WO2021003741A1

WO2021003741A1 - APPLICATION OF β-CASEIN A2 AND COMPOSITION THEREOF IN PROMOTING PROLIFERATION OF BIFIDOBACTERIUM

Info

Publication number: WO2021003741A1
Application number: PCT/CN2019/095620
Authority: WO
Inventors: 陈历俊; 刘斌; 赵军英; 姜铁民; 周伟明; 李建涛; 刘彦品; 乔为仓
Original assignee: 北京三元食品股份有限公司
Priority date: 2019-07-11
Filing date: 2019-07-11
Publication date: 2021-01-14
Also published as: ZA202201337B

Abstract

The application of β-casein A2 and composition thereof in promoting the proliferation of bifidobacterium. Compared with ordinary milk, milk containing β-casein A2 may significantly reduce the proportion of abdominal distension and borborygmus and increase the frequency of defecation, and may change stool characteristics. In addition, the milk containing β-casein A2 may significantly increase the relative abundance of bifidobacterium in intestinal bacteria.

Description

Application of β-casein A2 and its composition in promoting the proliferation of Bifidobacterium

Technical field

The present invention relates to the field of dairy products, in particular to the application of β-casein A2 and its composition in promoting the proliferation of Bifidobacterium.

Background technique

Milk is not only rich in nutrients, but also easy to digest and absorb and convenient to eat. It is the most ideal natural food. About 80% of the protein contained in milk is casein and 20% is whey protein. In the casein of milk, 30-35% is β-casein, which has many variants. The two most common variants are A1 and A2. The difference between β-casein A1 and β-casein A2 is that one of the 209 amino acids that make up this protein is different. The casein with proline at position 67 of the amino acid chain is β-casein A2, and the casein with histidine at position 67 is β-casein A1.

Beta-Casomorphin-7 (Beta-Casomorphin-7), also known as BCM-7, is a 7-amino acid polypeptide chain. In the digestion process, digestive enzymes cut A1 to produce BCM-7; while A2 cannot be cut by these digestive enzymes to produce BCM-7. Studies have shown that BCM-7 may interfere with the normal metabolism of the human body and increase the risk of some infants suffering from type I diabetes; in addition, it may also cause functional disorders of immune response, digestion and breathing.

It has been reported that the intestinal flora is closely related to human health. The intestinal flora includes microorganisms that are beneficial to the health of the host, such as Bifidobacterium, Lactobacillus, etc.; it also contains some harmful microorganisms, such as Escherichia coli and Proteus. Beneficial bacteria and harmful bacteria coexist together to maintain the balance of intestinal flora and maintain the health of the host. Once the intestinal flora is imbalanced, it will cause diarrhea and other clinical diseases.

The information disclosed in the background section is only intended to increase the understanding of the general background of the present invention, and should not be regarded as an acknowledgement or any form of suggestion that the information constitutes the prior art already known to those of ordinary skill in the art.

Summary of the invention

Purpose of invention

In order to solve the above technical problems, the object of the present invention is to provide the application of β-casein A2 and its composition in promoting the proliferation of Bifidobacterium. Compared with ordinary milk, milk containing β-casein A2 can significantly reduce the proportion of bloating and bowel movement, increase the frequency of bowel movements, and change stool characteristics; and milk containing β-casein A2 can significantly increase the double Relative abundance of Fidobacteria.

solution

To achieve the purpose of the present invention, the embodiment of the present invention provides the application of β-casein A2 in promoting the proliferation of Bifidobacterium.

The present invention also provides an application of a composition for promoting the proliferation of Bifidobacterium, the composition comprising β-casein; in the β-casein, the mass percentage of β-casein A2 is ≥75%.

In a possible implementation manner of the above application, in the β-casein, the mass percentage of β-casein A2 is ≥90%.

In a possible implementation manner of the above application, in the β-casein, the mass percentage of β-casein A2 is 100%.

In a possible implementation of the above application, the composition is milk or a dairy product.

