Breast milk oligosaccharide for regulating butyric acid and improving intestinal microenvironment health and application thereof

Abstract

The invention provides breast milk oligosaccharide for regulating butyric acid and improving intestinal microenvironment health and application thereof. Specifically, the invention provides an application of breast milk oligosaccharide in preparation of food for improving intestinal microenvironment health, and the breast milk oligosaccharide is one of fucosyl oligosaccharide, sialic acid oligosaccharide or lactose-N-tetrasaccharide. Wherein the improvement of the intestinal microenvironment health comprises the steps of regulating and controlling the production of butyric acid in an intestinal system, increasing the overall production of short-chain fatty acid, being utilized by intestinal flora as prebiotics in the intestinal system and producing gas, reducing the production of isobutyric acid and isovaleric acid and / or reducing the pH value to maintain the intestinal microenvironment health. The breast milk oligosaccharide is used for being added into infant food (including infant formula powder, complementary food and nutritional supplements) and nutritional supplements or food for children, teenagers and adults over 3 years old, and has a wide application prospect.

Description

[0001] This invention is a divisional application with application number 202011381158.X, application date November 30, 2020, and invention name “Human milk oligosaccharides and their applications for regulating butyric acid and improving the health of intestinal microenvironment”. Technical Field

[0002] The present invention relates to a new application of human milk oligosaccharides, specifically, to a new application of sialic acid-based oligosaccharides 3-SL or 6-SL in improving the health of the intestinal microenvironment, especially in regulating the production of butyric acid. Background Art

[0003] Human milk oligosaccharides (HMOs) are the third most abundant substance in breast milk after lactose and fat. Their total content varies at different stages of lactation, ranging from about 12-14 g / L in mature milk to about 20-24 g / L in colostrum. Each type of human milk oligosaccharide has a lactose at the reducing end, most of which have polylactosamine as the main chain, and contain fucose, sialic acid or both at the chain end. Human milk oligosaccharides are mainly composed of three categories: (1) fucosyl oligosaccharides, represented by 2'-fucosyl oligosaccharides and 3'-fucosyl oligosaccharides; (2) sialic acid oligosaccharides, represented by 3'-sialyllactose and 6'-sialyllactose; (3) oligosaccharides formed by core sugar chain structures without fucosyl or sialic acid groups, represented by lactose-N-tetraose and lactose-N-neotetraose. The presence and content of HMOs vary from person to person and are related to the Lewis secretory composition of the lactating mother. Since the raw material of infant formula is usually cow's milk, which usually contains no or very little of these oligosaccharides, HMOs have become a gap that infant formula must cross if it wants to be closer to the composition of breast milk.

[0004] Intestinal flora is an important component of the human intestinal micro-ecosystem and plays an important role in human health. Anaerobic Bacteroides, Bifidobacterium, Eubacteria, Streptococcus and Lactobacillus in the intestinal flora can release metabolites of short chain fatty acids (SCFA) by fermenting carbohydrates, proteins and lipids, mainly including acetic acid, propionic acid and butyric acid. SCFA can regulate various physiological functions of the body and play an important role in regulating the health of the intestinal microenvironment. For example, SCFA can provide energy and regulate electrolytes. Acetic acid is an important source of energy for the host. Propionic acid can participate in the process of reverse conversion of pyruvate into glucose. Butyric acid is taken up by epithelial cells and is the main energy source for epithelial cells. SCFA also has anti-inflammatory, intestinal barrier function and antibacterial effects. SCFA released by intestinal flora fermentation can reduce intestinal pH, thereby increasing the growth of beneficial bacteria in the intestine and reducing the proliferation of harmful bacteria. Other physiological functions of SCFA include reducing the risk of early-onset type 1 diabetes, reducing anxiety, and promoting bone growth. The production of butyric acid is significantly associated with a lower incidence of neonatal skin surface allergies, food allergies and respiratory allergies.

