protein powder

Abstract

The present invention provides a method for producing a protein powder with improved solubility and taste profile from brewers spent grain, which method includes nanofiltration at a specified applied pressure. The present invention also provides a protein powder produced from brewers spent grain, a method for producing a food or beverage product incorporating the protein powder, and a food or beverage product comprising the protein powder.

Description

[Technical Field]

[0001] The present invention relates to a method for producing a protein powder from brewers spent grain. The present invention also relates to a protein powder produced from brewers spent grain, a method for producing a food or beverage product incorporating the protein powder, and a food or beverage product comprising the protein powder. [Background technology]

[0002] The use of protein powders and supplements is well known in the art.For example, many people use protein powders to make drinks or other foods as part of their training regimen to provide additional protein for muscle growth.In addition, people may use protein supplements when their daily diet is insufficient to meet the daily protein needs of the human body.In addition, people with specific dietary habits, such as vegetarians and vegans, who do not allow the consumption of traditional meat-based protein sources, may supplement their diet with protein powders to meet their daily needs.

[0003] Traditionally, protein powders and supplements have generally been whey, soy, or casein-based products.Whey and casein proteins are generally recovered as by-products from dairy production, whey is separated from cheese production, and casein is separated from milk.Soy protein is separated from soybeans.Whey, soy, and casein-based protein powders and supplements are successfully used to provide beneficial amounts of protein, but the latter are not necessarily suitable for people with food intolerances or allergies, such as lactose intolerance.Although plant-based protein powders exist that produce lower immunogenic effects, these products are typically considered less palatable and, for example, less soluble than their whey counterparts.Therefore, consumers are less inclined to choose these alternatives.

[0004] Brewers' spent grain (BSG) is the most abundant by-product generated in the beer brewing process. This material comprises malt and husks obtained as the solid fraction after the mash or wort filtering steps. To date, brewery by-products have been primarily used for low-value purposes, especially as animal feed.

[0005] BSG is rich in nutrients, especially protein and fiber. Attempts have been made to use BSG to produce protein powders, as disclosed in U.S. Patent Application Publication Nos. 2018 / 0199593 and 2018 / 0199594.

[0006] It has been found that protein powders produced using BSG can have a bitter taste and a suboptimal solubility profile. Therefore, there is a need for a method of producing protein powders from BSG that improves the taste and solubility profile of the protein powder. Summary of the Invention

[0007] The present invention provides an improved method for producing protein powder from grain materials selected from brewer's spent grain, barley and malted barley, the method comprising: a) subjecting an aqueous slurry of grain material to enzymatic protein hydrolysis to produce a liquid protein stream; b) removing solids from the liquid protein stream; c) subjecting the liquid protein stream to microfiltration to obtain a microfiltration permeate comprising the protein and a microfiltration retentate; d) subjecting the microfiltration permeate to nanofiltration at an applied pressure of 1.0 bar (100 kPa) to 8.0 bar (800 kPa) to obtain a nanofiltration permeate and a nanofiltration retentate comprising the protein; and e) processing the nanofiltration retentate to produce a protein powder.

[0008] The grain material is preferably brewer's spent grain.

[0009] Nanofiltration is preferably carried out at an applied pressure of 1.3 bar (130 kPa) to 5.0 bar (500 kPa), preferably 1.3 bar (130 kPa) to 4.5 bar (450 kPa). More preferably, nanofiltration is carried out at an applied pressure of 1.3 bar (130 kPa) to 3.3 bar (330 kPa), preferably 1.4 bar (140 kPa) to 3.2 bar (320 kPa), preferably 1.5 bar (150 kPa) to 3 bar (300 kPa). Nanofiltration is preferably carried out using nanofiltration membranes having a molecular weight cut-off (MWCO) of 500 to 2,000 Da, preferably 800 to 2,000 Da, preferably 800 to 1,200 Da.

[0010] The microfiltration is preferably carried out using ceramic microfiltration membranes, which preferably have a pore size of 0.03 to 0.5 μm, preferably 0.05 to 0.25 μm, preferably 0.05 to 0.2 μm, preferably 0.07 to 0.13 μm. The microfiltration preferably comprises a diafiltration step.

[0011] The brewer's spent grain preferably comprises spent barley and optionally one or more other spent grains or other starchy materials selected from rice, maize, sorghum and cassava, preferably selected from rice and maize, preferably selected from rice. Preferably, the grain used in brewing is spent grain obtained from a brewing process comprising barley in an amount of at least 30% by weight, preferably at least 40% by weight, preferably at least 60% by weight, preferably at least 70% by weight, based on the total dry matter weight of the grain.

[0012] The ratio of cereal material (by dry weight) in the aqueous slurry is preferably from 8:1 to 12:1, preferably from 10:1 to 11:1.

[0013] The enzymatic protein hydrolysis preferably comprises treatment with a protease enzyme, preferably an alkaline protease. Preferably, prior to the enzymatic protein hydrolysis, the aqueous slurry is subjected to enzymatic starch hydrolysis. The enzymatic starch hydrolysis preferably comprises treatment with a glucoamylase enzyme.

[0014] Solids are preferably removed from the liquid protein stream by decantation, preferably by a decantation centrifuge.

[0015] The cereal material may be subjected to micronization before and / or during a).

[0016] The solids removed from the liquid protein stream are preferably washed with water and the resulting wash water is combined with the liquid protein stream. The solids removed from the liquid protein stream may be further processed to provide a fibrous product.

[0017] The microfiltration retentate may be subjected to enzymatic protein hydrolysis in a rehydrolysis step, and the liquid product of the rehydrolysis step may be combined with the liquid protein stream.

[0018] The nanofiltration retentate preferably has a total solids content of 10 to 30% by weight, preferably 12 to 25% by weight, and a protein content (% dry matter weight) of at least 80%, preferably at least 85%, as determined by AOAC 990.03 or AOAC 992.15. Treating the nanofiltration retentate to produce the protein powder preferably comprises evaporating to increase the total solids content to 20 to 55%, preferably 25 to 55%, preferably 35 to 55%, preferably 45 to 55%, preferably 48 to 52% by weight total solids content, followed by spray drying to produce the protein powder.

