Application of plant lactobacillus in preparation of product for relieving alcoholic liver disease

Abstract

The invention belongs to the technical field of probiotics, and particularly relates to application of plant lactobacillus in preparation of a product for relieving alcoholic liver diseases. The invention provides an application of plant lactobacillus LP-Only in preparation of a product for relieving alcoholic liver diseases, after the plant lactobacillus LP-Only is used for gavage of mice, it is found that the plant lactobacillus LP-Only can relieve alcoholic fatty liver diseases caused by chronic alcohol intake, liver lipid accumulation caused by long-term alcohol intake can be reduced, liver lipid consumption is increased, and the effect of relieving alcoholic liver diseases is achieved. The liver inflammation reaction is relieved. The invention further provides application of the plant lactobacillus LP-Only in preparation of functional food and / or health care products for protecting the liver. The prepared yoghourt and fermented soybean milk beverage can relieve the alcoholic fatty liver caused by chronic alcohol intake.

Description

Technical Field

[0001] The invention belongs to the technical field of probiotics, and particularly relates to application of a strain of Lactobacillus plantarum in preparing a product for alleviating alcoholic liver disease. Background Art

[0002] Long-term drinking can damage liver function and cause alcoholic liver disease, such as alcoholic fatty liver disease (AFLD). Alcoholic fatty liver disease is a liver disease caused by long-term excessive drinking, in which fat is deposited in liver cells. It is characterized by the accumulation of cholesterol and triglycerides in serum and liver. It is the early stage of alcoholic liver disease. It can be completely cured by quitting drinking and appropriate intervention treatment. If it is not effectively controlled, it can further develop into alcoholic hepatitis, liver fibrosis, cirrhosis and even hepatocellular carcinoma. With the rapid development of my country's economy in recent years, although no national and large-scale epidemiological surveys have been conducted, some regional epidemiological surveys have shown that the drinking rate and prevalence of Chinese residents have been increasing year by year. At present, drug treatment is still one of the main methods to control AFLD, but drugs have serious side effects and some people cannot tolerate them. Therefore, exploring natural active ingredients with significant auxiliary treatment effects and no side effects for AFLD has become a hot topic in recent years.

[0003] "Probiotics" are defined as products containing live microorganisms that, when given in adequate amounts, have beneficial effects on the host by changing the intestinal microbiota. Bifidobacterium and Lactobacillus are the most commonly used probiotics, which can inhibit the proliferation of pathogens, enhance the host's immune function, improve metabolic diseases, and reduce alcohol-induced liver damage. Therefore, the use of probiotics and their products to alleviate alcoholic fatty liver disease has very important research significance and broad application prospects.

[0004] Studies have shown that Lactobacillus rhamnosus CCFM1107 (L.rhamnosus CCFM1107) has high antioxidant activity. When taking L.rhamnosus CCFM1107 and taking alcohol orally for 3 months, it was found that L.rhamnosus CCFM1107 had a significant inhibitory effect on the increase of serum lipopolysaccharide content and cholesterol and triglyceride levels in serum and liver of mice. It can also increase the superoxide dismutase activity of mouse liver and reduce the level of malondialdehyde, while increasing the number of lactobacilli and bifidobacteria and reducing the number of enterococci and enterobacteria, and restoring the intestinal flora imbalance caused by alcohol. Therefore, L.rhamnosus CCFM1107 has a strong protective effect on alcoholic liver damage by reducing oxidative stress and restoring intestinal flora. The application of plant lactobacillus in relieving alcoholic liver disease has also been reported in the prior art, but plant lactobacillus needs to be compounded with other strains to play the corresponding effect, for example, the prior art has Lactobacillus plantarum LC27 (Lactobacillus plantarum LC27) and Bifidobacterium longum (Bifidobacterium longumLC67) gavage mice, which reduces the transaminase, cholesterol, and triglyceride levels in the blood and liver of ethanol-induced mice, inhibits the activation of alcohol-induced liver NF-κB, increases the activity of alcohol dehydrogenase and acetaldehyde dehydrogenase, and improves the inhibitory effect of ethanol on AMPK. There is no report in the prior art that a single application of plant lactobacillus can prevent and treat alcoholic liver disease. Summary of the invention

[0005] The invention provides an application of a Lactobacillus plantarum strain in preparing a product for alleviating alcoholic liver disease. Single application of Lactobacillus plantarum LP-Onlly can alleviate alcoholic fatty liver disease caused by chronic alcohol intake, with good effect.

[0006] In order to solve the above technical problems, the following technical solutions are proposed:

[0007] The invention provides application of Lactobacillus plantarum LP-Onlly in preparing a product for alleviating alcoholic liver disease.

[0008] Preferably, the number of viable Lactobacillus plantarum LP-Onlly in the product is ≥ 1×10 9 CFU / mL.