[0005] At present, in the fields of infant formula, complementary food and nutritional supplements, there is a need for solutions that can produce butyric acid and regulate the health of infant microecology. At the same time, in the fields of children over 3 years old, adolescents and adults, it is also necessary to maintain the stability and health of the intestinal microecology. Summary of the invention

[0006] One object of the present invention is to provide a new application of human milk oligosaccharides.

[0007] The present invention finds that some human milk oligosaccharides have the effect of significantly improving the health of the intestinal microenvironment, which is specifically manifested in that they can regulate the production of butyric acid in the intestinal system, increase the overall production of short-chain fatty acids, be utilized by intestinal flora in the intestinal system as prebiotics and produce gas, reduce the production of isobutyric acid and isovaleric acid and / or lower the pH to maintain the health of the intestinal microenvironment, thereby providing a new application of human milk oligosaccharides.

[0008] Specifically, the present invention provides the use of human milk oligosaccharides in the preparation of food for improving the health of the intestinal microenvironment, wherein the human milk oligosaccharide is one of fucosyl oligosaccharides, sialic acid oligosaccharides or lactose-N-tetraose.

[0009] It is known that human milk oligosaccharides include fucosyllactose, sialyl lactose, and the basic sugar chain structure of human milk oligosaccharides without fucosyl or sialic acid groups (typical representatives include lactose-N-tetraose and its isomer lactose-N-neotetraose).

[0010] Among them, 2'-fucosyllactose (2'-FL or 2-FL or 2FL) is a trisaccharide structure formed by fucose and lactose, and is a representative substance of fucosylated oligosaccharides. The commercially available substance is usually prepared by microbial fermentation and has the same structure as the oligosaccharides found in human milk.

[0011] 3-fucosyllactose (3'-FL or 3-FL or 3FL) is a trisaccharide structure formed by fucose and lactose, and is an isomer of 2'-fucosyllactose. It is a representative substance of fucosylated oligosaccharides. It is prepared by microbial fermentation and has the same structure as the oligosaccharides found in human milk.

[0012] Lacto-N-tetraose (LNT) is a representative substance of oligosaccharides with a core sugar chain as the basic structure and without fucosyl or sialic acid groups. It is prepared by microbial fermentation and has the same structure as the oligosaccharides found in human milk.

[0013] 3'-sialyllactose (3'-SL or 3-SL or 3SL) is a trisaccharide structure formed by sialic acid and lactose, and is a representative substance of sialic acid-based oligosaccharides. This substance is prepared by microbial fermentation and has the same structure as the oligosaccharides found in human milk.

[0014] 6'-sialyllactose (6'-SL or 6-SL or 6SL) is a trisaccharide structure formed by sialic acid and lactose, and is a representative substance of sialic acid-based oligosaccharides. This substance is prepared by microbial fermentation and has the same structure as the oligosaccharides found in human milk.

[0015] According to a specific embodiment of the present invention, the human milk oligosaccharides of the present invention are used to improve the health of the intestinal microenvironment, including: regulating the production of butyric acid in the intestinal system, increasing the overall production of short-chain fatty acids, being utilized by intestinal flora in the intestinal system as prebiotics and producing gas, reducing the production of isobutyric acid and / or isovaleric acid, and / or lowering the pH to maintain a healthy intestinal microenvironment.

[0016] According to a specific embodiment of the present invention, in the use of the human milk oligosaccharide of the present invention in preparing food for improving the health of the intestinal microenvironment, the fucosyl oligosaccharide is 2'-FL or 3-FL, and the sialic acid oligosaccharide is 3-SL or 6-SL.

[0017] According to some specific embodiments of the present invention, the human milk oligosaccharide of the present invention is used to regulate the production of butyric acid in the proximal colon, and the human milk oligosaccharide is 6-SL. The food can be milk powder or emulsion (liquid milk), preferably infant formula powder. In this application, taking the food as infant formula powder as an example, the application amount of 6-SL in the food is: the application amount in milk powder is 14.2-1515.3 mg / 100g powder, or 0.02-2.0 g / L when converted into milk; preferably 70.9-606.1 mg / 100g powder, or 0.1-0.8 g / L when converted into milk; more preferably 70.9-454.6 mg / 100g powder, or 0.1-0.6 g / L when converted into milk. When the food is other foods, the amount of each human milk oligosaccharide can be adjusted with reference to the above-mentioned dosage range.