[0019] The protein powder produced by this method preferably has a total solids content of at least 90% by weight, preferably at least 93% by weight, and a protein content (% dry matter) of at least 80%, preferably at least 85%, as determined by AOAC 990.03 or AOAC 992.15. Its molecular weight distribution is preferably 300 Da to 100 kDa, preferably 300 Da to 30 kDa, with a major peak of 500 Da to 4.5 kDa, preferably 2 kDa to 4.5 kDa. Its solubility in water at a pH of 3-8 and a temperature of 20°C is preferably at least 50%, preferably at least 75%, in water at a pH of 5-8 and a temperature of 20°C, preferably at least 80%, preferably at least 85%, in water at a pH of 5-8 and a temperature of 20°C, and preferably at least 90% in water at a pH of 5.5-8 and a temperature of 20°C.

[0020] The present invention also provides a protein powder produced from a grain material selected from brewers spent grain, barley and malted barley, the protein powder comprising: a total solids content of at least 90% by weight; A protein content (% by dry matter) of at least 80% as determined by AOAC 990.03 or AOAC 992.15; It has a pH of 3 to 8 and a solubility of at least 50% in water at a temperature of 20°C.

[0021] The protein powder is preferably produced from brewer's spent grain.

[0022] The protein powder preferably has a total solids content of at least 93% by weight, as determined by AOAC 990.03 or AOAC 992.15, and a protein content (% by dry matter) of at least 85%, and has a solubility of at least 75% in water at a pH of 3 to 8 and a temperature of 20° C. For example, the protein powder may have a solubility of at least 80%, preferably at least 85%, in water at a pH of 5 to 8 and a temperature of 20° C., and preferably at least 90% in water at a pH of 5.5 to 8 and a temperature of 20° C.

[0023] The protein powder preferably has a molecular weight distribution of from 300 Da to 100 kDa, preferably from 300 Da to 30 kDa, and a main peak of from 500 Da to 4.5 kDa, preferably from 2 kDa to 4.5 kDa.

[0024] The protein powder may have one or more of the following characteristics: Dispersibility of at least 95%; Turbiscan stability index (AU) less than 10, preferably less than 8; a surface tension of less than 50 mN / m and / or an interfacial tension of less than 15 mN / m; Water retention capacity less than 0.3g / g and / or oil retention capacity less than 3g / g; Temperature 25℃ and 0.1s -1 ~1000s -1 1.10 -1 Viscosity less than Pa.s; No gelling ability; Fat content less than 2%, total fiber content 1-5%, total carbohydrate content 0-7%, and total ash content 1-8%; a glutamine concentration of 15 to 25 g per 100 g of the composition; and / or A total essential amino acid concentration of 10 g to 50 g per 100 g of the protein powder, wherein the essential amino acids are histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.

[0025] The protein powder is preferably produced according to the method of the present invention.

[0026] The present invention also provides a method of producing a food or beverage product, the method comprising incorporating the protein powder of the present invention into the food or beverage product.The present invention also provides a food or beverage product comprising the protein powder according to the present invention. [Brief explanation of the drawings]

[0027] [Figure 1] Figure 1 shows the solubility profiles of compositions according to one embodiment of the present invention, with Figure 1A showing the results for barley and rice samples and Figure 1B showing the profiles for barley and corn samples. [Figure 2] Figure 2 shows the viscosity profiles of compositions according to one embodiment of the present invention. Figure 2A shows the results for barley and rice samples, and Figure 2B shows the profiles for barley and corn samples. Dotted line: viscosity of water. [Figure 3] FIG. 3 shows the molecular weight distribution of a protein powder produced according to the present invention. Detailed Description of the Invention

[0028] The present invention provides improved methods for producing protein powder (also referred to herein as "powdered protein composition") from brewer's spent grain, methods for producing the protein powder, food or beverage products incorporating the protein powder, and food or beverage products comprising the protein powder.

[0029] Unless otherwise defined, all terms, including technical and scientific terms, used in disclosing the present invention have the meanings commonly understood by one of ordinary skill in the art to which this invention belongs. By way of further guidance, term definitions are included to better understand the teachings of the present invention. As used herein, the following terms have the following meanings:

[0030] As used herein, "A," "an," and "the" refer to both singular and plural referents unless the context clearly dictates otherwise. By way of example, "a section" refers to one or more sections.

[0031] As used herein, "about" when referring to a measurable value such as a parameter, amount, time interval, etc., is meant to encompass a variation from the specified value of no more than + / - 20%, preferably no more than + / - 10%, more preferably no more than + / - 5%, even more preferably no more than + / - 1%, and even more preferably no more than + / - 0.1%, insofar as such variations are appropriate to practice the disclosed invention, provided that the value to which the modifier "about" refers is itself specifically disclosed.

[0032] As used herein, "comprise" and "comprising" are synonymous with "include," "including," "includes," or "contain," "containing," or "contains," and are inclusive or open-ended terms specifying the presence of what follows (e.g., components), but do not exclude or preclude the presence of additional, unrecited components, features, elements, materials, or steps that are known in the art or disclosed therein.

[0033] Furthermore, the terms first, second, third, etc. in the specification and claims are used to distinguish between similar elements, unless otherwise specified, and are not necessarily intended to describe a sequential or chronological order. The terms so used are interchangeable under appropriate circumstances, and it is understood that the embodiments of the invention described herein can be practiced in orders other than those described or illustrated herein.

[0034] The recitation of numerical ranges by endpoints includes all numbers and fractions subsumed within that range, as well as the recited endpoints.

[0035] In this specification and throughout the specification, the terms "wt %", "weight percent", "% wt" or "wt%" refer to the relative weight of each component based on the total weight of the formulation, unless otherwise defined.

[0036] The term "one or more" or "at least one," such as one or more or at least one member of a group of members, is clear in itself, but by way of further illustration, the term specifically encompasses reference to any one of said members, or any two or more of said members, such as >3, >4, >5, >6 or >7 of said members, and up to all of said members.

[0037] References throughout this specification to "one embodiment" or "an embodiment" mean that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrase "in one embodiment" or "in an embodiment" in various places throughout this specification do not necessarily all refer to the same embodiment, although they may. Furthermore, particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments, as would be apparent to one of ordinary skill in the art from this disclosure. Furthermore, while some embodiments described herein include some features and not other features included in other embodiments, combinations of features from different embodiments are meant to form different embodiments within the scope of the present invention, as would be understood by one of ordinary skill in the art.

