A composite microbial fermentation agent and its application in the preparation of metagenomic food with multi-cereal as substrate

Abstract

The application discloses a kind of composite microbial fermentation inoculant and its application in metagenomic food preparation with multi-grain as substrate, belong to the field of microbial fermentation technology.The application adds composite microbial fermentation inoculant into multi-grain substrate containing longan meat extract to obtain metagenomic food with multi-grain as substrate.The metagenomic food with multi-grain as substrate is used for animal experiment, and the blood pressure reduction rate of complex group is higher, the fasting blood glucose value and the area under curve are lower, the body weight and TC value are smaller, the proinflammatory cytokines IL-1β, IL-6 and TNF-α are smaller, and the chao and ace index are larger.It is shown that compared with the use of composite microbial fermentation inoculant, longan meat extract or multi-grain substrate alone, the metagenomic food with multi-grain as substrate obtained by the application has good effects of reducing blood pressure, reducing blood sugar and reducing blood lipid, can reduce inflammatory response, remove body toxins and regulate intestinal function.

Description

Technical Field

[0001] This invention relates to the field of microbial fermentation technology, specifically to a compound microbial fermentation agent and its application in the preparation of postbiotic foods based on multigrain substrates. Background Technology

[0002] The "three highs" refers to hypertension, hyperglycemia, and hyperlipidemia. These three conditions often coexist due to metabolic abnormalities and are common chronic diseases in modern society. They are "diseases of affluence" derived from modern life, closely related to unhealthy lifestyle habits such as poor diet, lack of exercise, and irregular sleep patterns. As a metabolic syndrome, the three highs are usually associated with other disease characteristics, such as gastrointestinal metabolic abnormalities or inflammatory responses in the body. In addition to taking medication on time, lifestyle changes are crucial in managing the three highs, such as the often-mentioned "eat less and exercise more," highlighting the importance of a healthy diet for improving bodily functions.

[0003] Compared to refined grains, whole grains are rich in dietary fiber, vitamins, and minerals, making them popular. However, for people with poor digestion or weak spleen and stomach, excessive consumption of whole grains, due to their lower processing level, can actually increase the burden on the digestive tract and is not conducive to alleviating conditions like high blood pressure, high cholesterol, and high blood sugar. Fermented foods are a type of food processed using beneficial microorganisms as fermenting agents. The fermentation process alters the structure of the food, promotes the decomposition of nutrients, and forms new nutritional components. These foods not only have a unique flavor but are also more easily absorbed by the body, making them suitable for people with weaker constitutions.

[0004] Longan ( Dimocarpus longan Lour. is a member of the Sapindaceae family and the genus *Lour.*. Dimocarpus Longan is a large evergreen tree, also known as longan, round-eyed longan, and sheep-eye fruit tree. The fruit is nearly spherical, usually yellowish-brown or sometimes grayish-yellow, with a slightly rough outer surface; the seeds are brownish-red, glossy, and completely covered by a fleshy aril. Flowering occurs in spring and summer, and fruiting in summer. Longan's native range extends from tropical and subtropical Asia to northern Queensland, Australia. Longan fruit is sweet and juicy, with high nutritional value, and is a prized specialty fruit of southern my country. In traditional Chinese medicine, longan pulp is believed to nourish the heart and spleen, replenish qi and blood, strengthen the spleen and stomach, and nourish muscles.

[0005] While there is some awareness of the effects of microbial fermentation agents, whole grains, and longan pulp, the specific effects of using these three substances together to lower blood pressure, blood sugar, and blood lipids, eliminate toxins from the body, and regulate intestinal function have long been unknown. Summary of the Invention

[0006] In view of the above-mentioned prior art, the purpose of this invention is to provide a compound microbial fermentation agent and its application in the preparation of postbiotic foods based on multigrain.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] In a first aspect, the present invention provides a composite microbial fermentation agent, wherein the composite microbial fermentation agent is prepared by the following method:

[0009] A compound microbial fermentation agent was obtained by mixing *Lactobacillus plantarum*, *Pediococcus pentosaceus*, *Bifidobacterium breve*, *Lactobacillus reuteri*, and *Saccharomyces cerevisiae* in a live cell ratio of (0.5-5): (0.5-5): (0.5-5): (0.5-5): (0.5-5).

