Beta-glucan-Ackermania muciniphila co-microcapsule and preparation method thereof

Abstract

The invention discloses a beta-glucan-Ackermania mucilaginosa co-microcapsule and a preparation method thereof, the beta-glucan-Ackermania mucilaginosa co-microcapsule comprises a core material and a wall material, and the volume ratio of the core material to the wall material is 1: (2-6); the core material comprises Akk bacteria and beta-glucan, the bacterial liquid concentration of the Akk bacteria is 108-1010 CFU / g, and the concentration of the beta-glucan is 10-40 g / L; the wall material comprises sodium carboxymethyl cellulose, whey protein, skimmed milk powder and trehalose, the mass fraction of the sodium carboxymethyl cellulose is 2%-3%, the mass fraction of the whey protein is 0.5%-1.0%, the mass fraction of the skimmed milk powder is 1%, and the mass fraction of the trehalose is 2.5%. The problem that the activity and colonization quantity of the Akk bacteria are reduced when the Akk bacteria are taken by a human body and pass through the gastrointestinal tract of the human body is solved.

Description

Technical Field

[0001] The present invention relates to the technical field of synbiotics, and in particular to a beta-glucan-Akkermansia muciniphila co-microcapsule and a preparation method thereof. Background Art

[0002] Akkermansia muciniphila (Akk bacteria for short), as a mucin-degrading bacterium, is widely distributed in the intestinal mucus layer of humans and animals, accounting for 3% to 5% of the total intestinal microorganisms. It is a new generation of star intestinal probiotics. Appropriate supplementation of Akk bacteria can regulate the blood sugar concentration of the human body, improve various symptoms of diabetes, and also relieve metabolic diseases such as obesity and hepatic fatty degeneration.

[0003] To exert the efficacy of Akk bacteria, two conditions must be met: first, it must maintain a high level of activity after being ingested by the human body; second, it must have a sufficient number of colonization in the intestine. Akk bacteria are susceptible to oxygen, high temperature, and gastric acid during processing, storage, and human digestion, which can reduce their activity, thereby reducing the activity and colonization of Akk bacteria in the intestine.

[0004] The information disclosed in this background technology section is only intended to enhance the understanding of the overall background of the invention and should not be regarded as an acknowledgment or any form of suggestion that the information constitutes the prior art already known to a person skilled in the art. Summary of the invention

[0005] In order to overcome the defects of the prior art, a β-glucan-Akkermansia muciniphila co-microcapsule and a preparation method thereof are provided to solve the problem that the activity and colonization number of Akk bacteria in the intestine are reduced due to the adverse effects of the environment on Akk bacteria.

[0006] To achieve the above object, a β-glucan-Akkermansia muciniphila co-microcapsule is provided, comprising a core material and a wall material, wherein the volume ratio of the core material to the wall material is 1:2-6;

[0007] The core material includes Akk bacteria and β-glucan, and the concentration of Akk bacteria liquid is 10 8 -10 10 CFU / g, the concentration of β-glucan is 10-40g / L;

[0008] The wall material comprises sodium carboxymethyl cellulose, whey protein, skim milk powder and trehalose, wherein the mass fraction of the sodium carboxymethyl cellulose is 2% to 3%, the mass fraction of the whey protein is 0.5 to 1.0%, the mass fraction of the skim milk powder is 1%, and the mass fraction of the trehalose is 2.5%.

[0009] Furthermore, the β-glucan is malt β-glucan.

[0010] The present invention provides a method for preparing β-glucan-Akkermansia muciniphila co-microcapsules, comprising the following steps:

[0011] The Akk bacteria are fermented and cultured to a stable stage, and the culture fluid in the Akk bacterial solution is separated to obtain concentrated bacterial mud;

[0012] The concentrated bacterial sludge is added to a β-glucan solution with a concentration of 10 to 40 g / L and mixed evenly to obtain a core material, so that the final bacterial solution concentration of the core material is 10 10 CFU / g;

[0013] Adding the wall material to the core material in a volume ratio of 1:2 to 6, and mixing them evenly to obtain a slurry;

[0014] The slurry is added dropwise into a CaCl2 solution, and wet microcapsules are obtained after solidification and washing;

[0015] The wet microcapsules are freeze-dried to prepare β-glucan-Ackermansia muciniphila co-microcapsules.