[0038] "Protein content" as used herein refers to protein content as measured according to the Dumas method (conversion factor 6.25), in particular AOAC 990.03 or AOAC 992.15. Other methods known in the art, such as the Kjeldahl method (conversion factor 6.25), can also be used to obtain essentially the same results.

[0039] Brewer's Spent Grain: The starting material for the process of the present invention is a grain material selected from brewer's spent grain, barley and barley malt, preferably brewer's spent grain.

[0040] Brewer's spent grain is a by-product of the brewing industry after the mashing process. At this point in the brewing process, the soluble fraction (known as "wort") is carried forward to further brewing steps while the insoluble fraction is removed. This insoluble fraction is the brewer's spent grain.

[0041] The brewer's spent grain used in the process of the present invention is preferably obtained after brewing with barley and optionally one or more other cereals or other starchy cereals, such as rice, oats, wheat, maize, sorghum, cassava and / or millet, particularly rice, maize, sorghum and / or cassava, more particularly rice and / or maize. Most preferably, the brewer's spent grain is obtained after brewing with barley, or a mixture of barley and rice, or maize, preferably rice.

[0042] The grain used for brewing (i.e. the grain mixture used at the start of the brewing process) preferably comprises barley in an amount of at least 30% by weight (e.g. at least 30, 35, 40, 45, 50, 55, 60, 65 or 70% by weight, or any intermediate value), preferably at least 40% by weight, preferably at least 60% by weight, preferably at least 70% by weight, based on the total dry matter weight of the grain.

[0043] method: The present invention provides an improved method for producing protein powder from grain materials selected from brewer's spent grain, barley and malted barley, the method comprising: a) subjecting an aqueous slurry of grain material to enzymatic protein hydrolysis to produce a liquid protein stream; b) removing solids from the liquid protein stream; c) subjecting the liquid protein stream to microfiltration to obtain a microfiltration permeate comprising the protein and a microfiltration retentate; d) subjecting the microfiltration permeate to nanofiltration at an applied pressure of 1.0 bar (100 kPa) to 8.0 bar (800 kPa) to obtain a nanofiltration permeate and a nanofiltration retentate comprising the protein; and e) processing the nanofiltration retentate to produce a protein powder.

[0044] The aqueous slurry is formed by mixing the grain material with water. The ratio of water to grain material (by dry weight) in the aqueous slurry is preferably 8:1 to 12:1, preferably 10:1 to 11:1. The aqueous slurry is preferably formed in a jacketed mixing tank, preferably using heating means.

[0045] The aqueous slurry is subjected to enzymatic proteolysis to produce a liquid protein stream. If desired, the grain material may be subjected to micronization prior to and / or during this step. Any suitable comminution technique may be used, such as milling.

[0046] Prior to enzymatic protein hydrolysis, the aqueous slurry is preferably subjected to enzymatic starch hydrolysis. The enzymatic starch hydrolysis preferably involves treatment with a glucoamylase enzyme. Suitable glucoamylase enzymes include those used in the brewing industry and are available, for example, from EDC (Enzyme Development Corporation, New York) or Novozymes.

[0047] The enzymatic protein hydrolysis is preferably carried out at the native pH of the aqueous slurry, which can be, for example, from about 4.5 to about 6.5 (e.g., 4.5, 5, 5.5, 6, or 6.5, or any intermediate value).

[0048] The enzymatic starch hydrolysis is preferably carried out at a temperature of about 50°C to about 65°C (e.g., 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, or 65°C, or any intermediate temperature).

[0049] The enzymatic starch hydrolysis is preferably carried out for at least about 15 minutes, preferably at least about 20 minutes, and up to about 60 minutes, preferably about 45 minutes. For example, the enzymatic starch hydrolysis can be carried out for a period of 15, 20, 25, 30, 35, 40, 45, 50, 55, or 60 minutes, or any intermediate period.

[0050] The enzymatic starch hydrolysis is preferably carried out until at least about 90% by weight, preferably at least about 95% by weight, of the initial starch content has been hydrolyzed to sugars (i.e., to glucose and / or other water-soluble sugars such as disaccharides and other short-chain oligosaccharides).

[0051] Enzymatic protein hydrolysis preferably involves treatment with a protease enzyme. The protease enzyme is preferably a food-grade protease enzyme, preferably a serine protease. It is preferably an alkaline protease, preferably an endopeptidase, preferably a serine endopeptidase. Suitable protease enzymes are available, for example, from Novozymes or EDC (Enzyme Development Corporation, New York).

[0052] Enzymatic protein hydrolysis is preferably carried out at a pH of about 7 to about 10 (e.g., 7, 7.5, 8, 8.5, 9, 9.5, or any intermediate value), preferably about pH 9. The target pH can be achieved by adding alkali such as sodium hydroxide and / or potassium hydroxide prior to enzymatic treatment.

[0053] The enzymatic protein hydrolysis is preferably carried out at a temperature of about 50°C to about 75°C (e.g., 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74 or 75°C, or any intermediate temperature), preferably about 55°C to about 68°C, preferably about 55°C to about 65°C.

[0054] The enzymatic proteolysis is preferably carried out for at least about 15 minutes, preferably at least about 20 minutes, and up to about 80 minutes, preferably about 60 minutes. For example, the enzymatic proteolysis can be carried out for a period of 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, or 80 minutes, or any intermediate period.

[0055] Enzymatic protein hydrolysis is preferably carried out to reach a degree of hydrolysis (dH) of 1 to 10 (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, or any intermediate value), preferably to reach a dH of 4 to 8. As used herein, dH may be determined using the pH-stat method by adding alkali (e.g., NaOH) and applying the following formula:

number

[0056] The enzymatic starch hydrolysis (if performed) and the enzymatic protein hydrolysis are preferably carried out in a jacketed mixing tank where an aqueous slurry is formed.

[0057] Following enzymatic proteolysis, the enzymes are preferably inactivated by raising the temperature, for example, to about 75 to about 90°C (e.g., about 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89 or 90°C, or any intermediate temperature), preferably to about 80°C, for any intermediate period, such as up to about 35 minutes, for example up to about 25 minutes, for example about 10, 15, 20 or 25 minutes.

