WO2020092306A1 - Protéine végétale mycéliée et compositions alimentaires comprenant celle-ci - Google Patents

Protéine végétale mycéliée et compositions alimentaires comprenant celle-ci Download PDF

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Publication number
WO2020092306A1
WO2020092306A1 PCT/US2019/058470 US2019058470W WO2020092306A1 WO 2020092306 A1 WO2020092306 A1 WO 2020092306A1 US 2019058470 W US2019058470 W US 2019058470W WO 2020092306 A1 WO2020092306 A1 WO 2020092306A1
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Prior art keywords
protein
myceliated
product
pea
food product
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PCT/US2019/058470
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English (en)
Inventor
Joseph George Akamittath
Alan D. HAHN
Anthony J. Clark
Todd MCDONALD
Savita Jensen
Lisa Schmidt
Bhupendra Kumar Soni
James Patrick Langan
Brooks John Kelly
Huntington DAVIS
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Mycotechnology, Inc.
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Publication of WO2020092306A1 publication Critical patent/WO2020092306A1/fr

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23P—SHAPING OR WORKING OF FOODSTUFFS, NOT FULLY COVERED BY A SINGLE OTHER SUBCLASS
    • A23P30/00—Shaping or working of foodstuffs characterised by the process or apparatus
    • A23P30/20—Extruding
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
    • A23J1/00—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites
    • A23J1/006—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites from vegetable materials
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
    • A23J3/00—Working-up of proteins for foodstuffs
    • A23J3/22—Working-up of proteins for foodstuffs by texturising
    • A23J3/225—Texturised simulated foods with high protein content
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L11/00—Pulses, i.e. fruits of leguminous plants, for production of food; Products from legumes; Preparation or treatment thereof
    • A23L11/50—Fermented pulses or legumes; Fermentation of pulses or legumes based on the addition of microorganisms
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
    • A23L3/16—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by heating loose unpacked materials
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L7/00—Cereal-derived products; Malt products; Preparation or treatment thereof
    • A23L7/10—Cereal-derived products
    • A23L7/104—Fermentation of farinaceous cereal or cereal material; Addition of enzymes or microorganisms

Definitions

  • the present inventors have previously disclosed a method to prepare a myceliated high-protein food product, which includes culturing a filamentous fungus in an aqueous media which has a high level of protein, for example at least 20% protein by dry weight (w/w) and at least 50 g/L protein.
  • the fungi can include Pleurotus ostreatus, Pleurotus eryngii, Lepista nuda, Hericium erinaceus, Lentinula edodes, Agaricus blazeii, Laetiporus sulfureus and combinations thereof.
  • the present invention discloses a texturized plant protein product comprising an extruded mixture comprising (a) a myceliated high-protein food product, wherein the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein food product is derived from pea and/or rice, wherein the myceliated high-protein product is myceliated by an aqueous fungal culture comprising Lentinula edodes, Agaricus spp., Pleurotus spp., Boletus spp., or Laetiporus spp ., in a media comprising at least 50 g/L protein in liquid culture, and wherein the myceliated high-protein food product has reduced undesirable flavor and reduced undesirable aroma compared with a non-myceliated food product; and (b) an additional high-protein material comprising at least 50% protein on a dry weight basis, wherein the myceliated high-
  • the food compositions of the invention comprise, without limitation, meat- structured plant protein which are meat extenders or meat analogs, comprising a textured plant-based protein as disclosed herein.
  • the inventors have previously disclosed culturing a filamentous fungus in a high protein media to provide a protein product, and also found that such treatment can also alter the taste, flavor or aroma of high protein food compositions in unexpected ways.
  • the processes of the invention enable the production of food compositions, protein concentrates, isolates and high protein foodstuffs that have been imbued with mycelial material, thereby altering aspects of the media used in the production of products according to the methods of the present invention.
  • the invention also presents the ability to stack protein sources to optimize amino acid profiles of products made according to the methods of the invention.
  • the present invention discloses a texturized plant protein product comprising an extruded mixture comprising (a) a myceliated high-protein food product, wherein the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein food product is derived from pea and/or rice, wherein the myceliated high-protein product is myceliated by an aqueous fungal culture comprising Lentinula edodes, Agaricus spp., Pleurotus spp., Boletus spp., or Laetiporus spp ., in a media comprising at least 50 g/L protein in liquid culture, and wherein the myceliated high-protein food product has reduced undesirable flavor and reduced undesirable aroma compared with a non-myceliated food product; and (b) an additional high-protein material comprising at least 50% protein on a dry weight basis, wherein the myceliated high-
  • methods to make the textured plant-based protein product includes a method step to prepare or provide a myceliated high-protein food product.
  • the method may optionally include the steps of providing an aqueous media comprising a high-protein material.
  • the aqueous media may comprise, consist of, or consist essentially of at least 50% (w/w) protein, on a dry weight basis and may also comprise at least 50 g/L protein.
  • the media may also comprise, consist of or consist essentially of optional additional excipients as identified herein below.
  • the aqueous media may be inoculated with a fungal culture, optionally comprising Lentinula edodes, Agaricus spp., Pleurotus spp., Boletus spp., or Laetiporus spp.
  • the inoculated media may then be cultured to produce a myceliated high-protein food product, and the myceliated high-protein food product’s taste, flavor, and/or aroma may be modulated compared to the high-protein material in the absence of the culturing step.
  • the method step to provide a myceliated high-protein food product comprises providing a food product that is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein food product is derived from pea and/or rice, wherein the myceliated high-protein product is myceliated by an aqueous fungal culture comprising Lentinula edodes, Agaricus spp., Pleurotus spp., Boletus spp., or Laetiporus spp ., in a media comprising at least 50 g/L protein in liquid culture, and wherein the myceliated high-protein food product has reduced undesirable flavor and reduced undesirable aroma compared with a non-myceliated food product.
  • the method steps to ferment and/or myceliate high protein materials to form a myceliated high protein material are described in more detail elsewhere herein.
  • the present invention also includes a method to prepare a textured plant-based protein product useful for products such as meat-structured meat analogs or meat extenders.
  • This textured plant-based meat analog or meat extender in one embodiment, has texture associated with meat.
  • the method optionally provides a“meat structured protein product” which can be made from the“texturized protein product” as disclosed herein.
  • Integral to a meat structured protein product is a texturized protein product which refers to a product comprising protein fiber networks and/or aligned protein fibers that produce meat-like textures.
  • Methods for determining the degree of protein fiber network formation and/or protein fiber alignment include visual determination based upon photographs and micrographic images, as exemplified in U.S. Utility application Ser. No. 14/687,803 filed Apr. 15, 2015.
  • at least about 55%, at least about 65%, at least about 75%, at least about 85%, or at least about 95% of the protein fibers are substantially aligned.
  • Protein fiber networks and/or protein fiber alignments may impart cohesion and firmness whereas open spaces in the protein fiber networks and/or protein fiber alignments may tenderize the meat structured protein products and provide pockets for capturing water, carbohydrates, salts, lipids, flavorings, and other materials that are slowly released during chewing to lubricate the shearing process and to impart other meat-like sensory characteristics.
  • the method to make a textured plant-based protein product includes the step of providing a myceliated high-protein food product and at least one additional high-protein material.
  • the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, and the myceliated high-protein food product is derived from pea and/or rice.
  • the myceliated high-protein product is a myceliated high-protein product that has been myceliated by an aqueous fungal culture, in a media comprising at least 50 g/L protein in liquid culture.
  • the myceliated high-protein food product has reduced undesirable flavor and reduced undesirable aroma compared with a non-myceliated food product.
  • the additional high- protein material comprises at least 50% protein on a dry weight basis.
  • the step of providing a textured plant-based protein product can include steps of preparing the myceliated high-protein food product by the following steps.
  • the myceliated high-protein food product is produced by culturing a fungus in an aqueous media which includes a high-protein material, with amounts of protein of at least 50% (w/w) protein, in a media comprising at least 50 g/L protein in liquid culture, resulting in a myceliated high-protein food product, whereby the flavor or taste of the myceliated high- protein food product is modulated compared to the high-protein material; providing an additional high-protein material; and mixing the myceliated high-protein food product and the additional high-protein material to form a mixture; optionally preconditioning the mixture, e.g., by adding steam and/or water to the mixture, and extruding the mixture under heat and pressure under conditions capable of forming a textured plant-based protein product useful for products such as meat-structured meat analogs or meat
  • the method may also include the steps of mixing the myceliated high-protein food product and an additional high-protein material, wherein the myceliated high-protein food product is present at between about 5% and 90% on a dry weight basis compared with the additional high-protein material.
  • the method may also include the steps of extruding the mixture.
  • the extrusion step(s) may include the step of adding steam and/or water to the mixture and extruding the mixture under heat and/or pressure to form the textured plant-based protein product.
  • the method to prepare a textured plant-based protein product may also include the step of providing at least one additional high-protein food product.
  • the additional high- protein food product can comprise, consist of, or consist essentially of at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95%, protein.
  • This additional high-protein food product in an embodiment of the method to make a textured plant-based protein product, has the purpose of providing gelating or other properties to supplement the formation of texturized protein from the myceliated high-protein food product, which in some embodiments has lower gelling properties.
  • the additional food protein comprises a gel-forming protein.
  • Meat is considered the highest quality protein source, in part because meat proteins impart specific functionalities.
  • the overall properties of meat and meat products, including appearance, texture, and mouthfeel, are dependent on protein functionality.
  • the principal protein functionalities in processed meats are gelation and related properties (for example, meat particle binding and adhesion), emulsification, and water-holding.
  • Gel network formation is important to the generation of texture within foods, providing structural matrices for holding moisture, flavors, sugars, and other food ingredients, and stabilizes the dispersed phase. Gelation involves protein-protein and protein-solvent interactions usually initiated by the unfolding of the protein’s native structure to fully extend the polypeptide chains followed by intermolecular interaction to form a three-dimensional network.
  • the protein in the method to make a textured plant-based protein product, includes as an additional high-protein material, a gel-forming vegetarian protein.
  • Suitable gel-forming proteins include any gel-forming protein known in the art.
  • the additional high-protein material is preferably a plant protein material.
  • Suitable additional high-protein materials include, without limitation, cereal proteins, including vital wheat gluten; legume (pulse) proteins, including, without limitation, pea protein, soy protein, peanut protein, and bean proteins.
  • Suitable gel-forming proteins may also include mycoproteins.
  • the additional high-protein material can include vegetarian sources (e.g., plant sources) as well as non-vegetarian sources, and can include a protein concentrate and/or isolate.
  • Vegetarian sources include meal, protein concentrates and isolates prepared from a vegetarian source such as legumes or grains, such as, including pea, rice, chickpea, soy, cyanobacteria, hemp, chia, corn gluten meal and other sources, or a combination thereof.
  • the ratio of the myceliated high-protein food product to the additional high-protein material is between about 1% myceliated high-protein food product to about 99% additional high- protein material, to about 99% myceliated high-protein food product to about 1% additional high-protein material.
  • Other ratios include about 5% myceliated high-protein food product to about 95% additional high-protein material.
  • ratios include about 10% to about 90%; about 15% to about 85%; about 20% to about 80%; about 25% to about 75%; about 30% to about 70%; about 35% to about 65%; about 40% to about 60%; about 45% to about 55%; about 50% to about 50%; about 55% to about 45%; about 60% to about 40%; about 65% to about 35%; about 70% to about 30%; about 75% to about 25%; about 80% to about 20%; about 85% to about 15%; about 90% to about 10%, respectively, myceliated high-protein food product to additional high-protein material, w/w.
