阅读说明：本技术 基于blg的速溶饮料粉末 (Instant beverage powder based on BLG ) 是由 K·B·劳里德森 H·贝特尔森 S·B·尼尔森 G·D·M·马西尔 K·桑德加 B·R·帕 于 2019-06-26 设计创作，主要内容包括：本发明涉及一种速溶饮料粉末产品及一种用于制备该速溶饮料粉末产品的方法、一种由该速溶饮料粉末生产的液体食品和一种用于制备该液体食品的方法、该液体食品的用途和一种包括该速溶饮料粉末产品的试剂盒。(The present invention relates to an instant beverage powder product and a method for preparing the instant beverage powder product, a liquid food product produced from the instant beverage powder and a method for preparing the liquid food product, the use of the liquid food product and a kit comprising the instant beverage powder product.)

1. Instant beverage powder comprising at least 1% w/w, preferably at least 5% w/w beta-lactoglobulin (BLG), wherein:

blg has a crystallinity of at least 20%, preferably at least 40%, and/or

Blg represents at least 85% w/w of the total amount of protein,

the powder further comprises at least one other ingredient selected from the group consisting of: vitamins, flavoring agents, coloring agents, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, antifoaming agents, and non-whey proteins.

2. Powder according to claim 1, wherein the instant beverage powder comprises BLG in the range of 1-90% w/w BLG.

3. The powder of any of claims 1-2, wherein the powder further comprises one or more of:

i. sweeteners, such as sugar sweeteners and/or non-sugar sweeteners,

(ii) a flavoring agent,

at least one food acid, such as citric acid or other suitable food acid,

the total amount of Na, K, Mg and Ca in the instant beverage is at most 10mmol/g protein,

wherein the pH of a 10% w/w solution of the powder in demineralised water is in the range 2-8.

4. Powder according to any one of the preceding claims, wherein the powder comprises at least 85% w/w of the total amount of protein of BLG, such as at least 86% w/w of the total amount of protein of BLG, at least 87% w/w of the total amount of protein of BLG, at least 88% w/w of the total amount of protein of BLG, at least 89% w/w of the total amount of protein of BLG.

5. Powder according to any one of the preceding claims, wherein the powder comprises at least 91% w/w of total protein BLG, such as at least 92% w/w of total protein BLG, at least 93% w/w of total protein BLG, at least 94% w/w of total protein BLG, at least 95% w/w of total protein BLG, at least 96% w/w of total protein BLG, at least 97% w/w of total protein BLG, at least 98% w/w of total protein BLG or at least 99% w/w of total protein BLG.

6. Powder according to any one of the preceding claims, wherein the water content comprised in the powder is at most 6% w/w, such as at most 5% w/w, preferably at most 4% w/w, more preferably at most 3% w/w, even more preferably at most 2% w/w.

7. The powder according to any of the preceding claims, wherein the powder comprises:

i. up to 6% w/w of water,

at least 15% w/w protein in total relative to the total solids,

at least 85% w/w BLG relative to the total amount of protein,

wherein the powder is a dry powder.

8. Powder according to any one of the preceding claims, wherein the bulk density of the powder is at least 0.30g/mL, preferably at least 0.4g/mL, more preferably 0.5g/mL or even more preferably 0.5 g/mL.

9. Powder according to any one of the preceding claims, wherein the total amount of ALA and Caseinomacropeptide (CMP) is at least 40% w/w, preferably at least 60% w/w, even more preferably at least 70% w/w, most preferably at least 85% w/w of the non-BLG proteins of the powder.

10. Powder according to any one of the preceding claims, wherein the total amount of Na, K, Mg and Ca is at most 10mmol/g protein.

11. Powder according to any of the preceding claims, wherein the energy content of the powder is in the range of about 200-.

12. Powder according to any one of the preceding claims, wherein the protein has an energy content of at least 7E%, preferably at least 25E%, more preferably at least 30E%, even more preferably at least 40E%.

13. A powder according to any preceding claim wherein the energy contribution from the lipid is in the range 0-60E%.

14. Powder according to any of the preceding claims, wherein the energy contribution from the carbohydrate is in the range of 0-90E%.

15. A powder according to any preceding claim wherein the pH of a 10% w/w solution of the powder in demineralised water is from 2 to 8 at room temperature.

16. Powder according to any of the preceding claims, wherein the powder is used as a food ingredient.

17. Powder according to any one of the preceding claims, wherein the powder is used in a method for treating a patient suffering from or at risk of malnutrition.

18. Powder according to any of the preceding claims, wherein the powder is for use in a method for treating kidney disease.

19. A liquid food product comprising a liquid and the instant beverage powder according to any of claims 1-18.

20. A food product according to claim 19, wherein the food product comprises at most 40 grams of the powder per 100 grams of the liquid, preferably at most 30 grams of the powder per 100 grams of the liquid.

21. The food product according to any one of claims 19-20, wherein the food product has a turbidity of at most 200 NTU.

22. The food product according to any one of claims 19-20, wherein the food product has a turbidity of greater than 200 NTU.

23. The food product of any of claims 19-22, wherein the liquid is selected from the group consisting of water, dairy products, fruit juices, vegetable juices, beverages, and combinations thereof.

24. A comestible according to any of claims 19-23 comprising water and the instant beverage powder according to any of claims 1-18, having an energy content in the range of 30-300kcal per 100g comestible.

25. Food product according to claim 24, wherein the food product has an energy content in the range of 30-100kcal per 100g of food product, preferably in the range of 30-100kcal per 100g of food product, more preferably in the range of 40-90kcal per 100g of food product, or even more preferably in the range of 40-70kcal per 100g of food product.

26. The comestible as claimed in claim 24, wherein the energy content of the comestible is in the range of 100-300kcal/100 g comestible, preferably in the range of 100-250kcal/100 g comestible or in the range of 125-225kcal/100 g comestible.

27. A food product according to any of claims 19-26, wherein the protein fraction of the liquid food product has a colour value Δ b, measured at room temperature, in the range-0.10 to +0.51 of the CIELAB colour scale, wherein Δ b ═ b_{Samples normalized to 6.0 w/w% protein}*-b_{Demineralized water}*。

28. The food product according to any one of claims 19-27, wherein the food product further comprises vegetables and/or fruits.

29. A food product as claimed in any of claims 19 to 28, wherein the food product is for use as a nutritional supplement.

30. The food product of any one of claims 19-29, wherein the food product is ingested prior to, during, or after exercise.

31. A food product as claimed in any of claims 19 to 30, wherein the food product is for use in the treatment of a patient suffering from or at risk of a malnutrition.

32. A food product as claimed in any of claims 19 to 31 for use in the treatment of kidney disease.

33. A kit comprising the powder of any one of claims 1-18, the kit comprising:

i. a tool for measuring said powder, and

a container having a lid for opening and closing the container,

wherein the container is for mixing the powder with a liquid to form a food product, the container being adapted for drinking the food product directly from the container.

34. A process for preparing a liquid food product according to any one of claims 19 to 32, the process comprising:

i. adding the powder of any one of claims 1-18 to a liquid,

optionally adding at least one other ingredient, and

mixing the obtained powder and liquid to form a homogeneous mixture.

35. The method of claim 34, wherein the other ingredient is selected from a fruit or a vegetable.

36. The method of any one of claims 33-34, wherein the mixing is performed by shaking.

37. A method for preparing an instant beverage powder comprising BLG and at least one optional ingredient, the method comprising: blending the dried BLG isolate with at least one other ingredient selected from the group consisting of: vitamins, flavoring agents, coloring agents, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, antifoaming agents, and non-whey proteins.

38. The method of claim 37 wherein the BLG in the dried BLG isolate is coated with an organic acid selected from the group consisting of: pyruvic acid (salt), aconitic acid (salt), citric acid (salt), isocitric acid (salt), ketoglutaric acid (salt), succinyl-coenzyme a, succinic acid (salt), fumaric acid (salt), malic acid (salt), oxaloacetic acid (salt), tartaric acid (salt), acetic acid (salt), tannic acid, benzoic acid, maleic acid and lactic acid (salt).

Technical Field

The present invention relates to an instant beverage powder product and a method for preparing the instant beverage powder product, a liquid food product produced from the instant beverage powder and a method for preparing the liquid food product, the use of the liquid food product and a kit (kit) comprising the instant beverage powder product.

Background

Nutritional supplements containing whey proteins are commonly used for muscle synthesis, weight control, and maintenance of muscle and body weight. Nutritional supplements are directed to different types of consumers, such as athletes/women, sports players, children, elderly people and patients suffering from or at risk of malnutrition and/or having an increased protein demand. Therefore, the perception of the nutritional supplement by the consumer is very important, as the consumer should feel like to drink the product.

Whey proteins may be separated from whey or whey. Whey typically comprises a mixture of beta-lactoglobulin (BLG), alpha-lactalbumin (ALA), serum albumin and immunoglobulins, with BLG being the predominant. Whey Protein Concentrate (WPC) therefore comprises a mixture of these proteins. Whey Protein Isolate (WPI) has a fat and lactose content lower than WPC. The isolation of beta-lactoglobulin (BLG) from milk serum or whey is the subject of many publications and typically involves multiple separation steps and common chromatographic techniques to obtain a purified beta-lactoglobulin product.

International patent application WO2002/056707 (nestle) relates to a balanced powder blend composition having at least one source of fat or oil, at least one source of carbohydrates and at least one source of proteins. The composition is conveniently added to food to supplement the nutritional value of the food, but does not substantially alter the mouthfeel of the food.

WO 2018/115520 a1 discloses a method for producing an edible isolated beta-lactoglobulin composition and/or a composition comprising crystallized beta-lactoglobulin based on crystallization of BLG in a salting-in mode. The crystallized BLG may then be separated from the remaining mother liquor.

WO 2011/112695 a1 discloses nutritional compositions and methods of making and using the nutritional compositions. The nutritional composition comprises whey protein micelles and leucine and provides a sufficient amount of leucine to improve protein synthesis in the human body while also maintaining a low viscosity fluid matrix and acceptable organoleptic properties.

WO2011/051436 a1 discloses at least partially transparent compositions designed for human or animal consumption and relates to packaging of such compositions. One embodiment of the present invention relates to an at least partially transparent container comprising an at least partially transparent aqueous non-alcoholic composition. The container includes at least one polarizer that makes visible the liquid crystals present in the composition.

WO2004/049819 a2 discloses a method for improving the functional properties of globular proteins, the method comprising the steps of: providing a solution of one or more globular proteins, wherein the one or more proteins are at least partially aggregated in fibrils, and performing one or more of the following steps in random order: increasing the pH; increasing the salt concentration; concentrating the solution; and changing the solvent quality of the solution. Preferably, the solution of one or more globulins is provided by heating at low pH or by addition of a denaturant. Also disclosed are the protein additives so obtained, their use in food and non-food applications and food and non-food products containing the protein additives.

WO 2010/037736 a1 discloses the isolation of whey proteins and the preparation of whey products and whey isolates. In particular, the invention relates to the isolation of a beta-lactoglobulin product and the isolation of an alpha-enriched whey protein isolate from whey obtained from animals. The alpha-enriched whey protein isolate provided by the present invention has high alpha-lactalbumin and immunoglobulin G in addition to low beta-lactoglobulin.

FR 2296428 discloses protein compositions for dietary and therapeutic use based on whey proteins (lactoserum proteins) obtained by any known separation method. The composition can be used for treating or preventing digestive disorders (e.g., diarrhea), increasing resistance to intestinal infections, and treating certain metabolic disorders (e.g., hyperphenylalaninemia) in infants and adults. They may also be used dermatologically or cosmetically and may form part of a low protein diet.

Disclosure of Invention

The present inventors provide an instant beverage powder product with a high content of BLG. The product is stable in storage, and can be used as food for treating sialism; i.e. the appearance and taste of the product is appealing to the consumer.

Accordingly, one aspect of the present invention relates to an instant beverage powder comprising at least 1% w/w BLG (preferably at least 5%), wherein:

blg has a crystallinity of at least 20%, preferably at least 40%, and/or

Blg represents at least 85% w/w of the total amount of protein,

the beverage further comprises at least one additional ingredient (additional ingredient) selected from the group consisting of: vitamins, flavoring agents, coloring agents, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, antifoaming agents, and non-whey proteins.

Another aspect of the invention relates to a method for preparing an instant beverage powder comprising BLG and at least one optional ingredient, the method comprising: blending the dried BLG isolate with at least one other ingredient selected from the group consisting of: vitamins, flavoring agents, coloring agents, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, antifoaming agents, and non-whey proteins.

A further aspect of the invention relates to a liquid food product comprising a liquid and a powder according to the invention.

Another aspect of the invention relates to a method for preparing a liquid food product according to the invention, the method comprising:

i. the addition of the instant beverage powder according to the invention,

optionally adding at least one other ingredient (further ingredient), and

mixing the resulting powder and liquid to form a homogeneous mixture.

Another aspect of the invention relates to the instant beverage powder of the invention for use as a nutritional supplement.

Another aspect of the invention relates to a kit comprising the powder of the invention, the kit comprising:

i. a tool for measuring said powder, and

a container having a lid for opening and closing the container,

wherein the container is for mixing the powder with a liquid to form a food product, the container being adapted to drink the food product directly from the container.

Drawings

FIG. 1 is a photomicrograph of BLG crystals recovered from feed 3 of example 3 of PCT application PCT/EP 2017/084553.

Fig. 2 shows a photomicrograph of the whole and fragments of BLG crystals obtained from feed 2 of example 3 of PCT application PCT/EP 2017/084553.

FIG. 3 is a tube photograph of a subsample containing six low phosphorous beverages prepared in example 5.

Detailed Description

Definition of

In the context of the present invention, the term "beta-lactoglobulin" or "BLG" relates to beta-lactoglobulin from a mammalian species, e.g. in native, unfolded and/or glycosylated form, and includes naturally occurring genetic variants. The term also includes aggregated BLG, precipitated BLG and crystallized BLG. When referring to the amount of BLG, reference is made to the total amount of BLG including aggregated BLG. The total amount of BLG was determined according to example 1.31. The term "aggregated BLG" relates to the following BLGs: the BLG is at least partially unfolded and further aggregated with other denatured BLG molecules and/or other denatured whey proteins, typically by hydrophobic interactions and/or covalent bonds.

BLG is the most predominant protein in bovine whey and is present in several genetic variants, the major variants in cow milk being labeled a and B. BLG is a lipocalin protein that binds to a number of hydrophobic molecules, suggesting a role in their transport. BLG has also been shown to be able to bind iron via siderophores and may play a role in combating pathogens. Human breast milk lacks homologs of BLG.

Bovine BLG is a relatively small protein having about 162 amino acid residues and a molecular weight of about 18.3-18.4 kDa. Under physiological conditions, it is mainly dimeric, but dissociates to monomers below about pH3, retaining its native state as determined using nmr spectroscopy. In contrast, BLG also exists as tetramers, octamers, and other multimeric aggregated forms under a variety of natural conditions.

In the context of the present invention, the term "non-aggregated β -lactoglobulin" or "non-aggregated BLG" also relates to β -lactoglobulin from a mammalian species, e.g. in native, expanded and/or glycosylated form, and includes naturally occurring genetic variants. However, the term does not include aggregated BLG, precipitated BLG, or crystallized BLG. The amount or concentration of unaggregated BLG is determined according to example 1.6.

By calculating (m)_{Total BLG}-m_{Non-aggregated BLG})/m_{Total BLG}X 100% to determine the percentage of non-aggregated BLG relative to the total BLG. m is_{Total BLG}Is the concentration or amount of BLG determined according to example 1.31, and m_{Non-aggregated BLG}Is the concentration or amount of non-aggregated BLG determined according to example 1.6.

In the context of the present invention, the term "crystal" refers to a solid material whose constituents (e.g. atoms, molecules or ions) are arranged in a highly ordered microstructure, forming a lattice that extends in all directions.

In the context of the present invention, the term "BLG crystals" refers to protein crystals arranged in a highly ordered microstructure forming an omni-directionally extended lattice, which protein crystals predominantly comprise non-aggregated and preferably native BLG. The BLG crystal may be, for example, monolithic or polycrystalline, and may be, for example, a whole crystal, a crystal fragment, or a combination thereof. The crystal fragments are formed, for example, when the intact crystals are subjected to mechanical shearing during processing. The crystal fragments also have a highly ordered crystal microstructure, but may lack uniform surfaces and/or uniform edges or corners of the intact crystals. See, for example, fig. 1 for an example of a number of complete BLG crystals and fig. 2 for an example of a BLG crystal fragment. In both cases, the BLG crystal or crystal fragment can be visually identified as a well-defined, compact and coherent structure using an optical microscope. The BLG crystals or crystal fragments are typically at least partially transparent. Furthermore, protein crystals are known to be birefringent, and this optical property can be used to identify unknown particles with a crystal structure. Amorphous BLG aggregates, on the other hand, typically appear as unsharpened, opaque, and as open or porous masses of irregular size.

In the context of the present invention, the term "crystallization" relates to the formation of protein crystals. Crystallization may occur, for example, spontaneously or by addition of seed crystals.

In the context of the present invention, the term "edible composition" refers to a composition that is safe for human consumption and use as a food ingredient and does not contain problematic amounts of toxic ingredients (e.g., toluene) or other unwanted organic solvents.

In the context of the present invention, the term "ALA" or "alpha-lactalbumin" refers to alpha-lactalbumin from a mammalian species, for example in native and/or glycosylated form, and includes naturally occurring genetic variants. The term also includes aggregated ALA and precipitated BLG. When referring to the amount of ALA it is meant the total amount of ALA including e.g. aggregated ALA. The total amount of ALA was determined according to example 1.31. The term "aggregated ALA" refers to ALA which is typically at least partially unfolded and further aggregated with other denatured ALA molecules and/or other denatured whey proteins, typically by hydrophobic interactions and/or covalent bonds.

Alpha-lactalbumin (ALA) is a protein that is present in the milk of almost all mammalian species. ALA forms the regulatory subunit of the Lactose Synthase (LS) heterodimer, and β -1, 4-galactosyltransferase (β 4Gal-T1) forms the catalytic component. These proteins together allow LS to produce lactose by transferring the galactose moiety to glucose. One of its main structural differences with β -lactoglobulin is that ALA does not have any free thiol groups that can act as starting points for covalent aggregation reactions.

In the context of the present invention, the term "non-aggregated ALA" also relates to ALA from a mammalian species, e.g. in native, expanded and/or glycosylated form, and includes naturally occurring genetic variants. However, the term does not include aggregated ALA or precipitated ALA. The amount or concentration of non-aggregated BLG was determined according to example 1.6.

By calculating (m)_{Total ALA}-m_{Non-aggregating ALA})/m_{Total ALA}X 100% to determine the percentage of non-aggregated ALA relative to total ALA. m is_{Total ALA}Is the concentration or amount of ALA determined according to example 1.31, and m_{Non-aggregating ALA}Is the concentration or amount of non-aggregated ALA determined according to example 1.6.

In the context of the present invention, the term "macrocasein" or "CMP" refers to the hydrophilic peptide, residue 106-169, which is derived from the hydrolysis of "kappa-CN" or "kappa-casein" in mammalian species by aspartic proteases (e.g., chymosin), e.g., in native and/or glycosylated form, and includes naturally occurring genetic variants.

In the context of the present invention, the term "BLG isolate" refers to a composition comprising at least 85% w/w BLG relative to the total amount of protein. The total amount of protein of the BLG isolate is preferably at least 30% w/w, preferably at least 80% w/w, relative to the total amount of solids.

In the context of the present invention, the term "BLG isolate powder" refers to BLG isolate in powder form, and preferably a free flowing powder.

In the context of the present invention, the term "BLG isolate liquid" refers to a BLG isolate in liquid form, preferably an aqueous liquid.

The term "whey" refers to the liquid phase remaining after casein in milk has been precipitated and removed. The casein precipitation may be accomplished, for example, by acidification of milk and/or the use of rennet. There are several types of whey, such as "sweet whey" which is a whey product produced by casein based precipitation of lectins and "acid whey" or "acid whey". Acid-based precipitation of casein may be achieved, for example, by addition of food acids or by bacterial culture.

The term "whey" refers to the liquid that remains when casein and milk fat globules are removed from milk, for example by microfiltration or macrofiltration. The whey may also be referred to as "ideal whey".

The term "milk albumin" or "serum protein" refers to the proteins present in milk albumin.

In the context of the present invention, the term "whey protein" refers to the protein found in whey or milk serum. The whey protein may be a subset of the protein species found in whey or milk serum, even a single whey protein species, or it may be a complete set of protein species found in whey or/and milk serum.

In the context of the present invention, the main non-BLG proteins of standard whey protein concentrates from sweet whey are ALA, CMP, bovine serum albumin (bovine serum albumin), immunoglobulins, osteopontin, lactoferrin and lactoperoxidase. In the context of the present invention, the weight percentage of each major non-BLG whey protein of a standard whey protein concentrate from sweet whey is:

the ALA content was 18% w/w relative to the total protein content,

the CMP content was 18% w/w relative to the total amount of protein,

the BSA content was 4% w/w relative to the total protein,

the content of casein species relative to the total amount of protein is 5% w/w,

the content of immunoglobulins is 6% w/w relative to the total amount of protein,

the osteopontin content was 0.5% w/w relative to the total amount of protein,

the content of lactoferrin is 0.1% w/w relative to the total amount of protein,

the amount of lactoperoxidase was 0.1% w/w relative to the total amount of protein.

The term casein relates to casein found in milk and includes native micellar casein, casein species alone and caseinate salts found in raw milk.

In the context of the present invention, a liquid that is "supersaturated" or "supersaturated with respect to BLG" comprises a concentration of dissolved, unaggregated BLG that is above the saturation point of unaggregated BLG in the liquid under given physical and chemical conditions. The term "supersaturation" is well known in the Crystallization art (see, e.g., Grard Coquerla, "Crystallization of molecular systems from solution: phase diagraphs, supersaturation and other basic concepts", review of the Chemical Society (Chemical Society Reviews), p.2286-. In the context of the present invention, the supersaturation with respect to BLG is determined by the following procedure.

