dispersion for 5 seconds, and then the excess dispersion was removed by briefly applying compressed air. The coated strawberries were dried at room temperature for 10 minutes. This procedure was repeated 5 times until a uniform and transparent coating was applied. The results are shown in Figure 2. Figure 4 shown.

[0257] Example 7: Using pea protein dispersion as a coating for pharmaceutical products

[0258] The colloidal pea protein dispersion obtained in Example 2 was sprayed onto the uncoated acetaminophen tablets using a spray gun. One layer was first applied and then dried in an oven at 80°C for 1 minute to evaporate the remaining solvent. This procedure was repeated 15 times until a uniform and transparent coating was applied. The results are shown in FIG. Figure 5 shown.

[0259] Example 8: Using pea protein dispersion as seed coating

[0260] The colloidal pea protein dispersion obtained from Example 2 was applied as a coating to wheat seeds by a dip coating procedure. The wheat seeds were first dipped into 50 ml of the pea protein dispersion for 5 seconds, and then the excess dispersion was removed by briefly applying compressed air. The coated wheat seeds were dried at room temperature for 10 minutes. This procedure was repeated 2 times until a uniform and transparent coating was applied. The results are shown in Figure 2. Figure 6 shown.

[0261] Terms:

[0262] 1. A method for preparing a plant-based protein hydrogel slurry, the method comprising:

[0263] (a) forming a solution comprising one or more plant-based proteins in a solvent system, wherein the solvent system comprises miscible co-solvents; wherein a first co-solvent increases the solubility of the plant-based protein and a second co-solvent decreases the solubility of the plant-based protein;

[0264] (b) inducing a sol-gel transition of the protein in the solution to form a plant-based protein hydrogel; and

[0265] (c) subjecting the plant-based protein hydrogel to a shearing treatment to form a plant-based protein hydrogel slurry.

[0266] 2. The method according to clause 1, wherein the plant protein is selected from soy protein, pea protein, rice protein, potato protein, wheat protein, corn zein protein or sorghum protein.

[0267] 3. A method according to clause 1 or clause 2, wherein the first co-solvent is an organic acid; preferably acetic acid, formic acid, propionic acid and / or α-hydroxy acid; wherein the α-hydroxy acid is preferably selected from glycolic acid, lactic acid, malic acid, citric acid and / or tartaric acid; particularly preferred organic acids are acetic acid and / or lactic acid.

[0268] 4. A method according to any one of clauses 1 to 3, wherein the second or additional co-solvent is an aqueous buffer solution; preferably selected from water, ethanol, methanol, acetone, acetonitrile, dimethyl sulfoxide, dimethylformamide, formamide, 2-propanol, 1-butanol, 1-propanol, hexanol, tert-butanol, ethyl acetate or hexafluoroisopropanol; particularly preferably water and / or ethanol; further particularly preferably water.

[0269] 5. A method according to any one of clauses 1 to 4, wherein the solvent system has a co-solvent ratio of the first co-solvent to the second co-solvent of about 20-80% v / v, preferably about 20-60% v / v, about 25-55% v / v, about 30-50% v / v, about 20%, about 30%, about 40%, about 50% or about 60% v / v, and most preferably about 30-50% v / v.

[0270] 6. A method according to any one of clauses 1 to 5, wherein the protein solution is heated to a first temperature above the sol-gel temperature of the one or more plant-based protein solutions and then reduced to a second temperature below the sol-gel temperature of the one or more plant-based protein solutions to form a hydrogel.

[0271] 7. A method according to any one of clauses 1 to 6, wherein the shear treatment comprises a high shear step.

[0272] 8. A method according to clause 7, wherein the high shear step comprises breaking the plant-based protein hydrogel into pieces.

[0273] 9. A method according to clause 8, wherein the fragments produced in the high shear step have a d value of less than 500 nm, preferably less than 300 nm, more preferably less than 200 nm, even more preferably less than 50 nm as determined by DLS. 50 .

[0274] 10. A process according to any one of clauses 7 to 9, wherein the high shear step comprises sonication, high shear mechanical agitation or cavitation.

[0275] 11. The method according to any one of clauses 1 to 6, wherein the shearing process comprises two steps.