[0018] According to some specific embodiments of the present invention, the human milk oligosaccharide of the present invention is used to regulate the production of butyric acid in the distal colon, and the human milk oligosaccharide is 3-SL or 6-SL. Preferably, the human milk oligosaccharide is further used to reduce the production of isobutyric acid and / or isovaleric acid in the distal colon, and in this application, the human milk oligosaccharide is further preferably 3-SL. The food can be milk powder or emulsion (liquid milk), preferably infant formula powder. In this application, taking the food as infant formula powder as an example, the amount of 3-SL used in the food is: the amount used in milk powder is 14.2-1515.3 mg / 100 g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-454.6 mg / 100 g powder, or 0.1-0.6 g / L when converted into milk liquid; more preferably 70.9-227.3 mg / 100 g powder, or 0.1-0.3 g / L when converted into milk liquid. Taking the food as infant formula powder as an example, the application amount of 6-SL in the food is: the application amount in milk powder is 14.2-1515.3 mg / 100g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-606.1 mg / 100g powder, or 0.1-0.8 g / L when converted into milk liquid; more preferably 70.9-454.6 mg / 100g powder, or 0.1-0.6 g / L when converted into milk liquid. When the food is other food, the amount of each human milk oligosaccharide can be adjusted with reference to the above dosage range.

[0019] According to some specific embodiments of the present invention, the human milk oligosaccharide of the present invention is used as a prebiotic in the proximal colon to be utilized by intestinal flora and produce gas, and the human milk oligosaccharide is 3-SL or 6-SL. The food can be milk powder or emulsion (liquid milk), preferably infant formula powder. In this application, taking the food as infant formula powder as an example, the application amount of 3-SL in the food is: the application amount in milk powder is 14.2-1515.3mg / 100g powder, or 0.02-2.0g / L when converted into milk; preferably 70.9-454.6mg / 100g powder, or 0.1-0.6g / L when converted into milk; more preferably 70.9-227.3mg / 100g powder, or 0.1-0.3g / L when converted into milk. Taking the food as infant formula powder as an example, the application amount of 6-SL in the food is: the application amount in milk powder is 14.2-1515.3 mg / 100g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-606.1 mg / 100g powder, or 0.1-0.8 g / L when converted into milk liquid; more preferably 70.9-454.6 mg / 100g powder, or 0.1-0.6 g / L when converted into milk liquid. When the food is other food, the amount of each human milk oligosaccharide can be adjusted with reference to the above dosage range.

[0020] According to some specific embodiments of the present invention, the human milk oligosaccharide of the present invention is used to increase the overall production of short-chain fatty acids in the proximal colon and / or distal colon, and the human milk oligosaccharide is 3-SL or 6-SL. The food can be milk powder or emulsion (liquid milk), preferably infant formula. In this application, taking the food as infant formula as an example, the amount of 3-SL used in the food is: the amount used in milk powder is 14.2-1515.3 mg / 100 g powder, or 0.02-2.0 g / L when converted to milk; preferably 70.9-454.6 mg / 100 g powder, or 0.1-0.6 g / L when converted to milk; more preferably 70.9-227.3 mg / 100 g powder, or 0.1-0.3 g / L when converted to milk. Taking the food as infant formula powder as an example, the application amount of 6-SL in the food is: the application amount in milk powder is 14.2-1515.3 mg / 100g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-606.1 mg / 100g powder, or 0.1-0.8 g / L when converted into milk liquid; more preferably 70.9-454.6 mg / 100g powder, or 0.1-0.6 g / L when converted into milk liquid. When the food is other food, the amount of each human milk oligosaccharide can be adjusted with reference to the above dosage range.