Procedure to test whether a liquid is supersaturated with respect to BLG under a specific set of conditions:

a) 50mL of the liquid sample to be tested was transferred into a centrifuge tube (VWR catalog No. 525-0402) having a height of 115mm, an inner diameter of 25mm and a capacity of 50 mL. In steps a) -h), care should be taken to keep the sample and its subsequent parts under the initial physical and chemical conditions of the liquid.

b) The samples were immediately centrifuged at 3000g for 3.0 minutes with a maximum acceleration of 30 seconds and a maximum deceleration of 30 seconds.

c) Immediately after centrifugation, as much supernatant as possible (without disturbing the pellet if it has formed) is transferred to a second centrifuge tube (of the same type as in step a)

d) A sub-sample of 0.05mL supernatant (sub-sample A) was taken

e) 10mg BLG crystals (at least 98% pure non-aggregated BLG relative to total solids) with a particle size of up to 200 microns were added to a second centrifuge tube and the mixture was stirred.

f) The second centrifuge tube was placed at the initial temperature for 60 minutes.

g) Immediately after step f), the second centrifuge tube was centrifuged at 500g for 10 minutes, and then another 0.05mL aliquot of the supernatant (subsample B) was taken.

h) Recovering the centrifuged pellet of step g), if any, resuspending it in milliQ water and immediately checking the suspension for the presence of crystals observable by microscope.

i) Example 1 was used.The method outlined in 6 determines the concentration of non-aggregated BLG in subsamples a and B-the results are expressed as% BLG w/w relative to the total weight of the subsample. The concentration of non-aggregated BLG of the subsample A is called C_BLG，AThe concentration of non-aggregated BLG of the subsample B is called C_BLG，B。

j) If C is present_BLG，BLower than C_BLG，AAnd crystals are observed in step i), the liquid from which the sample of step a) is taken is supersaturated (under certain conditions).

In the context of the present invention, the terms "liquid" and "solution" include both compositions that are free of particulate matter and compositions that comprise a combination of liquid and solid and/or semi-solid particles (e.g., protein crystals or other protein particles). Thus, a "liquid" or "solution" may be a suspension or even a slurry. However, "liquids" and "solutions" are preferably pumpable.

In the context of the present invention, the terms "whey protein concentrate" (WPC) and "serum protein concentrate" (SPC) relate to dry or aqueous compositions comprising a total amount of protein of 20-89% w/w relative to the total amount of solids.

The WPC or SPC preferably comprises:

20-89% w/w protein relative to the total solids,

15-70% w/w BLG relative to the total amount of protein,

(ii) 8-50% w/w ALA relative to total protein, and

0-40% w/w CMP relative to protein.

Optionally, but also preferably, the WPC or SPC may comprise:

20-89% w/w protein relative to the total solids,

15-90% w/w BLG relative to the total amount of protein,

4-50% w/w ALA relative to total protein, and

0-40% w/w CMP relative to protein.

Preferably, the WPC or SPC comprises:

20-89% w/w protein relative to the total solids,

15-80% w/w BLG relative to the total amount of protein,

4-50% w/w ALA relative to total protein, and

0-40% w/w CMP relative to protein.

More preferably, the WPC or SPC comprises:

70-89% w/w protein relative to the total solids,

30-90% w/w BLG relative to the total amount of protein,

4-35% w/w ALA relative to total protein, and

0-25% w/w CMP relative to protein.

Typically, SPC contains no CMP or only trace CMP.

The terms "whey protein isolate" (WPI) and "serum protein isolate" (SPI) relate to dry or aqueous compositions comprising a total amount of protein of 90-100% w/w relative to the total amount of solids.

The WPI or SPI preferably comprises:

90-100% w/w protein relative to the total solids,

15-70% w/w BLG relative to the total amount of protein,

(ii) 8-50% w/w ALA relative to total protein, and

0-40% w/w CMP relative to the total amount of protein.

Alternatively, but also preferably, the WPI or SPI may comprise:

90-100% w/w protein relative to the total solids,

30-95% w/w BLG relative to the total amount of protein,

4-35% w/w ALA relative to total protein, and

0-25% w/w CMP relative to the total amount of protein.

More preferably, the WPI or SPI may comprise:

90-100% w/w protein relative to the total solids,

30-90% w/w BLG relative to the total amount of protein,

4-35% w/w ALA relative to total protein, and

0-25% w/w CMP relative to the total amount of protein.

Typically, SPI contains no or only a minor amount of CMP.

In the context of the present invention, the term "other protein" refers to a protein that is not BLG. Other proteins present in whey protein solutions typically comprise one or more non-BLG proteins found in milk serum or whey. Non-limiting examples of such proteins are alpha-lactalbumin, bovine serum albumin, immunoglobulin, casein-macropeptide (CMP), osteopontin, lactoferrin, and milk fat globule membrane protein.

The term "consisting essentially of … …" means that the claim or feature in question covers the named material or step as well as those materials or steps that do not materially affect the basic and novel characteristics of the claimed invention.

In the context of the present invention, the phrase "Y and/or X" means "Y" or "X" or "Y and X". The phrase "n" according to the same logic₁，n₂，...，n_i-1And/or n_i"means" n₁"or" n₂"or_i-1"or" n_i", or the ingredients: n is₁，n₂，...n_i-1And n_iAny combination of (a).

In the context of the present invention, the term "dry" or "dried" means that the composition or product in question comprises at most 10% w/w water, preferably at most 6% w/w water, more preferably even less water.

In the context of the present invention, the term "physical microbial reduction" refers to a physical interaction with a composition which results in a reduction of the total amount of viable microorganisms of the composition. The term does not include the addition of chemicals that result in killing of microorganisms. The term also does not include the heat exposure to which the atomized droplets of liquid are exposed during spray drying, but includes possible preheating prior to spray drying.

In the context of the present invention, the pH of the powder refers to the pH of 10g of powder mixed into 90g of demineralized water and measured according to example 1.16.

In the context of the present invention, unless otherwise specified (e.g., total solids or total protein), the weight percent (% w/w) of a component of a certain composition, product or material refers to the weight percent of that component relative to the weight of the particular composition, product or material.

In the context of the present invention, the processing step "concentration" and the verb "concentration" relate to protein concentration, including protein concentration on a total solids basis and protein concentration on a total weight basis. This means that, for example, the concentration does not necessarily require an increase in the absolute concentration w/w of protein in the composition, as long as the content of protein relative to the total amount of solids is increased.

In the context of the present invention, the term "weight ratio" between component X and component Y means the ratio obtained by calculating m_X/m_YA value obtained wherein m_XIs the amount (by weight) of component X, and m_YIs the amount (by weight) of component Y.

In the context of the present invention, the term "at least pasteurization" relates to a heat treatment with a microbiological kill equal to or higher than a heat treatment at 70 ℃ for 10 seconds. The reference for determining the bactericidal effect was E.coli O157: H7(E.coli O157: H7).

In the context of the present invention, the term "whey protein feed" relates to a whey protein source from which the liquid BLG isolate originates. The BLG content of the whey protein feed, typically WPC, WPI, SPC or SPI, relative to the total amount of protein, is lower than that of the liquid BLG isolate.

In the context of the present invention, the term "BLG-enriched composition" relates to a BLG-enriched composition obtained by isolating BLG from a whey protein feed. BLG-enriched compositions typically comprise the same whey protein as the whey protein material, but BLG is present in significantly higher concentrations relative to the total amount of protein than in the whey protein material. BLG-enriched compositions may be prepared from whey protein materials, for example, by chromatography, protein crystallization, and/or membrane-based protein fractionation. The BLG enriched composition comprises BLG in an amount of at least 85% w/w, preferably at least 90% w/w, relative to the total amount of protein. In some cases, the BLG-enriched composition may be used directly as a liquid BLG isolate. However, additional processing is typically required to convert the BLG-enriched composition into a liquid BLG isolate.

In the context of the present invention, the term "whey protein solution" is used to describe a specific aqueous whey protein composition which is supersaturated with respect to BLG in a salt solution mode and which can be used to prepare BLG crystals.

In the context of the present invention, the term "sterile" means that the sterile composition or product in question does not contain any living microorganisms and therefore does not grow during storage at room temperature. The sterilized composition is sterile.

When a liquid such as a beverage product is sterilized and aseptically packaged in sterile containers, it typically has a shelf life of at least six months at room temperature. The sterilization process kills spores and microorganisms that may cause the liquid to deteriorate.

In the context of the present invention, the term "energy content" refers to the total content of energy contained in a food product. The energy content may be measured in kilojoules (kJ) or kilocalories (kcal) and is referred to as calories per serving, e.g., kcal per 100 grams serving. An example is an instant beverage powder with an energy content of 350kcal per 100g of instant beverage powder.

The total energy content of a food product includes energy contributions from all macronutrients (macronutrients) present in the food product, such as energy from proteins, lipids and carbohydrates. The energy distribution from macronutrients in a food product can be calculated based on the amount of macronutrients in the food product and the contribution of the macronutrients to the total energy content of the food product. The energy distribution can be expressed as a percentage of energy (E%) of the total energy content of the food product. For example, for an instant beverage powder comprising 20E% protein, 50E% carbohydrate and 30E% lipid, this means that 20% of the total energy is from protein, 50% of the total energy is from carbohydrate and 30% of the total energy is from fat (lipid).

In the context of the present invention, the term "nutritional supplement" relates to a food product comprising one or more macronutrients (e.g. proteins, lipids and/or carbohydrates), and optionally vitamins and minerals. The nutritional supplement may be complete or may be incomplete.

The term "nutritionally complete nutritional supplement" is understood to include food products comprising proteins, lipids and carbohydrates, and further comprising vitamins, minerals and trace elements (trace elements), wherein the food product has nutritional ingredients that are matched to a complete and healthy diet.

The term "nutritionally incomplete supplement" refers to a food product comprising one or more macronutrients and optionally also vitamins, minerals and trace elements. A nutritionally incomplete supplement may contain protein as the only nutrient, or may contain protein, lipids, and carbohydrates.

The term "Food for Special Medical Purposes (FSMP)" or "medical food" is a food for oral or tube feeding (tube feeding) for specific medical disorders, diseases or conditions having unique nutritional requirements and used under medical supervision. The medical food can be a supplement with complete nutrition or a supplement with incomplete nutrition.

The term "nutrient" refers to a substance used by an organism to survive, grow, and reproduce. The nutrient may be a macronutrient or a micronutrient. Macronutrients are nutrients that provide energy when consumed, such as proteins, lipids, and carbohydrates. Micronutrients are nutrients that are vitamins, minerals and trace elements.

The term "instant beverage powder" or "instant beverage powder product" refers to a powder that can be converted into a liquid beverage by the addition of a liquid, such as water.

In the context of the present invention, the terms "beverage product" and "product" as used as an entity (substentive) refer to any water-based liquid that can be ingested as a beverage, e.g. by pouring, sipping or tube feeding.

In the context of the present invention, the term "protein fraction" relates to the protein in the composition in question, for example in a powder or beverage preparation.

In the context of the present invention, the term "astringency" relates to mouthfeel. Astringency feels like contraction of the cheek muscles, resulting in increased salivation. Thus, astringency is not really a taste, but a physical mouth feel in the mouth and a feeling of change with time.

In the context of the present invention, the term "dry mouthfeel" relates to the sensation in the mouth, which is perceived as dry mouth and teeth and results in a minimization of salivary secretion. Thus, dry mouthfeel is not actually taste, but physical mouthfeel in the mouth and a time-varying sensation.

In the context of the present invention, the term "mineral" as used herein, unless otherwise indicated, refers to any of a major mineral, a trace or trace mineral, other minerals, and combinations thereof. The main minerals include calcium, phosphorus, potassium, sulfur, sodium, chlorine, and magnesium. Trace or trace minerals include iron, cobalt, copper, zinc, molybdenum, iodine, selenium, manganese, and other minerals include chromium, fluorine, boron, lithium, and strontium.

In the context of the present invention, unless otherwise indicated, the terms "lipid", "fat" and "oil" as used herein are used interchangeably and refer to a lipid material derived or processed from a plant or animal. These terms also include synthetic lipid materials, so long as such synthetic materials are suitable for human consumption.

In the context of the present invention, the term "transparent" includes beverage products that have a distinctly clear appearance and which allow light to pass through and display a clear image therethrough. The turbidity (turbidity) of the clear beverage was at most 200 NTU.

In the context of the present invention, the term "opaque" includes beverage products which have a distinctly unclear appearance and which have a turbidity of greater than 200 NTU.

In the context of the present invention, the term "mother liquor" relates to the whey protein solution remaining after the BLG has been crystallized and the BLG crystals have been at least partially removed. The mother liquor may still contain some BLG crystals, but typically only small BLG crystals that escape separation.

The term "instant beverage powder" or "instant beverage powder product" refers to a powder that can be converted into a liquid beverage by the addition of a liquid (e.g., water).

Detailed Description

Consumers are aware of the overall concept of nutritional supplements. The nutritional supplement should be salivating in taste and appearance, otherwise it will be rejected by the consumer. In addition, consumers appreciate natural products that do not contain additives. Another parameter that is important to consumers is the shelf life of the product.

One aspect of the present invention relates to an instant beverage powder comprising at least 1% w/w (preferably at least 5%) of BLG, wherein:

blg has a crystallinity of at least 20%, preferably at least 40%, and/or

Blg represents at least 85% w/w of the total amount of protein,

the beverage powder further comprises at least one additional ingredient selected from the group consisting of vitamins, flavorings, colorants, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, anti-foaming agents, and non-whey proteins.

As described in this patent application, the source of BLG used in the instant beverage powder may be a BLG isolate or a BLG isolate powder. In some preferred embodiments of the invention the BLG source comprises at least 90% w/w, more preferably at least 95% w/w, even more preferably at least 98% w/w of the total amount of protein in the instant beverage powder, most preferably all protein in the instant beverage powder.

In some preferred embodiments of the invention, the BLG source is a BLG isolate powder and it is the only protein source in the instant beverage powder.

In some preferred embodiments of the present invention, the instant beverage powder is prepared by dry blending the BLG isolate powder and other ingredients.

In a further preferred embodiment of the present invention, the instant beverage powder is prepared by using at least one ingredient in dissolved form followed by a drying step. The drying step may for example form part of a wet granulation process or a spray drying step.

The BLG of the instant beverage powder of the invention preferably has a low degree of denaturation, e.g. a degree of denaturation of at most 10%, preferably at most 4%, more preferably at most 1%, even more preferably at most 0.4%, even more preferably at most 0.1%. Most preferably, the BLG is not denatured at all. For instant beverage powders, it is advantageous that the BLG has a low degree of denaturation, as this reduces the tendency to foam when mixed with a liquid.

Preferably, the instant beverage powder has a degree of protein denaturation of at most 10%, preferably at most 4%, more preferably at most 1%, even more preferably at most 0.4%, even more preferably at most 0.1%. Most preferably, the protein is not denatured at all.

In one embodiment of the invention the instant beverage powder comprises 1-90% w/w BLG. In a preferred embodiment of the invention the instant beverage powder comprises 30-90% w/w BLG, more preferably in the range of 40-90% w/w BLG, or even more preferably in the range of 50-90% w/w BLG.

In a further preferred embodiment of the invention the instant beverage powder comprises 10-97% w/w BLG. In a preferred embodiment of the invention the instant beverage powder comprises 30-96% w/w BLG, more preferably in the range of 40-95% w/w BLG, or even more preferably in the range of 50-94% w/w BLG.

In one embodiment of the invention the instant beverage powder comprises 1-50% w/w BLG. In a preferred embodiment of the invention the instant beverage powder comprises 2-45% w/w BLG, more preferably in the range of 3-40% w/w BLG, or even more preferably in the range of 3-35% w/w BLG.

In a preferred embodiment of the invention the instant beverage powder comprises at least 85% w/w BLG based on the total amount of protein.

In one embodiment of the invention the instant beverage powder comprises at least 85% w/w of the total amount of protein of BLG, e.g. at least 86% w/w of the total amount of protein of BLG, at least 87% w/w of the total amount of protein of BLG, at least 88% w/w of the total amount of protein of BLG, at least 89% w/w of the total amount of protein of BLG.

In one embodiment of the invention the instant beverage powder comprises at least 91% w/w of total protein BLG, e.g. at least 92% w/w of total protein BLG, at least 93% w/w of total protein BLG, at least 94% w/w of total protein BLG, at least 95% w/w of total protein BLG, at least 96% w/w of total protein BLG, at least 97% w/w of total protein BLG, at least 98% w/w of total protein BLG or at least 99% w/w of total protein BLG.

In some preferred embodiments of the instant beverage powder according to the invention, at least 85% w/w of the protein is BLG. Preferably, at least 88% w/w of the protein is BLG, more preferably at least 90% w/w, even more preferably at least 91% w/w, most preferably at least 92% w/w of the protein is BLG.

Even higher relative amounts of BLG are both feasible and desirable, so in some preferred embodiments of the invention at least 94% w/w of the protein in the instant beverage powder is BLG, more preferably at least 96% w/w of the protein is BLG, even more preferably at least 98% w/w of the protein is BLG, most preferably about 100% w/w of the protein is BLG.

For example, the instant beverage powder preferably comprises BLG in an amount of at least 97.5% w/w of the total amount of protein, preferably in an amount of at least 98.0% w/w, more preferably at least 98.5% w/w, even more preferably at least 99% w/w, most preferably at least 99.5% w/w of the total amount of protein, e.g. in an amount of about 100.0% w/w of the total amount of protein.

The protein of the instant beverage powder is preferably made from mammalian milk, and preferably from ruminant milk, e.g., milk from cows, sheep, goats, buffalos, camels, llamas, horses and/or deer. Proteins derived from bovine milk are particularly preferred. Thus, the protein of the instant beverage powder is preferably a milk protein.

The protein of the instant beverage powder is preferably whey protein or milk albumin, even more preferably bovine whey protein or milk albumin.

Intrinsic tryptophan fluorescence emission rate (I330nm/I350nm) is a measure of the degree of BLG expansion, and the inventors found that intrinsic tryptophan fluorescence emission rate (I330nm/I350nm) was measured according to example 1.1 at high BLG tryptophan fluorescence emission rates, which correlate with low or no expansion of BLG.

In some preferred embodiments of the present invention, the instant beverage powder has an intrinsic tryptophan fluorescence emission rate (I330nm/I350nm) of at least 1.11.

In some preferred embodiments of the present invention, the instant beverage powder has an intrinsic tryptophan fluorescence emissivity (I330nm/I350nm) of at least 1.12, preferably at least 1.13, more preferably at least 1.15, even more preferably at least 1.17, most preferably at least 1.19.

If the instant beverage powder contains substantial amounts of non-protein material, it is preferred to separate the protein fraction before measuring the intrinsic tryptophan fluorescence emissivity. Thus, in some preferred embodiments of the present invention, the intrinsic tryptophan fluorescence emission rate of the protein fraction of the instant beverage powder is at least 1.11.

In some preferred embodiments of the present invention, the intrinsic tryptophan fluorescence emission (I330nm/I350nm) of the protein fraction of the instant beverage powder is at least 1.12, preferably at least 1.13, more preferably at least 1.15, even more preferably at least 1.17, most preferably at least 1.19.

The protein fraction can be separated from the instant beverage powder, for example, by dissolving the instant beverage powder in demineralized water and subjecting the solution to dialysis (dialysis) or ultrafiltration-based diafiltration (ultrafiltration) using a filter which retains the protein.

In some preferred embodiments of the present invention, the instant beverage powder has a crystallinity of BLG of at least 20%, preferably at least 40%, more preferably at least 60%, even more preferably at least 80%, most preferably at least 90%. The crystallinity of BLG of at least 20% means that a substantial amount of BLG is present in the instant beverage powder as dry BLG crystals.

The inventors have found that it is advantageous for the crystallinity of BLG to be at least 20%, as this means that the protein is present in a higher density form than in conventional WPI. This provides a higher overall bulk density of the instant beverage powder and makes it less dusty and easier to handle for the end user. The present inventors have also observed a reduced tendency for particle segregation in dry mix instant beverage powders (e.g., comprising a carbohydrate powder and/or a food acid powder in addition to a protein powder).

The invention makes it possible to provide a low carbohydrate instant beverage powder having both sweetness and high protein content.

Thus, in some preferred embodiments of the present invention, the instant beverage powder comprises:

an energy content in the range of 320-380kcal/100 g of powder, preferably in the range of 350-370kcal/100 g of powder,

-the energy contribution from the protein is in the range of 90-100E%, preferably in the range of 95-100E%,

-the amount of BLG is at least 85% w/w relative to the total amount of protein, preferably at least 90% w/w relative to the total amount of protein, more preferably at least 94% w/w relative to the total amount of protein,

-the contribution of energy from carbohydrates is in the range of 0-10E%, preferably in the range of 0-5E%,

-the total amount of high intensity sweeteners is in the range of 0.01-4% w/w, preferably in the range of 0.05-3%,

-a bulk density of at least 0.45g/mL, preferably at least 0.50g/mL, more preferably at least 0.6 g/mL.

The protein of the instant beverage powder is preferably provided by a BLG isolate powder which: has a pH in the range of i)2 to 4.9, ii)6.1 to 8.5 or iii)5.0 to 6.0 and comprises:

-protein in a total amount of at least 90% w/w, preferably at least 95% w/w,

-BLG at least 85% w/w of the total amount of protein, preferably at least 90% w/w of the total amount of protein, more preferably at least 94% w/w of the total amount of protein,

the BLG isolate powder has:

-a bulk density of at least 0.45g/mL, preferably at least 0.50g/mL, more preferably at least 0.6g/mL, and

-one or more of:

intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11, preferably at least 1.13, more preferably at least 1.15,

a degree of protein denaturation of at most 10%, preferably at least 5%,

a thermostability at pH 3.9 of at most 200NTU, and

-up to 1000 colony forming units/g.

