A tea extract, its preparation and use

Abstract

The present application relates to the field of tea processing and compound extraction preparation, in particular to a tea extract and its preparation method and application. The present application provides a preparation method of a tea extract, comprising the following steps: mixing a tea suspension and a bovine serum albumin solution to obtain a mixed solution, and extracting the mixed solution after reaction to obtain the tea extract; wherein the volume ratio of the tea suspension and the bovine serum albumin solution is (45-55):1; the tea concentration of the tea suspension is 0.18-0.22 g / ml, and the bovine serum albumin solution concentration is 45-50 mg / ml; the reaction temperature of the mixed solution is 36-38 DEG C, and the reaction time is 2-2.5 h. The tea extract obtained by the present application has high polyphenol and protein content, can directionally enrich g_Ileibacterium in the intestinal tract, can significantly improve intestinal disorders, and can promote intestinal health.

Description

Technical Field

[0001] This invention relates to the field of tea processing and complex extraction and preparation, specifically to a tea extract, its preparation method, and its application. Background Technology

[0002] The health benefits of tea are widely recognized. Active substances in tea (such as tea polyphenols and tea polysaccharides) can interact with the gut microbiota and participate in regulating human health. Existing research confirms that gut microbiota dysbiosis is closely related to digestive system diseases, metabolic diseases, immune system diseases, neurodegenerative diseases, and obesity. The establishment of mechanistic models such as the gut-microbiota-lung axis and the gut-microbiota-brain axis further reveals the bidirectional communication patterns between the gut microbiota and other organs. The regulatory effects of tea and tea extracts on gut microbiota may be an important mechanism influencing its health effects.

[0003] Existing research confirms that tea can regulate various microbial communities, including Bacteroidetes, Firmicutes, and Proteobacteria. However, due to the complex composition of tea, which includes tea polyphenols, proteins, sugars, and alkaloids, the response of microbial communities to different tea extracts is difficult to predict.

[0004] Bacteria in the order Erysipelotrichales of the phylum Firmicutes have been identified as butyrate-producing bacteria. Among them, *G. leibacterium* is a probiotic that exerts its beneficial effects in high-fat diet-induced fatty liver disease in mice by reducing serum lipopolysaccharide and branched-chain amino acid levels. *G. leibacterium* can also be used to regulate gastrointestinal and metabolic diseases. However, there are currently no tea extracts available that can specifically regulate *G. leibacterium*. Summary of the Invention

[0005] Therefore, the technical problem to be solved by the present invention is to overcome the deficiency of the prior art in that there is no tea extract with directional controllable g_Ileibacterium, thereby providing a tea extract, its preparation method and application.

[0006] This invention provides a method for preparing tea extract, comprising the following steps: mixing tea suspension and bovine serum albumin solution to obtain a mixture, and extracting the mixture after reaction to obtain the tea extract;

[0007] The volume ratio of the tea suspension to the bovine serum albumin solution is (45-55):1; the tea concentration of the tea suspension is 0.18-0.22 g / ml, and the concentration of the bovine serum albumin solution is 45-50 mg / ml.

[0008] The reaction temperature of the mixture is 36-38℃, and the reaction time is 2-2.5h.

[0009] Preferably, the tea suspension is prepared by grinding tea leaves in water to obtain the tea suspension.

[0010] Optionally, the grinding is performed using a grinding machine.

[0011] Preferably, the tea leaves are fresh tea leaves that have been withered;

[0012] Optionally, the temperature for withering fresh tea leaves is 25-30℃, the withering time is 8-10 hours, and the moisture content of the withered tea leaves is 55-60 wt%.

[0013] In this invention, fresh tea leaves refer to fresh tea leaves that have not been processed after being picked.

[0014] Preferably, the fresh tea leaves are fresh spring tea leaves;

[0015] More preferably, the tea leaves are fresh tea leaves picked in April or May.

[0016] Preferably, the fresh tea leaves are one bud with three to four leaves.

[0017] Preferably, the extraction step involves soaking and filtering the reaction mixture to obtain an extract, then concentrating the extract to obtain a concentrate, and finally subjecting the concentrate to alcohol precipitation, filtration, and drying to obtain the tea extract.

[0018] Optionally, the drying process includes freeze drying.

