CN107136295B

CN107136295B - Partially hydrolyzed whey protein powder and preparation method thereof

Info

Publication number: CN107136295B
Application number: CN201610118295.1A
Authority: CN
Inventors: 韩仁娇; 王彩云; 云战友; 罗述博
Original assignee: Inner Mongolia Yili Industrial Group Co Ltd; Inner Mongolia Dairy Technology Research Institute Co Ltd
Current assignee: Inner Mongolia Yili Industrial Group Co Ltd; Inner Mongolia Dairy Technology Research Institute Co Ltd
Priority date: 2016-03-01
Filing date: 2016-03-01
Publication date: 2021-07-27
Anticipated expiration: 2036-03-01
Also published as: CN107136295A

Abstract

The invention discloses partially hydrolyzed whey protein powder and a preparation method thereof. The protein powder has hydrolysis degree of 8-10%, contains at least 65% of components with molecular weight below 2000Da, contains free amino acid less than 5%, and has hydrolysis rate of beta-lactoglobulin up to 60%, lower bitter taste, and better dissolving property and appearance compared with similar products.

Description

Partially hydrolyzed whey protein powder and preparation method thereof

Technical Field

The invention relates to the technical field of food raw material processing. More particularly, relates to partially hydrolyzed whey protein powder and a preparation method thereof.

Background

Cow's milk is a particularly important substitute for breast milk and one of the earliest allergens to which newborns come into contact, and therefore cow's milk allergy is most common among infants. Milk Protein Allergic (CMPA) infants often develop symptoms such as vomiting, chronic diarrhea, malabsorption, and growth retardation. It has been shown by investigation that 2-3% of infants are allergic to cow's milk, of which about 85% develop immune tolerance with age to the young, while the other 15% are lifelong. More than 20 proteins are contained in cow milk, and the cow milk proteins have potential sensitization, but Beta-lactoglobulin (Beta-Lg) is generally considered as a main allergen at present. Therefore, reducing the content of beta-lactoglobulin in the infant formula powder is a key measure for improving and promoting the nutrition and growth and development of children suffering from CMPA.

Nowadays, in order to meet the nutritional requirements of infants fed with formula milk powder, the infant formula powder emulsified by mothers has become a trend, raw materials with low content of beta-lactoglobulin should be selected as much as possible when the infant formula powder is produced, and methods for reducing the content of beta-lactoglobulin include physical methods such as heat treatment and glycosylation, biological enzymolysis technology and the like. The Chinese patent application CN 102940126A discloses a method for reducing beta lactoglobulin in concentrated whey protein powder by biological compound protein enzymolysis, which comprises the steps of carrying out enzymolysis on whey protein concentrate by adopting compound protease consisting of trypsin, papain and neutral protease, then carrying out evaporation concentration and spray drying on hydrolysate to obtain the whey protein hydrolysate. The method uses trypsin to cause severe hydrolysis reaction, so that the flavor of the product is affected. In addition, the method adopts an evaporation concentration mode to improve the solid content of the feed liquid, the temperature is generally controlled at 60-70 ℃, the concentration time is about 1-2 hours, and the concentration time is even longer due to the influence of the performance of an evaporator. The above process is only suitable for preparing hydrolyzed whey protein with low protein content, for example, in the process of preparing hydrolyzed protein by using desalted whey powder and WPC34 as raw materials, but for the raw materials with protein content of more than 80%, the process cannot meet the actual requirements, the main reason is that the evaporation concentration adopts the process of heating vaporization and removing steam to achieve the purpose of improving the solute concentration in the solution, and after the raw materials with high protein content are subjected to the enzymolysis process, the viscosity of the hydrolysate can be greatly increased in the evaporation concentration process, the viscosity increase range is much larger than that of the feed liquid with the same concentration before enzymolysis, the mobility of the feed liquid with high viscosity in the evaporator is obviously reduced, so that the feed liquid is heated and denatured in the evaporator to form a film, the vaporization of the solution is hindered, and the evaporation efficiency is reduced, and the direct consequences are two types: firstly, the final solid content of the feed liquid is reduced in order to shorten the concentration time, and the reduction of the concentrated solid content of the feed liquid can directly influence the spray drying effect and finally influence the sensory state of the product; secondly, a large amount of denatured protein feed liquid is discarded in order to reach the predetermined solid content of the feed liquid, which directly results in the reduction of the yield of the final product. In addition, the organoleptic and functional properties of the products prepared by evaporative concentration are greatly affected and the product index is difficult to control by a large number of facts.