In one embodiment of the present invention, the instant beverage powder further comprises at least one further ingredient selected from the group consisting of: vitamins, flavoring agents, coloring agents, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, and non-whey proteins.

Other ingredients (further ingredients) may ensure that the instant beverage powder contains the required nutrients (i.e. nutrients particularly suitable for patients suffering from or at risk of malnutrition) for patients suffering from kidney disease, for weight gain, or it may be used as a nutritional supplement (e.g. by athletes or sports players).

In a preferred embodiment of the present invention, the instant beverage powder may comprise a substance selected from the group consisting of: vitamin a, vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), vitamin B7 (biotin), vitamin B9 (folic acid) and vitamin B12 (cobalamin), vitamin C, vitamin D, vitamin E, vitamin K, choline, inositol, salts thereof, derivatives thereof, and combinations thereof.

In one embodiment, the instant beverage powder may comprise a flavoring agent selected from the group consisting of salt, flavoring agent, taste enhancer and/or spice. In a preferred embodiment of the invention, the flavour attribute comprises chocolate, cocoa, lemon, orange, lime, strawberry, banana, forest fruit flavours or a combination thereof.

In one embodiment, the instant beverage powder may comprise minerals selected from the group consisting of: boron, calcium, chromium, copper, iodine, iron, magnesium, manganese, molybdenum, nickel, phosphorus, potassium, sodium, selenium, silicon, tin, vanadium, zinc, or combinations thereof.

The instant beverage powder may also comprise salts and minerals normally present in whey or milk-derived products. The mineral content of instant beverage powders is usually expressed as the ash content of the food ingredient or product. In a preferred embodiment, the instant beverage powder may comprise an antioxidant selected from the group consisting of: beta-carotene, vitamin C, vitamin E, selenium, or combinations thereof.

In one embodiment of the present invention, the instant beverage powder may comprise one or more sweeteners, such as carbohydrate sweeteners, polyols and/or high intensity sweeteners. The instant beverage powder may for example comprise a total amount of carbohydrate sweetener in the range of 0.001-20% w/w relative to the total weight of the instant beverage powder. Optionally, the instant beverage powder may comprise a total amount of carbohydrate sweetener in the range of 0.1-15% w/w relative to the total weight of the food product.

In one embodiment of the invention, the instant beverage powder comprises at least one high intensity sweetener. In one embodiment, the at least one high intensity sweetener is selected from the group consisting of: aspartame, cyclamate, sucralose, acesulfame salts, neotame, saccharin, stevia extracts, steviol glycosides (e.g., rebaudioside a), or combinations thereof. In some embodiments of the present invention, it is particularly preferred that the sweetener comprises or even consists of one or more High Intensity Sweeteners (HIS).

High intensity sweeteners are found in both natural and artificial sweeteners, and typically have a sweetness intensity at least ten times that of sucrose.

If used, the total amount of high intensity sweetener is typically in the range of 0.01 to 4% w/w. For example, the total amount of high intensity sweetener may be in the range of 0.05-3% w/w. Alternatively, the total amount of high intensity sweetener may be in the range of 0.1-2.0% w/w.

The choice of sweetener may depend on the beverage to be produced and the consumer of the product, for example it may be adjusted according to the specific diagnosis of the patient. High intensity sugar sweeteners (e.g., aspartame, acesulfame potassium, or sucralose) may be used in beverages that do not require the sweetener to provide energy, while natural sweeteners (e.g., steviol glycosides, sorbitol, or sucrose) may be used for beverages with natural characteristics.

Furthermore, it may be preferred that the sweetener comprises or even consists of one or more polyol sweeteners. Non-limiting examples of useful polyol sweeteners are maltitol, mannitol, lactitol, sorbitol, inositol, xylitol, threitol, galactitol, or combinations thereof. If used, the total amount of polyol sweetener is typically in the range of 1-40% w/w. For example, the total amount of polyol sweetener may be in the range of 2-30% w/w. Alternatively, the total amount of polyol sweetener may be in the range of 4-20% w/w.

In one embodiment of the invention, the instant beverage powder may comprise one or more of the following:

i. sweeteners, such as sugar sweeteners and/or non-sugar sweeteners,

(ii) a flavoring agent,

at least one food acid, such as citric acid or other suitable food acid,

the total amount of Na, K, Mg and Ca in the instant beverage is at most 10mmol/g protein,

wherein the pH of a 10% w/w solution of the powder in demineralised water is in the range 2-8.

The pH of the instant beverage powder can be measured by dissolving 10 grams of the instant beverage powder in 90mL of demineralized water at room temperature, as described in example 1.16.

The present inventors have found that the use of a low phosphorous/low potassium BLG isolate powder in an instant beverage powder is advantageous, for example, for an instant beverage powder that is particularly useful for patients with kidney disease.

By adding sweeteners, flavors and/or food acids, the taste of the product can be tailored to make the instant beverage powder appealing to the consumer. In one embodiment of the invention, the consumer may be a patient for whom the flavor, sweetener and acidity characteristics of the instant beverage powder are adjusted to suit the patient's needs and diagnosis.

In a preferred embodiment of the present invention, the instant beverage powder may comprise a high intensity sweetener and a flavoring agent. In an even more preferred embodiment of the invention the instant beverage powder comprises 0.001-0.05% w/w sucralose and 0.01-0.2% w/w flavour (which is selected from chocolate, cocoa, lemon, orange, lime, strawberry, banana, forest fruit flavour or a combination thereof).

In one embodiment of the invention, the instant beverage powder comprises an anti-foaming agent. The anti-foaming agent may be selected from anti-foaming agents suitable for use in food products. The defoamer may be selected from oil-based defoamers, water-based defoamers, (poly) silicone-based defoamers, EP/PO-based defoamers, or combinations thereof.

The water content of the instant beverage powder is at most 6% w/w. In one embodiment of the invention the instant beverage powder comprises at most 5% w/w water, preferably at most 4% w/w water, more preferably at most 3% w/w water, even more preferably at most 2% w/w water.

The storage stability of the instant beverage powder may be increased when the water content of the powder is reduced.

The inventors have found that it may be advantageous to control the mineral content to achieve certain desired properties of the instant beverage powder.

In some preferred embodiments of the invention the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein. Preferably, the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 6mmol/g protein, more preferably at most 4mmol/g protein, even more preferably at most 2mmol/g protein.

In a further preferred embodiment of the invention the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 1mmol/g protein. Preferably, the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 0.6mmol/g protein, more preferably at most 0.4mmol/g protein, even more preferably at most 0.2mmol/g protein, most preferably at most 0.1mmol/g protein.

In one embodiment of the invention, the instant beverage powder comprises dried BLG crystals (e.g. obtainable by one or more of the methods described in PCT/EP 2017/084553). The instant beverage powder comprising BLG crystals may have a bulk density of at least 0.30g/mL (preferably at least 0.4 g/mL).

The present inventors have observed that instant beverage powders in which at least some of the BLG is present in crystalline form have a higher density than comparable instant BLG compositions without BLG crystals. Thus, in some preferred embodiments of the present invention, the bulk density of the instant beverage powder is at least 0.30g/mL, preferably at least 0.40 g/mL. Preferably, the instant beverage powder has a bulk density of at least 0.45 g/mL. More preferably, the instant beverage powder has a bulk density of at least 0.50 g/mL. Even more preferably, the instant beverage powder has a bulk density of at least 0.6 g/mL. The instant beverage powder may for example have a bulk density of at least 0.7 g/mL.

The bulk density of the instant beverage powder according to the invention is preferably in the range of 0.3-1.0g/mL, preferably in the range of 0.40-0.9g/mL, more preferably in the range of 0.45-0.8g/mL, even more preferably in the range of 0.45-0.75g/mL, even more preferably in the range of 0.50-0.75g/mL, most preferably in the range of 0.6-0.75 g/mL.

The bulk density of the powder can be determined according to example 1.17.

The total amount of protein and the energy content in the instant beverage powder according to the invention depend on the intended use of the instant beverage powder. The energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder.

For instant beverage powders, the energy contribution from the protein may be at least 7E%, preferably at least 25E%, more preferably at least 30E%, even more preferably at least 40E%.

In a preferred embodiment of the invention, the energy contribution from the protein is in the range of 10-30E%, preferably in the range of 10-15E%, or even more preferably 11E%. Optionally, the energy contribution from the protein is in the range of 15-25E%, preferably in the range of 18-22E%.

In a preferred embodiment of the invention the energy contribution from the protein is in the range of 7-25E%, preferably in the range of 10-25E%, more preferably 15-20E%, or even more preferably the instant beverage powder comprises 15E% from protein or 20E% from protein, or alternatively the energy contribution from the protein is in the range of 8-15E%.

In another preferred embodiment of the invention, the energy contribution from the protein is at least 50E%, preferably at least 60E% or at least 70E% or even more preferably at least 80E%. In a preferred embodiment of the invention, the energy contribution from the protein is in the range of 80-100E%, preferably in the range of 90-100E%, or even more preferably in the range of 95-100E%.

In a preferred embodiment of the invention, the energy contribution from the protein is in the range of 30-80E%, preferably in the range of 60-80E%. In another embodiment of the invention the energy contribution from the protein is in the range of 30-40E%, or even more preferably the instant beverage powder comprises 33E% from the protein.

The instant beverage powder may be used as a nutritional supplement (e.g., for treating a patient suffering from or at risk of malnutrition, for a patient suffering from kidney disease, for weight gain), or may be used as a nutritional supplement (e.g., by an athlete or a sports player before, during, or after exercise).

The instant beverage powder of the present invention may comprise other macronutrients than protein. The instant beverage powder may comprise besides protein also carbohydrates and/or lipids. The total lipid content in the instant beverage powder according to the invention depends on the intended use of the instant beverage powder. In one embodiment of the invention, the energy contribution from the lipid is in the range of 0-60E%.

In a preferred embodiment of the invention, the energy contribution from the lipid is in the range of 0-5E%, preferably in the range of 0-3E%, or more preferably in the range of 0-2E% from the lipid.

Even less lipids may be preferred, and thus in a preferred embodiment of the invention the energy contribution from the lipids is in the range of 0-1E%, preferably in the range of 0-0.1E%, or more preferably in the range of 0-0.01E% from the lipids.

In a preferred embodiment of the invention, the energy contribution from the lipids is in the range of 30-60E%, preferably in the range of 30-50E%, or even more preferably the instant beverage powder comprises 35E%, 45E% or 50E% from the lipids. Optionally, the energy contribution from the lipid is in the range of 25-45E%.

In a preferred embodiment of the invention, the energy contribution from the lipids is in the range of 15-20E%, preferably in the range of 16-18E%, or even more preferably the instant beverage powder comprises 16E% from the lipids.

The instant beverage powder may be used as a nutritional supplement, e.g., for treating a patient suffering from or at risk of malnutrition, for a patient suffering from kidney disease, for weight gain, or may be used as a nutritional supplement (e.g., by an athlete or a sports player before, during, or after exercise).

In one embodiment of the invention, the instant beverage powder may comprise carbohydrates in addition to proteins. The energy contribution of carbohydrates to the total energy of the instant beverage powder may be in the range of 0-90E%.

The carbohydrate may be selected from a sugar, an oligosaccharide or a polysaccharide. Examples of sugars are monosaccharides, disaccharides and polyols (polyols). Examples of oligosaccharides are malto-oligosaccharides, such as maltodextrin or other oligosaccharides, such as raffinose, stachyose or fructo-oligosaccharides (fructo-oligosaccharide). Examples of polysaccharides are starches (e.g. amylose, amylopectin or modified starches) and non-starch polysaccharides (e.g. dietary fibres, cellulose, pectin and hydrocolloids). In a preferred embodiment of the invention, the carbohydrate is selected from maltodextrin, sucrose or glucose syrup.

The total carbohydrate content of the instant beverage powder according to the invention depends on the intended use of the instant beverage powder. For instant beverage powders for athletes or sportsmen, carbohydrates in the form of sugars may be added to enhance the immediate energy of the athlete, or carbohydrates in the form of slow carbohydrates or dietary fibers may be added to prolong satiety.

In a preferred embodiment of the invention, the energy contribution from carbohydrates is in the range of 70-90E%, preferably in the range of 75-85E%, or more preferably the instant beverage powder comprises 89E% from carbohydrates.

In a preferred embodiment of the invention, the energy contribution from carbohydrates is in the range of 30-50E%, preferably in the range of 35-45E%, or even more preferably the instant beverage powder comprises 35E%, 45E% or 50E% from carbohydrates. Alternatively, the energy contribution from carbohydrates is in the range of 40-60E%, such as in the range of 45-55E%.

In another preferred embodiment of the invention, the energy contribution from carbohydrates is in the range of 0-20E%. In a preferred embodiment of the invention, the energy contribution from carbohydrates is in the range of 0-10E%, preferably in the range of 0-5E%.

In yet another preferred embodiment of the invention, the energy contribution from the carbohydrate is in the range of 0-4E%, more preferably 0-1E%, even more preferably 0-0.2E%.

In another preferred embodiment of the invention, the energy contribution from carbohydrates is in the range of 3-20E%. In a preferred embodiment of the invention, the energy contribution from carbohydrates is in the range of 4-15E%. In another embodiment of the invention, the energy contribution from carbohydrates is in the range of 45-55E%.

The instant beverage powder may be used as a nutritional supplement, e.g., for treating a patient suffering from or at risk of malnutrition, for a patient suffering from kidney disease, for weight gain, or may be used as a nutritional supplement (e.g., by an athlete or a sports player before, during, or after exercise).

The instant beverage powder of the present invention comprises protein and may comprise lipids and/or carbohydrates in addition to protein depending on the intended use of the instant beverage powder.

In a preferred embodiment of the invention, the powder according to the invention further comprises vitamins, minerals and trace elements. The instant beverage powder may be used as a nutritional supplement, e.g. for treating patients suffering from or at risk of malnutrition, for patients suffering from kidney disease, for weight gain, or it may be used as a nutritional supplement (e.g. by athletes or sports players).

The energy content of the instant beverage powder may be in the range of 200-400kcal per 100g of powder. In a preferred embodiment of the invention the energy content of the instant beverage powder is in the range of 300-400kcal/100 g of powder, even more preferably the energy content of the instant beverage powder is in the range of 320-380kcal/100 g of powder, or most preferably in the range of 350-370kcal/100 g of powder.

In a preferred embodiment of the invention, the instant beverage supplement has an energy profile as follows: 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid. In a more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 10-15E% protein, 75-85E% carbohydrate and 0-1E% lipid. In a preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 11E% protein, 89E% carbohydrate and 0E% lipid.

In one embodiment of the invention, the instant beverage powder comprises protein, carbohydrates and lipids, and optionally vitamins, minerals and trace elements. The instant beverage powder may be designed such that the recommended daily intake of the beverage powder provides the recommended intake of vitamins, minerals and trace elements. However, this is not essential.

Such instant beverage powders may be used as nutritional supplements where the consumer is interested in nutritional supplements having a proportion of macronutrients (reflecting a healthy diet associated with energy distribution, macronutrients and micronutrients), for example where the nutritional supplement is administered under the supervision of a health care professional.

The energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder. In a preferred embodiment of the invention the energy content of the instant beverage powder is in the range of 410-480 kcal/100g powder, even more preferably the energy content of the instant beverage powder is in the range of 420-480kcal/100 g powder or most preferably in the range of 440-460kcal/100 g powder.

In a preferred embodiment of the present invention, the instant beverage supplement has an energy profile as follows; 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid. In a more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 10-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, or even more preferably an energy distribution of 15-20E% protein, 35-45E% carbohydrate and 35-50E% lipid. In an even more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 15E% protein, 35E% carbohydrate and 50E% lipid, an energy distribution of 20E% protein, 45E% carbohydrate and 35E% lipid or an energy distribution of 8-15E% protein, 40-47E% carbohydrate and 45E% lipid.

In a preferred embodiment of the invention, the powder according to the invention further comprises vitamins, minerals and trace elements.

The instant beverage powder may be used as a nutritional supplement, e.g., for treating a patient suffering from or at risk of malnutrition, for a patient suffering from kidney disease, for weight gain, or may be used as a nutritional supplement (e.g., by an athlete or a sports player before, during, or after exercise).

In one embodiment of the invention, the instant beverage powder comprises protein, carbohydrate and lipid. Such instant beverage powders may be used as nutritional supplements, wherein the intake of protein is the highest priority of the consumer, e.g. wherein the consumer wants to supplement a regular diet. The energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder. In a preferred embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal/100 g powder, or even more preferably the energy content of the instant beverage powder is in the range of 200-300kcal/100g powder.

In a preferred embodiment of the invention, the instant beverage supplement has the following energy profile: 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid. In a more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 90-98E% protein, 0-10E% carbohydrate and 0-3E% lipid. In a more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 95-98E% protein, 0-5E% carbohydrate and 0-2E% lipid.

In one embodiment of the invention, the instant powder further comprises vitamins, minerals and trace elements. In a preferred embodiment of the invention, the instant powder further comprises vitamins, minerals and trace elements. The instant beverage powder may be used as a nutritional supplement, e.g., for treating a patient suffering from or at risk of malnutrition, for a patient suffering from kidney disease, for weight gain, or may be used as a nutritional supplement (e.g., by athletes or sports players).

In one embodiment of the invention, the instant beverage powder comprises protein, carbohydrate and lipid. The energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder. In a preferred embodiment of the invention the energy content of the instant beverage powder is in the range of 300-420kcal/100 g of powder, or more preferably the energy content of the instant beverage powder is in the range of 320-380kcal/100 g of powder, or even more preferably the energy content of the instant beverage powder is in the range of 350-370kcal/100 g of powder.

In a preferred embodiment of the present invention, the instant beverage supplement has an energy profile as follows; 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid. In a more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 60-80E% protein, 4-15E% carbohydrate and 16-18E% lipid. In an even more preferred embodiment of the invention, the instant beverage supplement has an energy distribution of 30-40E% protein, 45-55E% carbohydrate and 12-18E% lipid, e.g. an energy distribution of 33E% protein, 46E% carbohydrate and 15E% lipid.

Alternatively, the energy content of the instant beverage powder may be in the range of 150-250kcal per 100g of powder, with an energy distribution as follows: 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, or preferably 15-25E% protein, 45-55E% carbohydrate and 30-40E% lipid.

In one embodiment of the invention, the instant powder further comprises vitamins, minerals and trace elements.

In a preferred embodiment of the invention, the instant powder further comprises vitamins, minerals and trace elements. The instant beverage powder may be used as a nutritional supplement, e.g. for treating patients suffering from or at risk of malnutrition, for patients suffering from kidney disease, for weight gain, or it may be used as a nutritional supplement (e.g. by athletes or sports players).

In a preferred embodiment of the present invention, the instant beverage powder may further comprise a flavoring agent, a coloring agent, a sweetener, an antioxidant, a food acid, a lipid, a carbohydrate, a prebiotic, a probiotic, or a non-whey protein.

For convenience, the instant beverage powder may be sold in the form of a kit (which kit comprises the instant powder of the invention, means for measuring said powder, and a container having a lid for opening and closing the container for mixing said powder with a liquid to form a food product, and said container being adapted to drink the food product directly from the container. Examples of useful containers are, for example, bottles, boxes, bricks, bags and/or packs.

A consumer purchasing the kit will obtain all the items for easily preparing the liquid food product of the present invention. The measuring means ensures that the consumer weighs the correct amount of instant powder for the amount of water in the container.

In one embodiment of the invention, the means for measuring the instant powder is a spoon and the container is a drinking bottle. In one embodiment of the invention, the container has an internal indication (indication) indicating how much liquid is to be filled into the container. In one embodiment of the invention, the lid has an opening adapted to drink the liquid food directly from the container and adapted to close when the liquid food is mixed.

The pH of the instant beverage powder is important because the taste of the product prepared from the instant beverage powder depends on the pH of the product. The pH of the instant beverage powder can be determined by measuring the pH of a 10% w/w solution of the powder in demineralized water at 25 ℃ as described in example 1.16. In one embodiment of the invention the pH of a 10% w/w solution of the instant beverage powder in demineralized water is between 2 and 8 at 25 ℃.

In one embodiment of the invention the pH is in the range of 2.0 to 4.9, such as in the range of 2.5 to 4.7, more preferably in the range of 2.8 to 4.3, even more preferably in the range of 3.2 to 4.0, most preferably 3.4 to 3.9. Optionally, but also preferably, the pH of the instant beverage powder may be in the range of 3.6-4.3.

Alternatively, the pH of a 10% w/w solution of the instant beverage powder in demineralized water is in the range of 5.0-6.0 at 25 ℃, preferably the pH of the powder is in the range of 5.1-5.9, more preferably in the range of 5.2-5.8, even more preferably in the range of 5.3-5.7, most preferably in the range of 5.4-5.6.

Optionally, the pH of the instant beverage powder is in the range of 6.1-8.5, more preferably in the range of 6.2-8.0, even more preferably in the range of 6.3-7.7, most preferably in the range of 6.5-7.5.

A further aspect of the invention relates to the use of the instant beverage as defined herein as an ingredient of a food product.

One aspect of the present invention relates to a packaged instant beverage powder product comprising a container comprising an instant beverage powder product as described herein.

In one embodiment of the invention, the instant beverage powder product is hermetically sealed in a container, optionally packaged with an inert gas.

A variety of different containers may be used to store the instant beverage powder product. For example, the container may be a container selected from the group consisting of a bottle, a can, a bag (bag), a pouch (pouch), and a sachet (sachet).