[0019] Preferably, the extraction process involves adding water for extraction;

[0020] And / or, the volume ratio of water added to the mixture is (1-1.2):1;

[0021] And / or, the extraction temperature is 70-75℃, and the time is 1-1.5h.

[0022] Preferably, the volume of the concentrate is (0.08-0.15) times the volume of the extract;

[0023] And / or, alcohol precipitation includes the process of first alcohol precipitation, filtration, dissolution, second concentration, and second alcohol precipitation of the concentrate;

[0024] Optionally, the volume ratio of ethanol solution to concentrate in the first alcohol precipitation is (4-5):1, the temperature of the first alcohol precipitation is 2-10℃, the time is 20-25h, and the concentration of ethanol solution is 90-100v / v%.

[0025] Optionally, the dissolution process involves dissolving the filtered product in water to obtain a solution, wherein the mass ratio of the filtered product to water is 1:(10-15); and the dissolution temperature is 70-75℃.

[0026] Optionally, the solution is concentrated a second time to obtain a second concentrate, and the volume ratio of the second concentrate to the solution is (0.08-0.15):1.

[0027] Optionally, the volume ratio of ethanol solution to the second concentrate in the second alcohol precipitation is (4-5):1, the temperature of the second alcohol precipitation is 2-10℃, the time is 20-25h, and the concentration of the ethanol solution is 90-100v / v.

[0028] The present invention also provides a tea extract, which is prepared by the preparation method described above.

[0029] Preferably, the tea extract comprises, by mass percentage, the following components: 20-25% polyphenols, 25-30% protein, 15-20% uronic acid, and 15-20% neutral sugars.

[0030] This invention provides a tea extract prepared by the above-described method, or the tea extract described above has any of the following applications:

[0031] (1) Application in targeted regulation of g_Ileibacterium bacteria;

[0032] (2) Application in the preparation of products for maintaining intestinal homeostasis, improving intestinal disorders, and preventing gastrointestinal diseases and / or metabolic diseases.

[0033] Preferably, the regulation of g_Ileibacterium bacteria includes increasing the abundance of g_Ileibacterium bacteria in the gut;

[0034] And / or, the gastrointestinal diseases include those caused by intestinal tissue damage, intestinal flora imbalance and / or decreased total short-chain fatty acid content; optionally, the intestinal flora imbalance includes a decrease in the abundance of g_Ileibacterium bacteria in the intestine;

[0035] And / or, the gastrointestinal disease includes alcohol-induced gastrointestinal disease; optionally, the gastrointestinal disease includes alcohol-induced acute gastrointestinal disease.

[0036] The technical solution of this invention has the following advantages:

[0037] This invention provides a method for preparing tea extract, comprising the following steps: mixing tea suspension and bovine serum albumin solution to obtain a mixture, and extracting the mixture after reaction to obtain the tea extract; wherein, the mass ratio of tea to bovine serum albumin in the mixture is (180-220):1; the reaction temperature of the mixture is 36-38℃, and the reaction time is 2-2.5h.

[0038] In this invention, tea suspension and bovine serum albumin solution are mixed to obtain a mixture, wherein the mass ratio of tea to bovine serum albumin in the mixture is defined as (180-220):1. The mixture is then reacted at 36-38°C for 2-2.5 hours. The addition of an appropriate amount of bovine serum albumin can better complex polyphenols and proteins, thereby obtaining a specific tea extract with high polyphenol and protein content. This extract can be selectively enriched in the intestines by g-Ileibacterium, which can significantly improve intestinal disorders and promote intestinal health. Attached Figure Description

[0039] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0040] Figure 1 This is a graph showing the relationship between body weight and time in healthy mice treated with different doses of tea extract from Example 1 of this invention in Experiment 1;

[0041] Figure 2 This is a graph showing the relationship between changes in gut microbial community composition at the genus level and time in healthy mice treated with different doses of tea extract from Example 1 of this invention.

[0042] Figure 3 This is a diagram showing the gut microbiota community composition at the genus level of healthy mice after being treated with physiological saline, tea extracts from Example 1 and Comparative Examples 1 and 2 for 2 weeks in Experimental Example 2 of the present invention.

