Composition for improving liver inflammation and preparation method thereof

Abstract

The present invention relates to a synbiotic health food containing green tea polyphenols extract and a preparation method thereof, wherein the synbiotic health food contains the following ingredients: prebiotics, probiotics and green tea polyphenols extract, wherein the prebiotics contain one or more of lactose and isomaltooligosaccharide; the probiotics contain one or more of rhamnosus Bacillus GG and Lactobacillus casei YRL577, and the probiotic strains are also cultured and amplified; the green tea polyphenols extract is obtained by extracting and separating green tea leaves, and its ingredients contain one or more of epigallocatechin gallate (EGCG), epicatechin gallate (ECG), gallic acid (EGC), and epicatechin (EC). The synbiotic health food containing tea polyphenols extract provided by the present invention can effectively improve the symptoms of hyperlipidemia, can regulate intestinal flora, help alleviate different liver inflammations and injuries, and improve physical health.

Description

Technical Field

[0001] The present invention relates to a composition for improving liver inflammation and a preparation method thereof. Background Art

[0002] Synbiotic products are a class of products that regulate human physiological functions, prevent diseases, and promote overall health. Common main ingredients include probiotics and prebiotics. While ensuring product safety, these ingredients can provide the human body with essential nutrients, enhance immunity, improve the balance of intestinal flora, and help prevent and improve various diseases. Combining natural plant extracts with prebiotic fermentation broth is a common R&D approach. Tea extract, as a natural compound with multiple biological activities, can synergize with other ingredients in synbiotics to enhance product efficacy. Green tea extract, compared to other teas, contains higher levels of tea polyphenols, such as tea polyphenols and epi-tea polyphenols. These tea polyphenols have the potential to lower blood lipids, regulate intestinal flora, and protect intestinal tissue, making them suitable for the development of functional products.

[0003] However, since the content of tea polyphenols in green tea extract is still low and the bioavailability is poor, it is not suitable for direct consumption and needs to be improved. Currently, there is no product with high-activity green tea polyphenols extract and synbiotics as the main raw materials. Summary of the Invention

[0004] One purpose of the present invention: In order to overcome the defects of the prior art, the present invention provides a composition for improving liver inflammation.

[0005] The technical solution of the present invention is a composition for improving liver inflammation, comprising the following ingredients: prebiotics, probiotics and green tea polyphenols extract,

[0006] The prebiotics include one or more of lactose and isomalto-oligosaccharides;

[0007] The probiotics include one or more of rhamnosus GG and Lactobacillus casei YRL577, and the probiotic strains are also cultured and amplified;

[0008] Green tea polyphenols extract is obtained by extracting and separating green tea leaves, and its components include one or more of epigallocatechin gallate (EGCG), epicatechin gallate (ECG), gallic acid (EGC), and epicatechin (EC).

[0009] Preferably, the green tea is high mountain green tea leaves.

[0010] Preferably, the extraction technique is ethanol immersion method, and the separation technique is resin adsorption method.

[0011] Preferably, the composition comprises the following percentages: 45-60% prebiotics, 25-35% probiotics, and 6-8% green tea polyphenols extract.

[0012] The specific percentage components are as follows:

[0013] Prebiotics: Lactose 30-40%, isomalto-oligosaccharides 15-20%, of which 10-20% lactose is used as a probiotic protector;

[0014] Probiotics: Lactobacillus rhamnosus GG 15-20%, Lactobacillus casei YRL577 10-15%;

[0015] Green tea polyphenols extract: epigallocatechin gallate (EGCG) 2-3%, epicatechin gallate (ECG) 1-1.5%, gallic acid (EGC) 1.5-2.5%, epicatechin (EC) 1.5-2.5%.

[0016] Preferably, the composition for improving liver inflammation further comprises a disintegrant and a lubricant, wherein the disintegrant is hydroxypropyl methylcellulose and the lubricant is magnesium stearate.

[0017] Preferably, the composition is in the form of liquid, powder or capsule for oral administration.

[0018] Another purpose of the present invention: In order to overcome the defects of the prior art, the present invention provides a method for preparing a composition for improving liver inflammation.