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprising at least 85% w/w of the total amount of protein of BLG, preferably at least 90% w/w of the total amount of protein of BLG.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w of the total amount of protein of BLG, preferably at least 90% w/w of the total amount of protein of BLG, and the pH of the powder is in the range of 2-8.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w of the total amount of protein of BLG, preferably at least 90% w/w of the total amount of protein of BLG, and the pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w of the total amount of protein of BLG, preferably at least 90% w/w of the total amount of protein of BLG, and the pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w of the total amount of protein of BLG, preferably at least 90% w/w of the total amount of protein of BLG, and the pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w BLG of the total amount of protein, preferably at least 90% w/w BLG of the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w BLG of the total amount of protein, preferably at least 90% w/w BLG of the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w BLG of the total amount of protein, preferably at least 90% w/w BLG of the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprises at least 85% w/w BLG of the total amount of protein, preferably at least 90% w/w BLG of the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprising at least 85% w/w BLG based on the total amount of protein, preferably at least 90% w/w BLG based on the total amount of protein, the pH of the powder being in the range of 2-8, the instant beverage powder further comprising vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprising at least 85% w/w BLG based on the total amount of protein, preferably at least 90% w/w BLG based on the total amount of protein, the pH of the powder being in the range of 2.0-4.9, the instant beverage powder further comprising vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprising at least 85% w/w BLG based on the total amount of protein, preferably at least 90% w/w BLG based on the total amount of protein, the pH of the powder being in the range of 5.0-6.0, the instant beverage powder further comprising vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-500kcal per 100g of powder, the instant beverage powder comprising at least 85% w/w BLG based on the total amount of protein, preferably at least 90% w/w BLG based on the total amount of protein, the pH of the powder being in the range of 6.1-8.5, the instant beverage powder further comprising vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder and the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG based on the total amount of protein, preferably at least 90% w/w BLG based on the total amount of protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG based on the total amount of protein, preferably at least 90% w/w BLG based on the total amount of protein, and the pH of the powder is in the range of 2-8.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 400-500kcal per 100g of powder, the energy distribution is in the range of 7-25E% protein, 30-50E% carbohydrate and 30-55E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder and the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2-8.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-400kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 70-90E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder and the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2-8.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 95% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal/100 g powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-350kcal per 100g of powder, the energy distribution is in the range of 80-98E% protein, 0-20E% carbohydrate and 0-5E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder and the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2-8.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrates and 15-20E% lipids, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrate and 15-20E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 200-420kcal per 100g of powder, the energy distribution is in the range of 30-80E% protein, 3-20E% carbohydrates and 15-20E% lipids, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder and the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2-8.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 2.0-4.9.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 5.0-6.0.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution being in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, and the pH of the powder is in the range of 6.1-8.5.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprising vitamins, minerals and trace elements.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2-8, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 2.0-4.9, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 5.0-6.0, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In one embodiment of the invention the energy content of the instant beverage powder is in the range of 150-250kcal per 100g of powder, the energy distribution is in the range of 10-30E% protein, 40-60E% carbohydrate and 25-45E% lipid, wherein the instant beverage powder comprises at least 85% w/w BLG relative to the total amount of protein, preferably at least 90% w/w BLG relative to the total amount of protein, the pH of the powder is in the range of 6.1-8.5, the instant beverage powder further comprises vitamins, minerals and trace elements, wherein the total amount of Na, K, Mg and Ca in the instant beverage powder is at most 10mmol/g protein.

In some preferred embodiments of the invention, the food product is a dry food product (e.g. a bar or an instant beverage powder) comprising carbohydrate and protein, said dry food product comprising at least 1% w/w BLG, preferably at least 5%, wherein:

i) the crystallinity of the BLG is at least 20%, preferably at least 40%, and/or

ii) BLG represents at least 90% w/w of the total amount of protein.

In some particularly preferred embodiments of the invention, the food product is a low-phosphorous food product comprising up to 40mg phosphorous per 100g protein.

Non-limiting examples of food products are e.g. dairy products, confectionery, beverages, instant beverages, protein bars, enteral nutritional compositions, bakery products.

In a further preferred embodiment of the invention, the food product is an instant beverage powder comprising or even consisting essentially of:

-an edible BLG composition as defined herein in powder form to provide a total amount of BLG of at least 1% w/w (preferably at least 5% w/w), the edible BLG composition having a crystallinity of BLG of at least 20%,

sweeteners in powder form, for example sugar sweeteners and/or non-sugar sweeteners,

-optionally, a flavoring agent,

at least one food acid in powder form, such as citric acid or other suitable food acid, and

-up to 80mg phosphorus per 100g protein, and

wherein the pH of a 10% solution of the instant beverage powder in demineralized water is in the range of 2.5-4.0.

In some preferred embodiments of the present invention, the edible BLG composition comprises:

-at most 6% w/w water,

-protein in a total amount of at least 80% relative to the total amount of solids,

-at least 95% BLG relative to the total amount of protein, and

the edible BLG composition:

is a dry powder, and

-a bulk density of at least 0.50g/mL, preferably at least 0.60 g/mL.

In other preferred embodiments of the present invention, the edible BLG composition comprises:

-at most 6% w/w water,

-protein in a total amount of at least 80% relative to the total amount of solids,

-at least 95% BLG relative to the total amount of protein, and

the edible BLG composition:

-is a dry powder, and the powder is,

-a bulk density of at least 0.50g/mL, preferably at least 0.60g/mL, and

-the crystallinity of BLG is at least 20%, preferably at least 40%.

In other preferred embodiments of the present invention, the edible BLG composition comprises:

-at most 6% w/w water,

-protein in a total amount of at least 80% relative to the total amount of solids,

-at least 95% BLG relative to the total amount of protein,

-up to 80mg phosphorus per 100g protein.

The edible BLG composition:

-is a dry powder.

In yet another preferred embodiment of the present invention, the edible BLG composition comprises:

-at most 6% w/w water,

-protein in a total amount of at least 90% relative to the total amount of solids,

-at least 97% BLG relative to the total amount of protein,

-up to 50mg phosphorus per 100g protein.

The edible BLG composition:

-is a dry powder.

In other preferred embodiments of the present invention, the edible BLG composition comprises:

-at most 6% w/w water,

-protein in a total amount of at least 80% relative to the total amount of solids, preferably in a total amount of at least 90% relative to the total amount of solids,

30-70% BLG relative to the total amount of protein,

-8-25% w/w ALA relative to the total amount of protein,

the edible BLG composition:

is a dry powder, and

-the crystallinity of BLG is at least 20%, preferably at least 40%.

In one aspect of the invention, the liquid food product is prepared from an instant beverage powder. By using the instant beverage powder according to the invention, a liquid food product can be obtained in a very short time.

Accordingly, one aspect of the present invention relates to a method for preparing a liquid food product according to the present invention, said method comprising:

i. the addition of the instant beverage powder according to the invention,

optionally adding at least one other ingredient (further ingredient), and

mixing the obtained powder and liquid to form a homogeneous mixture.

In one embodiment, the liquid is selected from the group consisting of water, dairy products, fruit juices, vegetable juices, beverages, and combinations thereof. In one embodiment, the other ingredient is selected from a fruit or a vegetable.

When mixing the powder and the liquid, the mixing may be performed by shaking. After shaking, the liquid food, instant beverage powder, may be allowed to stand 1/2-2 minutes to allow complete dissolution. One advantage of the instant beverage powder is that the instant beverage powder is readily soluble, forms a homogeneous solution and remains a homogeneous solution, i.e. substantially free of segregation.

One problem commonly associated with instant beverage powders is that when liquid food products are prepared from the powder, foaming occurs. The instant beverage powder of the present invention has a tendency not to foam when made from powder into a liquid food product.

A liquid food product comprising a liquid and a powder according to the invention can be prepared by mixing the instant beverage powder according to the invention with a liquid. In one embodiment of the invention, the instant beverage powder may comprise at most 40 grams of said powder per 100 grams of said liquid, such as at most 30 grams of said powder per 100 grams of said liquid. In a preferred embodiment of the invention, the liquid food product comprises 1-30 grams of said powder per 100 grams of said liquid, more preferably 1-20 grams of said powder per 100 grams of said liquid, even more preferably 1-10 grams of said powder per 100 grams of said liquid, or preferably 1-5 grams of said powder per 100 grams of said liquid, or even more preferably 2.5-5 grams of said powder per 100 grams of said liquid.

In one embodiment of the invention, the liquid food product comprises 5-25 grams of said powder per 100 grams of liquid, preferably 5-25 grams of said powder per 100 grams of said liquid, more preferably 10-15 grams of said powder per 100 grams of said liquid, even more preferably 11-14 grams of said powder per 100 grams of said liquid or more preferably 11-12 grams of said powder per 100 grams of said liquid.

In one embodiment of the invention, the energy content of the comestible is in the range of 30-300kcal/100 g comestible, preferably in the range of 30-100kcal/100 g comestible, more preferably in the range of 40-90kcal/100 g comestible, or even more preferably in the range of 40-70kcal/100 g comestible.

Optionally, the energy content of the comestible is in the range of 100-300kcal/100 g comestible, preferably in the range of 100-250kcal/100 g comestible, or more preferably in the range of 125-225kcal/100 g comestible.

The liquid food product may comprise a liquid selected from the group consisting of water, dairy products, fruit juices, vegetable juices, beverages, and combinations thereof.

The appearance of liquid food products, such as beverages made from instant beverage powders, is of great importance to consumers. Transparency is a parameter used by consumers to evaluate products. One way to determine the transparency of a liquid food product is by measuring the turbidity of the product as described in example 1.7.

In some embodiments of the beverage prepared from the instant beverage powder, it is beneficial that the beverage is transparent. This may be advantageous, for example, when the beverage is used as a sports drink or in "protein water", in which case it is beneficial for the beverage to have an appearance similar to water.

In a preferred embodiment of the invention, the beverage prepared from the instant beverage powder has a turbidity of at most 200NTU, and such beverage is transparent and/or translucent.

In some preferred embodiments of the invention, the beverage prepared from the instant beverage powder has a turbidity of at most 150NTU, or preferably at most 100NTU, or preferably at most 80NTU, or preferably at most 60NTU, or more preferably at most 40NTU, or preferably at most 30NTU, preferably at most 20NTU, more preferably at most 10NTU, more preferably at most 5NTU, even more preferably it has a turbidity of at most 2 NTU.

In a preferred embodiment of the invention, the beverage prepared from the instant beverage powder has a turbidity of more than 200NTU, which is opaque.

In some embodiments of the beverage prepared from the instant beverage powder, it is beneficial that the beverage is opaque. This is advantageous, for example, when the beverage should resemble milk and have a milky appearance. Nutritionally complete nutritional supplements are generally opaque in appearance.

In some preferred embodiments of the invention, the turbidity of the beverage prepared from the instant beverage powder is greater than 250 NTU. Preferably, the beverage has a turbidity of greater than 300NTU, more preferably, it has a turbidity of greater than 500NTU, more preferably, it has a turbidity of greater than 1000NTU, preferably a turbidity of greater than 1500NTU, even more preferably it has a turbidity of greater than 2000 NTU.

The color of the product is very important to the consumer. The instant beverage powder product comprising whey protein has a light yellow colour. However, when using instant beverage powders comprising BLG with a crystallinity of at least 20% or instant beverage powders with at least 90% BLG based on protein, the color of the product is substantially less yellow and the product looks whiter. Thus, there is no need to add colour to the instant beverage powder to mask the yellow colour.

One way to measure the color of a product is to use the CIELAB color space, which expresses the color as three values; l represents brightness, a and b represent green-red and blue-yellow color components (components). Example 1.9 describes how to measure L, a and b values of a liquid food product.

In one embodiment of the invention, the protein fraction of the liquid food product has a color value Δ b on the CIELAB scale in the range of-0.10 to +0.51, wherein Δ b ═ b_{Samples normalized to 6.0 w/w% protein}*-b_{Demineralized water}Measured at room temperature.

Another aspect of the invention relates to a method for preparing an instant beverage powder comprising BLG and at least one optional ingredient, the method comprising: blending the dried BLG isolate with at least one other ingredient selected from the group consisting of: vitamins, flavorings, colorants, minerals, sweeteners, antioxidants, food acids, lipids, carbohydrates, prebiotics, probiotics, antifoaming agents, and non-whey proteins to obtain an instant beverage powder.

In one embodiment of the present invention, BLG derived BLG is coated with an organic acid. If the BLG source is, for example, a powder, this means that the powder is coated with an organic acid. Lecithin is often used to improve the solubility of proteins when preparing instant beverage powders comprising proteins. However, lecithin is a source of phosphorus. It is therefore desirable to find another way of improving the solubility of an instant beverage powder comprising protein.

The present inventors have found that by coating BLG-derived BLG crystals or powder particles with one or more organic acids, the solubility of the instant beverage powder is improved. The organic acid or salt of an organic acid may be selected from the group consisting of: pyruvate, (cis) aconitic acid(s), citric acid(s), isocitric acid(s), ketoglutaric acid(s), succinyl-CoA, succinic acid(s), fumaric acid(s), malic acid(s), oxaloacetic acid(s), tartaric acid(s), acetic acid(s), tannic acid, benzoic acid, maleic acid and lactic acid(s). In a preferred embodiment of the present invention, the BLG crystals are coated with an organic acid or a salt of an organic acid selected from the group consisting of: pyruvate, citric acid (salts), isocitric acid (salts), ketoglutaric acid (salts), succinic acid (salts), fumaric acid (salts), malic acid (salts), oxaloacetic acid (salts), tartaric acid (salts), acetic acid (salts), maleic acid and lactic acid (salts) and their salts.

In a preferred embodiment of the present invention, BLG crystals of the BLG source are coated with a citrate salt (e.g., a citrate salt selected from the group consisting of trisodium citrate, potassium citrate, and calcium citrate).

In a preferred embodiment of the present invention, BLG derived BLG is coated with an organic acid by using spray drying or fluidized bed. It is particularly preferred to coat the dried BLG crystals with an organic acid or a salt thereof using, for example, spray drying or a fluidized bed.

The BLG source may be obtained from whey protein feed from which BLG is isolated as crystals. One method of making BLG isolates is described in international patent application No. PCT/EP2017/084553, which is incorporated herein by reference. BLG isolates may be prepared as described in filed PCT/EP2017/084553, page 6, lines 23-32, wherein an edible composition comprising BLG in crystalline and/or isolated form corresponds to the BLG isolate of the present invention. In a preferred embodiment, the BLG isolate is prepared by the method described in page 39, lines 15-34 of the filed PCT/EP 2017/084553. In another preferred embodiment, the BLG isolate is prepared by the method described in filed PCT/EP2017/084553, page 41, lines 1-24.

In some preferred embodiments of the invention, the instant beverage powder comprises or even consists of a BLG isolate powder comprising dried BLG crystals, said BLG isolate powder being coated with an organic acid and/or a salt of an organic acid. The weight ratio between the weight of the BLG isolate and the total weight of the sum of organic acid and deprotonated organic acid is preferably from 5 to 100, more preferably from 8 to 60, even more preferably from 10 to 40, and most preferably from 12 to 30.

One aspect of the present invention relates to a method of producing a BLG isolate powder coated with an organic acid and/or a salt of an organic acid, the method comprising the steps of:

-providing a powder of BLG isolate to be coated, preferably comprising or even consisting of dried BLG crystals, preferably obtained by the BLG crystallization process described herein, and

applying an organic acid and/or a salt of an organic acid to the BLG isolate powder to be coated, preferably in an amount of organic acid and/or salt of an organic acid sufficient to coat the BLG isolate powder but to avoid the BLG isolate powder being dissolved,

-optionally evaporating residual moisture from the coated BLG isolate powder.

The BLG isolate powder to be coated preferably has both a high protein content and a high BLG purity. The BLG isolate powder to be coated preferably has a BLG crystallinity of at least 20% (preferably at least 40%, more preferably at least 60%, even more preferably at least 80%).

The organic acid and/or salt of the organic acid is applied to the BLG isolate powder to be coated in an amount sufficient to provide a weight ratio between the weight of the BLG isolate powder and the total weight of the sum of the organic acid and the deprotonated organic acid of preferably 5 to 100, more preferably 8 to 60, even more preferably 10 to 40, most preferably 12 to 30.

The organic acid and/or salt of the organic acid is preferably applied to the BLG isolate powder in a fluidized bed system by spraying the organic acid and/or salt of the organic acid, preferably in dissolved form into the fluidized bed to coat the BLG isolate powder. The temperature during operation is preferably in the range of 5-70 deg.C, more preferably in the range of 50-65 deg.C, for example, preferably about 60 deg.C.

After application of the organic acid and/or the salt of the organic acid, the coated BLG isolate may be treated to evaporate additional water, preferably until the water content is at most 6% w/w, more preferably at most 5% w/w.

The organic acid is preferably an edible organic acid, so-called food acid.

In some preferred embodiments of the invention, the source of BLG used for preparing the instant beverage powder has a solids content of at least 20% w/w. Preferably, the solid content of the BLG source is at least 30% w/w, more preferably the solid content of the BLG source is at least 40% w/w, even more preferably the solid content of the BLG source is at least 50% w/w, e.g. at least 60% w/w.

In other preferred embodiments of the invention, the source of BLG used for preparing the instant beverage powder has a solids content of 20-80% w/w. Preferably, the solid content of the BLG source is in the range of 30-70% w/w. More preferably, the solid content of the BLG source is in the range of 40-65% w/w. Even more preferably, the solid content of the BLG source is in the range of 50-65% w/w, for example, about 60% w/w.

The BLG source is preferably a BLG isolate powder or an aqueous liquid BLG isolate, and the amount of solids of the BLG isolate powder is in the range of 1-50% w/w. It is particularly preferred that the source of BLG is BLG isolate powder.

A beta-lactoglobulin (BLG) isolated powder, preferably prepared by spray drying, having a pH in the range i)2-4.9, ii)6.1-8.5 or iii)5.0-6.0 and comprising:

-protein in a total amount of at least 30% w/w,

-at least 85% w/w BLG relative to the total amount of protein, and

-water in an amount of at most 10% w/w.

The BLG isolate powder preferably has one or more of the following:

-bulk density of at least 0.2g/cm³，

Intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11,

-a degree of protein denaturation of at most 10%,

a thermostability at pH 3.9 of at most 200NTU, and

-at most 1000 colony forming units/g.

The BLG isolate powder is preferably an edible composition. In some preferred embodiments of the invention, the BLG isolate powder is an edible BLG composition as defined herein.

In some preferred embodiments of the invention, the pH of the BLG isolate powder is in the range of 2-4.9. Such powders are particularly useful for acidic foods and especially acidic beverages.

In other preferred embodiments of the present invention, the pH of the BLG isolate powder is in the range of 6.1-8.5.

In some preferred embodiments of the invention the BLG isolate powder comprises a total amount of protein of at least 40% w/w (preferably at least 50% w/w, at least 60% w/w, more preferably at least 70% w/w, even more preferably at least 80% w/w) of the total amount of protein.

Even higher protein contents may be required and in some preferred embodiments of the invention the BLG isolate powder comprises a total amount of protein of at least 85% w/w, preferably at least 90% w/w, at least 92% w/w, more preferably at least 94% w/w, even more preferably at least 95% w/w of the total amount of protein.

The total amount of protein was measured according to example 1.5.

In some preferred embodiments of the invention the BLG isolate powder comprises BLG in an amount of at least 92% w/w, preferably at least 95% w/w, more preferably at least 97% w/w, even more preferably at least 98% of the total amount of protein, and most preferably in an amount of at least 99.5% w/w BLG, relative to the total amount of protein.

In some preferred embodiments of the invention the sum of alpha-lactalbumin (ALA) and Caseinomacropeptide (CMP) comprises at least 40% w/w, preferably at least 60% w/w, even more preferably 70% w/w, and most preferably at least 90% w/w of the non-BLG proteins of the powder.

In a further preferred embodiment of the invention each major non-BLG whey protein is present in a weight percentage relative to the total amount of protein of at most 25%, preferably at most 20%, more preferably at most 15%, even more preferably at most 10%, most preferably at most 6% by weight relative to the total amount of protein in a standard whey protein concentrate from sweet whey.

Even lower concentrations of each major non-BLG whey protein may be desired. Thus, in a further preferred embodiment of the invention, each major non-BLG whey protein is present in a weight percentage relative to the total amount of protein of at most 4%, preferably at most 3%, more preferably at most 2%, even more preferably at most 1% relative to the total amount of protein of a standard whey protein concentrate from sweet whey.

The inventors have seen evidence that the reduction of lactoferrin and/or lactoperoxidase is particularly advantageous for obtaining a color neutral whey protein product.

Thus, in some preferred embodiments of the invention, lactoferrin is present in a weight percentage relative to the total amount of protein which is at most 25%, preferably at most 20%, more preferably at most 15%, even more preferably at most 10%, most preferably at most 6% by weight relative to the total amount of protein in a standard whey protein concentrate from sweet whey. Even lower concentrations of lactoferrin may be desired. Thus, in a further preferred embodiment of the invention, lactoferrin is present in a weight percentage relative to the total amount of protein which is at most 4%, preferably at most 3%, more preferably at most 2%, even more preferably at most 1% by weight relative to the total amount of protein in a standard whey protein concentrate from sweet whey.

Similarly, in some preferred embodiments of the invention, the lactoperoxidase is present in a weight percentage relative to the total amount of protein of at most 25%, preferably at most 20%, more preferably at most 15%, even more preferably at most 10%, most preferably at most 6% relative to the total amount of protein in a standard whey protein concentrate from sweet whey. Even lower concentrations of lactoperoxidase may be desired. Thus, in a further preferred embodiment of the invention, the lactoperoxidase is present in a weight percentage relative to the total amount of protein which is at most 4%, preferably at most 3%, more preferably at most 2%, even more preferably at most 1% by weight relative to the total amount of protein in a standard whey protein concentrate from sweet whey.

Lactoferrin and lactoperoxidase were quantified according to example 1.29.