[0276] 12. The method according to clause 11, wherein the shear treatment comprises a low shear step followed by a high shear step.

[0277] 13. The method according to clause 12, wherein the low shear step comprises breaking the plant-based protein hydrogel into pieces.

[0278] 14. A process according to clause 13, wherein at least 80% of the fragments produced in the low shear step have a particle size in the range of 1 mm to 50 mm, preferably 1 mm to 30 mm, more preferably 10 mm to 30 mm, more preferably 15 mm to 30 mm, even more preferably 20 mm to 30 mm as determined by sieving.

[0279] 15. A process according to any one of clauses 12 to 14, wherein the low shear step comprises mechanical cutting.

[0280] 16. A method according to any one of clauses 13 to 15, wherein the high shear step comprises further fragmenting the plant-based protein hydrogel.

[0281] 17. A method according to clause 16, wherein the fragments produced in the high shear step have a D50 as determined by DLS of less than 500 nm, preferably less than 300 nm, more preferably less than 200 nm, even more preferably less than 50 nm.

[0282] 18. A method according to any one of clauses 12 to 17, wherein the high shear step comprises sonication, high shear mechanical agitation or cavitation.

[0283] 19. The method according to any one of clauses 12 to 18, wherein step (c) further comprises subjecting the plant-based protein hydrogel slurry to a solvent reduction step, preferably an organic solvent reduction step, between the low shear step and the high shear step.

[0284] 20. The method according to clause 19, wherein the solvent reduction step comprises the following steps:

[0285] (i) contacting the fragments of the plant-based hydrogel slurry with a non-solubilizing solvent;

[0286] (ii) separating the fragments of the plant-based hydrogel slurry from the non-solubilizing solvent to obtain a washed plant-based protein hydrogel; and

[0287] (iii) optionally repeating steps (i) and (ii).

[0288] 21. A method according to clause 20, wherein step (ii) involves screen filtration.

[0289] 22. A method according to clause 20 or 21, wherein before washing, the plant-based protein hydrogel has a storage modulus (G') at 10 rad / s greater than 1000 Pa, preferably greater than 2000 Pa, more preferably greater than 5000 Pa, even more preferably greater than 6000 Pa, most preferably greater than 8000 Pa.

[0290] 23. A method according to any one of clauses 20 or 22, wherein before washing, the plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of less than 20,000 Pa, preferably less than 15,000 Pa, more preferably less than 10,000 Pa.;

[0291] Optionally, before washing, the plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of about 1000 to 20,000 Pa, preferably about 2000 to 15,000 Pa, more preferably about 2000 to 10,000 Pa.

[0292] 24. A method according to any one of clauses 20 to 23, wherein the washed plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of greater than 500 Pa, preferably greater than 1000 Pa, more preferably greater than 2500 Pa, even more preferably greater than 3000 Pa, most preferably greater than 4000 Pa.

[0293] 25. A method according to any one of clauses 20 to 24, wherein the washed plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of less than 20,000 Pa, preferably less than 15,000 Pa, more preferably less than 10,000 Pa;

[0294] Optionally, the washed plant-based protein hydrogel has a storage modulus (G') at 10 rad / s of about 500 to 20,000 Pa, about 500 to 15,000 Pa, about 500 to 10,000 Pa, about 1000 to 20,000 Pa, about 1000 to 15,000 Pa, or about 1000 to 10,000 Pa.

[0295] 26. The method according to any one of clauses 1 to 25, further comprising the following steps:

[0296] (d) changing the pH of the plant-based protein hydrogel slurry so that it differs from the isoelectric point of the protein hydrogel by more than 1 pH unit.

[0297] 27. The method according to clause 26, wherein step (d) is performed after step (c).

[0298] 28. The method according to clause 26, wherein step (d) and step (c) are performed sequentially.

[0299] 29. A method according to any one of clauses 26 to 28, wherein step (d) involves adding a pH adjusting material to the plant-based protein hydrogel slurry.

[0300] 30. The method according to clause 29, wherein the pH adjusting material is a solution comprising monovalent metal ions, divalent metal ions or ammonium ions, preferably an alkaline aqueous solution comprising monovalent metal ions, divalent metal ions or ammonium ions.