[0016] Furthermore, when implementing the step of separating the culture solution in the Akk bacterial solution, the culture solution is separated and removed by centrifuging the Akk bacterial solution at 7500-8500 rpm for 10 minutes.

[0017] The invention provides an application of a beta-glucan-Akkermansia muciniphila co-microcapsule. The beta-glucan-Akkermansia muciniphila co-microcapsule is used for improving the structure of intestinal flora and regulating intestinal microecology.

[0018] The beneficial effects of the present invention are that the β-glucan-Akkermansia muciniphila co-microcapsules of the present invention can promote the proliferation of Akk bacteria and improve the tolerance and storage of Akk bacteria in the gastrointestinal tract. The β-glucan-Akkermansia muciniphila co-microcapsules of the present invention have the effects of improving the structure of intestinal flora, regulating intestinal microecology, and promoting the health of the body. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Other features, objects and advantages of the present application will become more apparent by reading the detailed description of non-limiting embodiments made with reference to the following drawings:

[0020] Figure 1 This is a schematic diagram of the proliferation of Akkermansia muciniphila under different carbon source conditions according to an embodiment of the present invention.

[0021] Figure 2 Schematic diagram of the growth curve and pH value of Akkermansia muciniphila according to an embodiment of the present invention.

[0022] Figure 3 Schematic diagram of the survival of bacteria under different treatments in simulated gastric fluid.

[0023] Figure 4 Schematic diagram of the survival of bacteria under different treatments in simulated intestinal fluid.

[0024] Figure 5 Graph showing the changes in survival rate under different treatments at 4°C.

[0025] Figure 6 Graph showing the changes in survival rate under different treatments at 25°C.

[0026] Figure 7 Schematic diagram of the Bate diversity index of intestinal flora in different treatments.

[0027] Figure 8 Schematic diagram of the species composition (phylum level) of intestinal flora in different treatments.

[0028] Fig. 9 Schematic diagram of the species composition (genus level) of intestinal flora in different treatments.

[0029] Fig.10 Schematic diagram of the concentrations of acetate, propionate, butyrate and total SCFAs in fecal fluids from different treatments.

[0030] Fig.11 Schematic diagram of the effect of the amount of sodium carboxymethyl cellulose added on the microcapsule embedding efficiency.

[0031] Fig.12 Schematic diagram of the effect of whey protein addition on microcapsule embedding efficiency.

[0032] Fig.13 Schematic diagram of the effect of the volume ratio of core material to wall material on the microcapsule embedding efficiency. DETAILED DESCRIPTION

[0033] The present application is further described in detail below in conjunction with the accompanying drawings and embodiments. It is to be understood that the specific embodiments described herein are only used to explain the relevant invention, rather than to limit the invention. It is also necessary to explain that, for ease of description, only the parts related to the invention are shown in the accompanying drawings.

[0034] It should be noted that, in the absence of conflict, the embodiments and features in the embodiments of the present application can be combined with each other. The present application will be described in detail below with reference to the accompanying drawings and in combination with the embodiments.

[0035] The invention provides a beta-glucan-Akkermansia muciniphila co-microcapsule, which comprises a core material and a wall material, wherein the volume ratio of the core material to the wall material is 1:2-6.

[0036] The core material includes Akk bacteria and β-glucan. The concentration of Akk bacteria solution is 10 8 -10 10 CFU / g. The concentration of β-glucan is 10-40g / L.

[0037] The wall material comprises sodium carboxymethyl cellulose, whey protein, skim milk powder and trehalose, wherein the mass fraction of sodium carboxymethyl cellulose in the wall material is 2% to 3%, the mass fraction of whey protein is 0.5 to 1.0%, the mass fraction of skim milk powder is 1%, and the mass fraction of trehalose is 2.5%.

[0038] The β-glucan is maltose β-glucan. The maltose β-glucan from maltose bran is more conducive to the proliferation of Eckermannia muciniphila.

[0039] The present invention provides a method for preparing β-glucan-Akkermansia muciniphila co-microcapsules, comprising the following steps:

[0040] S1. Add the concentrated bacterial slurry of Akk bacteria to β-glucan with a concentration of 10-40 g / L and mix them evenly to obtain the core material, so that the final concentration of the bacterial solution of the core material is 10 10 CFU / g.