  • the amount of myceliated high-protein food product, relative to the additional high-protein material can vary between about 1% and 99%, between about 5% and 95%, between about 10% and 90%, between about 15% and 85%, between about 20% and 80%, between about 25% and 75%, between about 30% and 70%, between about 35% and 65%, between about 40% and 60%, between about 45% and 55%, or about 48% to about 52%, or, about 40% and 85%, between about 45% and 80%, between about 50% and 75%.
  • the method to prepare a textured plant-based protein product may also include the step of providing an optional carbohydrate component.
  • the carbohydrate ingredients are typically classified as a starch, a flour, or an edible fiber and the carbohydrate component may comprise one or more types of starch, flour, edible fiber, and combinations thereof.
  • Starch is the primary carbohydrate source used to help the formation of the product texture in textured plant-based protein products.
  • Typical starches used include rice starch, wheat starch, oat starch, corn starch, potato starch, cassava starch, and tapioca starch, although starch from any source is contemplated.
  • starch from any source is contemplated.
  • the swelling ability of starch, solubility, amount of amylose leaching out during gelatinization, and the ability to produce a viscous paste have an effect on the textured plant-based protein product.
  • Chemical alterations occur due to structural changes of the macromolecules in the feed blend, such as starch gelatinization and protein denaturation, as well as incorporation of water into the molecular matrix, all of which convert the raw feed particles into a viscoelastic dough under a pressurized environment.
  • Physical changes are related to product expansion due to a drastic pressure drop and water evaporation during die exit.
  • the textured plant-based protein product can contain starch or flour from at least one cereal grain.
  • flour examples include wheat flour, rice flour, barley flour white com flour, oat flour, sorghum flour, rye flour, amaranth flour, quinoa flour, and combinations thereof.
  • the textured plant-based protein product includes an edible fiber.
  • suitable edible fiber include but are not limited to bamboo fiber, barley bran, carrot fiber, citrus fiber, com bran, soluble dietary fiber, insoluble dietary fiber, oat bran, pea fiber, soy fiber, soy polysaccharide, wheat bran, wood pulp cellulose, modified cellulose, seed husks, oat hulls, citrus fiber, carrot fiber, corn bran, soy polysaccharide, barley bran, and rice bran.
  • the fiber may be present in the dry pre-mix from about 0.1% to about 10% by weight. In one embodiment, the fiber is about 2% to about 8% by weight of the dry ingredients. In another embodiment the fiber is about 5% by weight of the dry ingredients.
  • Particularly desirable types of fiber are those that effectively bind water when the mixture of non-animal protein and fiber is extruded.
  • a textured plant-based protein product comprises where at least some of the carbohydrate is derived from pea.
  • the mixture comprises between about 0.1% and about 25%, between about 3% and about 20%, between about 5% and about 15%, between about 5% and about 10%, between about 4% and about 7%, or between about 3% and about 35% by weight of carbohydrate, such as pea fiber.
  • the textured plant-based protein product includes a textured plant-based protein product formed from a mixture that comprises about 45% pea protein, 5% pea fiber, and about 50% myceliated high-protein material, w/w, or a mixture that comprises about 70% myceliated high-protein material, about 25% pea protein, about 5% pea fiber, w/w.
  • the moisture of the textured plant-based protein product is between about 20% and 40% by weight, and may be dried to approximately between 3% and 5% moisture to allow for rehydration by finished food manufacturers.
  • the dried textured plant-based protein product rehydrates to between 0.5 and 3 times its weight in water within a period of time after water addition, for example, 10 minutes.
  • the textured plant protein has a reduced undesirable flavor, such as, for example, a pea flavor or a bitterness flavor, and a reduced undesirable aroma, such as, for example, a beany aroma or a rice aroma.
  • a reduced undesirable flavor such as, for example, a pea flavor or a bitterness flavor
  • a reduced undesirable aroma such as, for example, a beany aroma or a rice aroma.
  • pH modifiers Modifying pH above 8.0 also may result in the production of harmful lysinoalanines; and lowering the pH has the opposite effect and will decrease protein solubility, making the protein more difficult to process.
  • pH is considered between pH 6.5 and 7.5.
  • Other ingredients may include processing aids such as calcium salts, such as calcium chloride, which can increase the textural integrity of extruded vegetable proteins and aid in smoothing its surface. Dosing levels for CaCb range between 0.5 and 2.0%. Sulfur compounds are useful for their ability to aid in the cleavage of disulfide bonding, which assists the unraveling of long twisted protein molecules. This reaction with the protein molecules causes increased expansion, smooth product surface and adds stability to the extrusion process, however, off flavors and aromas can arise from their use.
  • Appropriate sulfur compounds to use in the present invention includes sodium metabi sulfite, sodium bisulfite, as well as cystine, can be used with effects similar to those from using sulfur.
  • the normal dosing levels for sulfur or sulfur derivatives - are in the 0.1 to 0.2% range.
  • Cystine is used at approximately 0.5 to 1% level.
  • the present invention also includes products such as meat-structured plant-based meat analogs or meat extenders, for example, a plant-based meat analog patty, a plant-based meat analog sausage, a plant-based meat analog frankfurter, and a plant-based meat analog meat extended patty, sausage, or frankfurter comprising the textured plant-based protein product of the present invention.
  • meat-structured plant-based meat analogs or meat extenders for example, a plant-based meat analog patty, a plant-based meat analog sausage, a plant-based meat analog frankfurter, and a plant-based meat analog meat extended patty, sausage, or frankfurter comprising the textured plant-based protein product of the present invention.
  • transglutaminase can be used as a processing aid in the present invention to improve textural attributes, water-holding capacity and appearance of the textured plant-based protein product.
  • transglutaminase may improve the firmness, viscoelasticity water binding and gelling capacity of the compositions or food products.
  • Transglutaminase is an enzyme that catalyzes the formation of cross-links both within a protein molecule and between molecules of different proteins.
  • transglutaminase catalyzes covalent cross-link formation between primary amines (such as e amino group of lysine) and the amide group of glutamine residues to form high molecular weight polymers, resulting in improvement of mechanical properties.
  • MTGase is used as a texturizing agent in several meat products, such as sausages, burgers and restructured meats.
  • transglutaminase inclusion facilitates strong cohesion of meat blocks or multi-meat compositions without the need for thermal processing or addition of salt or phosphates.
  • Use of transglutaminase in meat processing can significantly improve the texture of the final product, which results in, for example, an increase in its hardness. Moreover, it strengthens the texture of homogenized sausages made of pork, beef or poultry meat.
  • transglutaminase mediated textured plant-based protein product has the potential to offer new technological opportunities for producing restructured meats, fine and coarse-minced sausages, fresh and frozen hamburgers, hot dogs and similar products.
  • Transglutaminase may be used in the textured plant-based protein products of the present invention to improve gelation through the formation of covalent intramolecular and interm olecular bonds.
  • transglutaminase may increase hardness, springiness, chewiness, and cutting-force of the textured plant-based meat analog of the present invention.
  • Transglutaminase may be used at amounts known in the art, generally up to about 0.75%.
  • Microbial transglutaminase are active over a broad range of pH 4.5 to 8.0 and temperatures 0° C to 45° C and require calcium.
  • the myceliated high-protein product and/or additional high-protein material can be mixed with transglutaminase in the range of 0.1% to 0.7% to promote crosslinking and form a solid matrix/restructured form that can be further processed at elevated temperatures (> 75° C for 5 mins) to inactivate the enzyme.
  • the methods of the invention further include a method to make a restructured textured plant-based protein product comprising the steps of providing a proteinaceous material, e.g., a myceliated high-protein food product, and mixing with transglutaminase to form the restructured textured plant-based protein product.
  • a proteinaceous material e.g., a myceliated high-protein food product
  • the proteinaceous material includes an additional high-protein food product and other ingredients, excipients or additives as disclosed herein.
  • the invention further includes a restructured textured plant-based protein product meat analog product or extender.
  • Seasonings can be added before or after the extruding and/or cooking and/or puffing steps.
  • Seasonings include, but are not limited to, minerals such as salt, grain-based seasonings (such as, but not limited to, whole, cracked or ground wheat, corn, oats, rye, flax, barley, spelt and rice), plant-derived seasonings (such as, but not limited to, onion, garlic, pepper, capsicum pepper, herbs, spices, nuts, olives, fruits, vegetables, etc.), and other flavorings (such as, but not limited to, vanilla, sugar, cheese, yeast extract, whey), and combinations thereof.
  • minerals such as salt, grain-based seasonings (such as, but not limited to, whole, cracked or ground wheat, corn, oats, rye, flax, barley, spelt and rice), plant-derived seasonings (such as, but not limited to, onion, garlic, pepper, capsicum pepper, herbs, spices, nuts, olives, fruits, vegetables,
  • Vitamins can also be included such as, but not limited to, niacin, iron, zinc, thiamine mononitrate (vitamin Bl), riboflavin (vitamin B2), folic acid, tocopherol(s) (vitamin E), vitamin C, vitamin B6, vitamin B 12, vitamin A, vitamin D, pantothenic acid and copper.
  • Edible oil and fat can also be included. Oils such as, but not limited to, soy, com, canola, sesame, safflower, olive, sunflower, rapeseed, cottonseed, peanut, copra, palm kernel, palm, linseed, lupin, and combinations thereof can be used. Other fats such as butter or lecithin and their mixtures can also be used.
  • ingredients can be included such as emulsifiers (such as, but not limited to, lecithin, soy lecithin), leavening (such as, but not limited to, baking soda, calcium phosphate, yeast), natural and artificial sweeteners, preservatives (such as, but not limited to, BHT, BHA, and tocopherol), fiber (such as, but not limited to, insoluble fiber, soluble fiber (e.g., Fibersol®)), and any combinations of such ingredients.
  • emulsifiers such as, but not limited to, lecithin, soy lecithin
  • leavening such as, but not limited to, baking soda, calcium phosphate, yeast
  • preservatives such as, but not limited to, BHT, BHA, and tocopherol
  • fiber such as, but not limited to, insoluble fiber, soluble fiber (e.g., Fibersol®)
  • dry ingredients includes all the ingredients in the mixture to form the textured plant-based protein product except for added water and ingredients added with the
  • the ingredients utilized to make the textured plant- based protein product may include an edible lipid component that comprises one or more edible lipids.
  • an edible lipid component that comprises one or more edible lipids.
  • nearly any edible lipid material may be employed, including natural and synthetic oils, for example, rapeseed, canola, soybean, coconut, cottonseed, peanut, palm and corn oils and in either non-hydrogenated or hydrogenated form.
  • the edible lipid material is an edible vegetable oil, such as canola oil, coconut oil, cottonseed oil, peanut oil, and olive oil.
  • the total edible lipid content is no more than about 5% of the weight of the dry ingredients utilized the make the meat analog product.
  • the total edible lipid content is an amount of about 0.1% to about 1% by weight of the dry ingredients.
  • the total edible lipid content is an amount of about 0.2% to about 0.5% by weight of the dry ingredients.
  • the textured plant-based protein product can optionally include water at a relatively high amount.