[0043] Figure 4 This is a H&E staining image and HI score of the proximal colon of mice after alcohol exposure, which were treated with physiological saline, tea extract of Comparative Example 1, tea extract of Comparative Example 2, and tea extract of Example 1 for 4 weeks in Experiment 3 of this invention.

[0044] Figure 5This is a graph showing the relative abundance of gut microbiota at the genus level in mice treated with physiological saline, tea extract of Comparative Example 1, tea extract of Comparative Example 2, and tea extract of Example 1 for 4 weeks after alcohol exposure in Experimental Example 3 of this invention.

[0045] Figure 6 This is a graph showing the total short-chain fatty acid content in the intestines of mice after alcohol exposure in Experimental Example 3 of this invention, after being treated with physiological saline, tea extract of Comparative Example 1, tea extract of Comparative Example 2, and tea extract of Example 1 for 4 weeks. Detailed Implementation

[0046] The following embodiments are provided to better understand the present invention and are not limited to the preferred embodiments described. They do not constitute a limitation on the content and scope of protection of the present invention. Any product that is the same as or similar to the present invention, derived by any person under the guidance of the present invention or by combining the features of the present invention with other prior art, falls within the protection scope of the present invention.

[0047] For experiments not specifically described in the examples, the procedures or conditions should be followed according to the conventional experimental procedures described in the literature in this field. Reagents or instruments whose manufacturers are not specified are all commercially available conventional reagent products.

[0048] The fresh tea leaves mentioned in this embodiment of the invention are Longjing 43, with one bud and three or four leaves, and are harvested from Hangzhou City, Zhejiang Province.

[0049] Example 1

[0050] This embodiment provides a method for preparing tea extract, including the following steps:

[0051] (1) 100g of fresh Longjing 43 tea leaves with one bud and three leaves were spread out in a withering trough and withered at 26℃ for 8 hours to obtain tea leaves with a moisture content of 58.63%.

[0052] (2) The withered tea leaves from step (1) and 5 times their weight of water are mixed and ground in a grinder to obtain a tea suspension.

[0053] (3) Add 10 mL of bovine serum albumin solution with a concentration of 50 mg / mL to 500 mL of tea suspension obtained in step (2) to form a mixture. Place the mixture in a magnetically stirred water bath and react at 37°C for 2 h. Then add an equal volume of water to the solution after reaction and extract at 70°C in a water bath for 1 h. After filtration, take the filtrate, which is the extract.

[0054] (4) The extract obtained in step (3) is concentrated to 1 / 10 of its original volume using a rotary evaporator to obtain a concentrated solution. Four times its volume of 95% v / v ethanol aqueous solution is added to the concentrated solution and precipitated at 8°C for 22 hours. After filtration, the filtrate is dissolved in 10 times its weight of water at 70°C to obtain a solution. The solution is then concentrated a second time to 1 / 10 of its volume to obtain a second concentrated solution. Four times its volume of 95% ethanol aqueous solution is added to the second concentrated solution for precipitation (at 8°C for 22 hours). After filtration and freeze-drying, the tea extract is obtained.

[0055] Example 2

[0056] This embodiment provides a method for preparing tea extract, including the following steps:

[0057] (1) 100g of fresh Longjing 43 tea leaves with one bud and three leaves were spread out in a withering trough and withered at 28℃ for 10h to obtain tea leaves with a moisture content of 56.18%.

[0058] (2) The tea leaves after withering in step (1) and 4.5 times their weight of water are mixed and ground in a grinder to obtain a tea suspension;

[0059] (3) Add 10 mL of bovine serum albumin solution with a concentration of 50 mg / mL to 450 mL of tea suspension obtained in step (2) to form a mixture. Place the mixture in a magnetically stirred water bath and react at 36°C for 2.5 h. Then add an equal volume of water to the solution after reaction and extract at 70°C in a water bath for 1 h. After filtration, take the filtrate, which is the extract.

[0060] (4) The extract obtained in step (3) is concentrated to 0.08 times its original volume using a rotary evaporator to obtain a concentrated solution. Four times its volume of 100 v / v ethanol is added to the concentrated solution and precipitated at 2°C for 25 h. After filtration, the filtrate is dissolved in 15 times its weight of water at 75°C to obtain a solution. The solution is then concentrated a second time to 0.08 times its volume to obtain a second concentrated solution. Five times its volume of 100% ethanol is added to the second concentrated solution for precipitation (at 2°C for 25 h). After filtration and freeze-drying, the tea extract is obtained.