In addition, the chinese patent application CN 101297674 a discloses a low-sensitivity whey protein hydrolysate and a preparation method thereof, the method comprises the steps of firstly, denaturing protein at high temperature, then, carrying out enzymolysis on concentrated whey protein by using compound protease consisting of alkaline protease, papain and flavourzyme, and then, desalting and drying hydrolysate to obtain the whey protein hydrolysate. The alkaline protease adopted in the method can cause severe hydrolysis reaction, on one hand, a great amount of salt is introduced into the final product, so that a desalting treatment link needs to be added in the process, the production period is prolonged, and the energy consumption is increased; on the other hand, the flavor of the product is influenced by the violent hydrolysis reaction. In addition, the application only mentions the inhibition rate of beta-lactoglobulin and alpha-lactalbumin, does not describe the key physicochemical and sensory indexes of the hydrolyzed whey protein powder obtained by the method, and cannot comprehensively evaluate whether the method is suitable for preparing the whey protein hydrolysate.

Aiming at the defects of the method, the partially hydrolyzed whey protein which has lower sensitization component, reasonable proportion of small peptide and free amino acid and good flavor, dissolvability and appearance character is developed, and has important social value.

Disclosure of Invention

An object of the present invention is to provide a partially hydrolyzed whey protein powder having an effectively reduced content of beta-lactoglobulin, while having a bitterness and a free amino acid content significantly lower than those of similar products on the market.

The invention also aims to provide a preparation method of the partially hydrolyzed whey protein powder.

The invention also claims the application of the partially hydrolyzed whey protein powder in preparing food, in particular infant milk powder.

In order to achieve the purpose, the invention adopts the following technical scheme:

the invention provides a partially hydrolyzed whey protein powder, which contains at least 65% of components with molecular weight below 2000Da, the proportion of free amino acid components is lower than 5%, the hydrolysis rate of beta-lactoglobulin reaches more than 60%, the hydrolysis degree is 8% -10% by using an OPA method, compared with the similar products sold in the market, the hydrolysate has lower bitter taste and better reconstitution property and appearance state, and the partially hydrolyzed whey protein powder is prepared by the following method:

(1) preparing concentrated whey protein into whey protein aqueous solution, performing heat treatment to denature the protein, and then cooling the aqueous solution to a temperature suitable for enzymatic hydrolysis;

(2) adding compound protease into the cooled whey protein aqueous solution, hydrolyzing for 2-4 hours at 50-55 ℃, then adjusting the pH value of the solution to 6.7-6.9, and thermally inactivating the protease to stop the hydrolysis reaction, wherein the compound protease is formed by mixing neutral protease F and neutral protease G or neutral protease F and neutral protease P according to the mass ratio of 3-8:1, and the mass ratio of the compound protease to the whey protein in the aqueous solution is 0.4-0.9: 100;

the method adopts compound neutral protease to hydrolyze the concentrated whey protein for the first time, and reduces the content of beta-lactoglobulin in the concentrated whey protein to the maximum extent under the condition of moderate hydrolysis. In the conventional invention patents, in order to obtain a product with a low content of β -lactoglobulin, the hydrolysis rate of β -lactoglobulin is increased by increasing the amount of protease to be added or by extending the hydrolysis time, which results in increased bitterness of the final product, longer process time, and increased energy consumption.