  • the total moisture level of the mixture extruded to make the textured plant-based protein product is controlled such that the textured plant-based protein product has a moisture content that is at least about 50% by weight.
  • water is typically added to the ingredients.
  • a relatively high moisture content is desirable, it may not be desirable for the textured plant-based protein product to have a moisture content much greater than about 65%.
  • the amount of water added to the ingredients and the extrusion process parameters are controlled such that the textured plant-based protein product (following extrusion) has a moisture content that is from about 40% to about 65% by weight.
  • the textured plant-based protein product may be optionally dried to form the final commercial product, which is then rehydrated by the finished food product manufacturer.
  • a texturized plant protein product can include between about 1 - 99%, preferably 85% to 95%, of a combination of a myceliated high- protein food product and an additional high-protein material; in a vegan sausage type product; using a twin screw extruder, the mixture can be further blended with starch or fiber of 5-15% (all percentages w/w dry ingredients).
  • Such textured plant-based protein products can be used to create a number of new food compositions, including, without limitation meat analogs and extenders, which contain a myceliated high-protein food product.
  • the methods to prepare a food composition can include the additional, optional steps of cooking, extruding, and puffing the food composition according to methods known in the art to form the food compositions comprising the textured plant-based protein products of the invention.
  • Extrusion is a technology to produce texturized proteins, a unique product which can be produced from a wide range of raw ingredient specifications, while controlling the functional properties such as density, rate and time of rehydration, shape, product appearance and mouthfeel.
  • the general procedure is as follows.
  • the flour mix is prepared and typically the dry ingredients are blended together in the premixture stage.
  • the optional preconditioning step in a section of an extruder device known as preconditioner
  • the steam and water are usually added at this stage to wet/moisten and warm the flour mix.
  • the extruder the majority of the work happens.
  • the starch and protein are plasticized using heat, pressure and/or mechanical shear, then realigned and expanded as the mixture exits the extruder.
  • the material coming from the extruder moisture ranges from 25% to 30%.
  • this extruded material can be dried to about 3% - 5% moisture or less in the dryer portion. Cooling then optionally occurs to lower the temperature of the dried product to ambient conditions followed by an optional packaging step.
  • TPP textured plant protein
  • textured meat-like products are derived from texturization processes.
  • First is a meat extender that has been commonly produced by processing plant proteins through high pressure/extrusion step. Resulting texturized product shows distinct fiber formation and highly expanded. On rehydration the TPP can be used to extend ground meat or meat products.
  • Second type are meat analogs which are used in place of meat.
  • the TPP based meat analogs are dense, have a layered-fiber conformations and maintains meat- like character after extensive cooking or retorting. High moisture extrusion processes have been used to produce meat analogs.
  • a typical extruder can be made up of the following parts.
  • the Flour Mix
  • Feeder the bin/feeder provides a means of uniformly metering the raw materials (granular or floury in nature) into a preconditioner or mixing cylinder and subsequently into the extruder itself. Also controls the product rate or throughput of the entire system.
  • Preconditioner here steam and water are metered and injected into the raw material to control raw material temperature and moisture. Preconditioning promotes moisture and heat penetration of the individual particles, resulting in uniform moisture application and elevated raw material temperature. In the absence of a preconditioner, a longer extruder barrel with adequate mixing and hydration zones may serve the purpose of a preconditioner. Typical moisture content of the raw material exiting the preconditioner range from 10% to 25% and temperatures in the range of 1 l0°F to l20°F.
  • extruder portion itself is described as follows. Extruders are popularly classified as either being a single or twin-screw design. In both designs, the impact on final product texture is affected by screw and barrel profile, screw speed, processing conditions such as temperature, moisture, pressure, raw material characteristics and die selection.
  • the initial section of the extruder barrel is designed to act as a feeding or metering zone and simply conveys the raw or preconditioned vegetable protein away from the inlet portion of the barrel and into the extruder.
  • Product then enters a processing zone where the amorphous, free-flowing vegetable protein is worked into a colloidal dough.
  • the compression ratio of the screw profile is increased in this stage to assist in blending water or steam with the raw material.
  • a food composition of the invention can also include a texturized protein, such as a texturized plant protein.
  • Texturized plant protein comprising the myceliated high-protein product of the present invention include meat analog products and methods for making meat analog products comprising the myceliated high-protein product as disclosed within.
  • the meat analog products can be produced with high moisture content and provide a product that simulates the fibrous structure of animal meat and has a desirable meat-like moisture, texture, mouthfeel, flavor and color.
  • Meat analog products having qualities (for example, texture, moisture, mouthfeel, flavor, and color) similar to that of whole muscle animal meat may be produced using non-animal proteins formed using extrusion under conditions of relatively high moisture.
  • meat analog products may include myceliated high- protein products of the invention, an additional high-protein material, one or more of flour, starch, an edible lipid material and optionally a flavor modifier.
  • extrusion processing is widely used in the modern food industry. Extrusion processing is a multi-step and multifunctional operation, which leads to mixing, hydration, shear, homogenization, compression, deaeration pasteurization or sterilization, stream alignment, shaping, expansion and/or fiber formation.
  • the non-animal protein typically introduced to the extruder in the form of a dry blend, is processed to form a fibrous material.
  • Texturization of protein is the development of a texture or a structure via a process involving heat, and/or shear and the addition of water.
  • the texture or structure will be formed by protein fibers that will provide a meat-like appearance and perception when consumed.
  • the mechanism of texturization of proteins starts with the hydration and unfolding of a given protein by breaking intramolecular binding forces by heat and/or shear.
  • the unfolded proteins molecules are aligned and bound by shear, forming the characteristic fibers of a meat-like product.
  • texturization into fibrous meat analogs for example, through extrusion processing has been an accepted approach.
  • extrusion processes are well known in the art and appropriate techniques can be determined by one of skill.
  • "Extrusion” is a process used to create objects of a fixed cross-sectional profile. A material is pushed or pulled through a die of the desired cross-section. High-moisture extrusion is known as wet extrusion.
  • Extruders typically comprise an extruder barrel within which rotates a close-fitting screw. The screw is made up of screw elements, some of which are helical screw threads to move material through the extruder barrel. Material is introduced into the extruder barrel toward one end, moved along the extruder barrel by the action of the screw and is forced out of the extruder barrel through a nozzle or die at the other end.
  • the rotating screw mixes and works the material in the barrel and compresses it to force it through the die or nozzle.
  • the degree of mixing and work to which the material is subjected, the speed of movement of the material through the extruder barrel and thus the residence time in the extruder barrel and the pressure developed in the extruder barrel can be controlled by the pitch of the screw thread elements, the speed of rotation of the screw and the rate of introduction of material into the extruder barrel.
  • the extruder barrel comprises multiple extruder barrel sections which are joined end to end.
  • extruder barrel sections are required to carry out different processes involved in extrusion such as conveying, kneading, mixing, devolatilizing, metering and the like.
  • Each extruder barrel section comprises a liner which is press fit into an extruder barrel casing, and heating and cooling elements are provided to regulate temperature of extruder barrel section within permissible range.
  • the total length of an extrusion process can be defined by its modular extrusion barrel length.
  • An extruder barrel is described by its unit of diameter.
  • a "cooling die” is cooling the extruded product to a desired temperature.
  • Hot extrusion is used to thermomechanically transform raw materials in short time and high temperature conditions under pressure.
  • Steam-induced expansion means melt expansion at the die exit due to water flashing off, hence leading to highly expanded products. Subsequent processing then determines the textural attributes of extruded products such as crispness, crunchiness, hardness, etc.
  • the extruding can include, for example, melting and/or plasticization of the ingredients, gelatinization of starch and denaturation of proteins.
  • the heat can be applied either through, for example, steam injection, external heating of the barrel, or mechanical energy.
  • the material can be pumped, shaped and expanded, which forms the porous and fibrous texture, and partially dehydrates the product. The shape and size of the final product can be varied by using different die configurations.
  • Extruders can be used to make products with little expansion (such as pasta), moderate expansion (shaped breakfast cereal, meat substitutes, breading substitutes, modified starches, pet foods (soft, moist and dry)), or a great deal of expansion (puffed snacks, puffed curls and balls, etc.).
  • the material may be extruded by means of a ram or a piston.
  • Other extruders use one or more screws.
  • Variable pitch single screw extruders produce high product consistency by combining the ingredients to produce a homogeneous mixture and pushing it out of the machine at a rate that is highly controllable.
  • Twin screw extruders contain two screws that are either co-current (the screws rotate in the same direction) or are counter-current (the screws rotate in opposite directions).
  • Twin screw extruders can handle material with a wide range of moisture content and have greater control over the residence time and the amount of shear to which the material is exposed.
  • the ingredients may be fed into the extruder via a feeder, such as, but not limited to, a gravimetric or volumetric feeder.
  • a feeder such as, but not limited to, a gravimetric or volumetric feeder.
  • the type of feeder used depends on the type of ingredient, and different feeders are used for batch versus continuous feed.
  • the feeder also can direct the ingredients into a preconditioner, if desired.
  • the feed section of the screw may have deep flights to accept the ingredients and move the ingredients forward.
  • the ingredients move into the compression section of the screw, which is heated, and has either shallower or more frequent flights, which compresses the ingredients and works them into continuous dough.
  • the cooking section of the screw applies maximum heat, pressure and shear to the mixture in the barrel prior to the die. Within the screw barrel, the mixture is heated and pressurized.
  • the reduction in pressure to atmospheric pressure generally causes the mixture to expand. If the moist dough within the barrel is heated over 100° C., the sudden reduction in pressure to atmospheric pressure causes the moisture to convert to steam. The combination of sudden expansion and associated cooling yields a puffed, crisp product.
  • One suitable extrusion device is a double-barrel, twin screw extruder as described, for example, in U.S. Pat. No. 4,600,311.
  • Examples of commercially available double-barrel, twin screw extrusion apparatus include a CLEXTRAL Model BC-72 extruder manufactured by Clextral, Inc. (Tampa, Fla.); a WENGER Model TX-57 extruder manufactured by Wenger (Sabetha, Kans.); and a WENGER Model TX-52 extruder manufactured by Wenger (Sabetha, Kans.).
  • Other conventional extruders suitable for use in this disclosure are described, for example, in ET.S. Pat. Nos. 4,763,569, 4,118,164, and 3,117,006, which are hereby
  • the screws of a twin-screw extruder can rotate within the barrel in the same (co- rotating) or opposite directions (counter-rotating). Rotation of the screws in the same direction is referred to as single flow whereas rotation of the screws in opposite directions is referred to as double flow (counter-rotating).
  • co-rotating twin screw extruders are used in the manufacturing of puffed snack products.
  • the speed of the screw or screws of the extruder may vary depending on the particular apparatus. However, the screw speed is typically from about 200 to about 600 revolutions per minute (rpm). Generally, as the screw speed increases, the density of the extrudates decreases. Particularly, the screw speed of the twin screw extruder may effect residence time of the dough in the extruder, the amount of shear generated, and the degree of cooking of the dough; as the screw speed increases, residence time decreases, and the amount of shear increases.
  • conventional extruder systems generally act as an ingredient mixer (e.g., to form the dough), mixture cooker, and composition (extrudate) former. Each of these functions can be accomplished in the same cooking extruder. In some instances, however, it may be desirable to have at least two extruders arranged in a series.
  • the extrusion apparatus also can include one or more heating zones through which the dough is conveyed under mechanical pressure prior to exiting the extrusion apparatus through an extrusion die.