In a possible implementation of the above application, the milk includes fresh milk, milk powder, liquid milk reconstituted from powder, skimmed milk, homogenized milk, condensed milk, low-strength concentrated milk, pasteurized milk or non-pasteurized milk. At least one of the sterilized milk.

In a possible implementation of the above application, the milk is milk, and every 100g of the milk includes the following nutrients: protein 3.2g, fat 3.8g, carbohydrate 4.5g, sodium 70mg; wherein, the In the protein, the content of β-casein A2 is 0.9g, and the mass percentage of β-casein A2 in β-casein is 100%.

In a possible implementation of the above application, the milk is prepared by the following method: raw milk is collected, filtered, standardized, homogenized, sterilized, cooled, and canned to obtain milk.

In a possible implementation manner of the above application, the raw milk comes from a herd of A2 variant cows.

In a possible implementation manner of the above application, the milk is consumed within 1 hour after breakfast and/or dinner.

In a possible implementation manner of the above application, the dairy product includes at least one of cream, yogurt, quark food, cheese, butter, or ice cream.

Beneficial effect

The embodiment of the present invention carried out a cohort study of healthy adults. By comparing the dietary supplement intake, gastrointestinal comfort and fecal characteristics of A2 milk containing only β-casein A2 and ordinary milk, the intestinal flora was evaluated. 16S rRNA high-throughput gene sequencing to analyze changes in the composition of intestinal bacteria and probiotics. The results prove that milk containing β-casein A2 can significantly reduce the proportion of abdominal distension and bowel movement, increase the frequency of bowel movements, and change stool characteristics compared with ordinary milk; and milk containing β-casein A2 can significantly increase intestinal bacteria The relative abundance of the genus Bifidobacterium.

Description of the drawings

One or more embodiments are exemplified by pictures in the corresponding drawings, and these exemplified descriptions do not constitute a limitation on the embodiments. The dedicated word "exemplary" here means "serving as an example, embodiment, or illustration." Any embodiment described herein as "exemplary" need not be construed as being superior or better than other embodiments.

Figure 1 shows the changes of Bifidobacterium genus with the intervention stage under various age conditions in the A2 milk group and the ordinary milk group of the experimental example of the present invention.

Detailed ways

In order to make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of them.的实施例。 Example. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative work shall fall within the protection scope of the present invention. Unless otherwise expressly stated otherwise, throughout the specification and claims, the term "comprising" or its transformations such as "comprising" or "including" will be understood to include the stated elements or components, and not Other elements or other components are not excluded.

In addition, in order to better illustrate the present invention, numerous specific details are given in the following specific embodiments. Those skilled in the art should understand that the present invention can also be implemented without certain specific details. In some embodiments, raw materials, elements, methods, and means that are well known to those skilled in the art are not described in detail, so as to highlight the gist of the present invention.

Example 1

A type of A2 milk obtained by empirically harvesting, filtering, standardizing, homogenizing, sterilizing, cooling, and canning raw milk; the raw milk comes from a herd of A2 variant cows; the composition of the A2 milk is shown in Table 1:

Table 1

Test example

The milk used in the experiment was A2 milk and ordinary milk prepared in Example 1;

In ordinary milk, the mass ratio of β-casein A2 and β-casein A1 is 6:4.

To study the effect of A2 milk group and normal milk group on human intestinal health, the experimental method is as follows.

1. Research object:

There are 60 volunteers who are 20-65 years old and have no serious diseases, including 30 young people (20-44 years old), 20 middle-aged people (45-59 years old), and 10 elderly people (60-65).