[0019] The technical solution of the present invention is a method for preparing a composition for improving liver inflammation, comprising steps S, T and U.

[0020] Step S is the preparation of green tea polyphenols extract: fresh high mountain green tea leaves are used as raw materials, and the green tea polyphenols extract is prepared by ethanol soaking and resin adsorption methods;

[0021] Step T is the preparation of probiotic freeze-dried powder: a probiotic single-bacterial culture is cultured in a culture medium to a logarithmic growth phase, and then the probiotic single bacteria are collected by centrifugation or other methods; the collected probiotic single-bacterial suspension is freeze-dried and pulverized by a grinder or other equipment to obtain a uniform probiotic powder;

[0022] Step U is the preparation of a composition for improving liver inflammation: the green tea polyphenols extract prepared in step S is first mixed with prebiotics, wet granulated and ground into powder, and then mixed with lyophilized probiotic powder to prepare a finished product in the form of a liquid, powder granule or capsule.

[0023] The efficacy of the composition for improving liver inflammation prepared by the present invention is described as follows:

[0024] Lactose is a soluble cellulose that cannot be synthesized by lactose hydrolase on its own. β-lactosylase produced by Lactobacillus rhamnosus GG and α-lactosylase produced by Lactobacillus casei YRL577 can hydrolyze lactose into glucose and galactosonic acid, thereby utilizing lactose as a carbon and energy source for growth and reproduction.

[0025] Isomaltooligosaccharides are polysaccharide molecules that serve as a preferred energy source for Lactobacillus rhamnosus GG and Lactobacillus casei YRL577. They provide them with a rich source of carbon and energy, increasing the numbers of these two probiotics in the intestine. Furthermore, Lactobacillus rhamnosus GG and Lactobacillus casei YRL577 can ferment isomaltooligosaccharides to produce short-chain fatty acids, such as acetic acid and propionic acid, which maintain the balance of the intestinal microbiome. Isomaltooligosaccharides, Lactobacillus rhamnosus GG, and Lactobacillus casei YRL577 can mutually promote each other, forming a prebiotic network that works together to exert beneficial health effects.

[0026] Lactobacillus rhamnosus GG and Lactobacillus casei YRL577 play an important role in lowering blood lipids, promoting the proliferation of intestinal mucosal immune cells, and regulating the composition of the intestinal flora. Lactobacillus rhamnosus GG can also regulate liver inflammation, alleviate liver disease symptoms, and reduce the severity of liver damage. Spreading the dose across two probiotics reduces the risk of side effects and is safer than taking a single, large dose. Taking different strains simultaneously can promote intestinal microbial diversity and produce a wider range of beneficial effects, such as lowering blood lipids and controlling liver inflammation.

[0027] Tea polyphenols have antioxidant and cholesterol-lowering properties, scavenging free radicals, slowing cell aging, and boosting immunity, all contributing to overall health. Green tea contains the highest concentration of tea polyphenols, accounting for approximately 9%-13%, depending on the fermentation method used. The combined use of tea polyphenols and probiotics can synergistically lower blood lipids and maintain intestinal health. Furthermore, tea polyphenols stimulate the growth of probiotics, while probiotic metabolites facilitate their absorption and utilization, resulting in a mutually reinforcing effect.

[0028] Disintegrants (hydroxypropyl methylcellulose) can improve the viscosity, stability, mouthfeel and texture of the composition; lubricants (magnesium stearate) are used to reduce the viscosity and friction of the composition for easier processing, mixing and packaging.

[0029] The composition for improving liver inflammation provided by the present invention has high activity of tea polyphenol extracts, and the raw materials of each component cooperate with each other to achieve synergistic effect, so that the composition has the effects of improving inflammation and lowering blood lipid levels. It also has a therapeutic and regulatory effect on intestinal flora disorders and intestinal barrier damage, and has excellent health-preserving effects. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is a graph showing the extraction rate of tea polyphenols in the preparation examples under different extraction conditions;

[0031] Figure 2 This is a graph showing the extraction rate of tea polyphenols from preparation examples of green tea from different sources;

[0032] Figure 3 The chart shows the serum alanine aminotransferase levels of mice with steatohepatitis in each group treated with synbiotics of different composition ratios;

[0033] Figure 4 The graph shows the serum triglyceride levels of mice with steatohepatitis in each group treated with synbiotics of different composition ratios;

[0034] Figure 5 The graph shows the serum alanine aminotransferase levels of each group of autoimmune hepatitis mice treated with synbiotics of different composition ratios;

[0035] Figure 6 This is a graph showing serum transaminase and triglyceride levels in mice with steatohepatitis after adding tea polyphenol extract to synbiotics.