In some preferred embodiments of the invention the water content in the BLG isolate powder is at most 10% w/w, preferably at most 7% w/w, more preferably at most 6% w/w, even more preferably at most 4% w/w, most preferably at most 2% w/w.

In some preferred embodiments of the invention, the BLG isolate powder comprises carbohydrate in an amount of at most 60% w/w (preferably at most 50% w/w, more preferably at most 20% w/w, even more preferably at most 10% w/w, even more preferably at most 1% w/w, most preferably at most 0.1% w/w). The BLG isolate powder may, for example, comprise carbohydrates, such as lactose, oligosaccharides and/or hydrolysates of lactose (i.e. glucose and galactose), sucrose, and/or maltodextrin.

In some preferred embodiments of the invention the BLG isolate powder comprises lipids in an amount of at most 10% w/w (preferably at most 5% w/w, more preferably at most 2% w/w, even more preferably at most 0.1% w/w).

The inventors have found that it may be advantageous to control the mineral content to achieve certain desired properties of the BLG isolate powder.

In some preferred embodiments of the invention, the total amount of Na, K, Mg and Ca in the BLG isolate powder is at most 10mmol/g protein. Preferably, the total amount of Na, K, Mg and Ca in the BLG isolate powder is at most 6mmol/g protein, more preferably at most 4mmol/g protein, even more preferably at most 2mmol/g protein.

In other preferred embodiments of the invention, the total amount of Na, K, Mg and Ca in the BLG isolate powder is at most 1mmol/g protein. Preferably, the total amount of Na, K, Mg and Ca in the BLG isolate powder is at most 0.6mmol/g protein, more preferably at most 0.4mmol/g protein, even more preferably at most 0.2mmol/g protein, most preferably at most 0.1mmol/g protein.

In other preferred embodiments of the invention, the total amount of Mg and Ca in the BLG isolate powder is at most 5mmol/g protein. Preferably, the total amount of Mg and Ca in the BLG isolate powder is at most 3mmol/g protein, more preferably at most 1.0mmol/g protein, even more preferably at most 0.5mmol/g protein.

In other preferred embodiments of the invention, the total amount of Mg and Ca in the BLG isolate powder is at most 0.3mmol/g protein. Preferably, the total amount of Mg and Ca in the BLG isolate powder is at most 0.2mmol/g protein, more preferably at most 0.1mmol/g protein, even more preferably at most 0.03mmol/g protein, most preferably 0.01mmol/g protein.

The present inventors have found that a low phosphorous/low potassium variant (variant) of BLG isolate powder can be used, which is particularly useful for patients suffering from kidney disease. To produce such a product, the BLG isolate powder must have the same low phosphorus and potassium content.

Thus, in some preferred embodiments of the invention, the total phosphorus content of the BLG isolate powder is at most 100mg phosphorus per 100g protein. Preferably, the total phosphorus content of the BLG isolate powder is at most 80mg phosphorus per 100g protein. More preferably, the total phosphorus content of the BLG isolate powder is at most 50mg phosphorus per 100g protein. Even more preferably, the total phosphorus content of the BLG isolate powder is at most 20mg phosphorus per 100g protein. The total phosphorus content of the BLG isolate powder is at most 5mg phosphorus per 100g protein.

In some preferred embodiments of the invention, the BLG isolate powder comprises at most 600mg potassium per 100g protein. More preferably, the BLG isolate powder comprises at most 500mg potassium per 100g protein. More preferably, the BLG isolate powder comprises at most 400mg potassium per 100g protein. More preferably, the BLG isolate powder comprises at most 300mg potassium per 100g protein. Even more preferably, the BLG isolate powder comprises at most 200mg potassium per 100g protein. Even more preferably, the BLG isolate powder comprises at most 100mg potassium per 100g protein. Even more preferably, the BLG isolate powder comprises at most 50mg potassium per 100g protein, and even more preferably, the BLG isolate powder comprises at most 10mg potassium per 100g protein.

The phosphorus content is related to the total amount of elemental phosphorus in the composition in question and is determined according to example 1.19. Similarly, the content of potassium is related to the total amount of elemental potassium of the composition in question and is determined according to example 1.19.

In some preferred embodiments of the invention, the BLG isolate powder comprises at most 100mg phosphorus/100 g protein and at most 700mg potassium/100 g protein, preferably at most 80mg phosphorus/100 g protein and at most 600mg potassium/100 g protein, more preferably at most 60mg phosphorus/100 g protein and at most 500mg potassium/100 g protein, more preferably at most 50mg phosphorus/100 g protein and at most 400mg potassium/100 g protein, or more preferably at most 20mg phosphorus/100 g protein and at most 200mg potassium/100 g protein, or more preferably at most 10mg phosphorus/100 g protein and at most 50mg potassium/100 g protein. In some preferred embodiments of the invention, the BLG isolate powder comprises at most 100mg phosphorus per 100g protein and at most 340mg potassium per 100g protein.

The low phosphorus and/or low potassium compositions of the present invention can be used as food ingredients for the production of food products for a population of patients with renal dysfunction.

The present inventors have found that for certain applications (e.g. acidic food products, especially acidic beverages) it is particularly advantageous to have an acidic BLG isolate powder having a pH of at most 4.9 (even more preferably at most 4.3). This is especially true for high protein, clear acidic beverages.

In the context of the present invention, the turbidity of the clear liquid measured according to example 1.7 is at most 200 NTU.

Thus, in some preferred embodiments of the invention, the pH of the BLG isolate powder is in the range of 2-4.9. Preferably, the pH of the BLG isolate powder is from 2.5 to 4.7, more preferably from 2.8 to 4.3, even more preferably from 3.2 to 4.0, and most preferably from 3.4 to 3.9. Alternatively, but also preferably, the pH of the BLG isolate powder may be in the range of 3.6-4.3.

The present inventors have found that for certain applications (e.g., pH neutral foods, especially pH neutral beverages), a BLG isolate powder having a neutral pH is particularly advantageous. This is especially true for high protein, clear or opaque pH neutral beverages.

Thus, in some preferred embodiments of the invention, the pH of the BLG isolate powder is in the range of 6.1-8.5. Preferably, the pH of the powder is in the range of 6.1-8.5, more preferably 6.2-8.0, even more preferably 6.3-7.7, most preferably 6.5-7.5.

In other preferred embodiments of the present invention, the pH of the BLG isolate powder is in the range of 5.0-6.0. Preferably, the pH of the powder is in the range of 5.1-5.9, more preferably in the range of 5.2-5.8, even more preferably in the range of 5.3-5.7, most preferably in the range of 5.4-5.6.

Advantageously, the BLG isolate powder used in the present invention may have at least 0.20g/cm³(preferably at least 0.30 g/cm)³More preferably at least 0.40g/cm³Even more soMore preferably at least 0.45g/cm³And even more preferably at least 0.50g/cm³Most preferably at least 0.6g/cm³) The bulk density of (2).

Low density powders (e.g., lyophilized BLG isolate) are fluffy and are easily inhaled into the air at the manufacturing site during use. This is problematic because it increases the risk of cross-contamination of the lyophilized powder with other food products, and dusty environments are known to be responsible for hygiene problems. In extreme cases, dusty environments also increase the risk of dust explosions.

The high density version of the invention is easier to handle and less prone to flow into the surrounding air.

Another advantage of the high density variants of the present invention is that they take up less space during transportation, thereby increasing the weight of BLG isolate powder that can be transported in one volume unit.

Furthermore, one advantage of the high density variant of the present invention is when combined with other powdered food ingredients (e.g., powdered sugar (bulk density about 0.56 g/cm)³) Granulated sugar (bulk density about 0.71 g/cm)³) Powdered citric acid (bulk density about 0.77 g/cm)³) When used in a powdered mixture, they are less prone to segregation.

The bulk density of the BLG isolate powder of the present invention may be in the range of 0.2 to 1.0g/cm³In the range of 0.30 to 0.9g/cm, preferably³More preferably in the range of 0.40 to 0.8g/cm³In the range of from 0.45 to 0.75g/cm, even more preferably³In the range of from 0.50 to 0.75g/cm, even more preferably³Most preferably in the range of 0.6 to 0.75g/cm³Within the range of (1).

The bulk density of the powder was measured according to example 1.17.

The inventors found that it is advantageous to maintain the natural conformation of BLG, and have seen the following signs: when BLG is used in an acidic beverage, an increase in the spread of BLG results in an increase in the level of dry mouthfeel.

Intrinsic tryptophan fluorescence emissivity (I330/I350) is a measure of the extent of BLG expansion, and the inventors have found that at high intrinsic tryptophan fluorescence emissivity (associated with low or no expansion of BLG), less dry mouthfeel is observed. Intrinsic tryptophan fluorescence emission (I330/I350) was measured according to example 1.1.

In some preferred embodiments of the invention, the BLG isolate powder has an intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11.

In some preferred embodiments of the invention, the intrinsic tryptophan fluorescence emission (I330/I350) of the BLG isolate powder is at least 1.12, preferably at least 1.13, more preferably at least 1.15, even more preferably at least 1.17, and most preferably at least 1.19.

If the BLG isolate powder contains a large amount of non-protein material, it is preferred to separate the protein fraction before measuring the intrinsic tryptophan fluorescence emissivity. Thus, in some preferred embodiments of the invention, the intrinsic tryptophan fluorescence emission rate of the protein fraction of the BLG isolate powder is at least 1.11.

In some preferred embodiments of the invention, the intrinsic tryptophan fluorescence emissivity (I330/I350) of the protein fraction of the BLG isolate powder is at least 1.12, preferably at least 1.13, more preferably at least 1.15, even more preferably at least 1.17, most preferably at least 1.19.

The protein fraction may be separated from the BLG isolate powder, for example, by dissolving the BLG isolate powder in demineralized water and subjecting the solution to dialysis or ultrafiltration-based diafiltration using a filter that retains the protein. If the BLG isolate powder contains interfering levels of lipids, the lipids can be removed, for example, by microfiltration. The steps of microfiltration and ultrafiltration/diafiltration may be combined to remove lipids and small molecules from the protein fraction.

It is generally preferred that the bulk of the BLG isolate powder is non-aggregated BLG. Preferably, at least 50% of the BLGs are non-aggregated BLGs. More preferably, at least 80% of the BLGs are non-aggregated BLGs. Even more preferably, at least 90% of the BLGs are non-aggregated BLGs. Most preferably, at least 95% of the BLGs are non-aggregated BLGs. Even more preferably, about 100% of the BLG isolate powder is non-aggregated BLG.

In some preferred embodiments of the invention, the protein denaturation degree of the BLG isolate powder is at most 10%, preferably at most 8%, more preferably at most 6%, even more preferably at most 3%, even more preferably at most 1%, most preferably at most 0.2%.

However, it may also be preferred that the BLG isolate powder has a significant level of protein denaturation, for example if an opaque beverage is desired. Thus, in other preferred embodiments of the invention, the protein denaturation degree of the BLG isolate powder is at least 11%, preferably at least 20%, more preferably at least 40%, even more preferably at least 50%, even more preferably at least 75%, most preferably at least 90%.

If the BLG isolate powder has a significant level of protein denaturation, it is generally preferred to maintain a low level of insoluble protein material (i.e. precipitated protein material which will precipitate in the beverage during storage). The level of insolubles was determined according to example 1.10.

In some preferred embodiments of the invention, the BLG isolate powder comprises at most 20% w/w insoluble protein material (insoluble protein material), preferably at most 10% w/w insoluble protein material, more preferably at most 5% w/w insoluble protein material, even more preferably at most 3% w/w insoluble protein material, most preferably at most 1% w/w insoluble protein material. Even preferably, the BLG isolate powder does not comprise any insoluble proteinaceous material at all.

In some preferred embodiments of the present invention, the BLG crystallinity of the BLG isolate powder is at most 19%, preferably at most 10%, more preferably at most 5%, most preferably 0%.

In other preferred embodiments of the present invention, the BLG isolate powder has a BLG crystallinity of at least 20%, preferably at least 40%, more preferably at least 60%, most preferably at least 80%. These embodiments contain a large amount of dried BLG crystals and provide the benefit of having the protein source in a solid, dense form.

The present inventors have found that the thermal stability of BLG isolate powder at pH 3.9 is a good indicator of its usefulness for clear high protein beverages. The thermal stability at pH 3.9 was measured according to example 1.2.

Particularly preferably, the thermal stability of the BLG isolate powder at pH 3.9 is at most 200NTU, preferably at most 100NTU, more preferably at most 60NTU, even more preferably at most 40NTU, most preferably at most 20 NTU. Even better thermal stability is possible and the thermal stability of the BLG isolate powder at pH 3.9 is preferably at most 10NTU, preferably at most 8NTU, more preferably at most 4NTU, even more preferably at most 2 NTU.

The microbial content of the BLG isolate powder is preferably kept to a minimum. However, achieving a high degree of protein naturalness and a low content of microorganisms is challenging, as the process of microbial reduction often results in protein unfolding and denaturation. The present invention makes it possible to obtain very low levels of microorganisms while maintaining high levels of BLG naturalness.

Thus, in some preferred embodiments of the invention, the BLG isolate powder contains at most 15000 Colony Forming Units (CFU)/g. Preferably, the BLG isolate powder contains at most 10000 CFU/g. More preferably, the BLG isolate powder comprises at most 5000 CFU/g. Even more preferably, the BLG isolate powder comprises at most 1000 CFU/g. Even more preferably, the BLG isolate powder comprises at most 300 CFU/g. Most preferably, the BLG isolate powder comprises at most 100CFU/g, e.g., at most 10 CFU/g. In a particularly preferred embodiment, the powder is sterile. Sterile BLG isolate powder can be prepared, for example, by combining several physical microbial reduction processes (e.g., microfiltration and heat treatment at acidic pH) during the production of BLG isolate powder.

In some preferred embodiments of the invention, the pH of the BLG isolate powder is in the range i)2 to 4.9, ii)6.1 to 8.5 or iii)5.0 to 6.0, and comprises:

-protein in a total amount of at least 30% w/w, preferably at least 80% w/w, even more preferably at least 90% w/w,

-the amount of beta-lactoglobulin (BLG) relative to the total amount of protein is at least 85% w/w, preferably at least 90% w/w,

-the amount of water is at most 6% w/w,

-the amount of lipid is at most 2% w/w, preferably at most 0.5% w/w,

the BLG isolate powder has:

intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11,

-a degree of protein denaturation of at most 10%, and

-a thermostability at pH 3.9 of at most 200 NTU.

In some preferred embodiments of the invention, the pH of the BLG isolate powder is in the range of i)2 to 4.9 or ii)6.1 to 8.5, and comprises:

-protein in a total amount of at least 30% w/w, preferably at least 80% w/w, even more preferably at least 90% w/w,

-the amount of beta-lactoglobulin (BLG) is at least 85% w/w, preferably at least 90% w/w, relative to the total amount of protein, more preferably at least 94% w/w, relative to the total amount of protein,

-the amount of water is at most 6% w/w,

-the amount of lipid is at most 2% w/w, preferably at most 0.5% w/w,

the BLG isolate powder has:

intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11,

a degree of protein denaturation of at most 10%, preferably at most 5%, and

-a thermostability at pH 3.9 of at most 70NTU, preferably at most 50NTU, even more preferably at most 40 NTU.

In some preferred embodiments of the invention, the pH of the BLG isolate powder is in the range of i)2 to 4.9 or ii)6.1 to 8.5, and comprises:

-protein in a total amount of at least 30% w/w,

-the amount of beta-lactoglobulin (BLG protein) is at least 85% w/w, preferably at least 90% w/w,

-the amount of water is at most 6% w/w,

the BLG isolate powder has:

-bulk density of at least 0.2g/cm³，

Intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11,

-a degree of protein denaturation of at most 10%, and

-a thermostability at pH 3.9 of at most 200 NTU.

In other preferred embodiments of the present invention, the BLG isolate powder has a pH in the range of 2-4.9 and comprises:

-protein in a total amount of at least 80% w/w, preferably at least 90% w/w, even more preferably at least 94% w/w,

-the amount of beta-lactoglobulin (BLG) relative to the total amount of protein is at least 85% w/w, preferably at least 90% w/w relative to the total amount of protein, even more preferably at least 94% w/w relative to the total amount of protein,

-the amount of water is at most 6% w/w,

-the amount of lipid is at most 2% w/w, preferably at most 0.5% w/w,

the BLG isolate powder has:

-bulk density of at least 0.2g/cm³Preferably at least 0.3g/cm³More preferably at least 0.4g/cm³，

Intrinsic tryptophan fluorescence emissivity (I330/I350) of at least 1.11,

-a degree of protein denaturation of at most 10%, preferably at most 5%, more preferably at most 2%, and

-a thermostability at pH 3.9 of at most 50NTU, preferably at most 30NTU, even more preferably at most 10 NTU.

In other preferred embodiments of the present invention, the BLG isolate powder has a pH in the range of 6.1-8.5 and comprises:

-protein in a total amount of at least 80% w/w, preferably at least 90% w/w, even more preferably at least 94% w/w,

-the amount of beta-lactoglobulin (BLG) relative to the total amount of protein is at least 85% w/w, preferably at least 90% w/w relative to the total amount of protein, even more preferably at least 94% w/w relative to the total amount of protein,

-the amount of water is at most 6% w/w,

-the amount of lipid is at most 2% w/w, preferably at most 0.5% w/w,

the BLG isolate powder has:

-bulk density of at least 0.2g/cm³Preferably at least 0.3g/cm³More preferably at least 0.4g/cm³，

-a degree of protein denaturation of at most 10%, preferably at most 5%, more preferably at most 2%, and

-a thermostability at pH 3.9 of at most 50NTU, preferably at most 30NTU, even more preferably at most 10 NTU.

In other preferred embodiments of the present invention, the BLG isolate powder has a pH in the range of 6.1-8.5 and comprises:

-protein in a total amount of at least 80% w/w, preferably at least 90% w/w, even more preferably at least 94% w/w,

-the amount of beta-lactoglobulin (BLG) relative to the total amount of protein is at least 85% w/w, preferably at least 90% w/w relative to the total amount of protein, even more preferably at least 94% w/w relative to the total amount of protein,

-the amount of water is at most 6% w/w,

-the amount of lipid is at most 2% w/w, preferably at most 0.5% w/w,

the BLG isolate powder has:

-bulk density of at least 0.2g/cm³Preferably at least 0.3g/cm³More preferably at least 0.4g/cm³，

-a degree of protein denaturation of at most 10%, preferably at most 5%, more preferably at most 2%, and

-a thermostability at pH 3.9 of at most 50NTU, preferably at most 30NTU, even more preferably at most 10 NTU.

In other preferred embodiments of the present invention, the BLG isolate powder has a pH in the range of 5.0-6.0 and comprises:

-protein in a total amount of at least 80% w/w, preferably at least 90% w/w, even more preferably at least 94% w/w,

-the amount of beta-lactoglobulin (BLG) relative to the total amount of protein is at least 85% w/w, preferably at least 90% w/w relative to the total amount of protein, even more preferably at least 94% w/w relative to the total amount of protein,

-the amount of water is at most 6% w/w,

-the amount of lipid is at most 2% w/w, preferably at most 0.5% w/w,

the BLG isolate powder has:

-bulk density of at least 0.2g/cm³Preferably at least 0.3g/cm³More preferably at least 0.4g/cm³，

A degree of protein denaturation of at most 10%, preferably at most 5%, more preferably at most 2%,

-a thermostability at pH 3.9 of at most 50NTU, preferably at most 30NTU, even more preferably at most 10NTU, and

preferably, the BLG crystallinity is less than 10%.

A BLG isolate powder comprising at least 85% w/w BLG relative to the total amount of protein is typically provided by a process comprising the steps of:

a) providing a liquid BLG isolate having the following characteristics:

i) the pH value is within the range of 2-4.9,

ii) a pH value in the range of 6.1 to 8.5, or

iii) the pH value is in the range of 5.0 to 6.0,

said liquid BLG isolate containing at least 85% w/w BLG relative to the total amount of protein,

b) optionally, subjecting the liquid BLG isolate to physical microbial reduction,

c) the liquid BLG isolate is dried, preferably by spray drying.

BLG isolates are preferably made from mammalian milk, and preferably from ruminant milk (e.g., milk of cows, sheep, goats, buffalos, camels, llamas, mares and/or deer). Proteins derived from bovine milk are particularly preferred. Thus, the BLG is preferably bovine BLG.

The liquid BLG isolate can be provided in a number of different ways.

Typically, providing a liquid BLG isolate involves or even consists of: isolating BLG from a whey protein feed by one or more of the following methods, providing a BLG-enriched composition:

-crystallizing or precipitating BLG by salting-in,

crystallizing or precipitating the BLG in the BLG by salting out (salting-out),

-ion exchange chromatography (ion exchange chromatography), and

fractionation (fractionation) of whey proteins by ultrafiltration.

A particularly preferred way of providing a BLG-enriched composition is by crystallization of BLG, preferably by salt dissolution or alternatively by salting out.

The whey protein feed is preferably WPC, WPI, SPC, SPI or a combination thereof.

The term "whey protein feed" relates to a composition from which the BLG-enriched composition and subsequently the liquid BLG isolate are derived.

In some embodiments of the invention, the preparation of the BLG enriched composition according to US 2,790,790a1 comprises, or even consists of, high salt BLG crystals in the ph range 3.6-4.0.

In other embodiments of the invention, the preparation of the BLG-enriched composition comprises or even consists of the method described by de Jongh et al (Mill Isolation Procedure New Protein Structural Properties of β -Lactoglobulin, J Dairy Sci., vol.84(3),2001, pages 562 571) or by Vyas et al (Scale-Up of Native β -Lactogloblastic Affinity Process, J.Dairy Sci.85: 1639-.