[0301] 31. A method according to clause 30, wherein the pH adjusting material is an aqueous hydroxide solution, preferably sodium hydroxide, potassium hydroxide or ammonium hydroxide.

[0302] 32. A method according to any one of clauses 26 to 31, wherein the pH of the plant-based protein hydrogel slurry after step (d) is at least 1 pH unit below the isoelectric point of the plant-based protein.

[0303] 33. A method according to any one of clauses 26 to 31, wherein the pH of the plant-based protein hydrogel slurry after step (d) is at least 1 pH unit above the isoelectric point of the plant-based protein.

[0304] 34. The method according to any one of clauses 1 to 33, further comprising adding additional ingredients to the plant-based protein hydrogel slurry.

[0305] 35. A method according to clause 34, wherein the additional ingredient is selected from plasticizers, opacifiers, preservatives, pigments and nanoparticles, or mixtures thereof.

[0306] 36. A method according to clause 35, wherein the additional ingredient is a plasticizer.

[0307] 37. A method according to clause 36, wherein the plasticizer is selected from ethylene glycol, diethylene glycol, triethylene glycol, tetraethylene glycol, polyethylene glycol, propylene glycol, sorbitol, mannitol, xylitol, fatty acids, glucose, mannose, fructose, sucrose, ethanolamine, urea, triethanolamine, vegetable oils, lecithin, waxes and amino acids.

[0308] 38. A method according to any one of clauses 1 to 37, wherein the plant-based protein hydrogel slurry has a -1 The range of 10 to 10000 cps is preferably within 50 s -1 The viscosity is in the range of 15 to 5000 cps.

[0309] 39. A method according to any one of clauses 1 to 38, wherein the plant-based protein hydrogel slurry comprises protein aggregates having an average size of less than 200 nm, preferably less than 150 nm, less than 125 nm, less than 100 nm, less than 90 nm, less than 80 nm, less than 70 nm, less than 60 nm, less than 50 nm, less than 40 nm or less than 30 nm.

[0310] 40. A method according to any one of clauses 1 to 39, wherein the plant based protein hydrogel slurry has a protein solids content in the range of 5 wt% to 25 wt% based on the total weight of the plant based protein hydrogel slurry.

[0311] 41. A plant-based protein hydrogel slurry prepared according to the method of any one of clauses 1 to 40.

[0312] 42. A method for preparing a plant-based structured material, the method comprising:

[0313] (a) preparing a plant-based protein hydrogel slurry according to the method of any one of clauses 1 to 40; and

[0314] (b) subjecting the plant-based protein hydrogel slurry to one or more solvent level reduction steps to reduce the level of the first co-solvent and / or the second co-solvent to obtain the plant-based structuring material.

[0315] 43. The method according to clause 42, wherein step (b) involves placing the plant-based protein hydrogel slurry on a surface prior to forming the one or more solvent level reducing steps.

[0316] 44. A method according to clause 42 or clause 43, wherein the solvent level reducing step involves heating.

[0317] 45. A method according to clause 44, wherein the solvent level reducing step involves heating at a temperature in the range of 50°C to 100°C.

[0318] 46. ​​A method according to clause 42 or clause 43, wherein the solvent level reducing step involves forced convection of dry air.

[0319] 47. A method according to any one of clauses 42 to 46, wherein the plant-based structuring material is a film.

[0320] 48. A method according to any one of clauses 42 to 46, wherein the plant-based structured material is a casting.

[0321] 49. A method according to any one of clauses 42 to 46, wherein the plant-based structuring material is a coating.

[0322] 50. A method according to any one of clauses 42 to 49, wherein the plant-based structuring material comprises a plant-based protein having a secondary structure, wherein the secondary structure has at least 40% intermolecular β-sheets, at least 50% intermolecular β-sheets, at least 60% intermolecular β-sheets, at least 70% intermolecular β-sheets, at least 80% intermolecular β-sheets, or at least 90% intermolecular β-sheets.

[0323] 51. A method according to any one of clauses 42 to 50, wherein the plant-based structuring material has a Young's modulus greater than 20 MPa; preferably greater than 50 MPa, greater than 80 MPa, greater than 100 MPa, greater than 200 MPa, greater than 300 MPa, greater than 400 MPa, greater than 500 MPa or greater than 600 MPa.