The partial hydrolysis in the invention means that the substrate hydrolysis degree is controlled to be 8% -10% (measured by an OPA method), and a large number of studies prove that certain proteases in the intestinal tract of infants, such as pepsin in the stomach, trypsin, chymotrypsin and the like in the small intestine, have certain protein digestion capacity although the level is lower than that of adults, so that deep or complete hydrolysis of whey protein powder is not necessary for ordinary infants. The partially hydrolyzed whey protein powder is prepared by hydrolyzing part of macromolecular protein into small molecular weight peptide segments and free amino acids by a biological enzymolysis technology on the basis of whey protein powder, so that the burden of eating complete protein on the gastrointestinal tract of infants is avoided, and the digestibility of the protein is improved due to the increase of the small molecular weight peptide segments. In addition, the partially hydrolyzed whey protein powder has less severe hydrolysis degree, and the taste and flavor of the product are greatly improved.

(3) Nano-filtering and concentrating the solution after the hydrolysis reaction is stopped, wherein the feed flow is 1-3t/h, the feed pressure is 6-12bar, the membrane flux is 50-400L/h, and the solid content after concentration is 20-35%;

the method solves the concentration problem of high protein hydrolysate, greatly reduces the heating degree of the hydrolyzed protein in the processing process, and avoids the influence of long-time high temperature on the functional components and sensory properties of the hydrolyzed protein. In addition, as the nanofiltration concentration technology is adopted, the content of free amino acid in the hydrolysate is controlled to a certain degree, and researches show that: excessive free amino acids in the intestine can cause osmotic pressure to lose balance, affect moisture change and cause discomfort to the intestine.

It is particularly emphasized that the means for limiting the release of free amino acids in the present invention is not limited to the use of nanofiltration concentration technology, but the combined action and effect of the biological enzymolysis technology of step (2) and the nanofiltration technology of step (3).

(4) And drying the concentrated feed liquid to obtain the finished product.

Further, in a preferred embodiment of the present invention, in the step (1), the concentrated whey protein is formulated into a whey protein aqueous solution having a mass concentration of 5% to 10%. The heat treatment method for denaturing protein comprises keeping whey protein water solution at 55-85 deg.C for 5-10 min.

Further, in a preferred embodiment of the present invention, in the step (2), the temperature for heat-inactivating the protease is 80 ℃ to 95 ℃ for 5 to 10 min.

Further, in a preferred embodiment of the present invention, in the step (3), the nanofiltration concentration is performed at room temperature.

Further, in a preferred embodiment of the present invention, the drying method in step (4) is preferably spray drying. Specifically, the temperature of feed liquid during feeding is 20-25 ℃, the hot air inlet temperature is 135-175 ℃, the air outlet temperature is 70-85 ℃, and the atomization pressure is 3-5 bar.

In addition, in a preferred embodiment of the invention, the enzyme activity of the neutral protease F is 390000-430000HU/G, the enzyme activity of the neutral protease G is 276000PC/G, and the enzyme activity of the neutral protease P is 95000 PC/G.

The invention has the following beneficial effects:

firstly, composite neutral protease is adopted for enzymolysis, and endopeptidase and exopeptidase are combined, so that the hydrolysis condition is milder, more than 60% of beta-lactoglobulin in whey protein can be hydrolyzed under the moderate hydrolysis condition, and the probability of anaphylactic reaction of the infant due to eating of the hydrolyzed protein is reduced; simultaneously, the bitterness of hydrolysate is reduced; concentration treatment is carried out by adopting a nanofiltration technology, so that the concentration of univalent sodium ions and the content of free amino acid in the hydrolysate are reduced; thirdly, the damage of the functional components of the hydrolysate caused by the evaporation concentration treatment is avoided, and the protein denaturation caused by the evaporation concentration is avoided. The partially hydrolyzed whey protein obtained by the process has low bitterness; the components of the partially hydrolyzed whey protein prepared by the process, the molecular weight of which is distributed below 2000Da, at least reach more than 65 percent, and are more beneficial to the digestion and absorption of infants.