  • the pressure within the extruder barrel is not narrowly critical.
  • the extrusion mass is subjected to a pressure of at least about 400 psi (about 28 bar).
  • the barrel pressure is dependent on numerous factors including, for example, the extruder screw speed, feed rate of the mixture to the barrel, feed rate of water to the barrel, and the viscosity of the dough within the barrel.
  • Water can also be injected into the extruder barrel to further hydrate the dough. As an aid in forming the dough the water may also act as a plasticizing agent.
  • Water may be introduced to the extruder barrel via one or more injection jets in communication with a heating zone. Typically, water is injected at a rate of from about 2 kg/hr to about 7 kg/hr.
  • the mixture in the barrel typically contains from about 15% to about 30% (by weight) water.
  • the rate of introduction of water to any of the heating zones is generally controlled to promote production of an extrudate having desired characteristics. It has been observed that as the rate of introduction of water to the barrel decreases, the density of the extrudate decreases.
  • the dough in the extrusion apparatus is extruded through a die to produce an extrudate.
  • Extrusion conditions are generally such that the extrudate emerging from the extruder barrel typically has a moisture content of from about 5% to about 20% (by weight extrudate). In one embodiment, the moisture content of the extrudate is from about 5% (by weight extrudate) to about 15% (by weight extrudate). In another embodiment, the moisture content of the extrudate is about 10% (by weight extrudate).
  • the moisture content is derived from water present in the mixture introduced to the extruder, moisture added during preconditioning and/or any water injected into the extruder barrel during processing.
  • high moisture extrusion e.g., extrusion at higher moisture contents (> 40%), also known as wet extrusion
  • wet extrusion can be used to create a fresh (non-dried) product.
  • wet extrusion applications utilize twin screw extruders due to their efficient conveying
  • the dough Upon release of pressure, the dough exits the extruder barrel through the die, superheated water present in the mass flashes off as steam, causing simultaneous expansion (i.e., puffing) of the material.
  • the level of expansion of the extrudate upon exiting of the mixture from the extruder in terms of the ratio of the cross-sectional area of extrudate to the cross-sectional area of die openings is generally less than about 50: 1.
  • the ratio of the cross-sectional area of extrudate to the cross-sectional area of die openings is from about 2: 1 to about 50: 1.
  • the extrudate may be cut after exiting the die.
  • Suitable apparatus for cutting the extrudate include flexible knives manufactured by Wenger (Sabetha, Kans.) and Clextral (Tampa, Fla.).
  • Various die orifice designs will cause different expansions and geometric shapes of the extrudate.
  • the sliced or cut extrudate may be in the shape of a sheet, disc, pellet, rod, string, bar, and the like, or some other shape.
  • compositions made using textured plant-based protein products as disclosed herein include a burger, a patty, a sausage, a meatball, taco meat, or a frankfurter.
  • the invention includes methods to prepare a food composition or compositions which comprise or include a myceliated high-protein food product.
  • the method includes providing a myceliated high protein food product of the invention, e.g., a myceliated high-protein food product, wherein the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein is myceliated by an aqueous fungal culture, in a media comprising at least 50% protein in liquid culture; then, in another step, providing an edible material, and mixing the myceliated high protein food product of the invention and the edible material to form the food composition.
  • a myceliated high protein food product of the invention e.g., a myceliated high-protein food product, wherein the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein is myceliated by an aqueous fun
  • the fungal culture comprises Lentinula edodes, Agaricus spp., Pleurotus spp., Boletus spp., or Laetiporus spp:, and the myceliated high-protein food product is derived from a plant source and has reduced undesirable flavors and reduced undesirable aromas compared to the high-protein material that is not myceliated.
  • the food composition is a texturized plant protein which can be used in making, for example, meat- structured plant protein meat analog or meat extender products.
  • the present invention also includes food compositions made by the invention, and food compositions that comprise a myceliated high-protein food product, wherein the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein is myceliated by an aqueous fungal culture, in a media comprising at least 50% protein in liquid culture; and an edible material.
  • the method for preparing food compositions which comprise or include a myceliated high-protein food product wherein the step of providing a myceliated high protein food product can include the following steps.
  • the myceliated high protein food product is produced by culturing a fungus in an aqueous media which includes a high-protein material, with amounts of protein of at least 50% (w/w) protein on a dry weight basis, and wherein the media comprises at least 50 g/L protein, inoculating with a fungal culture, and culturing the medium to produce the myceliated high protein food products whereby the flavor or taste of the myceliated high-protein food product is modulated compared to the high-protein material; providing an edible material; and mixing the myceliated high protein food product and the edible material to form the food composition.
  • the fungal culture comprises Lentinula edodes, Agaricus spp., Pleurotus spp., Boletus spp., or Laetiporus spp:, and the myceliated high-protein food product is derived from a plant source and has reduced undesirable flavors and reduced undesirable aromas compared to the high-protein material that is not myceliated.
  • the food composition is a texturized plant protein which can be used in making, for example, meat- structured plant protein meat analog or meat extender products.
  • the present invention also includes food compositions made by the invention.
  • compositions of the invention can be used to create a number of new food compositions, including, without limitation, dairy alternative products, ready to mix beverages and beverage bases; extruded and extruded/puffed products; sheeted baked goods; meat analogs and extenders; bar products and granola products; baked goods and baking mixes; granola; and soups/soup bases.
  • the methods to prepare a food composition can include the additional, optional steps of cooking, extruding, and/or puffing the food composition according to methods known in the art to form the food compositions comprising the myceliated high protein food product of the invention.
  • the edible material can be, without limitation, a starch, a flour, a grain, a lipid, a colorant, a flavorant, an emulsifier, a sweetener, a vitamin, a mineral, a spice, a fiber, a protein isolate/concentrate or powder thereof, nutraceuticals, sterols, isoflavones, lignans, glucosamine, an herbal extract, xanthan, a gum, a hydrocolloid, a starch, a preservative, a legume product, a food particulate, and combinations thereof.
  • a food particulate can include cereal grains, cereal flakes, crisped rice, puffed rice, oats, crisped oats, granola, wheat cereals, protein nuggets, texturized plant protein ingredients, flavored nuggets, cookie pieces, cracker pieces, pretzel pieces, crisps, soy grits, nuts, fruit pieces, com cereals, seeds, popcorn, yogurt pieces, and combinations of any thereof.
  • the edible materials can be added to the myceliated high protein products of the invention using conventional processes to form the inventive food compositions, as described in more detail herein below.
  • the present invention can also include extruded and/or puffed products and/or cooked products comprising a myceliated high protein food product of the invention.
  • Extruded and/or puffed ready-to-eat breakfast cereals and snacks such as crisps or scoops and pasta noodles are known in the art. Extrusion processes are well known in the art and appropriate techniques can be determined by one of skill. "Extrusion” is a process used to create objects of a fixed cross-sectional profile. A material is pushed or pulled through a die of the desired cross-section. The two main advantages of this process over other
  • Extruders typically comprise an extruder barrel within which rotates a close-fitting screw. The screw is made up of screw elements, some of which are helical screw threads to move material through the extruder barrel.
  • the extruder barrel comprises multiple extruder barrel sections which are joined end to end.
  • extruder barrel sections are required to carry out different processes involved in extrusion such as conveying, kneading, mixing, devolatilizing, metering and the like.
  • Each extruder barrel section comprises a liner which is press fit into an extruder barrel casing, and heating and cooling elements are provided to regulate temperature of extruder barrel section within permissible range.
  • the total length of an extrusion process can be defined by its modular extrusion barrel length.
  • An extruder barrel is described by its unit of diameter.
  • a "cooling die” is cooling the extruded product to a desired temperature.
  • cold extrusion is used to gently mix and shape dough, without direct heating or cooking within the extruder.
  • food processing it is used mainly for producing pasta and dough.
  • These products can then be subsequently processed: dried, baked, vacuum- packed, frozen, etc.
  • Hot extrusion is used to thermomechanically transform raw materials in short time and high temperature conditions under pressure.
  • food processing it is used mainly to cook biopolymer-based raw materials to produce textured food and feed products, such as ready- to-eat breakfast cereals, snacks (savory and sweet), pet foods, feed pellets, etc.
  • the extruding can include, for example, melting and/or plasticization of the ingredients, gelatinization of starch and denaturation of proteins.
  • the heat can be applied either through, for example, steam injection, external heating of the barrel, or mechanical energy.
  • the material can be pumped, shaped and expanded, which forms the porous and fibrous texture, and partially dehydrates the product.
  • the shape and size of the final product can be varied by using different die configurations.
  • Extruders can be used to make products with little expansion (such as pasta), moderate expansion (shaped breakfast cereal, meat substitutes, breading substitutes, modified starches, pet foods (soft, moist and dry)), or a great deal of expansion (puffed snacks, puffed curls and balls, etc.).
  • the myceliated high protein food product of the invention may be used in formulating foods made by extrusion and/or puffing and/or cooking processes, such as ready to eat breakfast cereals and snack foods. These materials are formulated primarily with cereal grains and may contain flours from one or more cereal grains.
  • the textured plant protein products of the invention, as well as breakfast cereal and snack materials can obtain the desired flake structure by a process known as puffing.
  • a cereal is puffed by causing trapped moisture in the flake to change very rapidly from the liquid state to the vapor phase. Rapid heating or a rapid decrease in pressure are the methods commonly used throughout the industry. Gun puffing is an example of the principle of a rapid decrease in pressure. In this process the cereal flakes are first heated under high pressure and then the pressure is rapidly released to achieve the puffing effect.
  • the process disclosed in US. Pat. No. 3,253,533 is an example of a rapid heating puffing method.
  • the present invention includes a method step to prepare a myceliated high-protein food product.
  • the method may optionally include the steps of providing an aqueous media comprising a high-protein material.
  • the aqueous media may comprise, consist of, or consist essentially of at least 20% protein (w/w), on a dry weight basis, and is 50 g/L protein.
  • the media may also comprise, consist of or consist essentially of optional additional excipients as identified herein below.
  • the aqueous media may be inoculated with a fungal culture.
  • the inoculated media may then be cultured to produce a myceliated high-protein food product, and the myceliated high-protein food product taste, flavor, and/or aroma may be modulated compared to the high-protein material in the absence of the culturing step.
  • the aqueous media may comprise, consist of, or consist essentially of a high- protein material.
  • the high-protein material to include in the aqueous media can be obtained from a number of sources, including vegetarian sources (e.g., plant sources) as well as non vegetarian sources, and can include a protein concentrate and/or isolate.
  • Vegetarian sources include meal, protein concentrates and isolates prepared from a vegetarian source such as pea, rice, soy, cyanobacteria, grain, hemp, chia, chickpea, potato protein, algal protein and nettle protein or combinations of these.
  • the vegetarian source is pea, rice, chickpea or a combination thereof.
  • the vegetarian source is pea, chickpea or a combination thereof.
  • the vegetarian source is rice, chickpea, or a combination thereof.
  • a protein concentrate is made by removing the oil and most of the soluble sugars from a meal, such as soybean meal. Such a protein concentrate may still contain a significant portion of non-protein material, such as fiber. Typically, protein concentrations in such products are between 55 - 90%. The process for production of a protein isolate typically removes most of the non-protein material such as fiber and may contain up to about 90 - 99% protein.