[0041] When Akk bacteria fermentation culture reaches the stable period, the bacterial solution is centrifuged at 7500-8500 rpm for 10 minutes to separate and remove the culture solution to obtain concentrated bacterial sludge of Akk bacteria.

[0042] β-glucan was dissolved in 0.85% sterile physiological saline to obtain a β-glucan solution with a concentration of 10-40 g / L. The concentrated bacterial mud was added to the Yuanmai β-glucan solution and mixed evenly to obtain a core material. The final bacterial solution concentration of the core material was 10 10 CFU / g.

[0043] S2. Add wall material to the core material in a volume ratio of 1:2 to 6, and mix well to obtain slurry.

[0044] S3, adding the slurry dropwise into the CaCl2 solution, and obtaining wet microcapsules after solidification, filtering and washing.

[0045] Specifically, the slurry was uniformly added dropwise to the CaCl2 solution using a 5 mL sterile syringe (No. 6 needle), and after curing for 30 min, the wet microcapsules were collected after washing twice with sterile water.

[0046] S4. The wet microcapsules are freeze-dried to obtain β-glucan-Ackermansia muciniphila co-microcapsules.

[0047] Specifically, the prepared wet microcapsules were placed in a -80°C low-temperature refrigerator and pre-frozen for 2 hours, and then freeze-dried for 24 hours to obtain freeze-dried β-glucan-Ackermansia muciniphila co-microcapsules.

[0048] The invention provides an application of a beta-glucan-Akkermansia muciniphila co-microcapsule. The beta-glucan-Akkermansia muciniphila co-microcapsule is used for improving the structure of intestinal flora and regulating intestinal microecology.

[0049] The β-glucan-Akkermansia muciniphila co-microcapsules of the present invention can promote the proliferation of Akk bacteria, and can also improve the tolerance and storage of Akk bacteria in the gastrointestinal tract. The β-glucan-Akkermansia muciniphila co-microcapsules of the present invention have the effects of improving the structure of intestinal flora, regulating intestinal microecology, and promoting the health of the body.

[0050] In order to further illustrate the performance characteristics of the β-glucan-Akkermansia muciniphila co-microcapsules of the present invention, the following examples are given for detailed description.

[0051] Example 1

[0052] This example mainly focuses on the study of the effect of β-glucan in β-glucan-Akkermansia muciniphila co-microcapsules on promoting the proliferation of Akkermansia in vitro.

[0053] Specifically, wheat β-glucan, oat β-glucan and yeast β-glucan were added as carbon sources to 5 mL of brain heart infusion broth (BHI) liquid culture medium to make their final mass concentrations of 5 mg / mL, 10 mg / mL, 20 mg / mL and 40 mg / mL, respectively, sterilized and cooled, and 100 μL of Akkermansia muciniphila (bacterial solution concentration was 10 10 CFU / g), after anaerobic culture at 37℃ for 48h, the absorbance was measured at 600nm, and the sterile culture medium was used as the blank control. Three parallel experiments were performed in each group. The probiotic effect of β-glucan on Akk bacteria was analyzed by detecting the growth of the strain after fermentation ( Figure 1 ).

[0054] Depend on Figure 1 It can be seen that yeast β-glucan has no obvious growth-promoting effect on Akk bacteria, and when its concentration reaches 40 mg / mL, it will have an inhibitory effect on the bacteria.

[0055] Both wheat β-glucan and oat β-glucan have a probiotic effect on Akk, and wheat β-glucan has a greater growth-promoting effect on Akk than oat β-glucan. The probiotic effect of wheat β-glucan first increased and then decreased with the increase of mass concentration. When the mass concentration was 20 mg / mL, the probiotic effect was the best (P<0.05). The OD of the bacterial solution was measured at 0, 6, 12, 18, 24, 30, 36, 42 and 48 h. 600 And draw the growth curve and measure the pH change of the fermentation broth ( Figure 2 ), indicating that 20 mg / mL of Yuanmai β-glucan has an effective proliferation effect on Akk bacteria and can quickly reduce the pH in the intestinal environment during the fermentation process.