[0021] According to some specific embodiments of the present invention, the human milk oligosaccharide of the present invention is used to reduce the pH of the proximal colon to maintain the health of the intestinal microenvironment. The human milk oligosaccharide is 3-SL or 6-SL. In this application, taking the food as infant formula powder as an example, the application amount of 3-SL in the food is: the application amount in milk powder is 14.2-1515.3 mg / 100g powder, or 0.02-2.0 g / L when converted into milk; preferably 70.9-454.6 mg / 100g powder, or 0.1-0.6 g / L when converted into milk; more preferably 70.9-227.3 mg / 100g powder, or 0.1-0.3 g / L when converted into milk. Taking the food as infant formula powder as an example, the application amount of 6-SL in the food is: the application amount in milk powder is 14.2-1515.3 mg / 100g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-606.1 mg / 100g powder, or 0.1-0.8 g / L when converted into milk liquid; more preferably 70.9-454.6 mg / 100g powder, or 0.1-0.6 g / L when converted into milk liquid. When the food is other food, the amount of each human milk oligosaccharide can be adjusted with reference to the above dosage range.

[0022] According to a specific embodiment of the present invention, in the application of the human milk oligosaccharides of the present invention in the preparation of foods for improving the health of the intestinal microenvironment, the human milk oligosaccharides are also beneficial to promoting the production of short-chain fatty acids such as formic acid, propionic acid, acetic acid, etc. in the intestinal system that are beneficial to the human body.

[0023] According to a specific embodiment of the present invention, the human milk oligosaccharide of the present invention is used in the preparation of a food for improving the health of the intestinal microenvironment, and the food includes one or more of a nutritional supplement, an infant formula, and a complementary food. Specifically, the food can be an infant food (including infant formula, complementary food, nutritional supplement), or a nutritional supplement or food for children over 3 years old, adolescents and adults, for example, a fermented dairy product (such as fermented milk, flavored fermented milk, fermented milk beverage, etc.), cheese, milk-containing beverage, solid beverage or milk powder, etc.

[0024] In summary, the present invention found that human milk oligosaccharides can significantly improve the health of the intestinal microenvironment and can be added to infant food (including infant formula, complementary food, nutritional supplements), as well as nutritional supplements or foods for children over 3 years old, adolescents and adults, and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1A Schematic diagram of feces inoculation and culture in the SHIME device of the present invention.

[0026] Figure 1BA schematic diagram showing the SHIME fermentation grouping of the present invention.

[0027] Figure 2 Shown are the bacterial flora of the simulated proximal colon (left) and distal colon (right) after two weeks of culture in the SHIME device that simulates the infant colon.

[0028] Figure 3A The results of the pH change over time of the proximal colon of each HMO in Example 2 of the present invention in a small batch fermentation experiment are shown.

[0029] Figure 3B The results of the pH change over time of the distal colon in a small batch fermentation experiment of each HMO in Example 2 of the present invention are shown.

[0030] Figure 4A The results of the test of the change of the air pressure over time caused by each HMO in the proximal colon in a small batch fermentation experiment in Example 3 of the present invention are shown.

[0031] Figure 4B The results of the test of the change of air pressure over time caused by each HMO in Example 3 of the present invention in a small batch fermentation experiment in the distal colon are shown.

[0032] Figure 5 The overall detection results of short-chain fatty acids produced by small-batch fermentation of various HMOs in the simulated proximal colon environment in Example 4 of the present invention are shown.

[0033] Figure 6 The results of the detection of butyric acid produced in the batch fermentation experiment of feces simulating the proximal colon of infants by each monomer of human milk oligosaccharide in Example 4 of the present invention are shown.

[0034] Figure 7 The results of the detection of formic acid produced in the batch fermentation experiment of feces simulating the proximal colon of infants in Example 4 of the present invention are shown.

[0035] Figure 8 The results of the test of acetic acid produced in the batch fermentation experiment of feces simulating the proximal colon of infants by each monomer of human milk oligosaccharide in Example 4 of the present invention are shown.