Drawings

FIG. 1 electropherograms of standard protein and hydrolyzed samples.

FIG. 2 high performance liquid chromatogram of partially hydrolyzed whey protein powder prepared in example 1.

Detailed Description

In order to more clearly illustrate the invention, the invention is further described below in connection with preferred embodiments. It is to be understood by persons skilled in the art that the following detailed description is illustrative and not restrictive, and is not to be taken as limiting the scope of the invention.

Example 1

The partially hydrolyzed whey protein powder is obtained by the following preparation method:

preparing concentrated whey protein into a whey protein water solution with the mass concentration of 5%, preserving the heat of the solution at 60 ℃ for 10min to denature the protein, and then cooling to 55 ℃; adding compound protease (neutral protease F (validase FP concentrate)) and neutral protease G (C)

Protease GBW) in a ratio of 5:1) to a substrate whey protein in a mass ratio of 0.4:100, hydrolyzing at 55 ℃ for 4 hours while stirring, adjusting the pH value of the solution to 6.75-6.85 by using 1mol/L NaOH solution, heating the solution at 80 ℃ for 10min to inactivate the Protease, and then rapidly cooling to room temperature; nano-filtering and concentrating at room temperature, wherein the feed flow is 1-3t/h, the feed pressure is 6-12bar, the membrane flux is 50-400L/h, and the solid content after concentration is 25-35%; finally, spray drying is carried out on the concentrated feed liquid, the temperature of the feed liquid during feeding is 20-25 ℃, the hot air inlet temperature is 135-185 ℃, and the air outlet temperature is 70-85 ℃.

The detection method comprises the following steps: the molecular weight distribution of the experimental product is determined by high performance liquid chromatography (GB/T22729-:

TABLE 1

	Name (R)	Retention time	Mn	Mw	MP	Area of	% area
								1	Peak 11	11.539	14047	14622	12962	2910411	13.65
2	Peak 6	12.053	6948	7259	9983	1270558	5.96
								3	Peak 7	14.400	3726	3803	3035	1274563	5.98
4	Peak 8	15.206	2417	2450	2016	1364175	6.40
								5	Peak 9	16.568	1356	1414	1010	2964092	13.91
6	Peak 2	17.807	696	726	539	4023327	18.88
								7	Broad distribution	19.178	272	290	269	7007180	32.88
8	Peak 10	19.980	147	151	179	500085	2.35

TABLE 2

The partially hydrolyzed whey protein powder prepared by the method contains at least 65% of components with molecular weight below 2000Da, and the proportion of free amino acid components is lower than 5%.

The degree of hydrolysis of the partially hydrolyzed whey protein powder prepared by the method is 8-10% (measured by an OPA method).

Example 2

the difference from the embodiment 1 is that: preparing the concentrated whey protein into a whey protein water solution with the mass concentration of 10%, carrying out heat preservation treatment on the solution at 85 ℃ for 5min to denature the protein, and then cooling to 50 ℃; the mass ratio of the compound protease (the ratio of the neutral protease F to the neutral protease G is 5:1) to the substrate whey protein is 0.6:100, and the hydrolysis is carried out for 2 hours at the temperature of 50 ℃; heating at 85 deg.C for 8min to inactivate protease.

In addition, in this example, the hydrolysis rate of beta-lactoglobulin of the partially hydrolyzed whey protein powder was examined. The detection method comprises the following steps: the hydrolysis condition of beta-lactoglobulin in the hydrolysate is detected by SDS-dextran gel electrophoresis, and the specific detection result is shown in the bands of (fifthly) and (sixthly) in figure 1. The hydrolysis rate of beta-lactoglobulin was 73%.