[0010] Furthermore, the aforementioned *Lactobacillus plantarum* ( Lactiplantibacillus plantarum The strain is *Lactobacillus plantarum* JYZC-LP09, with accession number CGMCC NO.28057; *Pediococcus pentosaceus* ( Pediococcus pentosaceus The accession number for *Bifidobacterium breve* is CICC 23189; Bifidobacterium breve The bacteria identified are *Bifidobacterium breve* WZA-BB07, with accession number CCTCC NO: M 20241530; and *Lactobacillus reuteri* (…). Limosilactobacillus reuteri The strain is *Lactobacillus reuteri* WZA-LR30, with accession number CCTCC NO: M 20241390; *Saccharomyces cerevisiae* (…). Saccharomyces yeast The yeast is Saccharomyces cerevisiae JYZC-SC08, with preservation number CCTCC NO:M 20232528.

[0011] Furthermore, the viable cell count in the compound microbial fermentation agent is 1-9 × 10⁻⁶. 10 cfu / ml.

[0012] A second aspect of the present invention provides the application of the aforementioned compound microbial fermentation agent in the preparation of post-biotic foods based on multigrain substrates, wherein the compound microbial fermentation agent is added to the multigrain substrate for fermentation, as detailed below:

[0013] Take 0.8-1.2 kg of multigrain matrix, add 500 g of water and soak at room temperature for 10-20 h. Take the solid and steam for 20-60 min. During the 20 min steaming, add 200 g of water, spread it out and cool it to 30 ℃. Add 400 g of pure water, 15-30 g of longan pulp extract, and 45-70 g of compound microbial fermentation agent. Ferment at room temperature for 8-32 h. Control the temperature of the fermented material below 35 ℃ during fermentation. After fermentation, sterilize and dry to obtain post-biotic food with multigrain matrix.

[0014] The multigrain matrix includes rice, corn, millet, soybeans, sorghum, buckwheat, oats, red rice, adzuki beans, Job's tears, white lentils, and black sesame seeds.

[0015] Furthermore, the multigrain matrix is ​​prepared by the following method:

[0016] Take whole grains of rice, corn, millet, soybeans, sorghum, buckwheat, oats, red rice, adzuki beans, Job's tears, white lentils and black sesame seeds, dry them and grind them into powder to 200 mesh. Take the same mass of each grain and mix them to obtain a multigrain matrix.

[0017] Furthermore, the longan pulp extract is prepared by the following method:

[0018] Fresh longan pulp was taken, washed, and dried in an oven at 85℃. It was then pulverized and sieved through a 200-mesh sieve to obtain longan pulp powder. The powder was extracted at room temperature with 75% v / v ethanol at a mass-to-volume ratio of 1g:3ml for 12 hours to obtain an extract. The solid remaining after filtration to remove the extract was added to pure water at a mass-to-volume ratio of 1g:10ml and heated rapidly initially, maintaining a gentle boil for 2 hours. After heating was stopped, the mixture was allowed to stand for 40 minutes to remove the solid and aqueous phase, obtaining a secondary extract. The obtained extract and the secondary extract were combined and concentrated under reduced pressure at 60℃ to obtain the longan pulp extract.

[0019] The beneficial effects of this invention are:

[0020] (1) This invention utilizes *Lactobacillus plantarum*, *Pediococcus pentosaceus*, *Bifidobacterium breve*, *Lactobacillus reuteri*, and *Saccharomyces cerevisiae* to prepare a compound microbial fermentation agent, and also prepares longan pulp extract and a multigrain matrix. Experiments show that the compound microbial fermentation agent, longan pulp extract, and multigrain matrix have the effect of lowering blood pressure, blood sugar, and cholesterol, reducing inflammatory response, clearing toxins from the body, and regulating intestinal function.