[0036] Figure 7 The liver of mice with steatohepatitis was stained with eosin-hematoxylin and oil red O after adding tea polyphenol extract to synbiotics.

[0037] Figure 8 The serum transaminase levels and liver eosin-hematoxylin staining of autoimmune mice after adding tea polyphenols extract to synbiotics;

[0038] Figure 9 This is a graph showing the DPPH radical scavenging rate of the preparation example under different preparation conditions;

[0039] Figure 10 This is the distribution of intestinal flora in autoimmune mice after adding catechin extracts to synbiotics. DETAILED DESCRIPTION

[0040] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments:

[0041] 1. Preparation method of the composition for improving liver inflammation, step S (preparation of green tea polyphenols extract):

[0042] S1. Dissolve green tea powder in 60-100% ethanol at a ratio of 1g:10-50ml;

[0043] S2. Ultrasonic extraction for 20-60 min at an extraction temperature of 60-100 ° C and an ultrasonic extraction power of 40-80% (100-200 W);

[0044] S3. The upper tea solution was filtered through a cell sieve and rotary evaporated to obtain the primary green tea extract;

[0045] S4. The green tea primary extract was dissolved in distilled water at 1 g: 1 ml and passed through a resin column with edible ethanol as the equilibrium solution at a flow rate of 1 to 1.5 BV / h to collect the ethanol-water solution;

[0046] S5. The ethanol-water solution was concentrated by rotary evaporation and dried to obtain a green tea polyphenols extract powder;

[0047] Step T (preparation of probiotic freeze-dried powder):

[0048] T1. Single bacterial culture: Single strains of Lactobacillus rhamnosus GG and Lactobacillus casei YRL577 (viable count > 10 9 CFU / ml) were inoculated into MRS liquid medium (pH 6.5-7), cultured in an anaerobic incubator at 37-39°C and 160-180 rpm for 18-20 h, then expanded with an inoculum size of 1.5-2% (v / v) for 24-26 h, and then centrifuged at 4000-5000 rpm and 4-8°C for 15-30 min to collect the bacteria;

[0049] T2. Washing and resuspension: Wash the collected probiotics with sterile saline and then suspend them in an appropriate amount of saline to remove unused culture medium components;

[0050] T3. Add 10%-20% lactose;

[0051] T4 freeze-drying: The suspension containing Lactobacillus rhamnosus GG and the suspension containing Lactobacillus casei YRL577 are divided into containers, and then frozen at -80 to -100°C. The frozen bacterial suspension is placed in a freeze dryer, and the main vacuum is set to 0.1-0.5 mbar, the condenser temperature is -50°C to 80°C, and the freeze-drying time is 48-50 hours;

[0052] T5 crushing: crushing by grinder and other equipment to obtain uniform probiotic powder;

[0053] Step U (preparation of a composition for improving liver inflammation):

[0054] U1. 6-8% green tea polyphenol extract, prebiotic lactose, and 15-20% isomaltooligosaccharide were placed in a boiling granulator and thoroughly mixed for 15-20 minutes. Water was used as a wetting agent for spray granulation. The inlet air temperature was controlled at 60-100°C and the outlet air temperature at 30-50°C. The moisture content was regularly checked until it was 2-3%, and then ground into powder.

[0055] U2. Add 3-5% hydroxypropyl methylcellulose and 0.1-1% magnesium stearate and mix thoroughly in a multi-directional motion mixer;

[0056] U3. Add 20% Lactobacillus rhamnosus GG freeze-dried powder and 10% Lactobacillus casei YRL577 freeze-dried powder and mix well to prepare a composition for improving liver inflammation in the form of liquid, powder or capsule.