[0324] 52. A method according to any one of clauses 42 to 51, wherein the plant-based structuring material is a film having a thickness in the range of 1 to 1000 μm, preferably 1 to 100 μm, more preferably 10 to 100 μm, even more preferably 20 to 60 μm, most preferably 30 to 50 μm.

[0325] 53. A method according to any one of clauses 42 to 52, wherein the plant-based structuring material is a film having a tensile strength greater than 1 MPa, preferably greater than 5 MPa, preferably greater than 10 MPa and most preferably greater than 25 MPa.

[0326] 54. A method according to any one of clauses 42 to 53, wherein the plant-based structuring material is a film having an elongation at break percentage of 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 100% or more or more.

[0327] 55. A plant-based structured material prepared according to the method of any one of clauses 42 to 54.

[0328] 56. Use of a plant-based protein hydrogel slurry according to any one of clauses 1 to 40 for producing a plant-based structured material.

[0329] 57. The use according to clause 56, wherein the plant-based structured material is a film, a casting or a coating.

Claims

1. A method for preparing a plant-based protein hydrogel slurry, the method comprising: (a) forming a solution comprising one or more plant-based proteins in a solvent system, wherein the solvent system comprises a miscible co-solvent; wherein the first cosolvent increases the solubility of the plant-based protein and the second cosolvent decreases the solubility of the plant-based protein; (b) inducing a sol-gel transition of the protein in the solution to form a plant-based protein hydrogel; and (c) subjecting the plant-based protein hydrogel to a shearing treatment to form a plant-based protein hydrogel slurry, wherein (i) the shearing treatment comprises a high shearing step, and the fragments produced in the high shearing step have a d value of less than 500 nm as measured by DLS. 50 , or (ii) the shearing treatment comprises a high shear step and the fragments produced in the high shear step have a d of 0.5 to 150 microns as measured by laser diffraction 50 , The plant protein is selected from soy protein, pea protein, potato protein or rapeseed protein, and the plant-based protein hydrogel slurry can be formed into a film, a coating or a casting.

2. The method of claim 1, wherein the protein solution is heated to a first temperature above the sol-gel temperature of the one or more plant-based protein solutions and then cooled to a second temperature below the sol-gel temperature of the one or more plant-based protein solutions to form a hydrogel.

3. The method according to claim 1, wherein the high shear step involves breaking the plant-based protein hydrogel into fragments; and wherein the fragments produced in the high shear step have a d value of less than 300 nm as measured by DLS. 50 .

4. The method of claim 3, wherein the fragments produced in the high shear step have a d value of less than 200 nm as measured by DLS. 50 .

5. The method of claim 3, wherein the fragments produced in the high shear step have a d value of less than 50 nm as measured by DLS. 50 .

6. The method according to claim 1, wherein the high shear step involves breaking the plant-based protein hydrogel into fragments; and wherein the fragments produced in the high shear step have a d value of 0.6 to 100 microns as measured by laser diffraction. 50 .

7. The method of claim 6, wherein the fragments produced in the high shear step have a d value of 0.7 to 70 microns as measured by laser diffraction. 50 .

8. The method of claim 6, wherein the fragments produced in the high shear step have a d value of 0.8 to 50 microns as measured by laser diffraction. 50 .

9. The method of claim 6, wherein the fragments produced in the high shear step have a d value of 0.9 to 25 microns as measured by laser diffraction. 50 .

10. The method of claim 6, wherein the fragments produced in the high shear step have a d value of 1 to 20 microns as measured by laser diffraction. 50 .

11. The method of claim 6, wherein the fragments produced in the high shear step have a d value of 1 to 10 microns as measured by laser diffraction. 50 .

12. The method of claim 6, wherein the fragments produced in the high shear step have a d value of 1 to 5 microns as measured by laser diffraction. 50 .

13. The method according to claim 1, wherein the shear treatment further comprises a low shear step, and the high shear step is subsequent to the low shear step.

14. A process according to claim 13 wherein at least 80% of said fragments produced in said low shear step have a particle size in the range of 1 mm to 50 mm as determined by sieving.

15. A process according to claim 14 wherein at least 80% of said fragments produced in said low shear step have a particle size in the range of 1 mm to 30 mm as determined by sieving.