However, in a particularly preferred embodiment of the present invention, the BLG-enriched composition is prepared by crystallization at pH 5-6 under salt-dissolving conditions as described in PCT application PCT/EP2017/084553 (which is incorporated herein by reference).

In some preferred embodiments of the invention, the BLG enriched composition is an edible BLG composition according to PCT/EP2017/084553 comprising at least 90% BLG relative to the total amount of protein, and preferably comprising BLG crystals.

The BLG-enriched composition isolated from the whey protein feed may be subjected to one or more steps selected from the group consisting of:

demineralization (demineralization),

-the addition of minerals,

-dilution of the aqueous phase,

-a concentration of the organic phase,

-physical microbial reduction, and

-a pH adjustment of the aqueous phase,

as part of providing a liquid BLG isolate.

Non-limiting examples of demineralization include, for example, dialysis, gel filtration, UF/diafiltration, NF/diafiltration, and ion exchange chromatography.

Non-limiting examples of adding minerals include adding soluble, food acceptable salts, e.g., salts of Na, K, Ca, and/or Mg. Such salts may for example be phosphate salts, chloride salts or salts of edible acids, such as citrate or lactate. The minerals may be added in solid, suspended or dissolved form.

Non-limiting examples of dilution include, for example, the addition of a liquid diluent (e.g., water, demineralized water, or an aqueous solution of a mineral, acid, or base).

Non-limiting examples of concentration include, for example, evaporation, reverse osmosis, nanofiltration, ultrafiltration, and combinations thereof.

If concentration has to be increased with respect to the concentration of protein relative to the total amount of solids, it is preferred to use a concentration step, such as ultrafiltration or dialysis. Methods such as evaporation, nanofiltration and/or reverse osmosis are useful if concentration does not have to increase the concentration of protein relative to the total amount of solids.

Non-limiting examples of physical microbial reduction include, for example, heat treatment, bacterial filtration, Ultraviolet (UV) radiation, high pressure treatment, pulsed electric field treatment, and ultrasound. These methods are well known to those skilled in the art.

Non-limiting examples of pH adjustment include, for example, the addition of a base and/or acid, and preferably the addition of a food acceptable base and/or acid. Particular preference is given to using acids and/or bases which are capable of chelating divalent metal cations. Examples of such acids and/or bases are citric acid, citrate, EDTA, lactic acid, lactate, phosphoric acid, phosphate and combinations thereof.

In the following, a number of preferred embodiments of the liquid BLG isolate of step a) are provided from a BLG-enriched composition. The process steps mentioned herein are applied to the BLG-containing product stream following the BLG-enriched composition.

The invention has been described above with reference to a number of specific embodiments. However, other embodiments than the above described are equally possible within the scope of the invention. The different features and steps of the various embodiments and aspects of the present invention may be combined in other ways than those described herein, unless expressly stated otherwise.

Examples

Example 1: analytical method

Example 1.1: determination of protein Nature (protein Nature) by intrinsic Tryptophan fluorescence

Tryptophan (Trp) fluorescence spectroscopy is a well-described tool for monitoring protein folding and unfolding. Trp residues buried in native proteins typically show the highest fluorescence emission near 330nm than when present at more solvent-exposed positions (e.g., in unfolded proteins). In unfolded proteins, the wavelength used for Trp fluorescence emission is typically shifted to higher wavelengths and is typically measured around 350 nm. We here used this transition to monitor the thermally induced development by calculating the ratio between the fluorescence emissions at 330nm and 350nm to investigate the effect of heating temperature.

The analysis comprises the following steps:

dilute the beverage composition to 0.6mg/ml in MQ water.

Transfer 300. mu.l of sample to a white 96-well plate to avoid air bubbles, or transfer 3mL of sample to a 10mm quartz cuvette.

The tryptophan fluorescence emission intensity between 310 and 400nm was recorded from the top by excitation at 295 using a 5nm gap.

The samples were measured at 22 ℃ using a Cary Eclipse fluorescence spectrophotometer equipped with a plate reader attachment (G9810A) or a single cuvette holder.

The emission intensity ratio is calculated by dividing the fluorescence emission intensity measured at 330nm by the emission intensity at 350nm, R ═ I330/I350, and is used as a measure of protein naturalness.

R of at least 1.11 describes the predominant native BLG conformation, an

R reporting less than 1.11 involves at least partial unfolding and aggregation

Example 1.2: thermal stability at pH 3.9

Thermal stability at pH 3.9:

the heat stability at pH 3.9 is a measure of the ability of a protein composition to remain clear upon prolonged pasteurization at pH 3.9.

The thermal stability at pH 3.9 was carried out by the following method: a sample of the powder or liquid to be tested is mixed with water (or alternatively, if it is a dilute liquid, it is concentrated by low temperature evaporation) to form an aqueous solution having a pH of 3.9 and containing 6.0% w/w protein, the pH being adjusted to 3.9 with a minimum of the required 0.1M NaOH or 0.1M HCl.

The pH-adjusted mixture was allowed to stand for 30 minutes, and then 25mL of the mixture was transferred to a 30mL thin-walled glass tube. It was heated to 75.0 ℃ for 300 seconds by immersion in a water bath at 75.0 ℃. Immediately after heating, the glass tube was transferred to an ice bath, cooled to 1-5 ℃, and then the turbidity of the heat-treated sample was measured according to example 1.7.

Example 1.3: determination of protein denaturation degree of whey protein composition

It is known that the solubility of denatured whey protein at pH 4.6 is lower than at pH values below or above pH 4.6, and therefore, the denaturation of a whey protein composition is determined by measuring the amount of soluble protein at pH 4.6 relative to the total amount of protein at a certain pH at which the protein in solution is stable.

More specifically, for whey proteins, the whey protein composition (e.g. powder or aqueous solution) to be analyzed is converted into:

-a first aqueous solution comprising a total amount of protein of 5.0% (w/w) and a pH of 7.0 or 3.0, and

-a second aqueous solution comprising 5.0% (w/w) total protein and having a pH of 4.6.

The pH adjustment was carried out using 3% (w/w) NaOH (aq) or 5% (w/w) HCl (aq).

The total amount of protein (P) of the first aqueous solution was determined according to example 1.5_{pH 7.0 or 3.0})。

The second aqueous solution was stored at room temperature for 2 hours and then centrifuged at 3000g for 5 minutes. A sample of the supernatant was recovered and analyzed according to example 1.5 to obtain the protein concentration (S) in the supernatant_{pH 4.6})。

The protein denaturation degree D for the whey protein combination was calculated as follows:

D＝((P_{pH 7.0 or 3.0}-S_{pH 4.6})/P_{pH 7.0 or 3.0})×100％

Example 1.4: protein denaturation (precipitation with pH 4.6 acid) was determined using reverse phase UPLC analysis.

BLG samples (e.g., unheated reference and heated BLG beverage compositions) were diluted to 2% in MQ water. 5mL of the protein solution, 10mL of Milli-Q, 4mL of 10% acetic acid, and 6mL of 1.0M NaOAc were mixed and stirred for 20 minutes to allow the denatured protein to agglomerate at about pH 4.6. The solution was filtered through a 0.22 μm filter to remove agglomerates and non-native proteins.

All samples were diluted to the same extent by the addition of polishing water.

For each sample, the same volume of sample was loaded into a UPLC system with a UPLC chromatography column (Protein BEH C4;1.7 μm; 150x2.1mm) and detected at 214 nm.

The samples were run under the following conditions:

and (3) buffer solution A: Milli-Q water, 0.1% w/w TFA

And (3) buffer solution B: HPLC grade acetonitrile, 0.1% w/w TFA

Flow rate: 0.4ml/min

Gradient: 24-45% B at 0-6.00 min; 45-90% B in 6.00-6.50 min; 90% B at 6.50-7.00 min; 90-24% B in 7.00-7.50 min and 24% B in 7.50-10.00 min.

The area of the BLG peak against a protein standard (Sigma L0130) was used to determine the concentration of native BLG in a sample (5 th order calibration curve)

If the linear range is exceeded, the sample is further diluted and re-injected.

Example 1.5: determination of the Total amount of protein

The total amount of protein (true protein) of the sample was determined by:

1) total nitrogen in the sample was determined according to ISO 8968-1/2| IDF 020-1/2-Milk-Determination of nitrogen content-Part 1/2: Determination of nitrogen content using the Kjeldahl method.

2) The non-protein nitrogen in the samples was determined according to ISO 8968-4| IDF 020-4-Determination of nitrogen content-Part 4: Determination of non-protein-nitrogen content.

3) According to (m)_{Total nitrogen}-m_{Non-protein-nitrogen}) X 6.38 calculate total protein.

Example 1.6: determination of non-aggregated BLG, ALA and CMP

The content of non-aggregated alpha-aggregated whey protein (ALA), beta L lactoglobulin (BLG) and Caseinomacropeptide (CMP) was analyzed by HPLC analysis at 0.4mL/min, respectively. The 25microL filtered sample was injected onto 2 TSKgel3000PWxl (7.8 mm. times.30 cm, Tosohass, Japan) columns in tandem with an attached pre-column PWxl (6 mm. times.4 cm, Tosohass, Japan) equilibrated in an eluent consisting of 465g Milli-Q water, 417.3 g acetonitrile and 1mL trifluoroacetic acid and using a UV detector at 210 nm.

Native alpha-lactalbumin (C) was performed by comparing the peak areas obtained for the corresponding standard proteins with those of the samples_α) Beta l lactoglobulin (C)_β) And caseinomacropeptide (C)_CMP) Quantitative determination of the content of (c).

The total amount of additional protein (non-BLG protein) was determined by subtracting the amount of BLG from the total amount of protein (determined according to example 1.5).

Example 1.7: determination of turbidity

Turbidity is the turbidity (cloudiness) or turbidity (haziness) of a fluid caused by a large number of particles, which are generally invisible to the naked eye, similar to smoke in air.

Turbidity is measured in Nephelometric Turbidity Units (NTU).

20mL of beverage/sample was added to NTU-glass and put in3000 IR turbidimeter. NTU-values were measured after stabilization and repeated twice.

Example 1.8: measurement of viscosity

The viscosity of the beverage product was measured using a rheometer (Anton Paar, Physica MCR 301).

3.8mL of sample was added to cup DG 26.7. The sample was equilibrated to 22 ℃ and then at 50s^-1Pre-shearing under the conditions for 30 seconds, then balancing for 30 seconds, and performing for 1s^-1And 200s^-1And 1s^-1Shear rate sweep (shear rate sweep) between.

Unless otherwise stated, the viscosity is at 100s^-1Expressed in centipoise (cP) at shear rate of (d). The higher the measured cP value, the higher the viscosity.

Alternatively, the viscosity was estimated using the Viscoman of Gilson (Gilson) and taken at about 300s^-1Shear rate report of.

Example 1.9: determination of colour

The color was measured using a colorimeter (Konica Minolta, CR-400). 15g of sample was added to a small petri dish (55X 14.2mm, VWR Cat #391-0895) to avoid bubble formation. The protein content of the sample was normalized to 6.0 w/w% protein or less.

The colorimeter was calibrated to a white calibration plate (No. 19033177). The light source was set to D65 and the viewer was set to 2 degrees. The color (CIELAB color space, a-, b-, L-values) was measured with a lid covering the suspension as the average of three individual readings at different positions of the culture dish.

The demineralized water reference value had the following values:

L*39.97±0.3

a*0.00±0.06

b*-0.22±0.09

the measurement is converted to a delta/difference based on the demineralized water measurement.

ΔL*＝L_{Samples normalized to 6.0 w/w% protein}*-L_{Demineralized water}Measured at room temperature.

Δa*＝a_{Samples normalized to 6.0 w/w% protein}*-a_{Demineralized water}Measured at room temperature.

Δb*＝b_{Samples normalized to 6.0 w/w% protein}*-b_{Demineralized water}Measured at room temperature.

The samples were normalized to 6.0 w/w% protein or less.

L a b color space (also known as CIELAB space) is one of the unified color spaces defined by the international commission on illumination (CIE) in 1976 and is used for quantitative reporting of brightness (luminance) and hue (hue) (ISO 11664-4: 2008(E)/CIE S014-4/E: 2007).

In this space, L denotes luminance (a value from 0 to 100), and when L is 0, it is darkest black, and when L is 100, it is brightest white.

Color channels a and b represent true neutral gray values at a 0 and b 0. The a-axis represents the green-red component, with green in the negative direction and red in the positive direction. b-axis represents the blue-yellow component, with blue in the negative direction and yellow in the positive direction.

Example 1.10: beverage stability test/insoluble protein material

A whey protein beverage composition is considered stable if less than 15% of the total amount of protein in the heated sample precipitates after centrifugation for 5 minutes at 3000 g:

approximately 20g of sample was added to the centrifuge tube and centrifuged at 3000g for 5 minutes.

Keykig analysis (Kjeldahl analysis) of the protein before centrifugation and the supernatant after centrifugation was used for quantitative protein recovery, see example 1.5.

The loss of protein was calculated as follows:

this parameter, sometimes referred to as the level of insoluble proteinaceous material, can be used to analyze liquid and powder samples. If the sample is a powder, 10g of the powder is suspended in 90g of demineralized water and hydrated at 22 ℃ for 1 hour with gentle stirring. Approximately 20 grams of sample (e.g., liquid sample or suspended powder sample) is placed in a centrifuge tube and centrifuged at 3000g for 5 minutes. According to example 1.5, the protein before centrifugation (P) was used_{General assembly}) And supernatant after centrifugation (P)_3000xg) Kjeldahl analysis (Kjeldahl analysis) to quantify protein recovery.

The amount of insoluble proteinaceous material was calculated as follows:

example 1.11: sensory evaluation

Heat treated beverage products were subjected to descriptive sensory evaluations. The beverage product is heated using a plate heat exchanger.

A1 volume sample was mixed with 1 volume water and compared to the unheated whey protein isolate, and a profile list was also formed using lactic acid and citric acid prior to the final taste:

the mouth of the subjects between each sample was cleaned using crackers, white tea, melon and water.

A small cup is charged with 15mL of test sample at ambient temperature (20-25 ℃).

Test samples were provided to 10 persons three times in three different zones in a randomly assigned order.

Attributes (see table above) were scored on a 15cm scale, with 0 being low intensity and 15 being high intensity.

Statistical analysis was performed in "Panelcheck" software, using 3-way ANOVA testing for multiple replicates. Samples were fixed and the panel (panel) was set as random.

Significant differences between samples were assessed using Bonferroni correction meaning minimum significant difference values (pairwise comparison of groups associated with letters).

Example 1.12: determination of transparency by imaging

Photographs of beverage products were made by placing the sample into a measuring vial of turbidity NTU touching a piece of paper with the text "lorem ipsum". Using a smartphone to take a picture of the vial, the inventors evaluated whether the text could be clearly seen through the vial.

Example 1.13: determination of the ash content

The ash content of the food product was according to NMKL 173: 2005 ash in food, gravimetric.

Example 1.14: determination of the conductivity

The "conductivity" (sometimes referred to as "specific conductivity") of an aqueous solution is a measure of the conductive ability of the solution. Conductivity can be determined, for example, by measuring the Alternating Current (AC) resistance of the solution between two electrodes, the results of which are typically expressed in millisiemens per centimeter (mS/cm). Conductivity can be measured, for example, according to EPA (united states environmental protection agency) method 120.1.

Unless otherwise specifically stated, the conductivity values referred to herein have been normalized to 25 ℃.

Conductivity was measured on a conductivity meter (WTW Cond 3210 with tetracon 325 electrodes).

Before use, the system was calibrated as described in the manual. To avoid local dilution, the electrodes were rinsed thoroughly in the same medium as the measurements were performed. The electrodes are lowered into the medium so that the area where the measurement is made is completely submerged. The electrodes are then agitated to remove any air trapped on the electrodes. The electrodes are then held stationary until a stable value can be acquired and recorded from the display.

Example 1.15: determination of the Total solids of the solution

The Total solids of the solution can be determined according to NMKL 110, 2 nd edition, 2005 ((Total solids (Water) -weight determination in milk and milk products)) (Total solids (Water) -Gravimetric determination in milk and milk products). NMKL is "Nordisk MetodikkomLefor"abbreviation of.

The water content of the solution can be calculated as the relative amount (% w/w) of 100% minus the total amount of solids.

Example 1.16: determination of pH

All pH values were measured using pH glass electrodes and normalized to 25 ℃.

The pH glass electrode (with temperature compensation) was carefully rinsed and calibrated prior to use.

When the sample is in liquid form, the pH is measured directly in a liquid solution at 25 ℃.

When the sample was a powder, 10 grams of the powder was dissolved in 90ml of demineralized water at room temperature while vigorously stirring. The pH of the solution was then measured at 25 ℃.

Example 1.17: determination of bulk and bulk Density

The density of the dry powder is defined as the relationship between the weight and volume of the powder, which is analyzed under specified conditions using a special Stampf volumeter (i.e., graduated cylinder). The density is usually expressed in g/ml or kg/L.

In this method, a sample of dry powder is tamped into a measuring cylinder. After a specified number of taps (tapping), the volume of the product is read and the density is calculated.

Three types of densities can be defined by this method:

pour density (poured density), i.e. the mass divided by the volume of powder after transfer to the specified cylinder.

Loose bulk density, i.e. the mass divided by the volume of the powder after 100 taps according to the conditions specified in the present standard.

Bulk density, i.e. the mass divided by the volume of the powder after 625 taps according to the conditions specified in the present standard.

The method uses a special measuring cylinder (250ml, scale 0-250ml, weight 190 + -15 g) (J.Engelsmann A.G.67059Ludwigshafen/Rh)) and a Stampf volumeter (e.g.Engelsmann A.G.).

The bulk and bulk densities of the dried products were determined by the following procedure.

Pretreatment:

the samples to be tested were stored at room temperature.

The sample is then thoroughly mixed by repeatedly rotating and turning the container (to avoid breaking up the particles). The container is filled to a level not exceeding 2/3.

The procedure is as follows:

100.0. + -. 0.1 g of powder are weighed out and transferred into a measuring cylinder. Volume V₀Read in ml.

If the cartridge is not filled with 100 grams of powder, the amount of powder should be reduced to 50g or 25 g.

The cylinder was fixed to a Stampf volumeter and tapped 100 times. The surface is smoothed with a spatula and the volume V is read in ml₁₀₀。

The tap number was changed to 625 (including the 100 taps). After tapping, the surface is flattened and the volume V is read in ml₆₂₅。

And (3) calculating the density:

bulk density and bulk density in g/ml were calculated according to the following formulas:

bulk density of M/V

Where M represents a weighed sample in grams and V represents the volume in ml after 625 taps.

Example 1.18: determination of the Water content in the powder

The water content of the food product is according to ISO 5537: 2004 (dry milk-Determination of moisture content (Reference method)) (referred milk-Determination of motion content (Reference method)). NMKL is "Nordisk MetodikkomLefor"abbreviation of.

Example 1.19: determination of the content of calcium, magnesium, sodium, potassium, phosphorus (ICP-MS method)

The total amount of calcium, magnesium, sodium, potassium and phosphorus was determined using the following procedure: in this procedure, the sample is first decomposed using a microwave digestion method, and then the total amount of mineral(s) is determined using an ICP instrument.

The instrument comprises the following steps:

the microwave oven is from Anton Paar (Anton Paar) and the ICP is Optima 2000DV from Perkin Elmer Inc. (Perkinelmer Inc.).

Materials:

1M HNO₃

at 2% HNO₃Yttrium in (III)

For at 5% HNO₃Suitable standards of calcium, magnesium, sodium, potassium and phosphorus

Pretreatment:

a quantity of powder was weighed out and then transferred to a microwave digestion tube. 5mL of 1M HNO was added₃. The samples were digested in the microwave as per the microwave instructions. The digested tube was placed in a fume hood and the lid removed to allow the volatile smoke to evaporate.

Measurement procedure:

the pretreated sample was transferred to a digestion tube (DigiTUBE) using a known amount of Milli-Q water. 2% of HNO₃The yttrium solution in (1) was added to the digestion tube (about 0.25mL per 50mL diluted sample), and Milli-Q water was usedDilute to a known volume. Samples were analyzed on ICP using the procedure described by the manufacturer.

10mL of 1M HNO by using Milli-Q water₃And 0.5mL in 2% HNO₃The mixture of yttrium solutions in (a) was diluted to a final volume of 100mL to prepare blind samples.

At least 3 standard samples were prepared, the concentrations of which contained the expected sample concentrations.

Example 1.20: determination of the furoic acid value:

the amount of furcellinic acid was determined as described in "Maillard Reaction Evaluation by furinic acid assay During Infant grain Processing", Guerra-Hernandez et al, Journal of grain Science 29(1999) 171-176, and the total amount of protein was determined according to example 1.5. The furosine value is reported in units of mg furosine per 100g protein.