Example 3

the difference from the embodiment 1 is that: preparing the concentrated whey protein into a whey protein water solution with the mass concentration of 8%, carrying out heat preservation treatment on the solution at 80 ℃ for 8min to denature the protein, and then cooling to 55 ℃; complex protease (neutral protease F (Validase FP concentrate) and neutral protease P (

PPU 95000) in a ratio of 6:1) to the substrate whey protein in a mass ratio of 0.7:100, and hydrolyzed at 55 ℃ for 3 hours; heating at 90 deg.C for 5min to inactivate protease.

In this example, the bitter taste and sensory properties of the partially hydrolyzed whey protein powder were examined. The detection method comprises the following steps: and (3) evaluating bitterness, namely, preparing caffeine into 0, 0.025, 0.05, 0.1, 0.2 and 0.3 concentration respectively, and respectively obtaining a rating value and a bitterness degree of 1-no bitterness, 2-slight bitterness, 3-weak bitterness, 4-common bitterness, 5-heavy bitterness and 6-extremely heavy bitterness respectively, wherein according to the rating standard, a person tasting the protein hydrolysate tastes the bitterness and compares the bitterness with the standard to score, and finally obtaining an average value to express the bitterness degree.

The partially hydrolyzed whey protein powder prepared by the method has lower bitter taste and better sensory property.

Comparative test example 1

Detecting hydrolysis condition of beta-lactoglobulin in hydrolysate by SDS-dextran gel electrophoresis method (see figure 1), wherein molecular weight of the beta-lactoglobulin is about 18KDa, and the hydrolysis rate of the beta-lactoglobulin can be roughly estimated by detecting the proportion of 18KDa components in whey protein before and after hydrolysis, namely electrophoresis bands of samples before and after hydrolysis by using compound protease consisting of alkaline protease, papain and flavourzyme, namely electrophoresis bands of samples before and after hydrolysis by using compound protease consisting of trypsin, papain and neutral protease, and fifth, namely electrophoresis bands of samples before and after hydrolysis by using compound neutral protease, wherein the electrophoresis bands of the samples before hydrolysis, the electrophoresis bands of the samples after hydrolysis, and the hydrolysis rate of the beta-lactoglobulin is 65 percent by calculation, ③ 60 percent of beta-lactoglobulin, and fifthly 73 percent of beta-lactoglobulin, and the results show that the hydrolysis rate of the beta-lactoglobulin is the highest when the compound neutral protease is adopted for hydrolysis.

Comparative test example 2

The difference between the key physicochemical indexes of the hydrolyzed protein powder prepared in example 2 of the present invention and the conventional commercial hydrolyzed protein powder is specifically shown in table 3 below:

TABLE 3

Comparative test example 3

The digestion and absorption of the hydrolysate are evaluated by adopting an animal digestion experiment, and the specific test method is as follows:

experimental animals: the Kunming white mouse has the weight of 18-22g and is half male and half female.

Experiment design: 72 Kunming mice with an average body weight (20. + -. 0.05) g were selected and divided into 3 groups of 24 mice each, of which the males and females are each half. The 3 groups of mice were: WPC80 powder was fed to group I, hydrolyzed whey protein powder (commercially available) was fed to group II, and the hydrolyzed whey protein powder of example 1 was fed to group III.

Collecting samples: the experimental mice were fasted for 12h without water deprivation, during which time water was freely available. Then quantitatively pouring 40% of milk powder, each 0.5mL, slaughtering 6 of each group at time points of 0.5, 1, 2 and 4h, rapidly dissecting abdominal cavity, taking out stomach and duodenum, and storing in ice box.

Measurement indexes are as follows: (1) determination of TCA soluble amino nitrogen with mass fraction of 5%

The digestive tract was flushed with physiological saline, and intestinal fluid was collected and vacuum freeze-dried (-50 ℃, 48 h). 0.02g of the lyophilized sample was weighed, dissolved in distilled water, and centrifuged at high speed at low temperature (12000r/min, 10 min). Taking supernatant, and mixing the supernatant and the supernatant according to a volume ratio of 1: 1 adding TCA (concentration of 0.6mol/L), centrifuging at low temperature again (5000r/min, 15min) to remove impurity proteins, collecting secondary supernatant, and measuring light absorption value at 280 nm.