16. A process according to claim 14 wherein at least 80% of said fragments produced in said low shear step have a particle size in the range of 10 mm to 30 mm as determined by sieving.

17. A process according to claim 14 wherein at least 80% of said fragments produced in said low shear step have a diameter in the range of 15 mm to 30 mm as determined by sieving.

18. The method according to claim 14, wherein at least 80% of said fragments produced in said low shear step have a particle size in the range of 20 mm to 30 mm as determined by sieving.

19. The method according to claim 14, wherein step (c) further comprises subjecting the plant-based protein hydrogel slurry to a solvent reduction step between the low shear step and the high shear step; wherein the solvent reduction step comprises the steps of: (i) contacting the fragments of the plant-based hydrogel slurry with a non-solubilizing solvent; and (ii) separating the fragments of the plant-based hydrogel slurry from the non-solubilizing solvent to obtain a washed plant-based protein hydrogel.

20. The method of claim 19, wherein the solvent reduction step is a solubilizer reduction step.

21. The method of claim 19, wherein the method further comprises step (iii) repeating steps (i) and (ii).

22. The method according to claim 19, wherein before washing, the plant-based protein hydrogel has a storage modulus (G') of 1000 to 20,000 Pa at 20°C, 1% strain at an oscillation frequency of 1 Hz, and 10 rad / s.

23. The method of claim 22, wherein the plant-based protein hydrogel has a storage modulus (G') of 1000 to 15,000 Pa at 20°C, 1% strain at an oscillation frequency of 1 Hz, and 10 rad / s before washing.

24. The method of claim 22, wherein the plant-based protein hydrogel has a storage modulus (G') of 1000 to 10,000 Pa at 10 rad / s prior to washing.

25. The method of claim 22, wherein the plant-based protein hydrogel has a storage modulus (G') of 2000 to 20,000 Pa at 10 rad / s prior to washing.

26. The method of claim 22, wherein the plant-based protein hydrogel has a storage modulus (G') of 2000 to 15,000 Pa at 10 rad / s prior to washing.

27. The method of claim 22, wherein the plant-based protein hydrogel has a storage modulus (G') of 2000 to 10,000 Pa at 10 rad / s prior to washing.

28. The method according to claim 19, wherein the washed plant-based protein hydrogel has a storage modulus (G') of 500 to 20,000 Pa at 10 rad / s.

29. The method according to claim 28, wherein the washed plant-based protein hydrogel has a storage modulus (G') of 500 to 15,000 Pa at 10 rad / s.

30. The method according to claim 28, wherein the washed plant-based protein hydrogel has a storage modulus (G') of 500 to 10,000 Pa at 10 rad / s.

31. The method according to claim 28, wherein the washed plant-based protein hydrogel has a storage modulus (G') of 1000 to 20,000 Pa at 10 rad / s.

32. The method according to claim 28, wherein the washed plant-based protein hydrogel has a storage modulus (G') of 1000 to 15,000 Pa at 10 rad / s.

33. The method according to claim 28, wherein the washed plant-based protein hydrogel has a storage modulus (G') of 1000 to 10,000 Pa at 10 rad / s.

34. The method according to any one of claims 1 to 33, further comprising the steps of: d) changing the pH of the plant-based protein hydrogel slurry so that the pH is different from the isoelectric point of the protein hydrogel by more than 1 pH unit.

35. The method according to any one of claims 1-33, further comprising adding additional ingredients to the plant-based protein hydrogel slurry; wherein the additional ingredients are selected from plasticizers, opacifiers, preservatives, pigments, or mixtures thereof.

36. The method according to any one of claims 1-33, further comprising adding nanoparticles to the plant-based protein hydrogel slurry.

37. The method according to any one of claims 1 to 33, wherein the plant-based protein hydrogel slurry has a -1 and a viscosity in the range of 10 to 10000 cps at 20°C.

38. The method according to claim 37, wherein the plant-based protein hydrogel slurry has a -1 and a viscosity in the range of 10 to 8000 cps at 20°C.

39. The method according to claim 37, wherein the plant-based protein hydrogel slurry has a -1 and a viscosity in the range of 12 to 6000 cps at 20°C.