Example 1.21: determination of the crystallinity of BLG in liquids

The following method was used to determine the crystallinity of BLG in liquids having a pH in the range of 5-6.

a) 10mL of the liquid sample in question was transferred to a Maxi-Spin filter of a CA membrane with a pore size of 0.45 microns.

b) The filter was immediately rotated at 1500g for 5 minutes. The centrifuge was maintained at 2 ℃.

c) 2mL of cold Milli-Q water (2 ℃) was added to the retentate side of the spin filter and the filter was immediately spun at 1500g for 5 minutes while keeping the centrifuge cool at 2 ℃, the permeate (permeate A) was collected, the volume was measured by HPLC and the BLG concentration was determined using the method outlined in example 1.31.

d) 4mL of 2M NaCl was added to the retentate side of the filter, stirred rapidly and the mixture was allowed to stand at 25 ℃ for 15 minutes.

e) The filter was immediately spun at 1500g for 5 minutes and the permeate (permeate B) was collected.

f) The total weight of BLG in permeate a and permeate B was determined using the method outlined in example 1.31 and the results were converted to BLGThe total weight of (a) instead of the weight percentage. The weight of BLG in permeate A is called m_{Permeate A}The weight of BLG in permeate B is called m_{Permeate B}。

g) The crystallinity of the liquid with respect to BLG was determined as:

degree of crystallinity ═ m_{Permeate B}/(m_{Permeate A}+m_{Permeate B})×100％

Example 1.22: determination of the crystallinity of BLG in Dry powder

This method was used to determine the crystallinity of BLG in dry powders.

a) A5.0 gram sample of the powder was mixed with 20.0 grams of cold Milli-Q water (2 ℃) and allowed to stand at 2 ℃ for 5 minutes.

b) The liquid sample in question was transferred to a Maxi-Spin filter with a 0.45 micron CA membrane.

c) The filter was immediately spun at 1500g for 5 minutes. The centrifuge was maintained at 2 ℃.

d) 2mL of cold Milli-Q water (2 ℃) was added to the retentate side of the rotary filter, then immediately the filter was rotated at 1500g for 5 minutes, the permeate (permeate A) was collected, the volume was measured, and the BLG concentration was determined by HPLC using the method outlined in example 1.6, and the results were converted to the total weight of BLG instead of weight percent. The weight of BLG in permeate A is called m_{Permeate A}。

f) The crystallinity of BLG in the powder was then calculated using the following formula:

wherein m is_{BLG assembly}Is the total amount of BLG in the powder sample of step a).

If the total amount of BLG of the powder sample is unknown, this can be determined by the following method: another 5g sample of the powder (from the same powder source) was suspended in 20.0 g Milli-Q water, the pH adjusted to 7.0 by adding aqueous NaOH, the mixture was allowed to stand at 25 ℃ for 1 hour with stirring, and finally the total amount of BLG of the powder sample was determined using example 1.31.

Example 1.23: determination of UF permeate conductivity

15mL of sample was transferred to an Amicon Ultra-15 centrifugal filter unit with a cut-off value (cut off) of 3kDa (3000NMWL) and centrifuged at 4000g for 20-30 minutes, or until a sufficient volume of UF permeate for measuring conductivity had accumulated in the bottom of the filter unit. Conductivity was measured immediately after centrifugation. Sample processing and centrifugation are performed at the temperature of the sample source.

Example 1.24: detection of dried BLG crystals in powder

The presence of dry BLG crystals in the powder can be determined by the following method:

the powder sample to be analyzed was resuspended and gently mixed in demineralized water at a temperature of 4 ℃ in a weight ratio of 2 parts water/1 part powder, and then rehydrated at 4 ℃ for 1 hour.

The rehydrated sample is examined microscopically to determine the presence of crystals, preferably using plane polarized light to detect birefringence.

The crystalline material is isolated and subjected to X-ray crystallography to verify the presence of a crystalline structure and preferably also that the crystal lattice (space group and unit cell size) corresponds to that of a BLG crystal.

The chemical composition of the isolated crystalline material was analyzed to verify that its solids consisted primarily of BLG.

Example 1.25: determination of the Total amount of lactose

The total amount of lactose is according to ISO 5765-2: 2002(IDF 79-2: 2002) "milk powder, dry ice mix and processed cheese-determination of lactose content-part 2: an Enzymatic method (Dried milk, Dried rice-mixes and processed cheese-Determination of lactose content-Part 2: enzyme method of using the galactose mobility of the lactose) "Determination of galactose content.

Example 1.26: determination of total amount of carbohydrates:

the amount of carbohydrates was determined by using Sigma Aldrich total carbohydrate assay kit (Cat MAK104-1KT), where carbohydrates were hydrolyzed and converted to furfural and hydroxyfurfural (which were converted to chromophores (chromagen), which were monitored spectrophotometrically at 490 nm).

Example 1.27: determination of the Total amount of lipids

The amount of lipids is according to ISO 1211: 2010 (determination of fat content-Gottlieb gravimetric analysis (Determination of Fat Content--Gottlieb Gravimetric Method)).

Example 1.28: measurement of Brix

Brix measurements were performed using a PAL-alpha digital hand-held refractometer (Atago) (calibrated against polished water (water filtered by reverse osmosis to obtain a conductivity of up to 0.05 mS/cm)).

Approximately 500. mu.l of sample was transferred to the prism surface of the instrument and the measurement was started. The measured values are read and recorded.

The brix of a whey protein solution is proportional to the Total Solids (TS), which is approximately brix 0.85 (% w/w).

Example 1.29 determination of lactoferrin and lactoperoxidase

The concentration of lactoferrin was determined by an ELISA immunoassay as outlined by Soyeur 2012(Soyeur et al; Mid-infrared prediction of lactoferrin content in cow's milk: potential indicator of mastitis (Mid-infrared prediction of lactoferrin content in bovine milk); Animal (2012),6:11, page 1830-1838).

The concentration of lactoperoxidase was determined using a commercially available bovine lactoperoxidase kit.

Example 1.30: determination of the number of colony Forming units

According to ISO 4833-1: 2013 (E): microbiology of food and animal feed-horizontal method of microbial count-Colony counting technique at 30 ℃ (Microbiology of food and animal feeding effects-horizontal method for the environment of microorganisms-Colony-count technology at 30 ℃) the number of Colony forming units per gram of sample is determined.

Example 1.31: determination of the Total amount of BLG, ALA and CMP

The procedure is a liquid chromatography (HPLC) method for the quantitative analysis of proteins (e.g. ALA, BLG and CMP and optionally other proteins in the composition). In contrast to the method of example 1.6, the present method also measures the protein present in aggregation, thus providing a measure of the total amount of protein species in the composition in question.

The separation format was Size Exclusion Chromatography (SEC) using 6M guanidine hydrochloride buffer as sample solvent and HPLC mobile phase. Mercaptoethanol is used as a reducing agent to reduce disulfides (S-S) in proteins or protein aggregates to produce unfolded monomeric structures.

Sample preparation was easily achieved by dissolving 10mg of protein equivalent in the mobile phase.

Two TSK-GEL G3000SWXL (7.7mm x 30.0cm) columns (GPC columns) and one guard column were placed in series to achieve adequate separation of the major proteins in the feedstock.

The eluted analytes were detected and quantified by UV detection (280 nm).

Equipment/materials:

1. HPLC Pump 515 with manual gasket Wash (Waters)

HPLC Pump controller Module II (Waters)

3. Auto sampler 717 (Waters)

4. Double absorbance detector 2487 (Waters)

5. Computer software capable of generating quantitative reports (Empower 3, Waters)

6. And (3) analyzing the column: two TSK-GEL G3000SWXL (7.8X300mm, P/N: 08541).

Protection of the column: TSK-guard column SWxL (6.0X40mm, P/N: 08543).

7. Ultrasonic bath (Branson 5200)

8.25mm syringe filter with 0.2 μm cellulose acetate membrane. (514 0060, VWR)

The procedure is as follows:

mobile phase:

A. stock buffer solution (Stock buffer solution).

1. 56.6g Na were weighed in a 1000mL beaker₂HPO₄、3.5g NaH₂PO₄And 2.9g EDTA. Dissolved in 800mL of water.

2. The pH was measured and adjusted to 7.5. + -. 0.1 with HCl (lowering the pH) or NaOH (increasing the pH) if necessary.

3. Transferred to a 1000mL volumetric flask and diluted to volume with water.

B.6M guanidine hydrochloride mobile phase。

1. 1146g of guanidine hydrochloride are weighed into a 2000mL beaker, and 200mL of stock buffer (A) are added.

2. The solution was diluted with water to about 1600mL while mixing with a magnetic stir bar (50 ℃).

3. The pH was adjusted to 7.5. + -. 0.1 with NaOH.

4. Transferred to a 2000mL volumetric flask and diluted to volume with water.

5. Filtration was performed using a solvent filtration apparatus with a 0.22 μm membrane filter.

And (5) calibrating the standard.

Calibration standards for each protein to be quantified were prepared by:

1. accurately (to 0.01mg) approximately 25mg of the protein reference standard is weighed into a 10mL volumetric flask and dissolved in 10mL of water.

This is a protein stock standard solution of the protein (S1).

2. 200 μ l S1 was pipetted into a 20ml volumetric flask and diluted to volume with mobile phase.

This is the low working standard solution (low working standard solution) WS 1.

3. 500 μ L S1 was pipetted into a 10mL volumetric flask and diluted to volume with mobile phase.

This is standard solution WS 2.

4. 500 μ L S1 was pipetted into a 5mL volumetric flask and diluted to volume with mobile phase.

This is standard solution WS 3.

5. 750 μ L S1 was pipetted into a 5mL volumetric flask and diluted to volume with mobile phase.

This is standard solution WS 4.

6. 1.0mL of S1 was pipetted into a 5mL volumetric flask and diluted to volume with mobile phase.

This is the high working standard solution (high working standard solution) WS 5.

7. Using a calibrated disposable pipette, 1.5mL of WS1-5 was transferred to a separate vial.

Add 10. mu.L of 2-mercaptoethanol to each vial and cap. The solution was vortexed for 10 seconds.

The standards were allowed to stand at ambient temperature for about 1 hour.

8. The standard was filtered using a 0.22 μm cellulose acetate syringe filter.

The purity of the protein (N.times.6.38) was measured using Kjeldahl method (Kjeldahl) and the area% of WS5 as a standard solution was determined using HPLC.

Protein (mg) ═ protein standard weight (mg) × P1 × P2

P1＝P％(Kjeldahl)

Protein area% (HPLC) with P2 ═ protein

Sample preparation

1. The original sample, corresponding to 25mg of protein, was weighed into a 25ml volumetric flask.

2. About 20mL of mobile phase was added and the sample was dissolved for about 30 minutes.

3. The mobile phase was added to a constant volume and 167. mu.L of 2-mercaptoethanol was added to 25ml of the sample solution.

4. Sonicate for about 30 minutes, then place the sample at ambient temperature for about 1¹/₂And (4) hours.

5. The solution was mixed and filtered using a 0.22 μ l cellulose acetate syringe filter.

HPLC system/chromatographic column

Chromatographic column balance

1. A GPC protected column and two GPC analysis columns were connected in series.

The new column is typically transported in phosphate buffered saline.

2. The water was run through a new column stepwise from 0.1 to 0.5mL/min over 30 to 60 minutes.

Rinsing was continued for about 1 hour.

3. The flow rate was gradually reduced from 0.5mL/min to 0.1mL/min and replaced with the mobile phase in the reservoir.

4. The pump flow rate was gradually increased from 0.1mL/min to 0.5mL/min over 30 to 60 minutes to avoid pressure shock and was maintained at 0.5 mL/min.

5. Ten samples were injected to saturate the column and then wait for the peak to elute.

This will facilitate the adjustment of the column.

This step is accomplished without waiting for each implant to be completed before the next implant.

6. Equilibrating with mobile phase for at least 1 hour.

Calculation results

The quantitative determination of the content of the proteins to be quantified (for example. alpha. -lactalbumin,. beta. -lactoglobulin and caseinomacropeptide) is carried out by comparing the peak areas obtained for the corresponding standard proteins with those of the sample. The results are reported as g of the specific protein per 100g of the original sample or as a weight percentage of the specific protein relative to the weight of the original sample.

Example 2: crystallization of beta-lactoglobulin from crude whey protein concentrate

Protocol (protocol):

the lactose depleted UF retentate derived from sweet whey from the standard cheese production process and filtered through a 1.2 micron filter was used as feed for the BLG crystallization process. On an ultrafiltration unit using a Koch HFK-328 type membrane with a 46mil spacer (spacer), a feed pressure of 1.5-3.0 bar, a feed concentration of 21% TS (total solids) ± 5 and polishing water as diafiltration medium (water filtered by reverse osmosis to obtain a conductivity of at most 0.05mS/cm to adjust the sweet whey feed; the temperature of the feed and retentate during ultrafiltration is about 12 ℃ then the pH is adjusted by adding HCl, the diafiltration was continued to obtain pH. of about 5.40 until the conductivity of the retentate dropped below 0.03 mS/cm. over a 20min period and then the retentate was concentrated to about 30% TS (about 23.1% total protein relative to the total weight of the concentrated retentate). A sample of the concentrated retentate was centrifuged at 3000g for 5 min, no visible precipitate formed, HPLC analysis of the supernatant, the composition of the feed is shown in table 1.

The concentrated retentate was seeded with 0.5g/L of pure BLG crystal material obtained from spontaneous BLG crystallization (as described in example 3 in the context of feed 2). The seeded material was prepared by washing the slurry of BLG crystals 5 times in milliQ water, collecting the BLG crystals after each wash. After washing, the BLG crystals were freeze-dried, ground with a pestle and mortar, and then passed through a 200 micron sieve. Thus, the seed particles have a particle size of less than 200 microns.

The concentrated retentate was transferred to a 300L crystallization tank where it was cooled to about 4 ℃ with gentle stirring overnight. The next morning, a sample of the cooled concentrated retentate was transferred to a test tube, visually and microscopically examined. Rapidly deposited crystals formed clearly between nights. The laboratory sample containing the mixture of crystals and mother liquor was further cooled to 0 ℃ in an ice-water bath. The mother liquor and crystals were separated by centrifugation at 3000g for 5 minutes and samples of the supernatant and pellet were taken for HPLC analysis. The crystals were washed once in cold polishing water, then centrifuged again, and the precipitate was freeze-dried.

Table 1 the concentrations of selected feed components normalized to 95% w/w total solids.

Quantification of relative yield of BLG by HPLC:

all samples were diluted to the same extent by the addition of polishing water. The sample was filtered through a 0.22 micron filter. For each sample, the same volume was loaded with Phenomenex5μm C4LC column 250X4.6mm (part number: 00G-4167-E0) on an HPLC system and detected at 214 nm.

The samples were run using the following conditions:

and (3) buffer solution A: MilliQ water, 0.1% w/w TFA

And (3) buffer solution B: HPLC grade acetonitrile, 0.085% w/w TFA

Flow rate: 1ml/min

Gradient: 82-55% A and 18-45% B in 0-30 min; 55-10% of A and 45-90% of B in 30-32 min; 32.5-37.5 minutes 10% A and 90% B; 10-82% A and 90-18% B for 38-48 min.

Data processing:

since all samples were treated in the same manner, we could directly compare the areas of the BLG peaks to obtain relative yields. Since the crystals contained only BLG and all samples were treated in the same manner, the alpha-lactalbumin (ALA) concentration as well as the ALA area should be the same in all samples. Thus, the ALA area before and after crystallization was used as a correction factor (cf) when calculating the relative yield.

The relative yield was calculated by the following equation:

as a result:

as is evident from the chromatograms obtained before and after crystallization of BLG from sweet whey, the concentration of BLG has been greatly reduced and the yield of BLG removed was determined to be 64.5% w/w using the yield calculation as described previously.

The crystal slurry was examined by microscopy. The sample contained hexagonal crystals, many of which were significantly larger than 200 microns in size, indicating that the crystals observed were not only seeded crystals. The crystals are easily broken when pressed with a needle, which demonstrates that they are protein crystals.

The chromatogram of the washed crystal product showed that BLG accounted for 98.9% of the total area of the chromatogram. The purity of the BLG product can be further improved by additional washing.

And (4) conclusion:

this example surprisingly demonstrates that BLG can be selectively crystallized from a crude whey protein concentrate (which contains more than 48% non-BLG protein relative to the total amount of protein) and that the obtained BLG crystal isolate has an extremely high purity. This finding opens up a new route for industrial milk protein isolation, where BLG is separated from other protein components in a gentle manner, which preferably avoids prolonged exposure to high temperatures and problematic chemicals.

Example 3: crystallization of BLG in three types of whey protein solutions

The scheme is as follows:

three different types of whey protein-containing feedstock as crystallization feeds were tested using the same experimental analysis setup as in example 2. However, in the experiment with feed 2, no inoculation was used. Feed 1 and feed 2 were based on sweet whey and had been fat reduced by Synder FR membranes prior to treatment as described in example 2. Feed 3 was derived from acid whey.

The compositions of the three feeds are shown in tables 2, 3 and 4 below. Feed 3 crystallized at 21% TS (total protein 13.3% w/w relative to the total weight of the feed) which was significantly lower than the other two feed concentrations (total protein 26.3% w/w in feed 1 and 25.0% w/w in feed 2).

The slurry of crystallized feed 1 was centrifuged at 1500g for 5 minutes on a Maxi-Spin filter with 0.45 micron CA membrane. Then 2 volumes of MilliQ water were added to the filter cake, which was then centrifuged again. The resulting filter cake was analyzed by HPLC. The precipitate from feed 2 was washed with 2 volumes of MilliQ water and centrifuged again under standard conditions, then the precipitate was analyzed by HPLC. The precipitate from feed 3 was analyzed without washing.

The crystals made from feed 2 were diluted to 10% TS and the pH was adjusted to pH 7 using 1M NaOH to reverse the crystallization. NaCl was added to the crystal slurry (36% TS) from feed 2 to reverse the crystallization.

Table 2: concentration of selected components in feed 1 (sweet whey based whey protein concentrate). BDL-below the limit of detection in the wet sample

Table 3: concentration of selected components in feed 2 (sweet whey based ALA reduced whey protein concentrate). BDL ═ below the limit of detection in wet, non-standardized samples.

Table 4: concentration of selected components in feed 3 (whey protein concentrate based on acid whey).

As a result:

feeding 1:

as can be seen from the chromatograms of the protein composition of the feed and mother liquor, by this method most of the BLG is recovered as crystals. The yield of isolated BLG (calculated as described in example 2) was about 65% relative to the total amount of BLG in the feed.

As is clear from the sample photomicrograph taken during the early stage of the crystallization period and the sample photomicrograph taken at the end of crystallization, the BLG crystals are relatively fragile. Some crystals appeared to break during stirring and turned from hexagonal or rhombohedral to crystal fragments that still appeared very compact and well defined, but had a more irregular shape.

From the chromatogram of the BLG crystals separated and washed on the rotary filter, it can be seen that the purity thereof is very high and the removal of other whey proteins is very effective.

Feeding 2:

from the chromatogram of the protein composition of feed 2 and the mother liquor obtained, it is evident that most of the BLG has been removed and that the calculated yield is 82% with respect to the total amount of BLG in feed 2.

By studying feed 2 before and after crystallization, it can be seen that during crystallization the feed changed from a clear liquid (in which the stirring magnet was visible) to a milky opaque liquid.

Micrographs of BLG crystals show hexagonal shapes, although most of the crystals are broken.

The chromatogram of the precipitate of BLG crystals isolated after washing with 2 volumes of MilliQ water clearly shows that the crystals contain very high purity BLG.

Either the conductivity was increased (by addition of NaCl) or the pH was changed (pH adjusted to 7 by addition of NaOH) so that the environment no longer favoured the crystal structure, in both cases indicating that the milky white suspension became a transparent liquid when the BLG crystals dissolved.

Table 5 provides the mineral composition of the crystalline product obtained from feed 2. We note that the ratio of phosphorus to protein is very low, which makes the crystal preparation suitable as a protein source for patients with kidney disease.

Feeding 3:

as is evident from the chromatograms of the protein composition of feed 3 and the resulting mother liquor, most of the BLG was isolated (calculated yield 70.3% relative to the total amount of BLG in the feed). Higher yields are obtained if the protein content before crystallization is higher.

Photomicrographs of BLG crystals isolated from feed 3 (substantially free of CMP) show that the crystals have a rectangular shape rather than a hexagonal shape. Rectangular crystals appear to be more robust than hexagonal crystals. For chromatograms that are chromatograms of the separated crystal precipitates without washing; the chromatogram clearly shows that the crystal is a BLG crystal, although having a rectangular shape rather than a hexagonal shape.

Table 6: the concentration of the selected component in the crystalline article obtained from feed 3.

The crystalline preparation from feed 3 contained 45mg P/100g protein. We note that the ratio of phosphorus to protein is very low, which makes this crystalline preparation suitable as a protein source for patients with kidney disease.

And (4) conclusion:

all three feeds are suitable for BLG crystallization. The BLG crystals are easily dissolved by adding salt or raising pH or temperature. The new process makes it possible to prepare BLG preparations with very low phosphorus content, which makes them suitable as protein sources for patients suffering from kidney diseases.

Example 4: preparation of spray-dried BLG crystals and determination of bulk Density

A portion of the BLG crystals produced in example 3 (using feed 2) were separated on a decanter centrifuge at 1200g, 5180RPM, 110RPM Diff, with a 64mil spacer (mil means 1/1000 inches)) and a flow rate of 25-30L/h. The BLG crystalline phase is then mixed with finishing water at a ratio of 1: 1 and then separated again on a decanter centrifuge using the same setup. The BLG crystal phase was then mixed with the polishing water to make a slurry containing about 25% dry matter and having a BLG crystallinity of about 80, and subsequently dried on a pilot plant spray dryer (where the inlet temperature was 180 ℃ and the outlet temperature was 85 ℃) without any preheating. Until the temperature of the spray-dried stream was 10-12 ℃. The water content in the resulting powder sampled at the outlet was 4.37% w/w.

The crystallinity of BLG in the slurry was about 90%.

The inventors have also succeeded in separating a slurry of BLG crystals and mother liquor on a decanter centrifuge at 350g, 2750RPM, 150RPM Diff, with 64mil intervals and a flow rate of 75L/h. The BLG crystalline phase is then mixed with finishing water at a ratio of 1: 2 and mixing. The BLG crystal phase was then mixed with finishing water to make it a thinner slurry and then dried on a pilot plant spray dryer using the same parameters as above.

The bulk density of the spray-dried powder was then measured according to example 1.17 and compared to that of a standard WPI dried on the same equipment. The standard WPI was found to have a bulk density (based on 625 punches) of 0.39g/mL) at the high end of the normal range for WPI powders. However, the bulk density of the spray dried BLG crystal preparation was 0.68g/mL, which is more than 75% higher than the bulk density of the standard WPI. This is really surprising and offers many advantages in relation to logistics and applications.