[0021] (2) In this invention, a compound microbial fermentation agent is added to a multigrain matrix containing longan pulp extract for fermentation to obtain a postbiotic food based on a multigrain matrix. The postbiotic food based on a multigrain matrix was used in animal experiments: In a hypertension animal model experiment, the blood pressure reduction rate in the experimental group was significantly higher than that in the control group. Among them, the blood pressure reduction rate in the compound group was the highest, indicating that adding a postbiotic food based on a multigrain matrix to the feed is superior to using the multigrain matrix, longan pulp extract, or compound probiotics alone. The postbiotic food based on a multigrain matrix of this invention has better effects.

[0022] In the hyperlipidemia animal model experiment, the complex group had the lowest fasting blood glucose level and area under the curve, indicating that adding multigrain-based postbiotic foods to the diet can effectively control the increase in fasting blood glucose and maintain blood glucose homeostasis. The complex group also had the lowest body weight and total cholesterol (TC) value, indicating that adding multigrain-based postbiotic foods to the diet can effectively control the increase in body weight and TC value and maintain relatively stable body weight and blood lipids. The complex group also had the lowest levels of pro-inflammatory cytokines, indicating that adding multigrain-based postbiotic foods to the diet can effectively inhibit the increase of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α and improve the inflammatory response. The complex group also had the highest chao and ace indices, indicating that adding multigrain-based postbiotic foods to the diet can effectively regulate the overall gut microbiota structure of obese mice induced by a high-fat diet. In summary, compared with the use of compound microbial fermentation agents, longan pulp extract, or multigrain matrix alone, the post-biotic food obtained by the compound microbial fermentation agent of this invention with multigrain matrix has good effects on lowering blood pressure, blood sugar, and blood lipids, and can reduce inflammatory response, clear toxins from the body, and regulate intestinal function. Detailed Implementation

[0023] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of this application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0024] To enable those skilled in the art to better understand the technical solution of this application, the technical solution of this application will be described in detail below with reference to specific embodiments.

[0025] Unless otherwise specified, all experimental materials used in the embodiments of this invention are conventional experimental materials in the art and can be purchased through commercial channels. The ordinary feed (D12450J) and high-fat feed (D12492, 60 kcal%) used in this invention were purchased from Research Diets. The C57BL / 6 mice, hypertensive rats (SHR), and original strain Wistar rats used in this invention were provided by Beijing Vital River Laboratory Animal Technology Co., Ltd. Animal welfare was ensured throughout the process. The *Lactobacillus plantarum* used in this invention (… Lactiplantibacillus plantarum The strain is *Lactobacillus plantarum* JYZC-LP09, with accession number CGMCC NO.28057, and has been disclosed in patent "CN117343883B A strain of *Lactobacillus plantarum* and its application"; *Pediococcus pentosaceus* ( Pediococcus pentosaceus The accession number for *Bifidobacterium breve* is CICC 23189, purchased from the China Industrial Microbial Culture Collection Center; *Bifidobacterium breve* ( Bifidobacterium breveThe strain is *Bifidobacterium breve* WZA-BB07, with accession number CCTCC NO: M 20241530, and has been disclosed in patent "CN119193442B A strain of *Bifidobacterium breve* capable of clearing toxins from the body and its metabiotics"; *Lactobacillus reuteri* ( Limosilactobacillus reuteri The strain is *Lactobacillus reuteri* WZA-LR30, with accession number CCTCC NO: M 20241390, and has been disclosed in patent "CN118530915B A strain of *Lactobacillus reuteri* capable of regulating gastrointestinal digestion and its metabiotics"; *Saccharomyces cerevisiae* ( Saccharomyces yeast The yeast is Saccharomyces cerevisiae JYZC-SC08, with accession number CCTCC NO:M 20232528, which has been disclosed in the patent "CN118374421B A compound microbial fermentation agent and its application in improving pancake quality".