  • a typical protein isolate is typically subsequently dried and is available in a powdered form and may alternatively be called "protein powder.”
  • Non- vegetarian sources for the high-protein material may also be used in the present invention.
  • Such non- vegetarian sources include whey, casein, egg, meat (beef, chicken, pork sources, for example), isolates, concentrates, broths, or powders.
  • mixtures of any of the high-protein materials disclosed herein can be used to provide, for example, favorable qualities, such as a more complete (in terms of amino acid composition) high-protein material.
  • high-protein materials such as pea protein and rice protein can be combined.
  • the ratio of a mixture can be from 1 : 10 to 10: 1 pea protein: rice protein (on a dry basis).
  • the ratios can optionally be 5: 1 to 1 :5, 2: 1 to 1 :2, or in one embodiment, 1 : 1. In other embodiments, the ratio can be 65:35.
  • the high-protein material itself can be about 20% protein, 30% protein, 40% protein, 45% protein, 50% protein, 55% protein, 60% protein, 65% protein, 70% protein,
  • This invention discloses the use of concentrated media, which provides, for example, an economically viable economic process for production of an acceptably tasting and/or flavored high-protein food product.
  • the total media concentration is up to 150 g/L but can also be performed at lower levels, such as 5 g/L. Higher concentrations in media result in a thicker and/or more viscous media, and therefore are optionally processed by methods known in the art to avoid engineering issues during culturing or fermentation.
  • a greater amount of high-protein material per L media is used. The amount is used is chosen to maximize the amount of high- protein material that is cultured, while minimizing technical difficulties in processing that may arise during culturing such as viscosity, foaming and the like.
  • the amount of total protein in the aqueous media may comprise, consist of, or consist essentially of at least 20 g, 25 g, 30 g, 35 g, 40 g, 45 g, 50 g, 55 g, 60 g, 65 g, 70 g, 75 g, 80 g, 85 g, 90 g, 95 g, or 100 g, or more, of protein per 100 g total dry weight w/w, or per total all components on a dry weight basis.
  • the amount of protein comprises, consist of, or consist essentially of between 20 g to 90 g, between 30 g and 80 g, between 40 g and 70 g, between 50 g and 60 g, of protein per 100 g dry weight w/win the media.
  • the total protein in aqueous media is about 45 g to about 100 g, or about 80-100 g of protein per 100 g dry weight w/w.
  • the aqueous media comprises between about 1 g/L and 200 g/L, between about 5 g/L and 180 g/L, between about 20 g/L and 150 g/L, between about 25 g/L and about 140 g/L, between about 30 g/L and about 130 g/L, between about 35 g/L and about 120 g/L, between about 40 g/L and about 110 g/L, between about 45 g/L and about 105 g/L, between about 50 g/L and about 100 g/L, between about 55 g/L and about 90 g/L, or about 75 g/L protein; or between about 50 g/L-l50 g/L, or about 75 g/L and about 120 g/L, or about 85 g/L and about 100 g/L.
  • the aqueous media comprises at least about 10 g/L, at least about 15 g/L, at least about 20 g/L, at least about 25 g/L, at least about 30 g/L, at least about 35 g/L, at least about 40 g/L or at least about 45 g/L protein.
  • the amount to use includes between about 1 g/L and 150 g/L, between about 10 g/L and 140 g/L, between about 20 g/L and 130 g/L, between about 30 g/L and about 120 g/L, between about 40 g/L and about 110 g/L, between about 50 g/L and about 100 g/L, between about 60 g/L and about 90 g/L, between about 70 g/L and about 80 g/L, or at least about 20 g/L, at least about 30 g/L, at least about 40 g/L, at least about 50 g/L, at least about 60 g/L, at least about 70 g/L, at least about 80 g/L, at least about 90 g/L, at least about 100 g/L, at least about 110 g/L, at least about 120 g/L, at least about 130 g/L or at least about 140 g/L.
  • the aqueous media comprises between about 50 g/L and about 100 g/L, or about 80 g/L, about 85 g/L, about 90 g/L, about 95 g/L about 100 g/L, about 110 g/L, about 120 g/L, about 130 g/L, about 140 g/L, or about 150 g/L.
  • the high-protein material after preparing the aqueous media of the invention, is not completely dissolved in the aqueous media. Instead, the high- protein material may be partially dissolved, and/or partially suspended, and/or partially colloidal. However, even in the absence of complete dissolution of the high-protein material, positive changes may be affected during culturing of the high-protein material. In one embodiment, the high-protein material in the aqueous media is kept as homogenous as possible during culturing, such as by ensuring agitation and/or shaking.
  • the aqueous media further comprises, consists of, or consists essentially of materials other than the high-protein material, e.g., excipients as defined herein and/or in particular embodiments.
  • Excipients can comprise any other components known in the art to potentiate and/or support fungal growth, and can include, for example, nutrients, such as proteins/peptides, amino acids as known in the art and extracts, such as malt extracts, meat broths, peptones, yeast extracts and the like; energy sources known in the art, such as carbohydrates; essential metals and minerals as known in the art, which includes, for example, calcium, magnesium, iron, trace metals, phosphates, sulphates; buffering agents as known in the art, such as phosphates, acetates, and optionally pH indicators (phenol red, for example).
  • Excipients may include carbohydrates and/or sources of carbohydrates added to media at 5-10 g/L. It is usual to add pH indicators to such formulations.
  • Excipients may also include peptones/proteins/peptides, as is known in the art. These are usually added as a mixture of protein hydrolysate (peptone) and meat infusion, however, as used in the art, these ingredients are typically included at levels that result in much lower levels of protein in the media than is disclosed herein. Many media have, for example, between 1% and 5% peptone content, and between 0.1 and 5% yeast extract and the like.
  • excipients include for example, yeast extract, malt extract, maltodextrin, peptones, and salts such as diammonium phosphate and magnesium sulfate, as well as other defined and undefined components such as potato or carrot powder.
  • organic as determined according to the specification put forth by the National Organic Program as penned by the ETSDA) forms of these components may be used.
  • excipients comprise, consist of, or consist essentially of dry carrot powder, dry malt extract, diammonium phosphate, magnesium sulfate, and citric acid. In one embodiment, excipients comprise, consist of, or consist essentially of dry carrot powder between 0 1-10 g/L, dry malt extract between 0.1 and 20 g/L, diammonium phosphate between 0.1 and 10 g/L, and magnesium sulfate between 0.1 and 10 g/L.
  • Excipients may also optionally comprise, consist of, or consist essentially of citric acid and an anti-foam component.
  • the anti-foam component can any anti-foam component known in the art, such as a food-grade silicone anti-foam emulsion or an organic polymer anti-foam (such as a polypropylene-based polyether composition).
  • the medium comprises, consists of or consists essentially of the high protein material as defined herein and an anti-foam component, without any other excipients present.
  • the method may also comprise the optional step of sterilizing the aqueous media prior to inoculation by methods known in the art, including steam sterilization and all other known methods to allow for sterile procedure to be followed throughout the inoculation and culturing steps to enable culturing and myceliation by pure fungal strains.
  • the components of the media may be separately sterilized and the media may be prepared according to sterile procedure.
  • the method also includes inoculating the media with a fungal culture.
  • the fungal culture may be prepared by culturing by any methods known in the art.
  • the methods to culture may be found in, e.g., PCT/US 14/29989, filed March 15, 2014, PCT/US14/29998, filed March 15, 2014, all of which are incorporated by reference herein in their entireties.
  • the fungal cultures prior to the inoculation step, may be propagated and maintained as is known in the art.
  • the fungi discussed herein can be kept on 2 - 3% (v/v) fruit puree with 3 - 4% agar (m/v).
  • Such media is typically prepared in 21.6 L handled glass jars being filled with 1.4 - 1.5 L media.
  • Such a container pours for 50 -60 90 mm Petri plates.
  • the media is first sterilized by methods known in the art, typically with an autoclave. Conventional B. stearothermophilus and thermocouple methods are used to verify sterilization parameters. Some strains, such as L. sulfureus , grow better when supplemented with 1% yellow cornmeal.
  • Agar media can also be composed of high-protein material to sensitize the strain to the final culture. This technique may also be involved in strain selection of the organisms discussed herein. Agar media should be poured when it has cooled to the point where it can be touched by hand ( ⁇ 40 - 50 °C).
  • maintaining and propagating fungi for use for inoculating the high-protein material as disclosed in the present invention may be carried out as follows. For example, a propagation scheme that can be used to continuously produce material according to the methods is discussed herein.
  • Petri plate cultures can be used at any point to propagate mycelium into prepared liquid media.
  • plates can be propagated at any point during log phase or stationary phase but are encouraged to be used within three months and in another embodiment within 2 years, though if properly handled by those skilled in the art can generally be stored for as long as 10 years at 4°C and up to 6 years at room temperature.
  • liquid cultures used to maintain and propagate fungi for use for inoculating the high-protein material as disclosed in the present invention include undefined agricultural media with optional supplements as a motif to prepare culture for the purposes of inoculating solid-state material or larger volumes of liquid.
  • liquid media are typically inoculated with agar, liquid and other forms of culture.
  • Bioreactors provide the ability to monitor and control aeration, foam, temperature, and pH and other parameters of the culture and as such enables shorter myceliation times and the opportunity to make more concentrated media.
  • the fungi for use for inoculating the high-protein material as disclosed in the present invention may be prepared as a submerged liquid culture and agitated on a shaker table, or may be prepared in a shaker flask, by methods known in the art and according to media recipes disclosed in the present invention.
  • the fungal component may be prepared from a glycerol stock, by a simple propagation motif of Petri plate culture to 0.5 4 L Erlenmeyer shake flask to 50 glycerol stock. Petri plates can comprise agar in 10 - 35 g/L in addition to various media components.
  • Petri plates growing anywhere from 1— 3,652 days can be propagated into 0.5 4 L Erlenmeyer flasks (or 250 to 1,000 mL Wheaton jars, or any suitable glassware) for incubation on a shaker table or stationary incubation.
  • the shaking is anywhere from 40 160 RPM depending on container size and, with about a 1" swing radius.
  • Liquid- state fermentation agitation and swirling techniques as known in the art are also employed which include mechanical shearing using magnetic stir bars, stainless steel impellers, injection of sterile high-pressure air, the use of shaker tables and other methods such as lighting regimen, batch feeding or chemostatic culturing, as known in the art.
  • culturing step is carried out in a bioreactor which is ideally constructed with a torispherical dome, cylindrical body, and spherical cap base, jacketed about the body, equipped with a magnetic drive mixer, and ports to provide access for equipment comprising DO, pH, temperature, level and conductivity meters as is known in the art. Any vessel capable of executing the methods of the present invention may be used. In another embodiment the set-up provides 0.1 5.0 ACH. Other engineering schemes known to those skilled in the art may also be used.
  • the reactor can be outfitted to be filled with sterile water.
  • the entire media is sterilized in situ while in another embodiment concentrated media is sterilized and diluted into a vessel filled water that was filter and/or heat sterilized, or sufficiently treated so that it doesn’t encourage contamination over the colonizing fungus.
  • high temperature high pressure sterilizations are fast enough to be not detrimental to the media.
  • the entire media is sterilized in continuous mode by applying high temperature between 130° and l50°C for a residence time of 1 to 15 minutes.
  • the tank can be mildly agitated and inoculated.
  • the media can be heat sterilized by steaming either the jacket, chamber or both while the media is optionally agitated.