[0057] 1.1. Preparation of Green Tea Polyphenols Extract (25 preparation examples in total)

[0058] The green tea polyphenols extract in Preparation Example 1 is prepared according to the following steps:

[0059] S1. Dissolve West Lake Longjing (green tea) tea powder in 70% ethanol at a ratio of 1g:20ml;

[0060] S2. Ultrasonic extraction was performed for 30 min at an extraction temperature of 80°C and an ultrasonic power of 60% (150 W).

[0061] S3. The upper tea solution was filtered through a cell sieve and rotary evaporated to obtain the primary green tea extract;

[0062] S4. The primary extract was dissolved in distilled water at a rate of 1 g: 1 ml and passed through a resin column using edible ethanol as the equilibrium solution at a flow rate of 1 to 1.5 BV / h to collect the ethanol-water solution;

[0063] S5. The ethanol-water solution is concentrated by rotary evaporation and dried to obtain green tea extract powder.

[0064] The green tea polyphenols extracts of Preparation Examples 2-21 differ from those of Preparation Example 1 in that the extraction ethanol concentrations of Preparation Examples 2-5 are different, the material-liquid ratios of Preparation Examples 6-9 are different, the extraction temperatures of Preparation Examples 10-13 are different, the extraction powers of Preparation Examples 14-17 are different, and the extraction times of Preparation Examples 18-21 are different. The specific parameters are shown in Table 1 below.

[0065] Table 1. Factor levels of orthogonal experiment of tea polyphenols extract

[0066] Ethanol concentration Material-liquid ratio Extraction temperature Extraction power Extraction time 60％ 1g:10ml 60℃ 40％(100W) 20min 70％ 1g:20ml 70℃ 50％(125W) 30min 80％ 1g:30ml 80℃ 60％(150W) 40min 90％ 1g:40ml 90℃ 70％(175W) 50min 100％ 1g:50ml 100℃ 80％(200W) 60min

[0067] Weigh 1 gram of sample and dissolve it in 25 mL of distilled water. Pour the mixture into a 25 mL volumetric flask. Add 4 mL of distilled water and 5 mL of ferrous tartrate solution. Mix and adjust to volume with phosphate buffer. Measure the absorbance A at 540 nm using a 10 mm cuvette. Use the reagent blank as a reference. The percentage of the tea polyphenol content measured in the experimental sample to the total tea polyphenol content of tea specified in GB / T 8313-2002 is the tea polyphenol extraction rate. The results are as follows: Figure 1As shown, the extraction rate of tea polyphenols using the method of Preparation Example 1 is the highest.

[0068] The green tea polyphenol extracts in Preparation Examples 22-25 differ from those in Preparation Example 1 in that the types of green tea are different. Preparation Example 22 uses Tieguanyin, Preparation Example 23 uses Long Leaf Light Green Tea, Preparation Example 24 uses Huangshan Maofeng Green Tea, and Preparation Example 25 uses Jinjunmei Green Tea. Figure 2 As shown, the efficiency of extracting tea polyphenols from different types of green tea using the method of Preparation Example 1 is similar.

[0069] The content of tea phenols in the tea polyphenols extract of Preparation Example 1 was determined by ultraviolet spectrophotometry. The results are shown in Table 2.

[0070] Table 2. Theanine and theanine content of tea polyphenols extracts

[0071] project English abbreviation content Tea polyphenols (%) 0.64±0.05 Catechin (mg / L) C 318.35±8.37 Epicatechin (mg / L) EC 455.22±5.41 Gallic acid (mg / L) EGC 489.29±10.11 Epicatechin gallate (mg / L) ECG 420.32±12.72 Gallic acid ester (mg / L) EGCG 1090.23±28.46

[0072] 1.2. Preparation of probiotic freeze-dried powder:

[0073] T1. Single bacterial culture and washing suspension: Single strains of Lactobacillus rhamnosus GG and Lactobacillus casei YRL577 (viable cell count > 10 9 CFU / ml) were inoculated into MRS liquid medium (pH 6.5), cultured in an anaerobic incubator at 37°C and 160 rpm for 18 h, then amplified with 1.5% (v / v) inoculum for 24 h, and then centrifuged at 4000 rpm and 4°C for 15 min to collect the bacteria;

[0074] T2. Wash the collected probiotics 1-2 times with sterile saline, then suspend them in an appropriate amount of saline to remove unused culture medium components and improve the purity of the lyophilized powder.