Table 7: concentration of selected components in the spray dried BLG crystal preparation of example 7. BDL is lower than detection limit

Samples of the spray dried BLG crystal preparation were then resuspended in cold demineralised water and the BLG crystals were still clearly visible by microscopy. The BLG crystals are dissolved by adding citric acid or NaCl, and the opaque crystal suspension is converted into a transparent liquid.

The inventors have seen evidence that prolonged heating during the drying step reduces the amount of BLG in crystalline form. Therefore, it is preferable that the heat exposure of the BLG crystal product is as low as possible.

And (4) conclusion:

this example demonstrates that: the slurry containing the BLG crystals may be spray dried and, if the heating during the drying step is controlled, the BLG crystals are still present in the resuspended spray dried powder.

The present inventors have also found that the bulk density of whey protein powder containing BLG crystals significantly exceeds that obtained by normal spray drying of a stream of dissolved protein. High density powders allow for more cost effective packaging and logistics of powders because less packaging material is required per kilogram of powder and more powder (mass) can be transported by a given container or truck.

High density powders also appear to be easier to handle and less bulky and dusty during manufacture and use.

Example 5: low-phosphorus protein beverage

Six low phosphorous instant beverage powders were prepared using the purified BLG product from example 3 (crystalline product obtained from feed 3). All dry ingredients were blended to obtain an instant beverage powder, then mixed with demineralized water to obtain 10kg of each sample, and allowed to hydrate at 10 ℃ for 1 hour.

Table 8: composition of six beverage samples.

A sub-sample of the six samples isTurbidity was measured on a 3000 IR turbidimeter and viscosity was measured on vicom of Gilson (Gilson). The results are shown in the table below.

Table 9: measured viscosity and turbidity of six beverage samples.

Sample (I)	Viscosity (Cp)	NTU
			A	1.42	36.2
B	2.37	46.3
			C	2.69	4.9
D	2.70	5.0
			E	1.45	63.1
F	2.25	82.1

A tube photograph of a subsample containing six low phosphorous beverage samples is shown in figure 3. From left to right, the subsamples are samples A, B, C, D, E and F. Visual inspection of the test tubes verified the turbidity measurements and recorded that all beverage samples were clear, and in particular samples C and D (pH 3.0) were very clear. The low viscosity indicates that the beverage sample is ready for consumption.

All ingredients used to prepare the beverage are low in phosphorus and contain no unnecessary minerals. The obtained beverage therefore has a phosphorus content of about 45mg P/100g protein and usually has a very low mineral content. Thus, the six liquid food products prepared from the instant drink powder are suitable for use as an instant protein drink for patients with kidney disease.

Example 6: crystal separation by dynamic cross-flow filtration

The lactose-depleted UF retentate, derived from sweet whey in a standard cheese manufacturing process and filtered through a 1.2 micron filter, was used as feed for the crystallization process. Sweet whey feed was conditioned on an ultrafiltration unit using a Koch HFK-328 type membrane with a 46mil spacer, a feed pressure of 1.5-3.0 bar, using a feed concentration of 10% TS (total solids) ± 5 and polishing water as diafiltration medium (water filtered by reverse osmosis to obtain a conductivity of at most 0.05 mS/cm). The temperature of the feed and retentate during ultrafiltration was about 12 ℃. The pH was then adjusted by adding HCl to obtain a pH of about 5.60. Diafiltration was continued until the conductivity of the retentate was below 1.30 mS/cm. The feed was then heated to 25 ℃, and the retentate was then concentrated to about 27% TS (about 21% total protein, relative to the total weight of the concentrated retentate). At the end of concentration, the permeate conductivity was 0.33 mS/cm. A sample of the concentrated retentate was centrifuged at 3000g for 5 minutes, but no visible precipitate formed.

The concentrated retentate was transferred to a 300L crystallization tank where it was cooled to about 6 ℃ and held at that temperature overnight with slow stirring. The next morning, the retentate had crystallized. The mother liquor and crystals were separated by centrifugation at 3000g for 5 minutes and the supernatant and precipitated samples were removed for HPLC analysis. The BLG yield for this process was calculated to be 67%.

The crystal slurry from the 300L tank was used as feed in an Andritz DCF 152S system using a disc membrane with a pore size of 500 nm. The filtration was run at 8 ℃ with a rotation speed of 32Hz and a transmembrane pressure of 0.4 bar. The system works as a dead-end filtration, where the retentate accumulates in the filtration chamber, unlike in larger units where the retentate is continuously removed. Filtration was run in a stable manner for just over 40 minutes, at which point the solids accumulated in the filtration chamber began to affect filtration.

During DFC operation, the amount of crystal mass increases significantly.

And (4) conclusion: DCF provides a stable and efficient means for separating crystals from ML. If desired, a wash solution can be added to the DCF.

Example 7: degree of protein denaturation for different whey protein products

The commercial product was compared to the protein denaturation of the four BLG isolates. The BLG isolate is suitable for preparing the instant beverage powder of the present invention. The samples were as follows.

Sample (I)
	A: BiPro (commercially available WPI; Davisco, USA)
BLG Crystal slurry as received-No drying (inventive)
	BLG Crystal slurry Freeze drying (inventive)
Redissolving (pH 7) and freeze-drying the BLG crystals
	BLG Crystal slurry spray drying (inventive)

Samples B-E were prepared as follows:

a crystal slurry was prepared as described in example 6 and isolated as described in example 4. Some of the separated BLG slurry is withdrawn and divided into four portions.

Sample B: re-dissolving the first portion of the separated BLG crystal slurry without any drying by adjusting the pH of the BLG crystal slurry to 7.01 using 3% NaOH; the samples were then diluted to Brix 6(Brix 6) to make an approximately 5% protein solution.

Sample C: a second portion of the separated BLG crystal slurry is freeze-dried. The powder was then resuspended in finished water, the pH adjusted to 7.09 using 3% NaOH, and the sample was then diluted to Brix 6(Brix 6) to make an approximately 5% protein solution.

Sample D: the third portion of the separated BLG crystal slurry was redissolved by adjusting the pH to 7.0 using 3% NaOH, and then freeze-dried. The freeze-dried powder was then resuspended in finished water and the pH was measured to be 7.07. The samples were then diluted to Brix 6(Brix 6) to make an approximately 5% protein solution.

Sample E: the fourth portion of the separated BLG crystal slurry was treated and spray dried as described in example 4. The powder was then resuspended in finishing water and the pH adjusted to 7.04 using 3% NaOH. The samples were then diluted to Brix 6(Brix 6) to make an approximately 5% protein solution.

The degree of protein denaturation was determined for each sample according to example 1.3 and the results are listed in the table below.

Table: the commercially available WPI product (Bipro) was compared to the protein denaturation degree of the 4 BLG products that can be used in the instant beverage powder of the present invention.

And (4) conclusion:

regardless of the drying method, BLG isolates have an unexpectedly low degree of protein denaturation; only one tenth of the above was found in the commercially available WPI used for comparison. It is particularly surprising that the spray dried BLG crystal slurry product still has the lowest degree of denaturation of all products.

Example 8: production of spray-dried acidic BLG isolate powder

Whey protein feed

The lactose depleted UF retentate, derived from sweet whey in a standard cheese manufacturing process, was filtered through a 1.2 micron filter and already reduced in fat by a Synder FR membrane and then used as feed for the BLG crystallization process. The chemical composition of the feed is shown in Table 10. We note that all weight percentages of the specific proteins (e.g. BLG, ALA) mentioned in this example relate to the weight percentage of non-aggregated proteins relative to the total amount of protein.

Regulating

A Koch HFK-328 type membrane (70 m) with a 46mil spacer, a feed pressure of 1.5-3.0 bar was used²Membrane) at 20 ℃, using a feed concentration of 21% Total Solids (TS) ± 5 and polishing water (water filtered by reverse osmosis to obtain a conductivity of at most 0.05 mS/cm) as diafiltration medium. The pH was then adjusted by adding HCl to make the pH about 5.5. Diafiltration was continued until the conductivity of the retentate dropped below 0.1mS/cm over a period of 20 minutes. The retentate was then concentrated until the permeate flow was below 1.43L/h/m². A first sample of the concentrated retentate was taken and centrifuged at 3000g for 5 minutes. The supernatant of the first sample was used to determine the BLG yield.

Crystallization of

The concentrated retentate was transferred to a 300L crystallization tank and seeded therein with pure BLG crystal material made from rehydrated spray-dried BLG crystals. Subsequently, the inoculated whey protein solution was cooled from 20 ℃ to about 6 ℃ for about 10 hours to allow BLG crystals to form and grow.

After cooling, a sample of the whey protein solution containing crystals (second sample) was taken and the BLG crystals were isolated by centrifugation at 3000g for 5 minutes. The supernatant and crystal precipitate from the second sample were subjected to HPLC analysis as described below. The crystallization yield was calculated as described below and determined to be 57%.

Table 10: chemical composition of feed

BLG yield was determined using HPLC:

the supernatants of the first and second samples were diluted to the same extent by the addition of polishing water and the diluted supernatants were then filtered through a 0.22 μm filter. For each filtered and diluted supernatant, the same volume of supernatant was loaded in an HPLC system with Phenomenex and detected at 214nm5μm C4LC column 250x4.6mm, Ea.

The samples were run using the following conditions:

and (3) buffer solution A: MilliQ water, 0.1% w/w TFA

And (3) buffer solution B: HPLC grade acetonitrile, 0.085% w/w TFA

Flow rate: 1mL/min

Column temperature: 40 deg.C

Gradient: 82-55% A and 18-45% B in 0-30 min; 55-10% of A and 45-90% of B in 30-32 min; 32.5-37.5 minutes 10% A and 90% B; 10-82% A and 90-18% B for 38-48 min.

Data processing:

since both supernatants were treated in the same manner, the areas of the BLG peaks could be directly compared to calculate the relative yield. Since the crystals contain only BLG and all samples have been treated in the same way, the concentration of alpha-lactalbumin (ALA) and thus the ALA area should be the same in all samples. Thus, the ALA area before and after crystallization was used as a correction factor (cf) when calculating the relative yield.

The relative yield was calculated by the following equation:

acid dissolution of BLG crystals

The material remaining in the crystallizer was separated using a decanter at 350g, 2750RPM, 150RPM Diff with 64mil spacers and a feed flow of 75L/h, the feed being mixed with finishing water at a ratio of 1: 2 and mixing. The BLG crystals/solids from the decanter are then mixed with the polishing water to make it a thinner slurry, and then phosphoric acid is added to lower the pH to about 3.0 to rapidly dissolve the crystals.

After dissolution of the BLG crystals, the pure BLG protein liquid was concentrated to 15 brix at the same UF setting as used for the preparation of the crystallization feed and the pH was adjusted to a final pH of about 3.8. The liquid BLG isolate was then heated to 75 degrees for 5 minutes and then cooled to 10 ℃. It was found that after heat treatment the microbial load decreased from 137.000CFU/g before heat treatment to <1000CFU/g after heat treatment. Heat treatment did not cause any protein denaturation, and the intrinsic tryptophan fluorescence (330nm/350nm) was determined to be 1.20, indicating the native conformation of the BLG molecule.

The BLG was dried on a pilot plant spray dryer at an inlet temperature of 180 ℃ and an outlet temperature of 75 ℃. The water content in the resulting powder sampled at the outlet was about 4% w/w, the chemical composition of the powder being shown in Table 11. A sample of the dried powder was dissolved and the degree of protein denaturation was determined to be 1.5% and the intrinsic tryptophan fluorescence emission (I330/I350) was measured to be 1.20.

Table 11: composition of BLG isolate powder (BDL ═ below detection limit)

The bulk density (625 tap) of the spray-dried powder was estimated to be 0.2-0.3g/cm³。

And (4) conclusion: by using the above method, we can produce a high purity BLG product that can be heat treated without substantial protein denaturation or protein unfolding during processing. The heat treatment greatly reduced the level of bacteria without destroying the protein product.

The inventors have seen evidence that even higher bulk densities can be obtained by increasing the protein content prior to spray drying. In addition, the inventors have observed that an even lower degree of denaturation can be obtained if the inlet and/or outlet temperature for spray drying is reduced.

Example 9: production of spray-dried, pH neutral BLG isolate powder

The lactose reduced whey protein isolate shown in table 12 was adjusted and used as the crystallization feed when using the same protocol and experimental set-up as example 2. The crystallization yield was calculated to be 68%.

We note that all weight percentages of the specific proteins mentioned in this example (e.g.BLG and ALA) relate to the weight percentage of non-aggregated protein relative to the total amount of protein.

TABLE 12 composition of the feeds

The feed and finishing water were mixed in a 1: 2 mixing and separating the material remaining in the crystallizer on a decanter at 350g, 2750RPM, 150RPM Diff, with 64 intervals and a feed rate of 75L/h. The BLG crystals/solids from the decanter are then mixed with polishing water to make a thinner slurry, and then 0.1M potassium hydroxide is added to adjust the pH to about 7 to rapidly dissolve the crystals.

After dissolution of the crystals, the pure BLG protein liquid was concentrated to brix 15 on the same UF setup as used for preparing the whey protein solution for crystallization and the pH was adjusted to a final pH of 7.0. The BLG was dried on a pilot plant spray dryer at an inlet temperature of 180 ℃ and an outlet temperature of 75 ℃. The water content in the resulting powder sampled at the outlet was about 4% w/w. The composition of the powder is shown in table 13. After drying, some of the powder was dissolved in demineralized water, and the degree of protein denaturation was determined to be 9.0% and the intrinsic tryptophan fluorescence (330nm/350nm) was 1.16.

Table 13: chemical composition of BLG isolate powder

The bulk density (625 tap) of the spray-dried powder was estimated to be 0.2-0.3g/cm³。

And (4) conclusion: by using the above method we are able to produce high purity BLG products with neutral pH with little to no protein denaturation during processing. The inventors have seen evidence that even higher bulk densities can be obtained by increasing the protein content prior to spray drying. Furthermore, the inventors have observed that an even lower degree of denaturation is obtained if the inlet and/or outlet temperature for spray drying is lowered. The degree of denaturation can also be reduced by reducing the mineral content prior to spray drying.

Example 10: preparation of coated BLG isolate powder

Spray produced as described in example 4 or example 9 was usedThe fog-dried BLG isolate produced a coated BLG isolate in a fluidized bed (DIOSNA, MINIBLAB XP No. 365-. The inlet temperature was 60 ℃. The powder temperature is 40-50 deg.C, and the air flow is 25-35m³H is used as the reference value. The coating material was dissolved in 50g of demineralized water and then slowly injected into the fluidized bed, where atomization was carried out. For each batch, 500g of spray dried BLG isolate was used. After addition of the coating material, drying is continued until the moisture content of the coated BLG isolate is 4-5%. Using this setup, BLG isolates coated with 25g citric acid and BLG isolates coated with 30g trisodium citrate were prepared.

Test samples were prepared as shown in the table and analyzed for solubility as described below. In addition to the 10% w/w solution described, a 30% w/w solution of BLG isolate powder was prepared and tested. The wettability of the experimental samples was further analyzed as described below. In addition, the experimental samples were subjected to sensory evaluation by two trained test persons according to the parameters listed in example 1.11.

Using the BLG isolate powder of example 8, lecithin coated powders were prepared in the same fluidized bed as above. 500g of powder are added to the fluidized bed at an inlet temperature of 75 ℃ and an air flow of 25m³H is used as the reference value. When the powder temperature reached 38 ℃, 50mL of water was slowly added through the atomizer. The powder temperature was raised to 45 ℃ and 5mL of lecithin was injected through the atomizer. The powder was heated to 65 ℃ and dried until its water content contained less than 5%.

And (3) testing the solubility: the solubility and ease of solubility (readability) of instant powders can be measured by this test. In a sealable transparent tube, 10g of the powder was added to 90g demineralized water (8 ℃). The mixture was shaken vigorously by hand for 30 seconds. The mixture was immediately evaluated, then allowed to stand for 1 minute, and then the mixture was evaluated again. Evaluation was performed by visual inspection of the following parameters: transparency of the liquid phase, foam formation, color and to what extent the powder has dissolved.

Results

Sample #	Powder% w/w
		1	10% neutral uncoated BLG isolate, pH 7.02
2	10% lecithin coated WPI
		3	10% lecithin coated BLG isolate
4	10% citric acid coated BLG isolate
		5	10% BLG isolate coated with trisodium citrate
6	10% standard WPI, no coating
		7	30% BLG coated with trisodium citrate
8	30% neutral BLG isolate, no coating, pH 7.02
		9	30% standard WPI, no coating

And (4) conclusion: the obtained BLG isolate powder has good agglomeration structure and is easily soluble in water. In this test, all of the coated BLG isolate powders (samples 1, 3-5, and 7-8) performed equal to or better than the standard coated WPI (test sample 2). When testing the 30% version, coated and uncoated BLG isolates (test samples 1, 3-5 and 7-8) were surprisingly easily solubilized, thus it is believed that protein concentrations may even exceed 30%. Both the coated and uncoated BLG isolates (samples 7-8) in 30% solution were readily soluble compared to the standard WPI (sample 9), which had visible particles in the foam.

Wettability: this method is used to describe the wettability of the powder. Wettability is defined as the time taken before the entire sample is wetted. 0.5g of powder was measured out and placed on the surface of 100g of demineralized water (5 ℃) in a cylindrical container having a diameter of 5 cm. The time from placing the powder on the water surface until the powder dissolves or has passed the water surface is measured.

Results

And (4) conclusion: the coated BLG isolate powders (samples 10, 12, 14) were wetted in a similar manner to the standard fast dissolving WPI (samples 11, 15) except the BLG isolate powder coated with citric acid (sample 13). Surprisingly, the uncoated BLG isolate powder wets much better than the standard WPI (sample 6).

Example 11: wettability of uncoated BLG isolate

In this example, the wetability of uncoated acidic BLG isolate and uncoated neutral BLG isolate was compared to the wetability of uncoated Whey Protein Isolate (WPI). Wettability is defined as the time taken until the entire sample is wetted. 0.5g of powder was measured out and placed on the surface of 100g demineralized water (10 ℃) in a cylindrical container having a diameter of 5 cm. The time from placing the powder on the water surface until the powder dissolves or has passed the water surface is measured.

Results

And (4) conclusion: surprisingly, the uncoated BLG isolate powders (samples 2, 3) had much better wetting than the standard WPI (sample 1).

Example 12: preparation of instant beverage powder

Preparation of instant powder for use as nutritional supplement.

100g of instant powder was prepared by blending the following ingredients: 91 grams of whey protein concentrate comprising at least 85% w/w BLG prepared as described in example 4, and 4 grams of soy lecithin.

100g of instant powder contains 360Kcal, with an energy distribution as follows: 2E% from lipids, 1E% from carbohydrates, 97E% from proteins. For example, food products prepared from instant powders can be used as protein supplements (for treating patients suffering from or at risk of malnutrition) by mixing the instant powder with water to obtain a beverage or by adding the powder to a regular diet. 10-15 g of instant powder are stirred into water at a temperature of 15-25 ℃. The instant powder beverage obtained has pleasant taste, color and viscosity.

Example 13: preparation of instant beverage powder

Preparation of a nutritionally complete instant powder comprising BLG.

100g of instant powder was prepared by blending the following ingredients: 18.2 grams of vegetable oil (a mixture consisting of palm kernel oil, coconut oil, rapeseed oil and sunflower oil), 56.4 grams of glucose syrup and 20 grams of whey protein concentrate, which contains at least 90% w/w BLG prepared as described in example 4 and about 3 grams of soy lecithin.

Further adding: potassium citrate, sodium chloride, magnesium hydrogen phosphate, potassium hydrogen phosphate, magnesium chloride, choline chloride, calcium carbonate, calcium phosphate, sodium L-ascorbate, a fragrance, ferric sulfate, L-ascorbic acid, zinc sulfate, magnesium citrate, DL-alpha-tocopheryl acetate, manganese sulfate, nicotinamide, D-biotin, copper sulfate, D-calcium pantothenate, pteroylmonoglutamic acid, sodium fluoride, DL-alpha-tocopherol, thiamine hydrochloride, pyridoxine hydrochloride, carotenoid, retinyl palmitate (retinyl palmitate), riboflavin, cyanocobalamin, cholecalciferol (sterol), chromium chloride, sodium molybdate, potassium iodide, sodium selenite, and vitamin K₁To obtain an instant powder having the following nutritional ingredients: 381 μ g RE vitamin A, 0.91mg carotenoid, 3.3 μ g vitamin D, 5.9mg α -TE vitamin E, 24 μ g vitamin K, 0.69mg vitamin B1, 0.74mg vitamin B2, 8.3mg NE nicotinic acid, 2.5mg pantothenic acid, 0.79 μ g vitamin B6, 123 μ g folic acid, 0.98 μ g vitamin B12, 24 μ g biotin, 60mg vitamin C, 156mg choline, 1.17 grams salt, 469mg sodium, 705mg potassium, 578mg chloride (chloride), 371mg calcium, 337mg phosphorus, 107mg magnesium, 7.4mg iron, 5.6mg zinc, 0.83mg copper, 1.5mg manganese, 0.5g (fluoride), 48 μ g molybdenum, 26 μ g selenium, 25 μ g, 61 μ g fluoride (diioride).

The instant powder contained 462Kcal/100 g and had an energy distribution as follows: 35.6E% from lipid, 48.7E% from carbohydrate, 15.7E% from protein. For example, food products prepared from the instant powder can be used as nutritionally complete food products (which are used to treat patients suffering from or at risk of malnutrition) by mixing the instant powder with water to obtain a beverage or to be used for tube feeding. .22 g of instant powder was stirred into 85ml of cold water (boiled) to obtain a beverage containing 100 kcal. 33 g of instant powder was stirred into 78ml of cold water (boiled) to obtain a beverage containing 150 kcal. The instant powder beverage prepared has a pleasant taste, color and viscosity.