[0058] Following enzymatic protein hydrolysis, solids are removed from the liquid protein stream. Removal of the solids is preferably carried out by decantation, preferably using a decantation centrifuge. To maximize recovery of the liquid protein stream, pressure may be applied to the solids, for example using a screw press.

[0059] The solids removed from the liquid protein stream are preferably washed with water, and the resulting wash water is then combined with the liquid protein stream, again to maximize protein recovery.

[0060] The solids removed from the liquid protein stream may be further processed to provide a fibrous product.

[0061] The liquid protein stream is then subjected to microfiltration to obtain a microfiltration permeate containing the protein and a microfiltration retentate. Microfiltration is preferably carried out using a ceramic microfiltration membrane. Surprisingly, ceramic microfiltration membranes have been found to be more effective than polymeric membranes in the process of the present invention.

[0062] Microfiltration is preferably carried out using a microfiltration membrane having a pore size of 0.03 to 0.5 μm (e.g., 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, or 0.5 μm, or any intermediate value), preferably 0.03 to 0.25 μm, preferably 0.05 to 0.2 μm, preferably 0.07 to 0.13 μm (e.g., 0.07, 0.08, 0.09, 0.10, 0.11, 0.12, or 0.13 μm, or any intermediate value). Suitable microfiltration membranes are available from Pall Corporation. Microfiltration preferably includes a diafiltration step.

[0063] The microfiltration retentate can be subjected to enzymatic protein hydrolysis in a rehydrolysis step, and the liquid product of the rehydrolysis step can be combined with the liquid protein stream. Rehydrolysis of the microfiltration retentate can advantageously improve protein recovery.

[0064] The microfiltration permeate is subjected to nanofiltration at an applied pressure of 1.0 bar (100 kPa) to 8.0 bar (800 kPa) to obtain a nanofiltration permeate and a nanofiltration retentate containing the protein. Applied pressure is a well-known concept in the field of filtration and relates to the pressure at which the feed is delivered to the filtration membrane. It is usually controlled by a feed pump and regulated by a pressure sensor to ensure that a constant target feed pressure is maintained.

[0065] Nanofiltration is typically carried out at applied pressures significantly greater than those employed in accordance with the present invention, typically at applied pressures of at least about 10 bar (1,000 kPa) and up to about 40 bar (4,000 kPa). The inventors have found that carrying out nanofiltration at much lower applied pressures, from 1.0 bar (100 kPa) to 8.0 bar (800 kPa), can produce protein powders with better taste and solubility profiles.

[0066] Nanofiltration may be carried out at an applied pressure of 1.0 bar (100 kPa), preferably 1.3 bar (130 kPa), up to 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5 or 8 bar (up to 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, or 800 kPa), or any intermediate value. Nanofiltration is preferably carried out at an applied pressure of from 1.3 bar (130 kPa) to 5.0 bar (500 kPa), preferably from 1.3 bar (130 kPa) to 4.0 bar (400 kPa), e.g., 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3, The pressure may be 3.4, 3.5, 3.6, 3.7, 3.8, 3.9, or 4.0 bar (130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, or 400 kPa), or any intermediate value.

[0067] Nanofiltration is more preferably carried out at an applied pressure of from 1.3 bar (130 kPa) to 3.3 bar (330 kPa), preferably from 1.4 bar (140 kPa) to 3.2 bar (320 kPa), preferably from 1.5 bar (150 kPa) to 3 bar (300 kPa). For example, nanofiltration may be carried out at an applied pressure of 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, 2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2 or 3.3 bar (130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250, 260, 270, 280, 290, 300, 310, 320 or 330 kPa), or any intermediate value.

[0068] Nanofiltration is preferably carried out using a nanofiltration membrane having a molecular weight cut-off (MWCO) of 500 to 2,000 Da, preferably 800 to 2,000 Da, preferably 800 to 1,200 Da. For example, nanofiltration can be carried out using a nanofiltration membrane having a molecular weight cut-off (MWCO) of 500, 600, 700, 800, 900, 1,000, 1,100, 1,200, 1,300, 1,400, 1,500, 1,600, 1,700, 1,800, 1,900, or 2,000 Da, or any intermediate value. Suitable microfiltration membranes are available from MICRODYN-NADIR.

[0069] The nanofiltration retentate preferably has a total solids content of 15 to 25% by weight, preferably 18 to 22% by weight, and a protein content (% dry matter wt.) of at least 80%, preferably at least 85%, as determined by AOAC 990.03 or AOAC 992.15.

[0070] The nanofiltration retentate is treated to produce a protein powder. Treating the nanofiltration retentate to produce a protein powder preferably comprises evaporation to increase the total solids content to 20 to 55% (e.g., 20, 25, 30, 35, 40, 45, or 50%, or any intermediate value), preferably 25 to 55%, preferably 35 to 55%, preferably 45 to 55% by weight (e.g., 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, or 55%, or any intermediate value), preferably 48 to 52% by weight, followed by spray drying to produce the protein powder.

[0071] The protein powder produced by the present method preferably has a total solids content of at least 90% by weight, preferably at least 93% by weight (e.g., at least 90, 91, 92, 93, or 94%, or any intermediate value), and a protein content (% dry matter by weight) of at least 80%, preferably at least 85% (e.g., at least 80, 81, 82, 83, 84, or 85%, or any intermediate value), as determined by AOAC 990.03 or AOAC 992.15.

[0072] The molecular weight distribution of the protein powder produced by the present method is preferably 300 Da to 100 kDa (e.g., 300 Da to 30 kDa, 40 kDa, 50 kDa, 60 kDa, 70 kDa, 80 kDa, 90 kDa or 100 kDa), preferably 300 Da to 30 kDa, with a main peak of 500 Da to 4.5 kDa (e.g., 500 Da, 600 Da, 700 Da, 800 Da, 900 Da, 1 kDa, 1.1 kDa, 1.2 kDa, 1.3 kDa, 1.4 kDa, 1.5 kDa, 1.6 kDa, 1.7 kDa, 1.8 kDa, 1.9 kDa or 2.0 kDa to 4.5 kDa), preferably 2 kDa to 4.5 kDa.