  • the medium may optionally be pasteurized instead.
  • the reactor is used at a large volume, such as in 500,000 - 200,000 L working volume bioreactors.
  • a large volume such as in 500,000 - 200,000 L working volume bioreactors.
  • the culture must pass through a successive series of larger bioreactors, any bioreactor being inoculated at 0.5 - 15% of the working volume according to the parameters of the seed train.
  • a typical process would pass a culture from master culture, to Petri plates, to flasks, to seed bioreactors to the final main bioreactor when scaling the method of the present invention.
  • 3 - 4 seeds may be used.
  • the media of the seed can be the same or different as the media in the main.
  • the fungal culture for the seed is a protein
  • foaming is minimized by use of anti-foam on the order of 0.5 to 2.5 g/L of media, such as those known in the art, including insoluble oils, polydimethylsiloxanes and other silicones, certain alcohols, stearates and glycols.
  • lowering pH assists in culture growth, for example, for L. edodes pH may be adjusted by use of citric acid or by any other compound known in the art, but care must be taken to avoid a sour taste for the myceliated high-protein product. The pH may be adjusted to between about 4.5 and 5.5, for example, to assist in growth.
  • the pH does not change during processing.“pH does not change during processing” is understood to mean that the pH does not change in any significant way, taking into account variations in measured pH which are due to instrument variations and/or error.
  • the pH will stay within about plus or minus 0.3 pH units, plus or minus 0.25 pH units, plus or minus 0.2 pH units, plus or minus 0.15 pH units, or plus or minus 0.1 pH units of a starting pH of the culture during the myceliation, e.g. processing step.
  • a minor change in pH can be defined as a pH change of plus or minus 0.5 pH units or less, plus or minus 0.4 pH units or less, plus or minus 0.3 pH units or less, plus or minus 0.25 pH units or less, plus or minus 0.2 pH units or less, plus or minus 0.15 pH units or less, or plus or minus 0.1 pH units or less of a starting pH.
  • Fungi useful for the present invention are from the higher order Basidio- and
  • fungi effective for use in the present invention include, but are not limited to, Lentinula spp., such as L. edodes, Agaricus spp., such as A. blazei, A. bisporus, A. campestris, A. subrufescens, A. brasiliensis, or A. silvaticus; Pleurotus spp., Boletus spp., or Laetiporus spp.
  • the fungi for the invention include fungi from optionally, liquid culture of species generally known as oyster, porcini,‘chicken of the woods’ and shiitake mushrooms.
  • Pleurotus (oyster) species such as Pleurotus ostreatus, Pleurotus salmoneostramineus (Pleurotus djamor), Pleurotus eryngii, or Pleurotus citrinopileatus
  • Boletus (porcini) species such as Boletus edulis
  • Laetiporus (chicken of the woods) species such as Laetiporus sulfureus , and many others such as L. budonii, L.
  • L. flos-musae L. discolor
  • Lentinula (shiitake) species such as . edodes.
  • Lepista nuda Hericium erinaceus, Agaricus blazeii , and combinations thereof.
  • the fungi is Lentinula edodes.
  • the filamentous fungus may comprise, consist of, or consist essentially of Hericium erinaceus, Lentinula edodes, Cantharellus cibarius, Cordyceps sinensis, Ganoderma lucidum, Laetiporus sulphureus, Laetiporus multiplinnatus, Morchella angusticeps, Morchella importuna, Grifola frondosa, Grifola curtisii, Pleurotus ostreatus, Pleurotus umbellatus, Volvariella volvacea, Pleurotus salmoneostramineus (Pleurotus djamor), Pleurotus eryngii, Pleurotus citrinopileatus, Cantherellus cibarius (chanterelle), Fumaria officianalis, Fistulina hepatica, Sparassis crispa, Inonotus obliquus, Cordyce
  • Fungi may be obtained commercially, for example, from the Penn State
  • Mushroom Culture Collection Strains are typically received as "master culture” PDY slants in 50 mL test tubes and are stored at all, but for A. blazeii , stored at 4 °C until plated.
  • small pieces of culture are typically transferred into sterile shake flasks (e.g. 250 mL) so as not to contaminate the flask filled with a sterilized media (liquid media recipes are discussed below).
  • Inoculated flasks shake for approximately ten hours and aliquots of said flasks are then plated onto prepared Petri plates of a sterile agar media.
  • One flask can be used to prepare dozens to potentially hundreds of Petri plate cultures. There are other methods of propagating master culture though the inventors find these methods as disclosed to be simple and efficient.
  • Determining when to end the culturing step and to harvest the myceliated high- protein food product, which according to the present invention, to result in a myceliated high- protein food product with acceptable taste, flavor and/or aroma profiles can be determined in accordance with any one of a number of factors as defined herein, such as, for example, visual inspection of mycelia, microscope inspection of mycelia, pH changes, changes in dissolved oxygen content, changes in protein content, amount of biomass produced, and/or assessment of taste profile, flavor profile, or aroma profile.
  • harvest can be determined by tracking protein content during culturing and harvest before significant catabolism of protein occurs.
  • biomass may be measured by filtering, such through a filter of 10-1000 pm, and has a protein concentration between 0.1 and 25 g/L; or in one embodiment, about 0.2-0.4 g/L.
  • harvest can occur when the dissolved oxygen reaches about 10% to about 90% dissolved oxygen, or less than about 80% of the starting dissolved oxygen.
  • mycelial products may be measured as a proxy for mycelial growth, such as, total reducing sugars (usually a 40-95% reduction), b-glucan and/or chitin formation; harvest is indicated at 102 - 104 ppm.
  • Other indicators include small molecule metabolite production depending on the strain (e.g. eritadenine on the order of 0.1 - 20 ppm for L. edodes or erinacine on the order of 0.1 - 1,000 ppm for H. erinaceus) or nitrogen utilization (monitoring through the use of any nitrogenous salts or protein, cultures may be stopped just as protein starts to get utilized or may continue to culture to enhance the presence of mycelial metabolites).
  • the total protein yield in the myceliated high-protein food product after the culturing step is about 75% to about 95%.
  • Harvest includes obtaining the myceliated high-protein food product which is the result of the myceliation step.
  • cultures can be processed according to a variety of methods.
  • the myceliated high-protein food product is pasteurized or sterilized.
  • the myceliated high-protein food product is dried according to methods as known in the art. Additionally, concentrates and isolates of the material may be prepared using variety of solvents or other processing techniques known in the art.
  • the material is pasteurized or sterilized, dried and powdered by methods known in the art. Drying can be done in a desiccator, vacuum dryer, conical dryer, spray dryer, fluid bed or any method known in the art. Preferably, methods are chosen that yield a dried myeliated high-protein product (e.g., a powder) with the greatest digestibility and
  • the dried myeliated high-protein product can be optionally blended, pestled milled or pulverized, or other methods as known in the art.
  • the flavor, taste and/or aroma of high-protein materials as disclosed herein such as protein concentrates or isolates from vegetarian or nonvegetarian sources (e.g. egg, whey, casein, beef, soy, rice, hemp, pea, chickpea, soy, cyanobacteria, and chia) may have flavors, which are often perceived as unpleasant, having pungent aromas and bitter or astringent tastes. These undesirable flavors and tastes are associated with their source(s) and/or their processing, and these flavors or tastes can be difficult or impossible to mask or disguise with other flavoring agents.
  • the present invention works to modulate these tastes and/or flavors.
  • flavors and/or tastes of the myceliated high- protein food product or products are modulated as compared to the high-protein material (starting material).
  • both the sterilization and myceliation contribute to the modulation of the resultant myceliated high-protein food products’ taste.
  • the aromas of the resultant myceliated high-protein food products prepared according to the invention are reduced and/or improved as compared to the high-protein material (starting material).
  • undesired aromas are reduced and/or desired aromas are increased.
  • flavors and/or tastes may be reduced and/or improved.
  • desirable flavors and/or tastes may be increased or added to the high-protein material by the processes of the invention, resulting in myceliated high- protein food products that have added mushroom, meaty, umami, popcorn, buttery, and/or other flavors or tastes to the food product.
  • the increase in desirable flavors and/or tastes may be rated as an increase of 1 or more out of a scale of 5 (1 being no taste, 5 being a very strong taste.)
  • Flavors and/or tastes of myceliated high-protein food products may also be improved by processes of the current invention. For example, deflavoring can be achieved, resulting in a milder flavor and/or with the reduction of, for example, bitter and/or astringent tastes and/or beany and/or weedy and/or grassy tastes.
  • the decrease in undesirable flavors and/or tastes as disclosed herein may be rated as a decrease of 1 or more out of a scale of 5 (1 being no taste, 5 being a very strong taste.)
  • Culturing times and/or conditions can be adjusted to achieve the desired aroma, flavor and/or taste outcomes. For example, cultures grown for approximately 2 - 3 days can yield a deflavored product whereas cultures grown for longer may develop various aromas that can change/intensify as the culture grows. As compared to the control and/or high- protein material, and/or the pasteurized, dried and powdered medium not subjected to sterilization or myceliation, the resulting myceliated high-protein food product in some embodiments is less bitter and has a milder, less beany aroma.
  • the myceliated high-protein food products made by the methods of the invention have a complete amino acid profile (all amino acids in the required daily amount) because of the media from which it was made has such a profile. While amino acid and amino acid profile transformations are possible according to the methods of the present invention, many of the products made according to the methods of the present invention conserve the amino acid profile while at the same time, more often altering the molecular weight distribution of the proteome.
  • the oyster fungi Pleurotus ostreatus
  • the oyster fungi can convey a strong savory aroma that leaves after a few seconds at which point a mushroom flavor is noticeable.
  • the strains convey a savory meaty aroma and/or umami, savory or meaty flavor and/or taste.
  • L. edodes and A. blazeii in some embodiments are effective at deflavoring with shorter culturing times, such as 1.5 - 8 days, depending on whether the culture is in a shake flask or bioreactor.
  • L. edodes is particularly good for the deflavoring of pea and rice protein concentrate mixtures.
  • the present invention discloses production of a food composition comprising the myceliated food product made by any of the methods of as disclosed herein, which is then used to mix with other edible components to provide the food compositions as disclosed herein.
  • the invention comprises a food composition for human consumption, comprising a myceliated high-protein food product, myceliated high-protein food product, wherein the myceliated high-protein food product is at least 50% (w/w) protein on a dry weight basis, wherein the myceliated high-protein product is myceliated by an aqueous fungal culture, in a media comprising at least 50 g/L protein in liquid culture; and an edible material.
  • a myceliated high protein food product of the present invention is also referred to herein as PURETASTETM protein, PT, PTP, and the like.
  • the present invention also comprises a food composition
  • a food composition comprising a mixture of a myceliated high-protein food product as defined herein and an edible material.
  • the food composition can comprise, consist of, or consist essentially of at least 10%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95%, protein.
  • Myceliated as used herein, means a high-protein material as defined herein having been cultured with live fungi as defined herein and achieved at least a 1%, at least 2%, at least 3%, at least 4%, at least a 5%, at least a 10%, at least a 20%, at least a 30%, at least a 40%, at least a 50%, at least a 60%, at least a 70%, at least a 80%, at least a 90%, at least a 100%, at least a 120%, at least a 140%, at least a 160%, at least a 180%, at least a 200%, at least a 250%, at least a 300%, at least a 400%, at least a 500% increase in biomass or more, to result in a myceliated high-protein food product.