40. The method according to claim 37, wherein the plant-based protein hydrogel slurry has a -1 and a viscosity in the range of 15 to 5000 cps at 20°C.

41. A plant-based protein hydrogel slurry prepared according to the method of any one of claims 1 to 40.

42. A method for preparing a plant-based structured material, the method comprising: (a) preparing a plant-based protein hydrogel slurry according to the method described in any one of claims 1 to 40; and (b) subjecting the plant-based protein hydrogel slurry to one or more solvent level reduction steps to reduce the level of the first co-solvent and / or the second co-solvent to obtain the plant-based structuring material.

43. A method according to claim 42, wherein the plant-based structured material is a film, a casting or a coating.

44. A method according to claim 43, wherein the plant-based structuring material is a coating and the coating is a surface coating.

45. The method according to claim 43, wherein the plant-based structuring material is a coating, and the coating is a food coating, a seed coating or a pharmaceutical coating.

46. ​​The method of claim 44, wherein the surface coating is a paper coating.

47. A method according to any one of claims 42 to 46, wherein the plant based structuring material comprises a plant based protein having a secondary structure, wherein the secondary structure has at least 40% intermolecular beta-sheets.

48. The method of claim 47, wherein the secondary structure has at least 50% intermolecular [beta]-sheets.

49. The method of claim 47, wherein the secondary structure has at least 60% intermolecular [beta]-sheets.

50. The method of claim 47, wherein the secondary structure has at least 70% intermolecular [beta]-sheets.

51. The method of claim 47, wherein the secondary structure has at least 80% intermolecular [beta]-sheets.

52. The method of claim 47, wherein the secondary structure has at least 90% intermolecular [beta]-sheets.

53. A plant-based structured material prepared according to the method of any one of claims 42 to 52.

54. Use of the plant-based protein hydrogel slurry according to claim 41 for producing a plant-based structured material.

55. Use according to claim 54, wherein the plant-based structured material is a film, a casting, a molding or a coating.

56. Use according to claim 55, wherein the plant-based structuring material is a coating and the coating is a surface coating.

57. The use according to claim 55, wherein the plant-based structuring material is a coating, and the coating is a food coating, a seed coating or a pharmaceutical coating.

58. Use according to claim 56, wherein the surface coating is a paper coating.

59. A plant-based protein hydrogel slurry having a protein solid content of 5 wt% to 25 wt% based on the total weight of the plant-based protein hydrogel slurry, and -1 and a viscosity in the range of 10 to 10000 cps at 20° C., wherein the plant-based protein hydrogel slurry comprises a d 50 The particle size of the fragments is 0.5 to 150 microns, and wherein the vegetable protein is selected from soy protein, pea protein, potato protein or rapeseed protein.

60. The plant-based protein hydrogel slurry according to claim 59, wherein the plant-based protein hydrogel slurry comprises a d 50 Particle sizes range from 0.6 to 100 microns.

61. A plant-based protein hydrogel slurry having a protein solid content of 5 wt% to 25 wt% based on the total weight of the plant-based protein hydrogel slurry, and -1 and a viscosity in the range of 10 to 10,000 cps at 20° C., wherein the plant-based protein hydrogel slurry comprises a d 50 The fragments have a particle size of less than 500 nm, wherein the plant protein is selected from soy protein, pea protein, potato protein or rapeseed protein.

62. The plant-based protein hydrogel slurry according to claim 61, wherein the plant-based protein hydrogel slurry comprises a d 50 Particle size less than 300nm.

63. The plant-based protein hydrogel slurry according to any one of claims 59 to 62, wherein Based on the total weight of the plant-based protein hydrogel slurry, the protein solid content is 6 wt % to 20 wt %.

64. The plant-based protein hydrogel slurry according to any one of claims 59 to 62, wherein the -1 and a viscosity at 20°C in the range of 10 to 8000 cps.

65. A film comprising the plant based protein hydrogel slurry as claimed in any one of claims 59 to 64.

66. A membrane according to claim 65, wherein the membrane: (a) comprising a plant-based protein having a secondary structure comprising at least 40% intermolecular β-sheets; and / or (b) having a tensile strength of 4 to 20 MPa; and / or (c) having a percent elongation at break greater than 10%.

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