[0075] T3. To protect bacterial cells during the freeze-drying process, an appropriate amount of lactose needs to be added, with the final concentration of lactose being 10%-20%;

[0076] T4. Freeze-drying: Dispense the suspension containing Lactobacillus rhamnosus GG and Lactobacillus casei YRL577 into appropriate containers (e.g., freeze-drying tubes). Place the tubes in a -80°C ultra-low temperature freezer for rapid freezing until completely frozen. Place the frozen bacterial suspension in a freeze dryer, set the main vacuum to 0.1-0.5 mbar, and the condenser temperature to -50°C to -80°C. Freeze-dry for 48 hours. After freeze-drying, immediately seal the freeze-drying tubes and store them at 4°C in a dark environment.

[0077] T5 crushing: crushing through grinders and other equipment to obtain uniform probiotic powder.

[0078] 1.3 Comparison of the efficacy of different ratios of synbiotics on steatohepatitis and autoimmune hepatitis

[0079] Experimental Example 1: lactose 35%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%.

[0080] Experimental Example 2: Isomaltooligosaccharide 17%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%.

[0081] Experimental Example 3: lactose 40%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%.

[0082] Experimental Example 4: lactose 30%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%.

[0083] Experimental Example 5: Lactose 35%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%. Experimental Example 6: Lactose 35%, Isomaltooligosaccharide 15%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%.

[0084] Experimental Example 7: lactose 35%, isomalto-oligosaccharide 20%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 13%.

[0085] Experimental Example 8: lactose 35%, isomaltooligosaccharide 17%, Lactobacillus casei YRL577 13%.

[0086] Experimental Example 9: lactose 35%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 15%, Lactobacillus casei YRL577 13%.

[0087] Experimental Example 10: lactose 35%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 20%, Lactobacillus casei YRL577 13%.

[0088] Experimental Example 11 Lactose 35%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 17%.

[0089] Experimental Example 12: lactose 35%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 10%.

[0090] Experimental Example 13: Lactose 35%, isomalto-oligosaccharide 17%, Lactobacillus rhamnosus GG 17%, Lactobacillus casei YRL577 15%.

[0091] Experimental Example 14: lactose 40%, isomalto-oligosaccharide 20%, Lactobacillus rhamnosus GG 20%, Lactobacillus casei YRL577 15%.

[0092] Experimental Example 15: lactose 30%, isomalto-oligosaccharide 15%, Lactobacillus rhamnosus GG 15%, Lactobacillus casei YRL577 10%.

[0093] Experimental Examples 1-15 were prepared according to the above ratios for the treatment of high-fat diet-induced fatty liver disease and S100 protein-induced autoimmune hepatitis mouse models.

[0094] For steatohepatitis testing

[0095] Group 1, control group: mice were fed with normal feed until week 12. From week 12 onwards, 1 ml of sterile saline was gavaged every other day until week 16.

[0096] Group 2, steatohepatitis model group: Mice were fed a high-fat diet for 12 weeks to establish a steatohepatitis model. From the 12th week onward, 1 ml of sterile saline was gavage administered every other day until 16 weeks.

[0097] 3-17, Synbiotics treatment group: mice were fed a high-fat diet until week 12. Starting from week 12, mice were given the treatments of Experimental Examples 1-15 on the basis of the high-fat diet, with a gavage volume of 1 ml per mouse, once every other day, until week 16.

[0098] Serum alanine aminotransferase levels of mice in groups 1-17 were detected ( Figure 3 ) and triglyceride content ( Figure 4 ) Serum transaminase and triglyceride levels were significantly elevated in mice with steatohepatitis. However, after treatment, these levels decreased, indicating that fatty liver damage was alleviated and lipid deposition was improved. The most significant decreases in serum transaminase and triglyceride levels were observed in mice with steatohepatitis following treatment in Experiments 1, 14, and 15.