[0036] Fig. 9 The results of the detection of propionic acid produced in the batch fermentation experiment of feces simulating the proximal colon of infants in Example 4 of the present invention are shown.

[0037] Fig.10 The overall detection results of short-chain fatty acids produced in the batch fermentation experiment of feces simulating the distal colon of infants in each monomer of human milk oligosaccharide in Example 5 of the present invention are shown.

[0038] Fig.11The results of the detection of butyric acid produced in the batch fermentation experiment of feces simulating the distal colon of infants by each monomer of human milk oligosaccharide in Example 5 of the present invention are shown.

[0039] Fig.12 The results of the detection of formic acid produced in the batch fermentation experiment of feces simulating the distal colon of infants by each monomer of human milk oligosaccharide in Example 5 of the present invention are shown.

[0040] Fig.13 The results of the detection of acetic acid produced in the batch fermentation experiment of feces simulating the distal colon of infants by each monomer of human milk oligosaccharide in Example 5 of the present invention are shown.

[0041] Fig.14 The results of the detection of propionic acid produced in the batch fermentation experiment of feces simulating the distal colon of infants by each monomer of human milk oligosaccharide in Example 5 of the present invention are shown.

[0042] Fig.15A and Fig. 15B The results of the detection of isobutyric acid and isovaleric acid produced in the batch fermentation experiment of feces simulating the distal colon of infants in Example 5 of the present invention are shown. DETAILED DESCRIPTION

[0043] In order to have a clearer understanding of the technical features, purposes and beneficial effects of the present invention, the following detailed description is now given in conjunction with specific embodiments and the technical solutions of the present invention. It should be understood that these examples are only used to illustrate the present invention and are not used to limit the scope of the present invention. In the embodiments, each raw reagent material can be obtained commercially, and the experimental method without specifying specific conditions is a conventional method and conventional conditions well known in the art, or according to the conditions recommended by the instrument manufacturer.

[0044] In addition, to avoid repetition, the common steps required for the experiments in each embodiment are listed below, such as the inoculation and cultivation of fecal bacteria.

[0045] Fecal inoculation culture in SHIME device

[0046] Use the SHIME device (see Figure 1A As shown in the figure, a fresh stool sample containing bacteria was obtained from a healthy infant who was born naturally and was exclusively breastfed at 5 months old, and inoculated into the containers corresponding to the proximal colon and distal colon. Feed was fed to the stomach / small intestine end of the device three times a day for two weeks to support the growth and colonization of bacteria in the proximal and distal colons. Among them, the food that was digested in the small intestine and entered the colon end was a standard food provided by ProDigest, the manufacturer of the SHIME device, with the ratio of lactose, casein and whey protein adjusted. The standard food consisted of the following ingredients: pectin (1g / L), glucose (1g / L), starch (1g / L), cellobiose (1g / L), Peptone (2g / L), mucin (6g / L), lactose (2.1g / L), casein (0.2g / L), whey protein-lactalbumin (2.7g / L), L-cysteine ​​hydrochloride (0.2g / L). The food materials in each experiment of the present invention refer to Le Blay et al. (2010) to adjust the ratio of lactose, casein and whey protein to about 12:1:15 and ensure the stability and balance of nutrients to simulate the food composition that the intestinal microecology of infants may contact under conventional breast milk or infant formula feeding. After the infant fecal flora has been stable in the SHIME model for two weeks, the proximal and distal colons will be sampled, dissolved in glycerol to form a stock solution, and stored at -80°C under anaerobic conditions.

[0047] The analysis of the microbiota composition will focus on specific strains: Lactobacillus, Bifidobacterium, Roseburia, Eubacterium and Faecalibacterium, as they are known to be associated with (prebiotic) health benefits. The analysis is based on qPCR.