[0056] The composition of short-chain fatty acid products in the fermentation broth was determined and analyzed using an Agilent 7820 gas chromatograph and an Agilent 5977 quadrupole mass spectrometer (Table 1). The results confirmed that 20 mg / mL of β-glucan fermentation from wheat could promote the production of short-chain fatty acids, significantly increasing the content of acetic acid, butyric acid and total SCFAs in the culture medium (P<0.05), but had no significant effect on the content of propionic acid.

[0057] Table 1. Content and composition of SCFAs produced by fermentation of 20 mg / mL β-glucan from wheat and Akk bacteria

[0058]

[0059] Note: Data are expressed as (x±s), n=3; different lowercase letters indicate significant differences among samples (P<0.05).

[0060] Example 2

[0061] This example is a study on the preparation process of β-glucan-Akkermansia muciniphila co-microcapsules.

[0062] Specifically, this embodiment provides a β-glucan-mucophilic Akkermansia co-microcapsule, wherein the concentration of Akkermansia spp. in the core material is 10 10 CFU / g, the concentration of wheat β-glucan is 20g / L; the mass fraction of sodium carboxymethyl cellulose in the wall material is 2.5%, the mass fraction of whey protein is 0.7%, the mass fraction of skim milk powder is 1% and the mass fraction of trehalose is 2.5%.

[0063] The volume ratio of core material to wall material is 1:6.

[0064] The preparation method of β-glucan-Akkermansia muciniphila co-microcapsules in this embodiment comprises the following steps:

[0065] When Akk bacteria fermentation culture reaches the stable stage, the bacterial solution is centrifuged at 8000 rpm for 10 minutes to separate and remove the culture solution to obtain concentrated bacterial sludge;

[0066] The wheat β-glucan was dissolved in 0.85% sterile saline to obtain a 20 g / L wheat β-glucan solution. The concentrated bacterial sludge was added to the wheat β-glucan solution and mixed evenly to obtain a final bacterial solution concentration of 10 10 CFU / g;

[0067] Then, adding wall materials in a ratio of 1:6 (w / w), the wall materials comprising 0.7% whey protein by mass, 2.5% sodium carboxymethyl cellulose by mass, 1% skim milk powder by mass and 2.5% trehalose by mass, and mixing them evenly;

[0068] The mixture was uniformly added to the CaCl2 solution using a 5 mL sterile syringe (No. 6 needle). After solidification for 30 min, the mixture was filtered and washed twice with sterile water to collect the wet microcapsules.

[0069] The prepared wet microcapsules were placed in a -80°C low-temperature refrigerator and pre-frozen for 2 hours, followed by freeze drying for 24 hours to obtain freeze-dried β-glucan-Akkermansia muciniphila co-microcapsules.

[0070] Example 3

[0071] This example is a study on the preparation process of β-glucan-Akkermansia muciniphila co-microcapsules.

[0072] Specifically, this embodiment provides a β-glucan-mucophilic Akkermansia co-microcapsule, wherein the concentration of Akk bacterial solution in the core material is 10 8 CFU / g, the concentration of β-glucan in wheat is 30g / L; the mass fraction of sodium carboxymethyl cellulose in the wall material is 3.0%, the mass fraction of whey protein is 0.5%, the mass fraction of skim milk powder is 1% and the mass fraction of trehalose is 2.5%.

[0073] The volume ratio of core material to wall material is 1:4.

[0074] The preparation method of β-glucan-Akkermansia muciniphila co-microcapsules in this embodiment comprises the following steps:

[0075] When Akk bacteria fermentation culture reaches the stable stage, the bacterial solution is centrifuged at 8000 rpm for 10 min, the culture solution is separated and removed, and concentrated bacterial sludge is obtained;

[0076] The wheat β-glucan was dissolved in 0.85% sterile saline to obtain a 30 g / L wheat β-glucan solution. The concentrated bacterial sludge was added to the wheat β-glucan solution and mixed evenly to obtain a final bacterial solution concentration of 10 8 CFU / g;

[0077] Then, adding wall materials in a ratio of 1:4 (w / w), the wall materials comprising 0.5% whey protein by mass, 3.0% sodium carboxymethyl cellulose by mass, 1% skim milk powder by mass and 2.5% trehalose by mass, and mixing them evenly;

[0078] The mixture was uniformly added to the CaCl2 solution using a 5 mL sterile syringe (No. 6 needle). After solidification for 30 min, the mixture was filtered and washed twice with sterile water to collect the wet microcapsules.