[0026] Example 1: Preparation of compound microbial fermentation inoculant

[0027] Lactobacillus plantarum, Pediococcus pentosaceus, Bifidobacterium breve, Lactobacillus reuteri, and Saccharomyces cerevisiae were mixed at the same viable count to form a compound microbial fermentation agent. The viable count of the compound microbial fermentation agent was 8 × 10⁻⁶. 10 cfu / ml.

[0028] Example 2: Preparation of post-biotic foods based on multigrain substrate

[0029] 1. Preparation of multigrain substrate

[0030] Twelve whole grains, including rice, corn, millet, soybeans, sorghum, buckwheat, oats, red rice, adzuki beans, Job's tears, white lentils, and black sesame seeds, were dried and ground into powder to 200 mesh. Equal masses of each grain were mixed to obtain a multigrain matrix.

[0031] 2. Preparation of Longan Pulp Extract

[0032] Fresh longan pulp was washed, dried in an oven at 85℃, and pulverized to a 200-mesh sieve to obtain longan pulp powder. The longan pulp powder was extracted in 75% v / v ethanol at a mass-to-volume ratio of 1g:3ml at room temperature for 12 hours to obtain an extract. The solid remaining after filtration to remove the extract was added to pure water at a mass-to-volume ratio of 1g:10ml and heated rapidly initially, maintaining a gentle boil for 2 hours. After heating was stopped, the mixture was allowed to stand for 40 minutes to remove the solid and aqueous phase, obtaining a secondary extract. The obtained extract and the secondary extract were combined and concentrated under reduced pressure at 60℃ to obtain the longan pulp extract.

[0033] 3. Preparation of post-biotic foods

[0034] Take 1 kg of the multigrain matrix prepared in Example 2, add 500 g of water and soak at room temperature for 10 h, take the solid and cook for 40 min, add 200 g of water when cooking for 20 min, spread it out and cool it to 30 °C, add 400 g of pure water, add 20 g of longan pulp extract prepared in Example 2, add 50 g of compound microbial fermentation agent prepared in Example 1, ferment at room temperature for 16 h, control the temperature of the fermented material to be below 35 °C during fermentation, after fermentation, sterilize by heat treatment at 121 °C for 10 min and then dry to obtain post-biotic food with multigrain matrix.

[0035] Example 3: Preparation of post-biotic foods based on multigrain substrate

[0036] Take 1.2 kg of the multigrain matrix prepared in Example 2, add 500 g of water and soak at room temperature for 10 h, take the solid and cook for 40 min, add 200 g of water when cooking for 20 min, spread it out and cool it to 30 °C, add 400 g of pure water, add 30 g of longan pulp extract prepared in Example 2, add 70 g of compound microbial fermentation agent prepared in Example 1, ferment at room temperature for 16 h, control the temperature of the fermented material to be below 35 °C during fermentation, after fermentation, sterilize by heat treatment at 121 °C for 10 min and then dry to obtain post-biotic food with multigrain matrix.

[0037] Example 4: Preparation of post-biotic foods based on multigrain substrate

[0038] Take 0.8 kg of the multigrain matrix prepared in Example 2, add 500 g of water and soak at room temperature for 10 h, take the solid and steam for 40 min, add 200 g of water when steaming for 20 min, spread it out and cool it to 30 °C, add 400 g of pure water, add 15 g of longan pulp extract prepared in Example 2, add 45 g of compound microbial fermentation agent prepared in Example 1, ferment at room temperature for 16 h, control the temperature of the fermented material to be below 35 °C during fermentation, after fermentation, sterilize by heat treatment at 121 °C for 10 min and then dry to obtain post-biotic food with multigrain matrix.