[0048] Small batch fermentation

[0049] After the infant flora was inoculated into the SHIME model and grown stably for 2 weeks (as described in the “Inoculation and culture of feces in the SHIME device”), 10 mL of the flora from the proximal and distal colons were taken and transferred to fermentation bottles under anaerobic conditions for small-batch fermentation. On the basis of 43 mL of basal buffer (used to adjust the pH and simulate the corresponding colon environment), each fermentation bottle also contained 20 mL of PBS buffer (used to dissolve and bring in the HMO test substance) with different HMO contents, so that the final concentrations of various HMOs were 0.02 g / L, 0.2 g / L, and 2 g / L, and the pH of the proximal colon was set to 5.6 and the pH of the distal colon was set to 6.5. The vials were incubated at 37°C with shaking. During the incubation, the air pressure was tested at 0, 6, 24, and 48 hours, and then samples were taken to test the pH and short-chain fatty acids. The determination was repeated three times.

[0050] During the HMO intervention, gas production was compared between the groups by measuring changes in gas pressure. Short-chain fatty acids were analyzed for isobutyric acid, isovaleric acid, butyric acid, propionic acid, and acetic acid, and formic acid by HPLC.

[0051] SHIME Fermentation

[0052] The fecal flora of infants sampled and stored in the "Inoculation and Culture of Feces in SHIME Device" was inoculated into the SHIME model to investigate the fermentation of HMO in the SHIME device. Two batches of experiments were conducted, with three sample groups (or controls) in each batch. In the three experimental devices, the devices simulating the extreme and distal colon were inoculated separately (see Figure 1BSchematic diagram of the experiment. Feed was fed three times a day. After 4 days of culture, HMO was added to the feed (the control group did not add HMO). 280 mg HMO (concentration of 2 g / L) and 60 mL pancreatic juice were added to 140 mL of each feed. Each group of HMO and control was only tested once, so the biological replicate was 1. SHIME fermentation continued until the 14th day after HMO intervention, and samples were taken at different time periods during the fermentation.

[0053] Data analysis

[0054] Two-tailed, paired t-test was performed on the data results. If there is a significant difference between the two groups and p<0.05, it is indicated by an asterisk *. Two asterisks ** indicate p<0.01. Three asterisks *** indicate p<0.001.

[0055] Example 1: Simulating the bacterial flora in the proximal and distal colon environments

[0056] Please refer to the above paragraphs for the preparatory steps and specific experimental methods.

[0057] Please refer to the fecal flora identification after two weeks of stable inoculation and culture in the SHIME device. Figure 2 After two weeks of stabilization in the SHIME model, the bacterial flora was determined and it was found that the content of bifidobacteria and lactobacilli was very low and almost undetectable, which was consistent with previous literature reports (Laforest-Lapointe, 2017). It was proved that in the simulated infant intestinal environment, after feeding with a standard food containing lactose / casein / whey, the corresponding colon flora environment was more similar to that of infants fed with formula powder rather than breastfed infants.

[0058] Example 2: pH changes over time for each HMO in a small batch fermentation experiment

[0059] Please refer to the above paragraphs for the preparatory steps and specific experimental methods.

[0060] The results of the pH changes over time in the proximal colon of each HMO in a small-batch fermentation experiment can be found in Figure 3A The results of the pH changes of the distal colon over time in the small-batch fermentation experiments of each HMO can be found in Figure 3B It can be seen that in the proximal colon, the pH decrease at 6 hours was more significant than at other time points. It can be seen that as the fermentation time increases, HMO will be utilized by the flora in the infant feces to produce short-chain fatty acids, thereby lowering the pH.

[0061] Example 3: Changes in air pressure caused by each HMO in a small batch fermentation experiment over time

[0062] Please refer to the previous paragraphs for the preparatory steps and specific experimental methods.

[0063] The results of the test on the temporal changes of the gas pressure caused by each HMO in the proximal colon in the small-batch fermentation experiment can be found in Figure 4A The results of the test on the changes of gas pressure in the distal colon caused by each HMO in the small batch fermentation experiment over time can be found in Figure 4B It can be seen that the two sialic acid oligosaccharides 3-SL and 6-SL produced higher air pressure in the proximal colon at 2g / L, proving that under this condition, the two oligosaccharides can be well utilized by fecal flora. Under conditions simulating the distal colon, all HMOs can lead to an increase in air pressure at 2g / L.