[0079] The prepared wet microcapsules were placed in a -80°C low-temperature refrigerator for pre-freezing for 2 hours, and then freeze-dried for 24 hours to obtain freeze-dried microcapsules.

[0080] Comparative Example 1

[0081] This comparative example is the control group, which is free bacteria (Akk bacteria). The bacteria were resuspended in 0.85% sterile saline. The concentration of the concentrated bacterial solution was adjusted to 10 10 CFU / g.

[0082] Comparative Example 2

[0083] This embodiment provides a method for preparing Akkermansia muciniphila microcapsules, comprising the following steps:

[0084] When Akk bacteria fermentation culture reaches the stable stage, the bacterial solution is centrifuged at 8000 rpm to separate and remove the culture solution to obtain concentrated bacterial sludge;

[0085] The concentrated bacterial sludge was resuspended in 0.85% sterile saline and the concentration of the concentrated bacterial solution was adjusted to 10 10 CFU / g. Then, wall materials were added to the bacterial solution at a ratio of 1:6 (w / w), the wall materials including 0.5% whey protein by mass, 3.0% sodium carboxymethyl cellulose by mass, 1% skim milk powder by mass and 2.5% trehalose by mass, and mixed evenly;

[0086] The mixture was uniformly added to the CaCl2 solution using a 5 mL sterile syringe (No. 6 needle). After solidification for 30 min, the mixture was filtered and washed twice with sterile water to collect the wet microcapsules.

[0087] The prepared wet microcapsules were placed in a -80°C low-temperature refrigerator and pre-frozen for 2 hours, followed by freeze drying for 24 hours to obtain freeze-dried Akkermansia muciniphila microcapsules.

[0088] 0.5 mL of comparative example 1, 0.5 g of comparative example 2, and 0.5 g of examples 2 and 3 were added to 4.5 mL of simulated gastric juice and intestinal juice, and the former was cultured at 37°C and 100 r / min for 30, 60, 90, and 120 min, and the latter was cultured for 1, 2, 3, and 4 h. Then, cyst dissociation fluid was added, and the cells were shaken for 10 min until the cysts were completely dissected, and then the live bacteria were counted. Three parallel experiments were performed for each group.

[0089] See also Figure 3 As shown, the characteristics of the synbiotic microcapsules of Example 2, Example 3, Comparative Example 1 and Comparative Example 2 are analyzed. Figure 3 It can be seen that the number of live bacteria of Akk in gastric juice, whether embedded or not, decreased, and was negatively correlated with time. Under low pH conditions and the action of pepsin, the number of live bacteria of free bacteria decreased rapidly until it reached 0. The number of live bacteria of Akk microcapsules, Examples 2 and 3 at 60 min was greater than 61 g (CFU / g).

[0090] In Examples 2 and 3, 6.81 and 6.03 log (CFU / mL) of bacteria still survived until 120 min, and the number of viable bacteria was still greater than 61g (CFU / g), indicating that the embedded synbiotic microcapsules are more helpful for Akk bacteria to resist the acidic environment of the stomach. Example 2 is slightly better than Example 3.

[0091] See also Figure 4 It can be seen that: in the simulated intestinal fluid environment, the number of live bacteria in the free bacteria and Akk bacteria microcapsules decreased. After 180 minutes of culture, the number of live bacteria in both decreased to 7.57 and 8.04 log (CFU / mL), respectively; while in Examples 2 and 3, the number of Akk bacteria increased at 60 minutes. This is due to the proliferation of Akk bacteria in the synbiotic microcapsules after being released in the intestinal fluid and utilizing the Yuanmai β-glucan, indicating that the prepared synbiotic microcapsules have good enteric solubility. After 240 minutes in the intestinal fluid, the number of live bacteria in Examples 2 and 3 decreased by less than 1 order of magnitude and less than 2 orders of magnitude, and the survival rate was high.