[0073] Its solubility (determined according to the method provided further below) is preferably at least 50%, preferably at least 75% (e.g., at least 50, 55, 60, 65, 70, or 75%, or any intermediate value) in water at a pH of 3 to 8 and a temperature of 20° C. Its solubility is preferably at least 80% (e.g., at least 80, 81, 82, 83, 84, or 85%, or any intermediate value), preferably at least 85%, in water at a pH of 5 to 8 and a temperature of 20° C. Its solubility is preferably at least 90% in water at a pH of 5.5 to 8 and a temperature of 20° C.

[0074] Protein powder: The protein powder of the present invention is produced from a grain material selected from brewer's spent grain, barley, and malted barley. a total solids content of at least 90% by weight; A protein content (% by dry matter) of at least 80% as determined by AOAC 990.03 or AOAC 992.15; It has a pH of 3 to 8 and a solubility of at least 50% in water at a temperature of 20°C.

[0075] The protein powder is preferably produced from brewer's spent grain.

[0076] The protein powder of the present invention has a particularly favorable taste and solubility profile compared to prior art protein powders derived from brewer's spent grain, and exhibits a particularly improved bitterness profile, exhibiting a low bitter taste.

[0077] The protein powder of the present invention has a total solids content of at least 90% by weight, preferably at least 93% by weight (e.g., at least 90, 91, 92, 93, or 94%, or any intermediate value), and a protein content (% dry matter by weight) of at least 80%, preferably at least 85% (e.g., at least 80, 81, 82, 83, 84, or 85%, or any intermediate value), as determined by AOAC 990.03 or AOAC 992.15.

[0078] The solubility of the protein powder of the present invention (determined according to the method provided further below) is preferably at least 50%, preferably at least 75% (e.g., at least 50, 55, 60, 65, 70, or 75%, or any intermediate value) in water at a pH of 3 to 8 and a temperature of 20° C. The solubility is preferably at least 80% (e.g., at least 80, 81, 82, 83, 84, or 85%, or any intermediate value), preferably at least 85%, in water at a pH of 5 to 8 and a temperature of 20° C. The solubility is preferably at least 90% in water at a pH of 5.5 to 8 and a temperature of 20° C.

[0079] The molecular weight distribution of the protein powder is preferably from 300 Da to 100 kDa (e.g., from 300 Da to 30 kDa, 40 kDa, 50 kDa, 60 kDa, 70 kDa, 80 kDa, 90 kDa or 100 kDa), preferably from 300 Da to 30 kDa, with a main peak from 500 Da to 4.5 kDa (e.g., from 500 Da, 600 Da, 700 Da, 800 Da, 900 Da, 1 kDa, 1.1 kDa, 1.2 kDa, 1.3 kDa, 1.4 kDa, 1.5 kDa, 1.6 kDa, 1.7 kDa, 1.8 kDa, 1.9 kDa or 2.0 kDa to 4.5 kDa), preferably from 2 kDa to 4.5 kDa.

[0080] The protein powder may have one or more of the following characteristics: Dispersibility of at least 95%; Turbiscan stability index (AU) less than 10, preferably less than 8; a surface tension of less than 50 mN / m and / or an interfacial tension of less than 15 mN / m; Water retention capacity less than 0.3g / g and / or oil retention capacity less than 3g / g; Temperature 25℃ and 0.1s -1 ~1000s -1 1.10 -1 Viscosity less than Pa.s; No gelling ability; Fat content less than 2%, total fiber content 1-5%, total carbohydrate content 0-7%, and total ash content 1-8%; a glutamine concentration of 15 to 25 g per 100 g of the composition; and / or A total essential amino acid concentration of 10 g to 50 g per 100 g of the protein powder, wherein the essential amino acids are histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.

[0081] The protein powder is preferably produced according to the method of the present invention.

[0082] Further description of protein powders (also called "powdered protein compositions") is provided below.

[0083] In one exemplary embodiment of the present invention, the powdered protein composition obtained from brewer's spent grain has a protein content of at least 75%, at least 80%, more preferably at least 85%, on a dry matter basis, and a dry matter content of at least 90%, and a protein solubility of at least 50%, at least 60%, more preferably at least 70% in an aqueous environment at a pH of 3-8, and at least 75%, more preferably 80% at a pH of 5-8.

[0084] High protein solubility is advantageous for further processing and use of the composition, for example when used in beverages.

[0085] The test for measuring the water solubility of a protein involves preparing a 2% protein solution in a beaker, stirring the solution at 500 rpm for 15 minutes using a magnetic stirrer, adjusting the pH to the desired pH (pH 3-8), and stirring the solution for an additional 30 minutes. Finally, the solution is centrifuged at 15,000 g (15,000 times gravity) at 20°C for 10 minutes, and the soluble fraction is analyzed by the Kjeldahl method (conversion factor 6.25). The percentage of solubility is calculated as follows: Solubility % = Protein content in supernatant / Total protein content * 100

[0086] Protein powders according to exemplary embodiments of the present invention can be produced using brewer's spent grain from cereal sources such as rice, oats, wheat, corn, sorghum, millet, malt, and barley. For example, the brewer's spent grain can be obtained after brewing with barley and optionally one or more other cereals or other starchy grains, such as rice, oats, wheat, corn, sorghum, cassava, and / or millet, particularly rice, corn, sorghum, and / or cassava, more particularly rice and / or corn. Most preferably, the brewer's spent grain is obtained after brewing with barley, or a mixture of barley and rice, or corn, preferably rice.

[0087] In one embodiment, the brewer's spent grain is a combination of at least barley and rice. In another embodiment, the brewer's spent grain is a combination of at least barley and corn. In another embodiment, the protein composition is derived from barley or (barley) malt.

[0088] Protein powders not only provide an additional revenue stream for the brewing process, but also have various attributes imparted by the brewing and recovery process that make them advantageous for use as a protein supplement. The protein powders of the present invention have several characteristics that make them advantageous for use when used to prepare foods and feeds, such as mixed and blended liquid beverages, pourable foods, or foodstuffs.

[0089] In another or further embodiment, the powdered protein composition according to the present invention has a dispersibility of at least 95%, more preferably at least 96%, more preferably at least 97%, more preferably at least 98%, and more preferably at least 99%. Dispersibility is defined as the ability of a composition to dissolve during agitation. While low dispersibility is preferred for certain applications, such as fish feed, high dispersibility is advantageous when the protein composition is used in food applications, such as beverages. The dispersibility of a composition can be measured by adding a predetermined concentration of the composition to an aqueous medium, such as water, with mixing (e.g., vortexing at 500 rpm) for a period of time. The dispersion is then filtered, and the filter and its contents are dried. Dispersibility is calculated based on the percentage of material (undispersed material) retained by the filter per gram of sample.