[0061] Example 3

[0062] This embodiment provides a method for preparing tea extract, including the following steps:

[0063] (1) 100g of fresh Longjing 43 tea leaves with one bud and three leaves were spread out in a withering trough and withered at 30℃ for 9 hours to obtain tea leaves with a moisture content of 55.24%.

[0064] (2) The tea leaves after withering in step (1) and 5.5 times their weight of water are mixed and ground in a grinder to obtain a tea suspension;

[0065] (3) Add 10 mL of bovine serum albumin solution with a concentration of 45 mg / mL to 550 mL of tea suspension obtained in step (2) to form a mixture. Place the mixture in a magnetically stirred water bath and react at 38°C for 2.2 h. Then add 1.2 times the volume of water to the solution after reaction and extract in a water bath at 75°C for 1.5 h. After filtration, take the filtrate, which is the extract.

[0066] (4) The extract obtained in step (3) is concentrated to 0.15 times its original volume using a rotary evaporator to obtain a concentrated solution. Five times its volume of 90 v / v% ethanol aqueous solution is added to the concentrated solution and precipitated at 10°C for 20 h. After filtration, the filtrate is dissolved in 10 times its weight of water at 70°C to obtain a solution. The solution is then concentrated a second time to 0.15 times its volume to obtain a second concentrated solution. Five times its volume of 95% ethanol aqueous solution is added to the second concentrated solution for precipitation (precipitated at 8°C for 22 h). After filtration and freeze-drying, the tea extract is obtained.

[0067] Comparative Example 1

[0068] The difference between this comparative example and Example 1 is that in step (3), the mixture is placed in a magnetically stirred water bath and reacted at 37°C for 1 hour.

[0069] Comparative Example 2

[0070] The difference between this comparative example and Example 1 is that, in step (3), the volume of bovine serum albumin solution added is 5 ml.

[0071] Comparative Example 3

[0072] The difference between this comparative example and Example 1 is that bovine serum albumin solution is not added in step (3).

[0073] Test case

[0074] The composition of tea extracts prepared in Examples 1 and 2 and Comparative Examples 1, 2, and 3 was determined. Neutral sugar content was determined using anhydrous glucose as a standard via the anthrone-sulfuric acid method; uronic acid content was determined using galacturonic acid as a standard via the sulfuric acid-carbazole method; total phenolic content was determined using gallic acid (GA) as a standard according to GB / T 8313-2018 (ISO 14502-1:2005); and protein content was determined using bovine serum albumin as a standard via the Bradford method. The results are shown in Table 1.

[0075] Table 1 Test Results

[0076]

[0077] Experimental Example 1: The regulatory effect of the tea extract of the present invention on the intestinal flora of animals.

[0078] Animal experiments: Forty 3-week-old male C57BL / 6J mice were used and housed in a pathogen-free environment with a 12-hour light / dark cycle, relative humidity of 50-60%, and temperature of 22-26°C. After one week of acclimatization, the mice were randomly divided into four groups (n=10 per group): control group (saline), low-dose group (100 mg / kg / day), medium-dose group (200 mg / kg / day), and high-dose group (300 mg / kg / day). Each group was administered the corresponding dose of the tea extract from Example 1 by gavage daily, while the control group was administered an equal volume of saline by gavage. All groups were provided with normal drinking water and normal feed. Mouse body weight was monitored for 4 weeks, and fecal samples were collected weekly for 16S rDNA high-throughput sequencing.

[0079] Animal experiment results:

[0080] (1) Changes in mouse body weight after oral administration of different doses of tea extract for 4 weeks are as follows: Figure 1 The mouse body weight change curves showed no significant difference in body weight between the low-dose, medium-dose, and high-dose groups and the control group. Similarly, there were no significant differences in body weight between the corresponding low-dose, medium-dose, and high-dose groups. This indicates that tea extract does not affect the body weight change in mice.