[0092] Equal amounts of freeze-dried concentrated bacterial solution, Akk microcapsules and synbiotic microcapsules were placed in sterile vials, sealed and stored at 4°C and 25°C, respectively. Live bacteria were counted at regular intervals, and three parallel experiments were performed in each group.

[0093] Storage stability of different treatments at 4°C Figure 5As shown, the viable counts of the four groups were all greater than 6lg (CFU / g) at 21 days of storage; at 49 days of storage, the viable count of Akk bacteria that were not embedded was only 1.82log (CFU / mL). The viable counts of Akk bacteria microcapsules, Examples 2 and 3 were still > 6lg (CFU / g); at 63 days of storage, no viable bacteria were detected in the unencapsulated Akk bacteria, and the viable count of Akk bacteria microcapsules was < 6lg (CFU / g). The viable counts of Examples 2 and 3, respectively, were 8.87 and 7.81log (CFU / mL) of the bacteria survived, and Example 2 was slightly better than Example 3.

[0094] Storage stability of different treatments at 25°C Figure 6 As shown, within the first 14 days of storage, the number of viable bacteria in the four groups was greater than 6lg (CFU / g); at 21 days of storage, the number of viable bacteria of the unencapsulated Akk bacteria decreased from 9.27 to 4.21log (CFU / mL), and the number of viable bacteria decreased by 3 orders of magnitude. The number of viable bacteria of Akk bacteria microcapsules, Examples 2 and 3 was still > 6lg (CFU / g); at 35 days of storage, the number of viable bacteria of the unencapsulated Akk bacteria decreased from 9.27log (CFU / mL) to 1.11log (CFU / mL), and the number of viable bacteria decreased by 8 orders of magnitude. The number of viable bacteria of Akk bacteria microcapsules, Examples 2 and 3, respectively, 4.21, 6.23 and 5.81log (CFU / mL) of the bacteria survived, and the number of viable bacteria in Example 2 can still meet the requirements of FAO (not less than 6log CFU / g).

[0095] In order to explore the effect of synbiotic preparations on intestinal flora, the synbiotic preparations prepared in Example 2 were fermented with intestinal flora derived from human feces in an in vitro simulated intestinal anaerobic system, and the abundance changes of typical probiotics in the flora after fermentation were noted. Different synbiotics were prepared using the preparation process and the addition amount of each component in Example 2. The differences in the effects of the synbiotic combination in Example 2 and the two treatments in Comparative Example 1 on the flora were compared.

[0096] The BHI fermentation medium was deoxygenated with N2 and dispensed into anaerobic vials. The initial pH of the fermentation was 7.5 ± 0.2. Fresh human feces was diluted with PBS to form a suspension, and Akk bacteria or synbiotic microcapsules were inoculated into anaerobic vials, with a bacterial concentration of 10 10 CFU / g, fermented at 37℃ for 24h, and the group without addition was set as blank group ck. The 16SrRNA microbial diversity of the 24h fermentation broth was analyzed using the HiSeq high-throughput sequencing platform to compare the changes in the abundance of intestinal flora.

[0097] Chao index is used to indicate the richness of sample flora. The larger the Chao index, the higher the richness of the flora. Shannon is the community diversity index. The larger the value, the higher the diversity of the flora. Pielou's evenness, pielou uniformity, or Shannon's evenness, only reflects uniformity. The larger the value, the more uniform. As shown in Table 2, compared with the ck group, the Chao index, Shannon index, and Pielou of the Akk bacteria group were slightly reduced. Compared with the ck group and the Akk bacteria group, the Chao index, Shannon index, and Pielou of the synbiotic microcapsule group were all reduced, and Chao and Shannon were significantly reduced (P<0.05), indicating that synbiotic microcapsules would reduce the richness, diversity, and uniformity of the flora in the intestine. Good'sCoverage, calculates the coverage of low-abundance OTUs in the sample, reflects the amount of sequencing data, and the larger the value, the more sufficient the sequencing data. As shown in the table, the Good's Coverage in all samples is greater than 99%, indicating that the sequencing data is sufficient.