[0090] In another or further embodiment, the powdered protein composition has a Turbiscan Stability Index (AU) of less than 10, preferably less than 8, preferably less than 7, for example, between 1.5 and 6, more preferably between 2 and 5. The latter allows for a stable solution of the protein composition when the protein composition is dissolved in a solution, preferably an aqueous medium. Sedimentation tests were performed on a Turbiscan LAB (Formulaction). This instrument measures the percentage of light transmitted through a suspension (clear suspension) and backscattered (opaque suspension) over time. Sedimentation is indicated by an increase in transmission at the top of the tube (the top of the suspension becomes more transparent) and an increase in backscattered at the bottom of the tube (the bottom of the suspension becomes opaque). An overall stability factor is calculated after the test (Turbiscan Stability Index). A TSI of less than 10 is considered to be a very stable solution (no sedimentation).

[0091] In another or further embodiment, the powdered protein composition of the present invention has a surface tension of less than 50 mN / m and / or an interfacial tension of less than 15 mN / m. In a further embodiment, the surface tension is 30-50 mN / m, more preferably 40-45 mN / m. The interfacial tension may be 5-15 mN / m, more preferably 10-14 mN / m.

[0092] The ability of a composition to reduce surface tension (at the water / air interface) and interfacial tension (at the oil / water interface) can be measured using a Kruss tensiometer. It has been found that the protein composition according to the present invention reduces surface and interfacial tension significantly more than casein protein isolate. As a result, the composition has good surfactant properties.

[0093] In another or further embodiment, the powdered protein composition has a water-holding capacity of less than 0.3 g / g, more preferably between 0.05 g / g and 0.3 g / g, and / or an oil-holding capacity of less than 3 g / g, or less than 2.5 g / g, more preferably between 0.5 and 2.5 g / g. In the context of the present invention, water-holding capacity (WHC) is defined as the ability of a composition to retain its own or added water during the application of force, pressure, centrifugation, or heat. The present compositions have been found to have little or no water-holding capacity. In contrast, the compositions have good oil-holding capacity.

[0094] In another or further embodiment, the powdered protein composition is 1.10 -1 Pa.s or less, more preferably 1.10 -1 ~0.5.10 -1 Viscosity may be measured by conventional means in the art. In one embodiment, a 10% aqueous solution of the composition is tested and has a viscosity of 0.1 s at a temperature of 25°C. -1 ~1000s -1 The viscosity profile was measured over a shear rate range of 1000 rpm, ...

[0095] In one embodiment, the protein compositions of the present invention lack gelling properties or gelling capacity and do not form gels when heated and cooled. Therefore, the compositions of the present invention can be advantageously used in the preparation of protein-enriched foods without adversely affecting taste, mouthfeel, and / or aesthetic appearance. Gelling capacity can be assessed with a rheometer by preparing a 10% solution at pH 7, heating the solution to 90°C, and then cooling. If a gel forms under these conditions, a strong and sharp increase in storage modulus G' is observed, with the final storage modulus ("solid behavior") being higher than the loss modulus G'' ("liquid behavior").

[0096] In the context of the present invention, the storage modulus of a 10% solution at pH 7 remains the same (within the same log range) before and after heating said solution to 90°C (for at least 10 minutes) and cooling it to 25°C.

[0097] In another or further embodiment, the powdered protein has a fat content of less than 0.2%, a total fiber content of 1-5%, a total carbohydrate content of 0-7%, and a total ash content of 1-8%.

[0098] In another or further embodiment, the powdered protein composition has a glutamine concentration of 15-25 g per 100 mg of the composition. Glutamine is known to be a conditionally essential amino acid commonly found in meats, such as beef or chicken, and dairy products. Glutamine can be used as a supplement during intense exercise or during illness. Several studies support the positive effects of long-term oral administration of the supplement on damage and inflammation induced by intense aerobic and exhaustive exercise.

[0099] In another or further embodiment, the powdered protein composition has a total essential amino acid concentration of 10 g to 50 g per 100 g of the composition, the essential amino acids being histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine. Thus, the protein powder may provide a good source of the daily amino acid requirement.

[0100] Combined with beneficial organoleptic properties such as a pleasant mouthfeel and mild flavor that allow brewers spent grain based protein powder to be used alone or as a protein fortifier in foods intended for human consumption, companion pet foods and commercial livestock feeds, brewers spent grain based protein powder is a highly advantageous protein supplement.

[0101] The present invention also provides food or beverage products comprising 1-99%, more preferably 10-95%, more preferably at least 15%, more preferably at least 20%, more preferably at least 30%, 40%, or 50% of the powdered protein composition according to any of the above embodiments. In some embodiments, the protein composition can comprise up to 50% by weight of the food or beverage product, preferably in an amount of 20-40% by weight of the food or beverage product, without affecting the flavor profile of the food or beverage product. In some embodiments, the food or beverage product is a beverage or infusible food product, such as, for example, energy drinks, shakes, smoothies, coffee and coffee-based beverages (i.e., lattes, mochas, etc.), and tea. In other embodiments, the food or beverage product can comprise a muscle-building supplement, such as a meal replacement bar or a training drink. In some embodiments, the food or beverage product can comprise a meat substitute, such as, for example, a meat-meat binder replacer and an extruded meat substitute. In some embodiments, the (protein-enriched) food or beverage product can comprise coatings and / or bindings for granola, nutrition bars, and muesli. In other embodiments, the protein-enriched food or beverage product can include seasonings for the preparation of bases, gravies, soups, and sauces. In some embodiments, the (protein-enriched) food or beverage product can include baked goods, such as brownies, cakes, cookies, breads, crackers, etc. In still other embodiments, the (protein-enriched) food or beverage product can include breakfast products, such as waffles, pancakes, quickbreads, and pastries. In some embodiments, the protein-enriched food or beverage product can include dairy products, such as yogurt, cheese spreads, and cheese-based products. In some embodiments, the (protein-enriched) food or beverage product can include cocoa powder bulking agents. In some embodiments, the protein-enriched food or beverage product can include chocolate, candy, and confectionery. In some embodiments, the (protein-enriched) food product can include carbohydrate-based entrees, such as pasta (macaroni and cheese), rice, and grains.In some embodiments, the protein-enriched food or beverage products can include dips, spreads and toppings (hummus).