[0081] (2) The trends in the composition of the microbial community of mice in the control group, low-dose group, medium-dose group and high-dose group over 4 weeks are as follows: Figure 2(The relative abundance of g_Ileibacterium is shown in the dashed box in the figure.) The microbial communities of mice corresponding to the control group, low-dose group, medium-dose group, and high-dose group were significantly enriched with g_Ileibacterium. The enrichment level in the low-dose group was significantly lower than that in the medium-dose and high-dose groups. There was no significant difference between the medium-dose and high-dose groups. Furthermore, the relative abundance of g_Ileibacterium tended to stabilize after two weeks of gavage. This indicates that the tea extract of Example 1 can directionally enrich g_Ileibacterium, with an optimal concentration of 200 mg / kg / day and a duration of action exceeding two weeks.

[0082] Experimental Example 2: Application of the tea extracts of the present invention and comparative examples in the regulation of g-Ileibacterium in the animal intestine.

[0083] Animal experiments: Forty 3-week-old male C57BL / 6J mice were used and housed in a pathogen-free environment with a 12-hour light / dark cycle, relative humidity of 50-60%, and temperature of 22-26°C. After one week of acclimatization, the mice were randomly divided into four groups (n=10 per group): control group (saline), comparative example group 1 (200 mg / kg / d), comparative example group 2 (200 mg / kg / d), and example group 1 (200 mg / kg / d). Each group was administered the corresponding tea extract by gavage daily, while the control group was administered an equal volume of saline by gavage. All groups were provided with normal drinking water and regular feed. Two weeks after gavage, mouse feces were collected for 16S rDNA high-throughput sequencing.

[0084] Animal experiment results: The trend of changes in the composition of the gut microbiota in mice after 2 weeks is as follows: Figure 3 As shown (the relative abundance of g_Ileibacterium is indicated by the dashed box in the figure), by Figure 3 It can be seen that g_Ileibacterium was significantly enriched in the mice of Example 1 group, confirming that the tea extract with a specific composition ratio provided by the present invention can directionally enrich g_Ileibacterium.

[0085] Experimental Example 3: Application of the tea extracts of the present invention and comparative examples in protecting animal intestinal homeostasis.

[0086] Animal Experiment: Fifty 3-week-old male C57BL / 6J mice were used and housed in a pathogen-free environment with a 12-hour light / dark cycle, relative humidity of 50-60%, and temperature of 22-26°C. After one week of acclimatization, the mice were randomly divided into 5 groups (n=10 per group): control group (saline), alcohol exposure group (saline), comparative example 1 (200 mg / kg / d), comparative example 2 (200 mg / kg / d), and example 1 (200 mg / kg / d). Each treatment group was administered the corresponding tea extract by gavage daily, while the control group and alcohol exposure group were administered an equal volume of saline by gavage. All groups were provided with normal drinking water and normal feed. After 4 weeks of pretreatment, the alcohol exposure group and the three tea extract treatment groups were administered 50% alcohol (13 mL / kg) by gavage once, while the control group was administered an equal volume of saline by gavage as a control. After the alcohol was administered via gavage, behavioral observations were conducted on the mice in each group. The righting reflex test was used to determine the mice's level of intoxication. The number of intoxicated mice, the number of deaths, and the duration and recovery time of the intoxicated mice were recorded. Water and feed were provided as usual during this period, and changes in mouse body weight after alcohol administration were monitored. All mice were sacrificed 24 hours after alcohol administration, and feces were collected before sacrifice for 16S rDNA high-throughput sequencing. After sacrifice, the proximal colon was fixed in 4% formalin for 24 hours, and the tissue was then placed in paraffin for hematoxylin and eosin (H&E) staining. Tissue sections were observed under a microscope. The remaining colonic tissue was washed with PBS, and colonic contents were collected for short-chain fatty acid determination.

[0087] Experimental results:

[0088] (1) After alcohol gavage treatment, the intoxication status of mice in each group was observed. Among the 50 mice treated with alcohol, 42 were intoxicated and 2 died. The average intoxication time of the intoxicated mice was 32 min, and the average sobering time was 6.5 h. The body weight of the mice in the alcohol exposure group was significantly reduced by 12.6% compared with the control group. The weight loss of each tea extract treatment group recovered to varying degrees, but there was no significant difference between the treatment groups.

[0089] (2) Histopathological sections of the proximal colon of mice, as shown in... Figure 4 As shown, tea extract significantly restored the structure of the colonic mucosa and glands, resulting in reduced inflammatory cell infiltration. Histological damage index (HI) assessment results showed that all tea extract treatment groups exhibited significant efficacy in restoring colonic tissue integrity, with Example 1 showing the best efficacy.