[0064] Example 4: Production of short-chain fatty acids by small-batch fermentation of HMOs in a simulated proximal colon environment

[0065] The results of the total amount of short-chain fatty acids produced by small-batch fermentation of each HMO in a simulated proximal colon environment are shown in Figure 5 It can be seen that after 48 hours of fermentation, the short-chain fatty acids produced by all HMOs were higher than those at other time points, especially 3-SL and 6-SL produced significantly more SCFA under the condition of 2g / L, among which 2g / L 3-SL also produced more SCFA after 24 hours of fermentation.

[0066] The results of the small-batch fermentation of each HMO to produce butyrate under the simulated proximal colon environment are shown in Figure 6 It can be seen that in the simulated proximal colon environment, only 6-SL can significantly increase the production of butyrate after 48 hours of fermentation.

[0067] The results of the small-batch fermentation of various HMOs to produce formic acid under simulated proximal colon conditions are shown in Figure 7 It can be seen that only 3-SL and 6-SL significantly increased formic acid after 48 hours of simulated proximal colon fermentation, and the effect of 6-SL was more significant.

[0068] The results of the small-batch fermentation of each HMO to produce acetic acid under the simulated proximal colon environment are shown in Figure 8 It can be seen that only 3-SL and 6-SL significantly increased acetic acid after 48 hours of simulated proximal colon fermentation, and the effect of 3-SL was more significant.

[0069] The results of the test on propionic acid produced by small batch fermentation of each HMO in the simulated proximal colon environment are shown in Fig. 9 It can be seen that only 6-SL significantly increased propionate after 48 h of simulated proximal colon fermentation.

[0070] Example 5: Production of short-chain fatty acids by small-batch fermentation of HMOs in a simulated distal colon environment

[0071] The results of the total amount of short-chain fatty acids produced by small-batch fermentation of each HMO in a simulated distal colon environment are shown in Fig.10 . It can be seen that in the distal colon, each HMO is more active in producing SCFA than in the proximal colon, which also confirms the recognized scientific view that most of the fermentation of intestinal flora occurs in the distal colon. As time extends from 0, 6, 24, to 48 hours, the production of SCFA also increases. At the 6-hour time point, 0.02g / L 3-SL and 0.02g / L LNT produce relatively more short-chain fatty acids; at the 24-hour time point, 2g / L 6-SL and 2g / L 3-FL produce relatively more short-chain fatty acids; and at the 48-hour time point, all HMOs can produce more short-chain fatty acids under the condition of 2g / L.

[0072] The results of the small-batch fermentation of each HMO to produce butyrate in a simulated distal colon environment are shown in Fig.11 As can be seen, only 3-SL and 6-SL significantly increased butyrate after 48 h of simulated distal colon fermentation.

[0073] The results of the small-batch fermentation of various HMOs to produce formic acid under simulated distal colon conditions are shown in Fig.12 It can be seen that all HMOs significantly increased formic acid after 48 hours of simulated distal colon fermentation, among which 3-SL, 6-SL, LNT, and 3-FL had more significant effects.

[0074] The results of the small-batch fermentation of each HMO to produce acetic acid under simulated distal colon conditions are shown in Fig.13 It can be seen that all HMOs significantly increased acetic acid after 48 hours of simulated distal colon fermentation, with 3-SL, 6-SL and LNT having more significant effects.

[0075] The results of the test on propionic acid produced by small batch fermentation of each HMO in the simulated distal colon environment are shown in Fig.14 It can be seen that all HMOs significantly increased propionic acid after 48 hours of simulated distal colon fermentation, with 3-SL and 3-FL having the most significant effects.

[0076] The results of the small-batch fermentation of each HMO to produce isobutyric acid and isovaleric acid under simulated distal colon conditions are shown in Fig.15A and Fig. 15B It can be seen that in the distal colon, 2'-FL, 3-FL, 3-SL and LNT can significantly reduce the production of isobutyric acid. The two sialic acid oligosaccharides and the two fucosyl oligosaccharides can reduce the production of isovaleric acid.