[0099] For autoimmune hepatitis liver injury test

[0100] Group 18, control group: mice were intraperitoneally injected with 1 ml of normal saline on day 1 and day 7. From day 1, mice were gavaged with 1 ml of sterile normal saline every other day for up to 4 weeks.

[0101] Group 19, Autoimmune Hepatitis Model Group: Livers from mice from the same batch were removed, rinsed in clean, pre-chilled PBS, cut into small pieces, and thoroughly homogenized on ice. After centrifugation at 150g for 10 minutes at 4°C to remove nuclei, the supernatant was collected into an ultracentrifuge tube and centrifuged at 100,000g for 1 hour. The resulting supernatant was S100. The supernatant was concentrated using an Amicon Ultra-15 filter and then separated using a pre-equilibrated 90cm CL-6B Sepharose column. The attenuated first peak protein was collected. Freshly prepared isogenic S100 antigen from the first peak was emulsified at a concentration of 1 mg / mL in an equal volume of complete Freund's adjuvant, mixed thoroughly, and set aside. 1 ml of this mixture was injected intraperitoneally into mice on days 1 and 7, respectively. Starting on day 1, mice were gavaged with 1 ml of sterile saline every other day for up to 4 weeks.

[0102] Groups 20-34, synbiotic treatment groups: mice were fed a high-fat diet until week 12. Starting from week 12, mice were given the treatments of Experimental Examples 1-15 on the basis of the high-fat diet, with a gavage volume of 1 ml per mouse, once every other day, for a period of 4 weeks.

[0103] Serum alanine aminotransferase levels of mice in groups 18-34 were detected ( Figure 5 ), the serum transaminase of mice with autoimmune hepatitis was significantly elevated, but after treatment, the serum transaminase of each group decreased, among which the decrease in groups 20, 33, and 34 was the most obvious, indicating that the treatment of experimental examples 1, 14, and 15 was more effective in alleviating liver damage caused by autoimmune hepatitis.

[0104] Comparison of the therapeutic effects of biostime supplemented with tea polyphenols extract on steatohepatitis

[0105] Twenty 10-week-old healthy male C57BL / 6 mice were randomly divided into the following 4 groups:

[0106] Group 1, control group: mice were fed with normal feed until week 12. From week 12 onwards, 1 ml of sterile saline was gavaged every other day until week 16.

[0107] Group 2, steatohepatitis model group: Mice were fed a high-fat diet for 12 weeks to establish a steatohepatitis model. From the 12th week onward, 1 ml of sterile saline was gavage administered every other day until 16 weeks.

[0108] Group 3, synbiotic treatment group: mice were fed a high-fat diet until week 12. Starting from week 12, synbiotic treatment according to Experimental Example 1 was added to the high-fat diet, with a gavage volume of 1 ml per mouse, once every other day, until week 16.

[0109] Group 4, Synbiotics combined with tea polyphenols extract group: Mice were fed a high-fat diet until week 12. Starting from week 12, mice were treated with 1 ml of Synbiotics liquid (Example 1) dissolved in 200 mg / kg body weight of tea polyphenols extract, once every other day, on the basis of the high-fat diet, and fed until week 16.

[0110] The serum transaminase levels and triglyceride content of mice in each group were detected. Figure 6 ), the serum transaminase and triglyceride levels of mice with fatty liver hepatitis were significantly increased, while after treatment, the serum transaminase and triglyceride levels decreased, especially the lipid-lowering effect of the postbiotics combined with tea polyphenols extract group was the most obvious.

[0111] The liver was stained with oil red and eosin-hematoxylin ( Figure 7 ) and found that the normal structure of the liver lobules of mice in the fatty liver hepatitis model group was destroyed, a large number of inflammatory cells infiltrated the portal area, the liver cells were swollen, the cytoplasm was loose, and most of the intracellular vacuoles were microvesicular fatty degeneration; after treatment, the liver inflammation and microvesicular fatty degeneration were alleviated.

[0112] Comparison of the therapeutic effects of biostime supplemented with tea polyphenols extract on autoimmune hepatitis Twenty 10-week-old healthy male C57BL / 6 mice were randomly divided into the following 4 groups:

[0113] Group 1, control group: mice were intraperitoneally injected with 1 ml of normal saline on day 1 and day 7. From day 1, mice were gavaged with 1 ml of sterile normal saline every other day for up to 4 weeks.