[0102] The food or drink product may be suitable for both human and animal consumption. In one embodiment, the composition is suitable for use as a pet food or pet food formulation.

[0103] The protein powders of the present invention can also be described with reference to the following numbered paragraphs:

[0104] 1. A powdered protein composition obtained from brewer's spent grain, barley or barley malt, having a protein content of at least 80% based on dry matter and a dry matter content of at least 90%, characterized in that the powdered protein composition has a solubility of at least 50% in an aqueous environment at a pH of 3 to 8.

[0105] 2. The powdered protein composition according to claim 1, characterized in that it has a solubility of at least 75% in an aqueous environment at a pH of 2.3 to 8.

[0106] 3. The powdery protein composition according to item 1 or 2, characterized in that it has a dispersibility of at least 95%.

[0107] 4. The powdery protein composition according to any one of items 1 to 3, characterized in that it has a Turbiscan stability index (AU) of less than 4.10, preferably less than 8.

[0108] 5. The powdery protein composition according to any one of items 1 to 4, having a surface tension of less than 5.50 mN / m and / or an interfacial tension of less than 15 mN / m.

[0109] 6. The powdered protein composition according to any one of items 1 to 5, having a water retention capacity of less than 0.3 g / g and / or an oil retention capacity of less than 3 g / g.

[0110] 7.25℃ temperature and 0.1s -1 ~1000s -1 1.10, measured over a shear rate range of -1 7. The powdery protein composition according to any one of items 1 to 6, having a viscosity of less than Pa.s.

[0111] 8. The powdery protein composition according to any one of items 1 to 7, which does not have gelling ability.

[0112] 9. The powdered protein composition according to any one of items 1 to 8, having a fat content of less than 9.2%, a total fiber content of 1 to 5%, a total carbohydrate content of 0 to 7%, and a total ash content of 1 to 8%.

[0113] 10. The powdered protein composition according to any one of items 1 to 9, having a glutamine concentration of 15 to 25 g per 100 g of the composition.

[0114] 11. The powdered protein composition according to any one of items 1 to 10, having a total essential amino acid concentration of 10 g to 50 g per 100 g of the composition, the essential amino acids being histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan, and valine.

[0115] 12. A food product comprising 1 to 99% of the powdered protein composition according to any one of items 1 to 11.

[0116] 13. A food product according to paragraph 12, suitable for humans and / or animals, such as pets.

[0117] 14. Use of the powdered protein composition according to any one of paragraphs 1 to 11 as a supplement in food. [Example]

[0118] The present invention will now be further illustrated with reference to the following examples. The present invention is in no way limited to the examples given or to the embodiments presented in the figures.

[0119] Example 1: Brewers spent grains, including spent barley and either spent corn or spent rice, were used to prepare protein powders according to the following general method.

[0120] Incoming grain was received in a jacketed mixing tank along with water to a water to dry weight ratio of 10.5:1. The resulting slurry was heated to 55°C and treated with glucoamylase enzyme (EDC Enzeco® Glucoamylase) for 45 minutes to hydrolyze the starch. The pH was then raised to 9 with alkali and maintained for 45 minutes.

[0121] The protein components were then hydrolyzed by treatment with a food-grade protease enzyme (EDC Enzeco® Alkaline Protease L-660) at 60°C for 20 to 60 minutes, after which the mixture was heated to 80°C and held for up to 25 minutes to inactivate the enzyme.

[0122] The solids were separated from the liquid protein stream by decanting the centrifuge. The liquid protein stream was fed to a microfiltration system (0.1 μm membrane, 70-80° C., suitable membrane available from Pall Corporation).

[0123] The permeate from the microfiltration was treated in a nanofiltration system (MWCO of c. 1000 Da, applied pressure of 1.5 to 3 bar, suitable membranes available from MICRODYN-NADIR), and the resulting excess was then subjected to vacuum evaporation to remove water, followed by spray drying.

[0124] Example 2: A sample of the protein composition obtained from barley and rice brewer's spent grain (Sample A) or barley and corn brewer's spent grain (Sample B) was analyzed as follows according to Example 1. Similar values were obtained for the samples from barley alone or from barley malt (data not shown).

[0125] Moisture and protein content Moisture content was determined using a Prepash apparatus (Precisa) based on oven drying (drying to constant weight at 105°C). Sample moisture was measured at 105°C for 12 hours. Protein content was determined using an automated apparatus (Foss) based on the Dumas method (AOAC 992.15). A conversion factor of 6.25 was used.

[0126] Sample A had a dry matter content of 95.5% and a protein content of 87.9% (N x 6.25) (db). Similar results were obtained for sample B.

[0127] [Table 1]

[0128] Protein solubility Protein solubility was tested in a composition suspension with a protein content of 2%. That is, to obtain a 2% protein solution, a predetermined amount of protein powder was mixed with an aqueous medium, preferably water. The protein solution was stirred at 500 rpm for 15 minutes using a magnetic stirrer, and the pH was adjusted. The solution was stirred for another 30 minutes, and finally centrifuged at 15,000 g for 10 minutes at 20 ° C. Then, the soluble fraction was analyzed by the Kjeldahl method. Protein solubility was calculated by dividing the supernatant protein content by the total protein content and multiplying by a factor of 100.

[0129] The protein solubility profiles of the samples are shown in Figures 1A (Sample A) and 1B (Sample B). The protein fraction of the samples is highly soluble (>75%) between pH 5 and pH 8.

[0130] dispersibility Powder dispersibility was measured using the inside method. 5 g of sample was added to 100 ml of water while mixing at 500 rpm (vortex). The dispersion was mixed for 5 minutes. The dispersion was filtered through a 30 μm filter. The filter and its contents were dried at 105°C for 4 hours and weighed. The percentage of material retained on the filter (undispersed) per gram of sample was calculated.