[0077] Example 6: Fermentation of butyrate by each HMO in the SHIME model under simulated distal colon environment The test results of butyrate production by each HMO in the SHIME model under the simulated distal colon environment are shown in Table 1.

[0078] Table 1

[0079]

[0080] Under the SHIME model fermentation, after 14 days of fermentation, the butyric acid produced by various HMOs was reduced compared with the first day of fermentation, but the control group (without HMO) had the largest reduction; after HMO intervention, the reduction on the 14th day was significantly improved compared with the first day, among which 3-SL had the lowest reduction, followed by 3-FL, and 6-SL, 2'-FL and LNT also had a good improvement compared with the control group. It can be seen that each HMO has certain advantages in regulating butyric acid in fermentation products compared with the control group without HMO.

Claims

1. Application of human milk oligosaccharides in the preparation of food for improving the health of intestinal microenvironment, wherein: The human milk oligosaccharide is one of fucosyl oligosaccharide, sialic acid oligosaccharide or lactose-N-tetraose, the fucosyl oligosaccharide is 2'-FL or 3-FL, and the sialic acid oligosaccharide is 3-SL or 6-SL.

2. The use according to claim 1, wherein: The improvement of the intestinal microenvironment health includes: regulating the production of butyric acid in the intestinal system, increasing the overall production of short-chain fatty acids, which are used as prebiotics by intestinal flora in the intestinal system and produce gas, reducing the production of isobutyric acid and / or isovaleric acid, and / or lowering the pH to maintain the health of the intestinal microenvironment.

3. The use according to claim 1 or 2, wherein: The human milk oligosaccharide is 2'-FL or 3-FL.

4. The use according to claim 1, wherein: The food includes one or more of nutritional supplements, infant formula powder, and complementary foods.

5. The use according to claim 1, wherein: Human milk oligosaccharide is used to regulate the production of butyrate in the proximal colon, and the human milk oligosaccharide is 6-SL.

6. The use according to claim 5, wherein: The food is milk powder or emulsion, preferably infant formula powder; The usage amount of 6-SL in food is: the usage amount in milk powder is 14.2-1515.3 mg / 100 g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-606.1 mg / 100 g powder, or 0.1-0.8 g / L when converted into milk liquid; more preferably 70.9-454.6 mg / 100 g powder, or 0.1-0.6 g / L when converted into milk liquid.

7. The use according to claim 1, wherein: Human milk oligosaccharides are used to regulate the production of butyric acid in the distal colon, preferably to further reduce the production of isobutyric acid and / or isovaleric acid, and the human milk oligosaccharides are 3-SL or 6-SL.

8. The use according to claim 1, wherein: Human milk oligosaccharides are used as prebiotics by intestinal flora in the proximal colon to produce gas, and / or reduce the pH value of the proximal colon. The human milk oligosaccharides are 3-SL or 6-SL.

9. The use according to claim 1, wherein: Human milk oligosaccharides are used to increase the overall production of short-chain fatty acids in the proximal colon and / or distal colon, and the human milk oligosaccharides are 3-SL or 6-SL.

10. The use according to claim 7, 8 or 9, wherein: The food is milk powder or emulsion, preferably infant formula powder; The application amount of 3-SL in food is: the application amount in milk powder is 14.2-1515.3 mg / 100 g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-454.6 mg / 100 g powder, or 0.1-0.6 g / L when converted into milk liquid; more preferably 70.9-227.3 mg / 100 g powder, or 0.1-0.3 g / L when converted into milk liquid; The usage amount of 6-SL in food is: the usage amount in milk powder is 14.2-1515.3 mg / 100 g powder, or 0.02-2.0 g / L when converted into milk liquid; preferably 70.9-606.1 mg / 100 g powder, or 0.1-0.8 g / L when converted into milk liquid; more preferably 70.9-454.6 mg / 100 g powder, or 0.1-0.6 g / L when converted into milk liquid.

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