Eligibility criteria: (1) 20-65 years old (including 20 and 60 years old); (2) Both men and women (except breastfeeding and pregnancy); (3) No history of major diseases, depression and pathological gastrointestinal diseases (HIV, cancer, low immunity, gastritis, colitis, etc.); (4) Irregular or occasional gastrointestinal symptoms (flatulence, abnormal bowel sounds, heaviness after eating, abdominal pain), recent gastrointestinal function It is especially good for those who are unwell; (5) slow or irregular intestinal motility (abnormal stool hardness, reduced bowel movements, such as 1 bowel movement for 2-3 days, or less than 3 bowel movements per week); (6) Volunteers can Understand the purpose and requirements of this trial, including possible risks and adverse reactions.

Exclusion conditions: (1) Patients with diarrhea and influenza; (2) Those who have used probiotics, prebiotics, antibiotics, and gastrointestinal digestive substances in the past 15 days; (3) Long-term use of drugs that inhibit or prevent influenza-like symptoms , Such as antihistamines, cough suppressants, high-dose vitamin C, etc.; (4) have received influenza vaccine in the past 6 months, or have received other vaccines within 15 days; (5) have alcohol or drug addicts; (6) Those who have participated in other clinical trials in the past 3 months.

Conditions for withdrawal: (1) Volunteers who the researcher thinks cannot fully cooperate with the test arrangement; (2) Those who use antibiotics, other probiotic products and dairy products during the observation period; (3) Those who have pathological diseases, diarrhea, and flu during the observation period .

2. Experimental design

This experiment was reviewed by the medical ethics committee of Luoyang North Enterprise Group Co., Ltd. Staff Hospital (No. 2018032).

(1) Intervention methods of A2 milk group and ordinary milk group

A2 milk group: drink one bottle of A2 milk (250mL) within 1 hour after breakfast and dinner for 15 days, stop taking observation for 15 days, continue to drink a bottle of ordinary milk (250mL) within 1 hour after breakfast and dinner, continue for 15 days, stop Observation for 7 days, 44 days in total;

Ordinary milk group: drink a bottle of ordinary milk (250mL) within 1 hour after breakfast and dinner for 15 days, stop taking observation for 15 days, continue to drink a bottle of A2 milk (250mL) within 1 hour after breakfast and dinner, continue for 15 days, stop Observation for 7 days, 44 days in total.

(2) Experimental method

Sample collection: The first stool sample of the day was collected after one week of enrollment, two weeks of intervention, two weeks of washout, and two weeks of second intervention.

Health status monitoring: The gastrointestinal health status (abdominal distension, belching, abdominal pain, defecation satisfaction, etc.) during the test period was filled out by the subjects three days before sampling.

High-throughput sequencing method: Use Qiagen Stool Kits (Qiagen, Valencia, CA) to extract total bacterial genome DNA from 100 mg of feces;

Further Quantitative NanoDrop, PCR amplification of the V3-V4 gene fragments on the bacterial genome (PCR primer sequence 341F: CCTAC GGGGN GGCWGC AG, 805R: GACTA CHVGG GTATC TAATC C) to construct a sequencing library;

Then perform Illumina HiSeq sequencing;

The diversity and richness of the flora is quantified as the number of operable taxa (OTUs), and the RDP 16s database is used to annotate flora species.

3. Experimental results:

(1) Enrollment baseline conditions: see Table 2; from Table 2, it can be seen that there is no significant statistical difference in baseline information such as gender, age, height, and weight of volunteers in the A2 milk group and the normal milk group, indicating that the experimental design is better Various variable factors are balanced to avoid interference from differences in baseline levels.

Table 2

x±s means mean±standard deviation; difference test method: Pearson's chi-square test (χ2); Wilcoxon rank test.

(2) Gastrointestinal comfort and stool characteristics:

After two weeks of intervention, compared with the regular milk group (drinking regular milk for 2 weeks), the A2 milk group (drinking A2 milk for 2 weeks) had a 6% lower proportion of occasional abdominal distension and a 4% lower rate of frequent abdominal distension; In terms of symptoms, the rate of reducing bowel sounds in the A2 milk group was 1% lower, and the rate of frequent bowel sounds decreased by 5%; the rate of defecation frequency twice a day in the A2 milk group increased by 6%, and the rate of once every two days decreased by 6%; The difference in defecation traits is relatively small. Compared between the two groups, it is mainly reflected in the 6% reduction of "3-intestinal lump with cracks on the surface" in the A2 milk group, and "4-type four, intestinal or snake-like and soft" "Smooth" has a 3% increase.