  • “myceliated” may refer to the distribution of a previously-grown biomass from a filamentous fungus as disclosed herein through the high-protein material.
  • the high-protein material is a protein concentrate or a protein isolate, which may be obtained from vegetarian or nonvegetarian source as defined herein, including pea, rice, chickpea, or combinations thereof.
  • the myceliated high-protein food product can be myceliated by a fungal culture as defined herein.
  • the myceliated high-protein food product can have enhanced meaty, savory, umami, popcorn, and/or mushroom flavors, aromas and/or tastes as compared to the high-protein material.
  • the myceliated high-protein food product has decreased flavors, tastes and/or aromas (deflavoring) leading to a milder and/or an improved flavor, taste or aroma.
  • deflavoring leading to a milder and/or an improved flavor, taste or aroma.
  • reduced bitterness, astringency and/or beany, grassy or weedy tastes are observed.
  • a 7 L bioreactor was filled with 4.5 L of a medium consisting of 5 g/L pea protein concentrate (labeled as 80% protein), 5 g/L rice protein concentrate (labeled as 80% protein),
  • a 250 L bioreactor was filled with 150 L of a medium consisting of 45 g/L pea protein concentrate (labeled as 80% protein), 45 g/L rice protein concentrate (labeled as 80% protein), 1 g/L carrot powder, 1.8 g/L diammonium phosphate, 0.7 g/L magnesium sulfate heptahydrate, 1 g/L anti-foam and 1.5 g/L citric acid and sterilized in place by methods known in the art, being held at 120 - 121 °C for 100 minutes.
  • the bioreactor was inoculated with 5 L of inoculant from two bioreactors as prepared in Example 3. The bioreactor was held at 26 °C.
  • the culture was harvested in 4 days upon successful visible (mycelial pellets) and microscope checks.
  • the culture was plated on LB media to ascertain the extent of any bacterial contamination and none was observed.
  • the culture was then pasteurized at 82 °C for 30 minutes with a ramp up time of 30 minutes and a cool down time of 45 minutes to 17 °C.
  • the culture was finally spray dried and tasted.
  • the final product was noted to have a mild aroma with no perceptible taste at concentrations up to 10%.
  • the product was -75% protein on a dry weight basis.
  • a 250 L bioreactor was filled with 150 L of a medium consisting of 45 g/L pea protein concentrate (labeled as 80% protein), 45 g/L rice protein concentrate (labeled as 80% protein), 1 g/L carrot powder, 1.8 g/L diammonium phosphate, 0.7 g/L magnesium sulfate heptahydrate, 1 g/L anti-foam and 1.5 g/L citric acid and sterilized in place by methods known in the art, being held at 120 - 121 °C for 100 minutes.
  • the bioreactor was inoculated with 5 L of inoculant from two bioreactors as prepared in Example 3. The bioreactor was held at 26 °C.
  • the culture was harvested in 2 days upon successful visible (mycelial pellets) and microscope checks.
  • the culture was plated on LB media to ascertain the extent of any bacterial contamination and none was observed.
  • the culture was then pasteurized at 82 °C for 30 minutes with a ramp up time of 30 minutes and a cool down time of 90 minutes to 10 °C.
  • the culture was finally concentrated to 20% solids, spray dried and tasted.
  • the final product was noted to have a mild aroma with no perceptible taste at concentrations up to 10%.
  • the product was -75% protein on a dry weight basis.
  • the flasks were shaken at 26 °C at 120 RPM for 8-15 days, at which point they were pasteurized as according to the parameters discussed in Example 5, desiccated, pestled and tasted.
  • the G. lucidum product contained a typical‘reishi’ aroma, which most of the tasters found pleasant. The other samples were deemed pleasant as well but had more typical mushroom aromas.
  • the resulting myceliated food products was thought to be much less bitter and to have had a more mild, less beany aroma that was more cereal in character than beany by 5 tasters.
  • the sterilized but not myceliated product was thought to have less bitterness than the non-sterilized control but still had a strong beany aroma. The preference was for the myceliated food product.
  • a 4,000 L bioreactor was filled with 2,500 L of a sterilized medium similar to Example 4, consisting of 45 g/L pea protein concentrate (labeled as 80% protein), 45 g/L rice protein concentrate (labeled as 80% protein), 3.6 g/l maltodextrin, 1.8 g/L carrot powder, 1.8 g/L diammonium phosphate, 0.7 g/L magnesium sulfate heptahydrate, 1.5 g/L anti-foam and 0.6g/L citric acid. Seed reactor was also prepared in 200 L bioreactor with medium volume of 100L with the following medium components: pea protein 5 g/l, rice protein 5 g/l,
  • the medium was inoculated with flask process developed the same way as shown in Example 2.
  • the 200 L bioreactor was harvested 40 to 70 hours post-inoculation.
  • the flasks were harvested 11 days post-inoculation.
  • the organism was Lentinula edodes sourced from the Penn State mushroom culture collection. [00136] Once the main fermenter was cooled it was inoculated with the 100 L inoculum from the 200 L fermenter. Fermenter was held at 26 °C.
  • the culture in the 4,000 L vessel was harvested at 48 hours post-inoculation upon successful visible (mycelial pellets) and microscope checks. Material was pasteurized in the bioreactor at 65° C for 60 minutes.
  • Fermenter was then cooled down and material was harvested in sanitized 55 gallon drums and sent to spray drying facility.
  • the raw pea product prior to myceliation has a pea aroma with no rice or mushroom aroma.
  • the rice samples prior to myceliation have rice aroma with no pea or mushroom aroma. After myceliation, these samples have mushroom aroma and no pea or rice aroma, respectively. There is also increased umami flavor in the myceliated samples.
  • the flasks were covered with a stainless-steel cap and sterilized in an autoclave.
  • the flasks were carefully transferred to a clean HEPA laminar flow hood where they cooled for 4 hours and inoculated with 5% of lO-day old submerged aliquots of each species.
  • All 8 flasks were placed on a shaker table at 150 rpm with a swing radius of 1" at room temperature and incubated for 3 days at which point pH was measured and is essentially the same.
  • a 7 L bioreactor was filled with 4.5 L of a medium consisting of the medium as described in following table (see Table 4): [00153] Table 4.
  • excipients other than an anti-foam were omitted from the fermentation medium, and only rice protein, pea protein, and anti-foam were used as the medium.
  • excipients such as magnesium sulfate, diammonium phosphate (which functions at least in part as a buffer), citric acid, carrot powder, were used and were omitted here. It was theorized that omission of these excipients will encourage the culture to convert protein metabolically and not proliferate. Open ports on the bioreactor were wrapped in foil and the vessel was subsequently sterilized in an autoclave. The bioreactors were carefully transferred to a clean bench in a cleanroom, setup and cooled for 4-6 hours.
  • the bioreactor was inoculated with 5%, 10% and 7.5% of inoculant of L. edodes from a l2-day old flask. Fermentation for these batches was completed in 44 hours, 24 hours and 30 hours respectively for medium 1, medium 2 and medium 3. A microscope check was done to ensure the presence of mycelium (mycelial pellets were visible by the naked eye) and the culture was plated on LB media to ascertain the extent of any bacterial contamination and none was observed. These cultures were pasteurized for 60 minutes at 65 °C.
  • PureTaste® protein (PTP) (made as described in Example 11) were processed to form extruded crisps ranging in protein content with varying degrees of expansion, color and texture.
  • PureTaste® protein crisps flavor ranged from light malted, toasted, grainy notes to very light earthy notes at elevated levels of PureTaste® protein.
  • High-protein (62% Protein) PureTaste® protein extruded scoops produced were tan in color, toasted brown notes, latent slight earthy notes.
  • Scoop PTP (79.25%) + 20% Tapioca Starch. Protein content was 60%. Density was 109 g/L, color was brown, texture was light, crunchy, flavor was toasted.
  • Scoop PTP (85%) + 5% Tapioca Starch + 10% pea protein isolate. Protein content was 72%. There was only limited expansion, color was brown, texture was harder, crunchy, flavor was toasted.
  • Raw material information Dry recipe density 490-530 kg/m 3 ; dry recipe rate 140-160 kg/hr; feeder speed 30-45 rpm; live bin weight 20-25 kg.
  • Preconditioner information Small side speed 750-800 rpm; large side speed, 100- 150 rpm; mixing intensity 70-90%; cylinder steam 0.05-0.2 kg/hr; cylinder water 40-50 kg/hr; cylinder discharge temp, 25-35 °C.
  • Extruder information Extruder speed, 300-500 rpm.
  • the zone 1 temp was 50-70 °C (SP) and 50-70 °C(PV)
  • zone 2 temp was 90-110 °C(SP) and 90-110 °C (PV)
  • zone 3 temp was 110 -130 °C(SP) and 110-130 °C(PV)
  • zone 4 temp was 110-130 °C(SP) and 110- 130 °C(PV)
  • zone 5 temp was 80-100 °C(SP).
  • the die pressure was 1500-2500 kPA.
  • PureTaste® protein (PT) in combination with starch, wheat gluten, pea protein and mineral salts can be processed to produce texturized vegetable protein with final protein content ranging from 60% to 77%, with varying degrees of expansion volume, color and flavor. Flavor was acceptable with malted, grainy and latent bitter notes. Successfully produced texturized PT protein with good to medium hydration capability, soft and harder texture.
  • Procedure used a modified extruder setting, with the addition of a 10-inch spacer or a l2-inch spacer with a Teflon nozzle (Venturi system) additional barrel coated on the inside with Teflon, followed by a narrow exit through a Teflon coated die. This set up provided successful texturization.
  • Test 1 Fine flours of PureTaste® protein, 60%, vital wheat gluten 20%, tapioca starch 20%, were mixed with water to result in 25 to 30% water content. Density was 200 kg/m 3 . After extrusion, the texturized pieces were in chunk form. The extruded material was a white/tan color and was crunchy when dried. When rehydrated, the extruded material had acceptable chewable characteristics, and was softer (not as chewable) compared to a control prepared with soy and had a clean, non-beany flavor and aroma.
  • Test 2 Fine flours of pea protein, 100%. Density was 98 kg/m 3 . After extrusion, the texturized pieces were in chunk form. The extruded material was a medium tan color and was crunchy when dried. When rehydrated, the extruded material was as chewy compared to a control prepared with soy and had a light pea flavor.
  • Test 3 Fine flours of pea protein isolate, 100%. Density was 141 kg/m 3 . After extrusion, the texturized pieces were in chunk form. The extruded material was a medium tan color and was crunchy when dried. When rehydrated, the extruded material was as chewy compared to a control prepared with soy and had a light pea flavor.
  • Test 4 Fine flours of pea flour, 100%. Density was 113 kg/m 3 . After extrusion, the texturized pieces were in chunk form. The extruded material was a medium tan color and was crunchy when dried. When rehydrated, the extruded material was as chewy compared to a control prepared with soy and had a light pea flavor.
  • Test 5 Fine flours of PureTaste® protein, 100%. Density was heavy. After extrusion, the texturized pieces were in crumble form. The extruded material was a medium tan color and had a clean flavor.
  • Test 6 Fine flours of PureTaste® protein, 50% and pea protein isolate 50% were mixed with water to result in 30% water content. Density was 280 kg/m 3 . Following extrusion, the texturized pieces were in shreds form. The extruded material was a light to medium tan color and was crunchy when dried. When rehydrated, the extruded material had acceptable chewable characteristics, and was softer (not as firm) compared to a control prepared with soy and has a clean, non-beany flavor and aroma.