[0098] Table 2. Comparison of alpha diversity of intestinal flora in different treatments

[0099]

[0100] See also Figure 7 As shown, Figure 7 is the Bate diversity index of intestinal flora in different treatments. The horizontal and vertical coordinates are PCo1 and PCo2, and their contribution rates are 22.33% and 30.34%, respectively, indicating that these two principal components are the main factors for the differences in the structure and composition of the sample flora. In the PCo1 and PCo2 directions, the samples of ck and Akk bacteria are relatively close, indicating that the flora characteristics of the two treatments are relatively similar at this principal component level, but there are large differences compared with the samples treated with synbiotic microcapsules, indicating that the flora structure of the samples treated with synbiotic microcapsules has undergone significant changes.

[0101] See also Figure 8 As shown in the figure, at the phylum level, the microorganisms in the fermentation broth of ck and the fermentation broth with only Akk bacteria added were mainly Bacteroidetes, Proteobacteria, Firmbacteria and Fusobacteria; the microorganisms in the fermentation broth with the addition of synbiotic microcapsules were mainly Bacteroidetes, Firmbacteria, Verrucomicrobiota and Proteobacteria.

[0102] See also Fig. 9 As shown in the figure, at the genus level, the microbial composition of the blank group and the fermentation broth with only Akk bacteria added was similar, but the proportion was different, and the Akk bacteria content was low. The proportion of Akk bacteria in the microbial composition of the fermentation broth of synbiotics increased, and the proportion of Escherichia-Shigella decreased. The results showed that synbiotics microcapsules promoted the abundance of beneficial bacteria in the intestinal flora and inhibited the abundance of harmful bacteria.

[0103] See also Fig.10 As shown, the production of short-chain fatty acids in the Akk bacteria group was slightly higher than that in the ck group, but there was no significant difference (p>0.05). Compared with the Akk bacteria group and the ck group, the synbiotic microcapsule group had significantly increased acetic acid, propionic acid, butyric acid and total short-chain fatty acids (p<0.05). It can be seen that the addition of the synbiotic microcapsules of the present invention can effectively promote the production of short-chain fatty acids.

[0104] The effect of Example 3 was verified according to the above method, and the application effect was the same as that of Example 2.

[0105] Example 4

[0106] This example is a study on different concentrations of sodium carboxymethyl cellulose in β-glucan-Akkermansia muciniphila co-microcapsules.

[0107] Specifically, this embodiment provides a β-glucan-Akkermansia muciniphila co-microcapsule, which is similar to the formula provided in Example 2, except that the proportion of sodium carboxymethyl cellulose used in this embodiment is 1%, 1.5%, 2.0%, 2.5% or 3.0%.

[0108] See also Fig.11 It can be seen that as the mass fraction of CMC increases, the embedding rate of microcapsules also gradually increases. When the mass fraction of CMC is 2.5%, the embedding rate of microcapsules is the largest, but as the mass fraction of CMC continues to increase, the embedding rate of microcapsules decreases. Therefore, 2.0% to 3.0% is selected as the appropriate concentration range of carboxymethyl cellulose.

[0109] Example 5

[0110] This example is a study on different concentrations of whey protein in β-glucan-Akkermansia muciniphila co-microcapsules.

[0111] This embodiment provides a β-glucan-Akkermansia muciniphila co-microcapsule, which is similar to the formula provided in Example 2, except that the proportion of whey protein used in this embodiment is 0.25%, 0.5%, 0.7%, 1.0% or 1.5%.

[0112] See also Fig.12It can be seen that the embedding rate of microcapsules increases with the increase of whey protein mass fraction. This may be because the addition of whey protein fills the pore structure of CMC gel, effectively traps Akk bacteria in microcapsules, and enhances the protective effect; when the whey protein mass fraction is 0.7%, the microcapsule embedding rate is the largest; but its mass fraction continues to increase, and the embedding rate is reduced due to the excessive viscosity of whey protein and the poor mixing effect of probiotics. Therefore, 0.5% to 1.0% is selected as the appropriate concentration range of whey protein.

[0113] Example 6

[0114] This example is a study on different volume ratios of core material to wall material in β-glucan-Akkermansia muciniphila co-microcapsules.

[0115] This embodiment provides a β-glucan-Akkermansia muciniphila co-microcapsule, which is similar to the formulation provided in Example 2, except that the volume ratio of the core material:wall material used in this embodiment is 1:1, 1:2, 1:4, 1:6 or 1:8.