[0114] Group 2, autoimmune hepatitis model group: Mice were intraperitoneally injected with 1 ml of S100-complete Freund's adjuvant mixture on day 1 and day 7. From day 1, they were gavaged with 1 ml of sterile saline every other day for up to 4 weeks.

[0115] Group 3, synbiotic treatment group: Mice were intraperitoneally injected with 1 ml of S100-complete Freund's adjuvant mixture on day 1 and day 7. From day 1, mice were gavaged with 1 ml of the synbiotic solution of Experimental Example 1 every other day for up to 4 weeks.

[0116] Group 4, synbiotics combined with tea polyphenols extract group: Mice were intraperitoneally injected with 1 ml of S100-complete Freund's adjuvant mixture on day 1 and day 7. From day 1, mice were gavaged with 1 ml of synbiotic solution (Example 1) dissolved in 200 mg / kg body weight of tea polyphenols extract every other day for up to 4 weeks.

[0117] Serum transaminase levels of mice in each group were detected and the livers were stained with oil red and eosin-hematoxylin. Figure 8) found that compared with the control group, mice injected intraperitoneally with S100 had significantly higher serum ALT, AST, and IgG levels. Liver pathology showed interface hepatitis, focal necrosis, and lymphocyte and plasma cell infiltration. After treatment, serum transaminase and immunoglobulin G levels decreased, and liver inflammatory infiltration was alleviated. The lipid-lowering effect was particularly pronounced in the postbiotics combined with tea polyphenols extract group.

[0118] The above examples demonstrate that 30-40% lactose, 15-20% isomalto-oligosaccharide, 15-20% Lactobacillus rhamnosus GG, 10-15% Lactobacillus casei YRL577, and 6-8% tea polyphenol extract have good functions of improving liver inflammation and lowering blood lipid levels.

[0119] Combination and preparation of tea polyphenols extract and composition for improving liver inflammation Preparation Example 1: 2-3 parts of lactose, 1.5-2 parts of isomalto-oligosaccharides, and 0.6-0.8 parts of tea polyphenols extract are put into a boiling granulator and mixed thoroughly for 10 minutes, then spray granulated with water as a wetting agent, controlling the inlet air temperature at 80°C and the outlet air temperature at 40°C, stopping spraying and drying, and checking the moisture every hour until the moisture content is 2-3%.

[0120] Preparation Example 2: Same as Example 1, except that the inlet air temperature was changed to 60°C, and other parameters remained unchanged. Preparation Example 3: Same as Example 1, except that the inlet air temperature was changed to 100°C, and other parameters remained unchanged. Preparation Example 4: Same as Example 1, except that the outlet air temperature was changed to 30°C, and other parameters remained unchanged.

[0121] Preparation Example 5: Same as Example 1, except that the air outlet temperature was changed to 50°C, and other parameters remained unchanged. Preparation Example 6: Same as Example 1, except that the mixing time was changed to 5 minutes, and other parameters remained unchanged. Preparation Example 7: Same as Example 1, except that the mixing time was changed to 20 minutes, and other parameters remained unchanged.

[0122] Weigh the preparation examples 1-7 and prepare them into 2.5 mg / mL solutions, then dilute them to 0.5, 1.0, 1.5, and 2.0 mg / mL in sequence, then add 3 mL of 0.004% DPPH methanol solution, shake well, shield from light and set aside for 20 minutes, use methanol solution as blank comparison solution, detect absorbance at 517 nm, use distilled water as negative comparison solution, and use VC solution as positive comparison solution, and calculate DPPH free radical scavenging rate. Figure 9 As shown, Preparation Example 1 has a higher DPPH free radical scavenging rate than other Preparation Examples, which means that the tea polyphenols in Preparation Example 1 are more active.

[0123] 0.3-0.5 parts of a disintegrant (hydroxypropyl methylcellulose) and 0.01-0.1 parts of a lubricant (magnesium stearate) are added to the obtained granules and thoroughly mixed in a multi-directional motion mixer to obtain a powder, which is then mixed with 2 parts of Lactobacillus rhamnosus GG freeze-dried powder and 1 part of Lactobacillus casei YRL577 freeze-dried powder to prepare a packaged composition for improving liver inflammation.