[0039] Experimental Example 1: Animal Model of Hypertension

[0040] Male, hereditary hypertensive rats (SHR), 11 weeks old and weighing 190–210 g, were selected and randomly divided into 5 groups of 3 rats each. Four groups were experimental groups, and the other group was a control group. Three normal Wistar rats were also selected as the control group. After one week of acclimatization in the same environment, blood pressure was measured and recorded as initial data. The experimental groups were divided into a complex group, a cereal group, an extract group, and a probiotic group. The feed additives for each group were as follows: the complex group was fed the multigrain-based postbiotic food prepared in Example 2; the cereal group was fed the multigrain matrix prepared in Example 2; the extract group was fed the longan pulp extract prepared in Example 2; and the probiotic group was fed the compound microbial fermentation agent prepared in Example 1. Feed additives were added at a rate of 20 mg / kg to the normal diet for each group. The control group was fed the normal diet. Under otherwise identical and suitable conditions, the animals were allowed free access to food. After 15 days of continuous feeding, blood pressure was measured again, and the rate of decrease in blood pressure (%), including the rate of decrease in systolic and diastolic blood pressure, was calculated. The experimental results are shown in Table 1.

[0041] Table 1. Results of the blood pressure lowering experiment

[0042]

[0043] Table 1 shows that the blood pressure reduction rate in the experimental group was significantly higher than that in the control group. The complex group exhibited the highest blood pressure reduction rate, indicating that adding a multigrain-based post-biotic food to the diet is superior to using multigrain-based food, longan pulp extract, or compound probiotics alone. The multigrain-based post-biotic food of this invention has a better blood pressure-lowering effect. Furthermore, normal rats in the control group were fed the same diet as the complex group, and no deaths or other adverse reactions occurred, demonstrating the safety of the multigrain-based post-biotic food of this invention.

[0044] Experimental Example 2: Animal Model Experiment of Hyperlipidemia

[0045] 1. Experimental Methods

[0046] The test animals were 53 male C57BL / 6 mice, SPF grade, 18-22g. After acclimatization for 1 week, the animals were randomly numbered and divided into a normal group (5 mice) and a model group (48 mice). The normal group was fed a normal diet, while the model group was induced to develop NAFLD model by a high-fat diet for 8 weeks. Three mice were randomly selected from each of the normal group and the model group. The success of the model was judged by weight. The judgment criterion was that the weight of the mice in the model group was 20% higher than that in the normal group.

[0047] C57BL / 6 mice that successfully developed the model were selected for subsequent experiments. The experimental groups were divided into a complex group, a cereal group, an extract group, a probiotic group, and a postbiotic group, with three mice in each group and three replicates. The feed additives for each group were as follows: the complex group was fed the postbiotic food prepared in Example 2 with a multi-grain matrix; the cereal group was fed the multi-grain matrix prepared in Example 2; the extract group was fed the longan pulp extract prepared in Example 2; and the probiotic group was fed the compound microbial fermentation agent prepared in Example 1. Feed additives were added at a rate of 20 mg / kg to the high-fat diet for each group. The control group was fed a high-fat diet. All other conditions were the same and suitable, and mice had free access to food.

[0048] Blood glucose was measured in mice at week 15 of drug administration, and body weight was measured at week 16. Samples were collected (blood from the apex of the heart, liver, and cecal contents were collected and fixed in 4% paraformaldehyde or flash-frozen in liquid nitrogen and stored at -80°C as needed).

[0049] 2. Indicator Measurement

[0050] Blood glucose, total cholesterol (TC), and pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) in mice were measured using a kit according to the manufacturer's instructions. Total bacterial DNA was extracted from the cecal contents of mice using a DNA extraction kit, and amplification was performed targeting the V3-V4 region of the 16S rRNA gene. The resulting PCR amplifications were quantified, homogenized, and mixed before library construction. Sequencing was performed using the Illumina Miseq platform. Optimized sequences were classified into OTUs based on 97% similarity, and the abundance of gut microbiota was analyzed at each taxonomic level.

[0051] 3. Experimental Results

[0052] 3.1 Measurement of blood glucose in mice

[0053] Fasting blood glucose levels were measured by collecting blood from the tail vein. A model of diabetes was considered successfully established when the blood glucose level was >11.1 mmol / L, and an oral glucose tolerance test was then conducted. All mice were fasted for 12 hours the night before the test, but allowed free access to water, and fasting blood glucose was measured. Subsequently, each group was administered 1.0 g / kg of glucose according to their body weight, and blood glucose levels were measured at 30 min, 60 min, and 120 min, with the area under the curve (AUC) calculated.