Two weeks after the washout, compared with the normal milk group (without taking ordinary milk for 2 weeks), the A2 milk group (without taking A2 milk for 2 weeks) had a significant increase of 13% without gas symptoms, and a decrease in normal exhaust. 6%. There is no significant difference in other indicators.

After alternate intervention for two weeks, there was a statistically significant difference in the distribution of abdominal pain scores between the normal milk group (continuing to drink A2 milk for 2 weeks) and the A2 milk group (continuing to drink ordinary milk for 2 weeks). Specifically, compared with the normal milk group who continued to drink A2 milk, the A2 milk group who continued to drink normal milk had a 9% higher rate of occasional abdominal distension.

table 3

The above results indicate that the cohort of volunteers after two weeks of drinking A2 milk, compared with drinking regular milk, significantly reduced the volunteers’ abdominal distension and bowel movements, increased the frequency of bowel movements, and slightly changed the stool characteristics; and this The improvement basically disappeared after two weeks of washing and changing the milk type.

(3) Changes in the composition of intestinal bacteria and probiotics:

The changes of Bifidobacterium in the A2 milk group and the ordinary milk group with the intervention stage at each age are shown in Figure 1. It can be seen from Figure 1 that age significantly affects the diversity of intestinal flora. In the A2 milk group, after two weeks of drinking A2 milk, the abundance of bifidobacteria in the intestinal tract of volunteers was significantly higher than when they were admitted into the group. In the normal milk group, volunteers eluted for two weeks and continued to drink A2 milk for two more weeks. The abundance of intestinal bifidobacteria was significantly higher than that in the elution phase.

Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that: The technical solutions recorded in the foregoing embodiments are modified, or some of the technical features are equivalently replaced; these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Industrial applicability

The application of β-casein A2 in the promotion of the proliferation of Bifidobacterium provided by the embodiments of the present invention. Compared with ordinary milk, milk containing β-casein A2 can significantly reduce the proportion of abdominal distension and bowel movement, increase the frequency of defecation, and It can change stool traits; and milk containing β-casein A2 can significantly increase the relative abundance of Bifidobacterium in intestinal bacteria.

Claims

Application of β-casein A2 in promoting the proliferation of Bifidobacterium.
An application of a composition for promoting the proliferation of Bifidobacterium, the composition comprising β-casein; in the β-casein, the mass percentage of β-casein A2 is ≥75%.
The application according to claim 2, characterized in that: in the β-casein, the mass percentage of β-casein A2 is ≥90%.
The application according to claim 2, wherein the mass percentage of β-casein A2 in the β-casein is 100%.
The use according to claim 2, wherein the composition is milk or dairy products.
The application according to claim 5, wherein the milk comprises fresh milk, milk powder, liquid milk reconstituted from powder, skimmed milk, homogenized milk, condensed milk, low-strength concentrated milk, pasteurized milk or At least one of non-pasteurized milk.
The application according to claim 5, characterized in that: the milk is milk, and every 100g of the milk contains the following nutrients: protein 3.2g, fat 3.8g, carbohydrate 4.5g, sodium 70mg; wherein, In the protein, the content of β-casein A2 is 0.9 g, and the mass percentage of β-casein A2 in β-casein is 100%.
The application according to claim 7, characterized in that the milk is prepared by the following method: raw milk is empirically collected, filtered, standardized, homogenized, sterilized, cooled, and canned to obtain milk.
The application according to claim 7, characterized in that the way of drinking the milk is within 1 hour after breakfast and/or dinner.
The application according to claim 5, wherein the dairy product comprises at least one of cream, yogurt, quark food, cheese, butter or ice cream.