[0124] Case Experiment

[0125] Oral animal toxicology test

[0126] Forty mice weighing 18-20 g were randomly divided into a control group and a test group, 20 mice per group, half male and half female. After a 6-hour fast (without water), mice were gavage-administered a synbiotic powder containing green tea polyphenols extract at a dose of 5000 mg / kg body weight. The control group received an equal volume of untreated purified water. After gavage, the onset and disappearance of signs of toxicity and the time of death were observed and recorded. The observation period was 14 days. Animals that died from poisoning during the observation period or were sacrificed at the end of the observation period were subjected to gross autopsy and macroscopic examination. Any tissue or organ abnormalities were further examined by histopathological examination. Weights were also recorded on days 0, 1, 3, 7, and 14 of gavage. No abnormalities were observed during the administration of the synbiotic powder containing green tea polyphenols extract or during the 14-day observation period. No animals showed any abnormalities, weight gain, or mortality. At the end of the experiment, all animals were subjected to gross autopsy, and no abnormalities were observed. The specific weight changes are shown in Table 2. Under the test conditions, the acute oral toxicity LD50 of the sample submitted for inspection to KM mice was greater than 5000 mg / kg body weight, and the oral toxicity classification of the sample submitted for inspection was slightly toxic (actually non-toxic).

[0127] Table 3. Oral toxicology test of synbiotic powder containing green tea polyphenols extract

[0128]

[0129] Comparison of the effects of adding tea polyphenols extract to synbiotics on the distribution of intestinal flora in patients with autoimmune hepatitis

[0130] like Figure 10 As shown in the results, the Chao1 richness value of the intestinal flora of mice with autoimmune hepatitis was significantly reduced compared with the control group, while after treatment, the Chao1 richness value was significantly increased. At the same time, the treatment of synbiotics combined with tea polyphenols extract increased the proportion of Lactobacillus and decreased the proportion of Enterococcus.

[0131] The observation results show that the liver inflammation-improving composition containing tea polyphenols extracts provided by the present invention can effectively improve hyperlipidemia symptoms, regulate intestinal flora, help alleviate different liver inflammations and injuries, and improve physical health.

[0132] In summary, the composition for improving liver inflammation provided by the embodiment of the present invention comprises 30-40% lactose (10-20% of which is used as a probiotic protective agent), 15-20% isomaltooligosaccharide, 15-20% Lactobacillus rhamnosus GG, 10-15% Lactobacillus casei YRL577, and 6-8% tea extract. The components and raw materials cooperate with each other and synergize to enhance the effect, so that the composition has the effects of improving inflammation and lowering blood lipid levels. At the same time, it also has a therapeutic and regulating effect on intestinal flora disorders and intestinal barrier damage, and has excellent health-preserving effects.

[0133] The embodiments described above are some, but not all, of the embodiments of the present invention. The detailed description of the embodiments of the present invention is not intended to limit the scope of the invention as claimed, but rather merely represents selected embodiments of the present invention. All other embodiments derived by persons of ordinary skill in the art based on the embodiments of the present invention without inventive effort are also within the scope of protection of the present invention.

Claims

1. A composition for improving liver inflammation, characterized in that: Made from the following components: Lactose 30-40%, isomaltooligosaccharide 15-20%, Lactobacillus rhamnosus GG 15-20%, Lactobacillus casei YRL577 10-15%, green tea polyphenols extract 6-8% and excipients, The green tea polyphenols extract is prepared by extracting and separating green tea leaves with 70% ethanol, and the green tea polyphenols extract includes 2-3% gallate, 1-1.5% epicatechin gallate, 1.5-2.5% gallic acid, and 1.5-2.5% epicatechin; The viable count of Lactobacillus rhamnosus GG and Lactobacillus casei YRL577 was 10 9 -10 12 CFU / mL; Inflammation of the liver is either steatohepatitis or immune hepatitis.

2. The composition for improving liver inflammation according to claim 1, characterized in that: Green tea leaves are selected from high mountain green tea leaves.

Images