[0054] Area under the curve (mmol / L) = (0.5(A+B)+1.5(B+C)) / 2;

[0055] In the formula: A, B, and C represent the blood glucose values ​​at 0 min, 30 min, and 120 min, respectively.

[0056] Higher fasting blood glucose levels and larger area under the curve indicate poorer blood glucose regulation. The effects of fasting blood glucose levels and area under the curve in the oral glucose tolerance test on the five groups of mice are shown in Table 2.

[0057] Table 2. Results of fasting blood glucose levels and area under the curve.

[0058]

[0059] Table 2 shows that the fasting blood glucose level and area under the curve in the experimental group were significantly lower than those in the control group. Among them, the fasting blood glucose level and area under the curve in the complex group were the lowest. This indicates that adding a multigrain-based post-biotic food to the feed is superior to using multigrain-based food, longan extract, or compound probiotics alone. It can effectively control the rise in fasting blood glucose and maintain blood glucose homeostasis. The multigrain-based post-biotic food of this invention has a better hypoglycemic effect.

[0060] 3.2 Effects on mouse body weight and blood lipids

[0061] A high-fat diet typically resulted in a significant increase in body weight and elevated serum total cholesterol (TC) levels in mice. Serum TC is the sum of all cholesterol in the blood; higher serum TC levels correlate with greater obesity. Body weight and blood lipid results are shown in Table 3.

[0062] Table 3. Weight and blood lipid results

[0063]

[0064] Table 3 shows that the body weight and TC value of the experimental group were significantly lower than those of the control group. Among them, the body weight and TC value of the complex group were the lowest. This indicates that adding post-biotic foods with multigrain as the matrix to the feed is better than using multigrain matrix, longan pulp extract or compound probiotics alone. It can effectively control the increase of body weight and TC value and maintain the relative stability of body weight and blood lipids. The post-biotic foods with multigrain as the matrix of this invention have a better blood lipid-lowering effect.

[0065] 3.3 Effects on chronic inflammation

[0066] Inflammatory cytokines are small protein molecules secreted by immune and non-immune cells, and are classified into pro-inflammatory and anti-inflammatory types. Pro-inflammatory cytokines mediate inflammatory responses by activating immune cells. A high-fat diet easily leads to liver damage such as fatty liver, triggering inflammatory responses in the liver or systemic system of mice, affecting the body's health. IL-1β, IL-6, and TNF-α are pro-inflammatory cytokines; the higher their levels, the more severe the inflammatory response. Inflammatory markers in mice are shown in Table 4.

[0067] Table 4 Inflammatory markers in mice

[0068]

[0069] Table 4 shows that the levels of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α in the experimental group were significantly lower than those in the control group. Among them, the complex group had the lowest levels of pro-inflammatory cytokines. This indicates that adding a multigrain-based postbiotic food to the feed is superior to using multigrain-based food, longan pulp extract, or compound probiotics alone. It can effectively inhibit the increase of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α and improve the inflammatory response. The multigrain-based postbiotic food of this invention has a better effect on reducing the inflammatory response and clearing toxins from the body.

[0070] 3.4 Effects on the gut microbiota of mice

[0071] Alpha diversity is an analysis of species diversity in a single sample, including the chao index and ace index. Both are used to assess the richness of the community in a sample; the higher the index, the richer the species content of the sample. The effects of each group on the alpha diversity of the gut microbiota are shown in Table 5.