  • Test 7 PureTaste® protein 50%, vital wheat gluten 50%, results in density of 313 kg/m 3 , protein content of 75%, consistency of chunks. When rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and has a clean, non-beany flavor and aroma.
  • Test 8 PureTaste® protein 50%, vital wheat gluten 40%, tapioca starch 10%, results in density of 204 kg/m 3 , protein content of 68%, consistency of chunks. When rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and has a clean, non-beany flavor and aroma.
  • Test 9 PureTaste® protein 60%, vital wheat gluten 20%, tapioca starch 20%, results in density of 177 kg/m 3 , protein content of 60%, consistency of chunks. When rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and has a clean, non-beany flavor and aroma.
  • Test 10 PureTaste® protein 50%, vital wheat gluten 35%, tapioca starch 15%, results in density of 179 kg/m 3 , protein content of 64%, consistency of chunks. When rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and has a clean, non-beany flavor and aroma.
  • Test 11 PureTaste® protein 25%, pea protein isolate, results in density of 132 kg/m 3 , protein content of 79%, consistency of chunks. When rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and has a clean, non-beany flavor and aroma.
  • Test 14 PureTaste® protein 25.75%, vital wheat gluten 33%, tapioca starch 5%, pea protein 15%, pea fiber 20%, with trisodium phosphate 0.5%, results in material that when rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and had a clean, non-beany flavor and aroma.
  • Test 15 PureTaste® protein 32.92%, vital wheat gluten 16.46%, tapioca starch 19.75%, pea protein 29.62%, with trisodium phosphate 0.5%, results in material that when rehydrated, the extruded material had acceptable chewable characteristics, and was almost identical in chewiness and firmness compared to a control prepared with soy (but softer) and had a clean, non-beany flavor and aroma.
  • A. Raw material information Dry recipe density 550-600 kg/m 3 ; dry recipe rate 90-110 kg/hr; feeder speed 20-25 rpm; live bin weight 20-35 kg.
  • Preconditioner information Small side speed 150-800 rpm; large side speed, 100- 130 rpm; mixing intensity 70-90%; cylinder steam 1.8-2.5 kg/hr; cylinder water 25-35 kg/hr; cylinder discharge temp, 40-50 °C.
  • Extruder information Extruder speed, 300-700 rpm; The zone 1 temp was 50-70 °C (SP) and 50-70 °C(PV), zone 2 temp was 90-110 °C(SP) and 90-110 °C (PV); zone 3 temp was 120 - l40°C(SP) and 120-130 °C(PV); zone 4 temp was 125-135 °C(SP) and 115-135 °C(PV); zone 5 temp was 80-100 °C(SP).
  • the die pressure was 2000-2300 kPA.
  • Dryer information zone 1 temperature 120-130 °C; zone 2 temperature, 80-100 °C, zone 3 temperature, 80-100 °C; retention time, Pass 1, 8-12 minutes; Retention time, Pass 2, 7-10 minutes, exhaust 1 temperature 100-130 °C; exhaust 2 temperature 80-100 °C.
  • PureTasteTM texturized protein compared with soy texturized protein.
  • PureTasteTM texturized protein both formulations with pea fiber, by sensory testing, have more spring and the particle size is larger; increased savory flavor and less sweet flavor, and had similar water adsorption. See Table 5 and 6.
  • Flavor attributes Compared with soy texturized protein and pea texturized protein, the flavors of the formulations in this Example had an improvement over pea texturized protein in that both formulations lacked a beany flavor (less pea flavor) and had increased cereal and malty notes, and less metallic and chalky flavor notes. Bitterness for the PureTaste was reduced compared to pea texturized protein. [00214] Texturized Protein Water Adsorption Study; Table 6.
  • Meat analog patty Using the successful PT texturized protein materials made in Example 14, a meat analogue patty was made. The ingredients in wt percent were PureTaste texturized protein 10.42%, PureTaste Protein 6.01%, Pea Protein texturized protein 8.42%, Vital Wheat Gluten 7.62%, Methylcellulose 2.00%, Beef Flavor 2.20%, Grill Flavor, 2.61%, Chicken Flavor 1.80%, Beet Powder 0.70%, Unrefined Coconut Oil 2.00%, brown flavor 0.1%, water 56.11%. After grilling the patty, the tasters agreed that the patty had good flavor and texture.
  • Meat extender patty Using the successful PT texturized protein materials made in Example 14, a meat extender patty was made. The ingredients in wt percent were PureTaste texturized protein 3.27%, PureTaste Protein 2.84%, Pea Protein texturized protein 2.98%, Vital Wheat Gluten 1.99%, Methylcellulose 0.71%, Beef Flavor 1.85%, Beet Powder 0.28%, water 11.08%, and ground beef 80:20 (protein Tat), 75%. After grilling the patty, the tasters agreed that the patty had good flavor and texture.
  • Meat extender patty Using the successful PT texturized protein materials made in Example 14, to a beef mixture, PT texturized protein was added at amounts up to 2% and water was increased up to 7% over the control’s 8.5%. Compared with control, the additional moisture was held in the product, indicating a cost savings obtained with PT texturized protein. Taste changes compared with a pure beef patty were very small.
  • Meatless meatballs Meatless meatballs.
  • PT texturized protein Example 15; 5-45-5 formulation was added as the main ingredient to a vegetarian meatball (other than water for rehydration).
  • the other ingredients included PureTasteTM protein powder, canola oil, coconut oil, contains less than 2% each of the following: rice, methylcellulose, natural flavors, beet powder, yeast extract, sea salt, modified food starch, garlic powder, onion powder, black pepper, beta carotene.
  • Meatless tacos PT texturized protein (Example 15; 5-45-5 formulation) was added as the main ingredient to a vegetarian meatball (other than water for rehydration).
  • the other ingredients included canola oil, vital wheat gluten, methylcellulose, coconut oil, natural flavors, spices (paprika, salt, chili pepper, oregano, com starch, dried onion, dried garlic).
  • High fat meat analog Ingredients: water for hydration, PT texturized protein (made per Example 14), coconut fat, coconut oil, flavors, methylcellulose, 2% or less of spices, beet powder, salt. 24 g protein per serving, 18 g fat per serving, 2 g carbohydrate per serving.

Abstract

La présente invention concerne une composition alimentaire qui comprend un produit alimentaire riche en protéines mycélié et des procédés de fabrication de telles compositions, qui sont des mélanges de produits alimentaires riches en protéines mycéliés et d'autres matériaux comestibles. Une composition alimentaire comprend des produits protéiques à base de plantes texturés qui sont utiles pour fabriquer des analogues et des succédanés de viande ayant une structure de viande. Les compositions alimentaires ont des saveurs indésirables réduites et des arômes indésirables réduits grâce à l'utilisation de produits alimentaires riches en protéines mycéliés par rapport à l'utilisation d'un matériau riche en protéines similaire qui n'est pas mycélié.
PCT/US2019/058470 2018-10-29 2019-10-29 Protéine végétale mycéliée et compositions alimentaires comprenant celle-ci WO2020092306A1 (fr)

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CN111631243A (zh) * 2020-06-24 2020-09-08 上海应用技术大学 一种即食混合蘑菇素肉饼及其制备方法
IT202000031427A1 (it) * 2020-12-18 2022-06-18 Joy S R L Procedimento per la fabbricazione di un insaccato a base vegetale
CN114907984A (zh) * 2021-09-24 2022-08-16 南京高新工大生物技术研究院有限公司 一种红平菇菌丝体的生产方法、生产的红平菇菌丝体及其应用
WO2022184317A1 (fr) * 2021-03-02 2022-09-09 Roquette Freres Substitut de viande d'origine végétale
WO2022197816A1 (fr) * 2021-03-16 2022-09-22 Terramino, Inc. Matière protéique texturée contenant des champignons, procédés de fabrication de ladite matière et utilisations correspondantes
CN115119898A (zh) * 2022-07-26 2022-09-30 福建农林大学 一种植物基人造肉肠的加工方法
CN115444111A (zh) * 2022-08-26 2022-12-09 中国科学院天津工业生物技术研究所 一种食用菌丝体蛋白在海产品中的应用
EP3965584A4 (fr) * 2019-05-08 2023-01-25 Mycotechnology, Inc. Procédés de production de compositions gonflantes mycéliées
CN115666270A (zh) * 2020-05-22 2023-01-31 马洛食品有限公司 可食用真菌
WO2023111033A1 (fr) * 2021-12-15 2023-06-22 Mycorena Ab Produit alimentaire à base de biomasse fongique ou ingrédient alimentaire à base de biomasse fongique
WO2023186977A1 (fr) * 2022-03-30 2023-10-05 Unilever Ip Holdings B.V. Procédé de production d'un produit protéique structuré
WO2023247051A1 (fr) * 2022-06-23 2023-12-28 Clonbio Group Ltd. Composition d'aliments pour animaux
WO2024009084A1 (fr) * 2022-07-08 2024-01-11 Marlow Foods Limited Produit alimentaire

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EP3965584A4 (fr) * 2019-05-08 2023-01-25 Mycotechnology, Inc. Procédés de production de compositions gonflantes mycéliées
CN115666270A (zh) * 2020-05-22 2023-01-31 马洛食品有限公司 可食用真菌
CN111631243A (zh) * 2020-06-24 2020-09-08 上海应用技术大学 一种即食混合蘑菇素肉饼及其制备方法
IT202000031427A1 (it) * 2020-12-18 2022-06-18 Joy S R L Procedimento per la fabbricazione di un insaccato a base vegetale
WO2022130329A1 (fr) * 2020-12-18 2022-06-23 Joy S.R.L. Procédé de fabrication d'une saucisse à base de plante
WO2022184317A1 (fr) * 2021-03-02 2022-09-09 Roquette Freres Substitut de viande d'origine végétale
WO2022197816A1 (fr) * 2021-03-16 2022-09-22 Terramino, Inc. Matière protéique texturée contenant des champignons, procédés de fabrication de ladite matière et utilisations correspondantes
CN114907984A (zh) * 2021-09-24 2022-08-16 南京高新工大生物技术研究院有限公司 一种红平菇菌丝体的生产方法、生产的红平菇菌丝体及其应用
WO2023111033A1 (fr) * 2021-12-15 2023-06-22 Mycorena Ab Produit alimentaire à base de biomasse fongique ou ingrédient alimentaire à base de biomasse fongique
WO2023186977A1 (fr) * 2022-03-30 2023-10-05 Unilever Ip Holdings B.V. Procédé de production d'un produit protéique structuré
WO2023247051A1 (fr) * 2022-06-23 2023-12-28 Clonbio Group Ltd. Composition d'aliments pour animaux
WO2024009084A1 (fr) * 2022-07-08 2024-01-11 Marlow Foods Limited Produit alimentaire
CN115119898A (zh) * 2022-07-26 2022-09-30 福建农林大学 一种植物基人造肉肠的加工方法
CN115119898B (zh) * 2022-07-26 2023-08-22 福建农林大学 一种植物基人造肉肠的加工方法
CN115444111A (zh) * 2022-08-26 2022-12-09 中国科学院天津工业生物技术研究所 一种食用菌丝体蛋白在海产品中的应用

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