Measurement indexes are as follows: (2) determination of free amino acids in mouse digestive juices

Adding 1.0mL of 5-sulfosalicylic acid of 0.2g/mL into 1.0mL of intestinal fluid of a mouse, shaking up, adding 8.0mL of distilled water, centrifuging for 15min at 4000r/min, taking supernatant, filtering by a 0.22m microporous filter membrane, and using filtrate for measuring the content of amino acid. And directly loading the processed sample to an automatic amino acid analyzer for determination.

The results are shown in tables 4 and 5:

TABLE 4 mouse intestinal juice absorbance value test results

TABLE 5 Total amino acid content in intestinal tract of mice digested for 1 hour

Sample (I)	Total amount of amino acids g/100g
		WPC80 powder	0.23
Commercial partially hydrolyzed whey protein powder	0.36
		Example 1 hydrolyzed whey protein powder of the invention	0.38

It should be understood that the above-mentioned embodiments of the present invention are only examples for clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention, and it will be obvious to those skilled in the art that other variations or modifications may be made on the basis of the above description, and all embodiments may not be exhaustive, and all obvious variations or modifications may be included within the scope of the present invention.

Claims

1. The partially hydrolyzed whey protein powder is characterized in that the degree of hydrolysis of the protein powder is 8% -10%, the protein powder contains at least 65% of components with molecular weight below 2000Da, the proportion of free amino acid components is lower than 5%, the hydrolysis rate of beta-lactoglobulin in the protein powder reaches more than 60%, and the partially hydrolyzed whey protein powder is prepared by the following method:

(1) preparing concentrated whey protein into a whey protein aqueous solution with the mass concentration of 5% -10%, performing heat treatment to denature the protein, and then cooling the aqueous solution to a temperature suitable for protease hydrolysis;

(2) adding compound protease into the cooled whey protein aqueous solution, hydrolyzing for 2-4 hours at 50-55 ℃, then adjusting the pH value of the solution to 6.7-6.9, and thermally inactivating the protease to stop the hydrolysis reaction, wherein the compound protease is formed by mixing neutral protease F and neutral protease G or neutral protease F and neutral protease P according to the mass ratio of 3-8:1, and the mass ratio of the compound protease to the whey protein in the aqueous solution is 0.4-0.9: 100; the neutral Protease F is Validase FP Concentrate, the neutral Protease G is Bakezyme Protease GBW, and the neutral Protease P is Bakezyme PPU 95000;

(4) and drying the concentrated feed liquid to obtain the finished product.

2. The partially hydrolyzed whey protein powder of claim 1, wherein in step (1), the heat treatment is performed to denature the whey protein by incubating the aqueous whey protein solution at 55-85 ℃ for 5-10 min.

3. The partially hydrolyzed whey protein powder of claim 1, wherein in step (2), the temperature of the heat-inactivated protease is 80-95 ℃ for 5-10 min.

4. The partially hydrolyzed whey protein powder of claim 1, wherein in step (3), the nanofiltration concentration is performed at room temperature.

5. The partially hydrolyzed whey protein powder of claim 1, wherein the drying method in step (4) is spray drying.

6. The partially hydrolyzed whey protein powder of claim 5, wherein the spray drying conditions are: the temperature of feed liquid is 20-25 ℃, the temperature of hot air inlet is 135-175 ℃, the temperature of air outlet is 70-85 ℃, and the atomization pressure is 3-5 bar.

7. A method of preparing partially hydrolyzed whey protein powder of any of claims 1-6, comprising:

(4) and drying the concentrated feed liquid to obtain the finished product.

8. Use of the partially hydrolyzed whey protein powder of any of claims 1-6 in the preparation of a food product.

9. Use according to claim 8, wherein the food product is an infant formula.