[0072] Table 5. Effects of each group on the Alpha diversity of gut microbiota

[0073]

[0074] Table 5 shows that the control group had lower chao and ace indices, indicating that a high-fat diet significantly reduced the species richness of the gut microbiota in mice. The experimental groups had significantly higher chao and ace indices than the control group, with the complex group exhibiting the highest chao and ace indices. This indicates that adding a multigrain-based postbiotic food to the diet is superior to using multigrain-based foods, longan extract, or compound probiotics alone in regulating the overall gut microbiota structure of obese mice induced by a high-fat diet. The multigrain-based postbiotic food of this invention has a better effect on regulating intestinal function.

[0075] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. The application of compound microbial fermentation agents in the preparation of post-biotic foods based on multigrain substrates, characterized in that, The compound microbial fermentation agent is added to a multigrain substrate for fermentation, as detailed below: Take 0.8-1.2 kg of multigrain matrix, add 500 g of water and soak at room temperature for 10-20 h. Take the solid and steam for 20-60 min. During the 20 min steaming, add 200 g of water, spread it out and cool it to 30 ℃. Add 400 g of pure water, 15-30 g of longan pulp extract, and 45-70 g of compound microbial fermentation agent. Ferment at room temperature for 8-32 h. Control the temperature of the fermented material below 35 ℃ during fermentation. After fermentation, sterilize and dry to obtain post-biotic food with multigrain matrix. The multigrain matrix includes rice, corn, millet, soybeans, sorghum, buckwheat, oats, red rice, adzuki beans, Job's tears, white lentils, and black sesame seeds; The aforementioned compound microbial fermentation agent is prepared by the following method: A compound microbial fermentation agent was obtained by mixing *Lactobacillus plantarum*, *Pediococcus pentosaceus*, *Bifidobacterium breve*, *Lactobacillus reuteri*, and *Saccharomyces cerevisiae* at a live cell count ratio of (0.5-5): (0.5-5): (0.5-5): (0.5-5): (0.5-5). The aforementioned Lactobacillus plantarum ( Lactiplantibacillus plantarum The strain is *Lactobacillus plantarum* JYZC-LP09, with accession number CGMCC NO.28057; *Pediococcus pentosaceus* ( Pediococcus pentosaceus The accession number for *Bifidobacterium breve* is CICC23189; Bifidobacterium breve The bacteria identified are *Bifidobacterium breve* WZA-BB07, with accession number CCTCCNO:M 20241530; and *Lactobacillus reuteri* (…). Limosilactobacillus reuteri The strain is *Lactobacillus reuteri* WZA-LR30, with accession number CCTCC NO: M 20241390; *Saccharomyces cerevisiae* (…). Saccharomyces cerevisiae The yeast is Saccharomyces cerevisiae JYZC-SC08, with preservation number CCTCC NO:M 20232528; The longan pulp extract is prepared by the following method: Fresh longan pulp was taken, washed, and dried in an oven at 85℃. It was then pulverized and sieved through a 200-mesh sieve to obtain longan pulp powder. The powder was extracted at room temperature with 75% v / v ethanol at a mass-to-volume ratio of 1g:3ml for 12 hours to obtain an extract. The solid remaining after filtration to remove the extract was added to pure water at a mass-to-volume ratio of 1g:10ml and heated rapidly initially, maintaining a gentle boil for 2 hours. After heating was stopped, the mixture was allowed to stand for 40 minutes to remove the solid and aqueous phase, obtaining a secondary extract. The obtained extract and the secondary extract were combined and concentrated under reduced pressure at 60℃ to obtain the longan pulp extract.

2. The application of the compound microbial fermentation agent according to claim 1 in the preparation of post-biotic foods based on multigrain substrates, characterized in that, The viable cell count in the compound microbial fermentation agent is 1-9 × 10⁻⁶. 10 cfu / ml.

3. The application of the compound microbial fermentation agent according to claim 1 in the preparation of post-biotic foods based on multigrain substrates, characterized in that, The multigrain matrix is ​​prepared by the following method: Take whole grains of rice, corn, millet, soybeans, sorghum, buckwheat, oats, red rice, adzuki beans, Job's tears, white lentils and black sesame seeds, dry them and grind them into powder to 200 mesh. Take the same mass of each grain and mix them to obtain a multigrain matrix.

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