[0090] (3) Figure 5(The relative abundance of g_Ileibacterium is shown in the dashed box in the figure.) This illustrates the relative abundance of microbial species at the genus level in each group. Compared to the control group, the alcohol exposure group significantly reduced the abundance of Lachnospiraceae_NK4A136_group and significantly enriched the pathogenic bacteria Helicobacter and Escherichia-Shigella. Comparative Example 1, Comparative Example 2, and Example 1 significantly increased the relative abundance of norank_f_Muribaculaceae and significantly inhibited the enrichment of Helicobacter and Escherichia-Shigella. Among them, Example 1 significantly enriched g_Ileibacterium.

[0091] (4) Figure 6 The total short-chain fatty acid (SCFA) content in the intestines of mice in each group was calculated from... Figure 6 It was found that tea extract pretreatment significantly restored the decrease in total SCFAs in the intestine of mice caused by acute alcohol exposure. Among them, the total SCFAs in the Example 1 group were the best, and even significantly better than those in the control group.

[0092] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A method to enhance gut microbiota g_Ileibacterium A method for preparing tea extract with high bacterial abundance, characterized in that, The process includes the following steps: mixing tea suspension and bovine serum albumin solution to obtain a mixture, and then extracting the mixture after the reaction to obtain the tea extract; The volume ratio of the tea suspension to the bovine serum albumin solution is (45-55):1; the tea concentration of the tea suspension is 0.18-0.22 g / ml, and the concentration of the bovine serum albumin solution is 45-50 mg / ml. The reaction temperature of the mixture is 36-38℃, and the reaction time is 2-2.5 h; The extraction steps are as follows: the mixture after reaction is extracted and filtered to obtain an extract, the extract is then concentrated to obtain a concentrate, and the concentrate is subjected to alcohol precipitation, filtration, and drying to obtain the tea extract. The tea extract comprises, by weight percentage, the following components: 20-25% polyphenols, 25-30% protein, 15-20% uronic acid, and 15-20% neutral sugars. The tea leaves referred to are fresh tea leaves that have undergone withering. The extraction process involves adding water for extraction.

2. The preparation method according to claim 1, characterized in that, The tea suspension is prepared by grinding tea leaves in water to obtain the tea suspension.

3. The preparation method according to claim 1, characterized in that, The temperature for withering fresh tea leaves is 25-30℃, the withering time is 8-10 hours, and the moisture content of the withered tea leaves is 55-60 wt%.

4. The preparation method according to claim 1, characterized in that, The drying process includes freeze drying.

5. The preparation method according to claim 1, characterized in that, The volume ratio of water added to the mixture is 1-1.2:1; And / or, the extraction temperature is 70-75℃, and the time is 1-1.5 h.

6. The preparation method according to claim 1, characterized in that, The volume of the concentrate is (0.08-0.15) times the volume of the extract; And / or, alcohol precipitation includes the process of first alcohol precipitation, filtration, dissolution, second concentration, and second alcohol precipitation of the concentrate.

7. The preparation method according to claim 6, characterized in that, The volume ratio of ethanol solution to concentrate in the first alcohol precipitation is (4-5):1, the temperature of the first alcohol precipitation is 2-10℃, the time is 20-25h, and the concentration of ethanol solution is 90-100v / v.

8. The preparation method according to claim 6, characterized in that, The dissolution process involves dissolving the filtered product in water to obtain a solution, wherein the mass ratio of the filtered product to water is 1:(10-15); the dissolution temperature is 70-75℃.

9. The preparation method according to claim 8, characterized in that, The solution was concentrated a second time to obtain a second concentrate, and the volume ratio of the second concentrate to the solution was (0.08-0.15):

1.

10. The preparation method according to claim 6, characterized in that, The volume ratio of ethanol solution to the second concentrate in the second alcohol precipitation is (4-5):

1. The temperature of the second alcohol precipitation is 2-10℃, the time is 20-25h, and the concentration of the ethanol solution is 90-100 v / v.

11. A method to enhance gut microbiota g_Ileibacterium The abundance of bacteria in tea extract, characterized by: The tea extract is prepared by the preparation method according to any one of claims 1-10.

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