[0116] See also Fig.13 As shown in the figure, as the volume ratio of the core material to the wall material decreases, the microcapsule embedding rate increases. When the volume ratio of the two is >1:6, although the embedding rate is high, the number of Akk bacteria per unit is small, and the particle size of the formed particles is large; when the volume ratio of the two is <1:2, due to the low colloid concentration, it is not easy to form particles, resulting in a decrease in the embedding rate. Therefore, 1:2 to 1:6 is selected as the volume ratio range of the core material to the wall material.

[0117] In the above examples and comparative examples, oat β-glucan and yeast β-glucan were prepared by Jiangsu Yanjiang Agricultural Science Institute. Akk bacteria were sourced from Guangdong Microbial Culture Collection Center. Brain heart infusion (BHI) broth was purchased from Beijing Solebow Technology Co., Ltd. Mucin was purchased from Sigma-Aldrich Company, USA.

[0118] Anhydrous CaCl2, sterile PBS, porcine bile salt, sodium carboxymethylcellulose, whey protein, pepsin (1:3000), and pancreatic protein (1:250) were purchased from Beijing Solebow Technology Co., Ltd. Hydrochloric acid and sodium hydroxide, both of analytical grade, were purchased from Sinopharm Chemical Reagent Co., Ltd.

[0119] Yuanmai β-glucan was prepared by the inventors themselves. For the preparation method, please refer to Song Juyi, Liu Jian, Wei Yafeng, Chen Hui. Bioenzymatic preparation process of β-glucan from Yuanmai bran [J]. Packaging and Food Machinery, 2019, 37(04): 10-14.

[0120] The above description is only a preferred embodiment of the present application and an explanation of the technical principles used. Those skilled in the art should understand that the scope of the invention involved in the present application is not limited to the technical solution formed by a specific combination of the above technical features, but should also cover other technical solutions formed by any combination of the above technical features or their equivalent features without departing from the inventive concept. For example, the above features are replaced with the technical features with similar functions disclosed in this application (but not limited to) by each other.

Claims

1. A β-glucan-Akkermansia muciniphila co-microcapsule, characterized in that: It comprises a core material and a wall material, wherein the volume ratio of the core material to the wall material is 1:2-6; The core material includes Akk bacteria and β-glucan, and the concentration of Akk bacteria liquid is 10 8 -10 10 CFU / g, the concentration of β-glucan is 10-40g / L; The wall material comprises sodium carboxymethyl cellulose, whey protein, skim milk powder and trehalose, wherein the mass fraction of the sodium carboxymethyl cellulose is 2% to 3%, the mass fraction of the whey protein is 0.5 to 1.0%, the mass fraction of the skim milk powder is 1%, and the mass fraction of the trehalose is 2.5%.

2. The β-glucan-Akkermansia muciniphila co-microcapsules according to claim 1, characterized in that: The β-glucan is malt β-glucan.

3. A method for preparing β-glucan-Akkermansia muciniphila co-microcapsules according to any one of claims 1 to 2, characterized in that: The following steps are involved: The Akk bacteria are fermented and cultured to a stable stage, and the culture fluid in the Akk bacterial solution is separated to obtain concentrated bacterial mud; The concentrated bacterial sludge is added to a β-glucan solution with a concentration of 10 to 40 g / L and mixed evenly to obtain a core material, so that the final bacterial solution concentration of the core material is 10 10 CFU / g; Adding the wall material to the core material in a volume ratio of 1:2 to 6, and mixing them evenly to obtain a slurry; The slurry is added dropwise into a CaCl2 solution, and wet microcapsules are obtained after solidification and washing; The wet microcapsules are freeze-dried to prepare β-glucan-Ackermansia muciniphila co-microcapsules.

4. The β-glucan-Akkermansia muciniphila co-microcapsules according to claim 3, characterized in that: When the step of separating the culture solution from the Akk bacterial solution is performed, the culture solution is separated and removed by centrifuging the Akk bacterial solution at 7500 to 8500 rpm for 10 minutes.

5. A use of the β-glucan-Akkermansia muciniphila co-microcapsules according to any one of claims 1 to 2, characterized in that: The β-glucan-Akkermansia muciniphila co-microcapsule is used for improving the intestinal flora structure and regulating the intestinal microecology.

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