[0131] [Table 2]

[0132] Sedimentation Sedimentation tests were performed on a Turbiscan. This instrument measures the percentage of light transmitted through a suspension (clear suspensions) and backscattered (opaque suspensions) over time. Sedimentation is indicated by an increase in transmission at the top of the tube (the top of the suspension becomes more transparent) and an increase in backscattered at the bottom of the tube (the bottom of the suspension becomes opaque). An overall stability factor is calculated after the test (Turbiscan Stability Index). In general, the Turbiscan Stability Index (TSI) is a scale developed by Turbiscan itself. A reading close to 0 indicates that the sample is very stable with no settling, a reading around 10 indicates that some settling is observed, while a reading above 30 indicates strong settling.

[0133] Therefore, a stable powder without sedimentation has a TSI index close to 0. A 1% solution (db) was prepared and placed in a glass cell. A laser beam was scanned vertically across the sample every minute for 30 minutes, and the light transmission and back diffusion along the glass cell were measured. The stability of the dispersion (sedimentation, creaming) was measured for 30 minutes.

[0134] The analyzed samples were very stable to sedimentation (TSI less than 10). Starch was used as a control.

[0135] [Table 3]

[0136] Surface and interfacial tension The ability of the samples to reduce surface tension (at the water / air interface) and interfacial tension (at the oil / water interface) was measured using a Kruss tensiometer. Solutions with 1% and 0.1% protein content were used to measure interfacial and surface tension, respectively. Surface tension was measured using a Wilhemy plate. Interfacial tension was measured using a DuNouy ring. The samples of the present invention reduced surface tension, and more importantly, interfacial tension, more than the control (casein protein isolate). The samples have good surface active properties.

[0137] [Table 4]

[0138] Water and oil retention capacity The water and oil retention capacities were measured by adding the samples to oil and water at a concentration of 20 mg / ml dry matter. The suspension was blended for 1 hour under stirring. After centrifugation at 15,000 g for 10 minutes, the water or oil content in the pellet was measured and compared with the initial weight of the material. The results are expressed as the number of times the sample can retain its weight in water or oil. The analyzed sample has no water retention capacity. It has an oil retention capacity of 1.9 g / g.

[0139] [Table 5]

[0140] viscosity Rheological analysis was carried out at 25°C using a DHR-2 rheometer (TA) in vane cup geometry. A 10% solution (dry basis) was used. The viscosity profile was measured at 0.1 s -1 ~1000s -1The viscosity profile of the sample in a 10% protein solution is shown in Figures 2A and 2B. The measured viscosity is very low (approximately 10-2 Pa.s), slightly higher than that of water alone. The viscosity is more or less independent of shear rate, which corresponds to Newtonian behavior.

[0141] Minimum gelling concentration The minimum gelling concentration was determined by preparing solutions containing 2% to 20% sample in test tubes. After solubilization, the solutions were heated in a water bath at 85°C for 1 hour and then cooled at 4°C for 2 hours. The solution was considered to have formed a gel if it behaved like a liquid (i.e., free-flowing) before heating and did not flow when the test tube was inverted after heating. Samples did not gel at 85°C in the 2% to 20% conditions tested.

[0142] gelling ability Gelation ability was assessed using a DHR-2 rheometer (TA) in a 40 mm plate / plate configuration by preparing a 10% protein solution, heating it to 90 °C, and cooling it to 25 °C. If the sample is capable of forming a gel at these concentrations and pH conditions, a strong and sharp increase in the storage modulus G' is observed, with the final storage modulus ("solid behavior") being higher than the loss modulus G'' ("liquid behavior"). For the analyzed samples, the storage modulus G' was stable during heating and only increased slightly during cooling from 40 °C to 25 °C. Furthermore, after cooling, G' ≈ G''. This suggests that the sample does not have gelation ability under the conditions tested.

Claims

1. 1. A method for producing a protein powder from a grain material selected from brewer's spent grain, barley, and barley malt, comprising: a) subjecting an aqueous slurry of said grain material to enzymatic proteolysis to produce a liquid protein stream; b) removing solids from said liquid protein stream; c) subjecting said liquid protein stream to microfiltration to obtain a protein-containing microfiltration permeate and a microfiltration retentate; d) subjecting the microfiltration permeate to nanofiltration at an applied pressure of 1.0 bar (100 kPa) to 8.0 bar (800 kPa) to obtain a nanofiltration permeate and a nanofiltration retentate comprising the protein; and e) processing said nanofiltration retentate to produce said protein powder.

2. the grain material is brewer's spent grain; and / or 10. The method of claim 1, wherein the brewer's spent grain is spent grain obtained from a brewing process in which the grain used in brewing includes barley in an amount of at least 30% by weight, based on the total dry matter weight of the grain.

3. 3. The method of claim 1 or 2, wherein the nanofiltration is carried out at an applied pressure of 1.3 bar (130 kPa) to 5.0 bar (500 kPa).

4. 4. The method of claim 3, wherein the nanofiltration is carried out at an applied pressure of 1.3 bar (130 kPa) to 3.3 bar (330 kPa).

5. 5. The method according to any one of claims 1 to 4, wherein the nanofiltration is carried out using a nanofiltration membrane having a molecular weight cut-off (MWCO) of 500 to 2,000 Da.

6. 6. The method according to any one of claims 1 to 5, wherein the microfiltration is carried out using a microfiltration membrane having a pore size of 0.03 to 0.5 μm.

7. The method according to any one of claims 1 to 6, wherein the enzymatic protein hydrolysis comprises treatment with a protease enzyme.

8. the nanofiltration retentate has a total solids content of 10 to 30% by weight and a protein content (% dry matter) of at least 80% as determined by AOAC 990.03 or AOAC 992.15; and / or 8. The method of any one of claims 1 to 7, wherein treating the nanofiltration retentate to produce the protein powder comprises evaporating to increase the total solids content to between 20 and 55% total solids content, followed by spray drying to produce the protein powder.

9. 9. The method of any one of claims 1 to 8, wherein the protein powder has a total solids content of at least 90% by weight and a protein content (% dry matter wt) of at least 80% as determined by AOAC 990.03 or AOAC 992.

15.

10. 10. The method of any one of claims 1 to 9, wherein the protein powder has a molecular weight distribution of from 300 Da to 100 kDa and a main peak of from 500 Da to 4.5 kDa.

11. 11. The method according to any one of claims 1 to 10, wherein the protein powder has a solubility of at least 50% in water at a pH of 3 to 8 and a temperature of 20°C.

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