[0009] Preferably, the Lactobacillus plantarum LP-Onlly is used for any one or two or more of the following (1) to (9):

[0010] (1) Reduce liver index;

[0011] (2) Reduce triglyceride levels in the liver;

[0012] (3) Reduce cholesterol levels in the liver;

[0013] (4) Reduce the activity of alanine aminotransferase and / or aspartate aminotransferase in the liver, and increase the activity of alcohol dehydrogenase and / or acetaldehyde dehydrogenase in the liver;

[0014] (5) increasing superoxide dismutase and / or catalase activity in the liver;

[0015] (6) Increase liver glutathione content;

[0016] (7) Reduce liver malondialdehyde levels;

[0017] (8) Reduce liver inflammation;

[0018] (9) Increase the mRNA expression levels of liver lipid metabolism-related genes Lipin-1α, PPAR-α and PGC-1α.

[0019] Preferably, the alcoholic liver disease includes alcoholic fatty liver.

[0020] Preferably, the product comprises a pharmaceutical product.

[0021] The invention provides application of Lactobacillus plantarum LP-Onlly in preparing functional food and / or health care product for protecting or assisting in protecting the liver.

[0022] Preferably, the functional food includes yogurt and / or fermented soy milk beverage, and the health product includes probiotic powder.

[0023] Preferably, the preparation method of the probiotic powder comprises: mixing the Lactobacillus plantarum LP-Onlly and a protective agent, and then freeze-drying to obtain the probiotic powder; the protective agent comprises one or more of skim milk, gelatin, maltodextrin, modified starch, gum arabic, sucrose, lactose, trehalose, sorbitol and xylitol.

[0024] Preferably, the method for preparing the yogurt comprises: inoculating the Lactobacillus plantarum LP-Onlly into dairy products for fermentation to obtain yogurt;

[0025] The method for preparing the fermented soybean milk beverage comprises: inoculating the Lactobacillus plantarum LP-Onlly into a base material for fermentation to obtain the fermented soybean milk beverage.

[0026] The invention provides a product for alleviating alcoholic liver disease, wherein the effective component comprises Lactobacillus plantarum LP-Onlly.

[0027] Beneficial effects of the present invention: The present invention provides an application of Lactobacillus plantarum LP-Onlly in the preparation of a product for alleviating alcoholic liver disease. After gavage of the product to mice, it was found that Lactobacillus plantarum LP-Onlly can alleviate alcoholic fatty liver caused by chronic alcohol intake, reduce liver lipid accumulation caused by long-term alcohol intake, increase liver lipid consumption, and alleviate liver inflammatory response.

[0028] The present invention also provides the use of Lactobacillus plantarum LP-Onlly in preparing functional food and / or health products for protecting or assisting in protecting the liver. The present invention uses Lactobacillus plantarum LP-Onlly to prepare functional food yogurt and fermented soybean milk beverage. Since Lactobacillus plantarum LP-Onlly can relieve alcoholic fatty liver, the prepared functional food yogurt and fermented soybean milk beverage can still relieve alcoholic fatty liver. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 This is an Oil Red O-stained section of mouse liver tissue (200×). Figure 1 The scale bar in the figure is 100 μm;

[0030] Figure 2 This is a H&E stained section of mouse liver tissue (200×). Figure 2 The scale bar in the figure is 100 μm;

[0031] Figure 3 The data of TG and TC content in mouse liver, a is the data of TG content in liver; b is the data of TC content in liver; data are expressed as Mean±SD (n=5-6), different letters represent significant differences among different groups, P<0.05;

[0032] Figure 4 The activity data of liver alcohol metabolism-related enzymes in mice, where a is liver ADH activity; b is liver ALDH activity; c is liver CYP2E1 activity; data are expressed as Mean±SD (n=5-6), different letters represent significant differences between different groups, P<0.05;

[0033] Figure 5 The data of mouse liver oxidative stress level, a is liver SOD activity; b is liver CAT activity; c is liver GSH content; d is liver MDA content; data are expressed as Mean±SD (n=5-6), different letters represent significant differences between different groups, P<0.05;

[0034] Figure 6 The data of mouse liver inflammatory factors and serum LPS content are shown in Figure 1, where a is liver TNF-α; b is liver IL-6; c is serum LPS; data are expressed as Mean±SD (n=5-6), and different letters represent significant differences between different groups, P<0.05;

[0035] Figure 7 The expression diagram of lipid oxidation-related genes in mouse liver; data are expressed as Mean±SD (n=4), and different letters represent significant differences among different groups, P<0.05. DETAILED DESCRIPTION

[0036] The present invention provides an application of Lactobacillus plantarum LP-Onlly in the preparation of a product for alleviating alcoholic liver disease, and the preservation number of Lactobacillus plantarum LP-Onlly is CGMCC No.1258. The Lactobacillus plantarum LP-Onlly described in the present invention is Lactobacillus plantarum LP-Onlly, and Lactobacillus plantarum LP-Onlly refers to Lactobacillus plantarum LP-Onlly in a composition containing Lactobacillus plantarum in Chinese patent CN1660300A. The reason why it is written as "Lactobacillus plantarum LP-Onlly" is that the strain has been renamed according to relevant national laws and regulations. For relevant laws and regulations, please refer to the announcement on the update of the strain list of "List of strains that can be used for food" and "List of strains that can be used for infant food" issued by the National Health Commission on August 18, 2022.

[0037] At present, many studies on probiotics are still limited to the laboratory stage and lack a large amount of human trial data. However, the Lactobacillus plantarum LP-Onlly involved in the present invention has a history of safe consumption for more than 30 years, with good feedback, no side effects and other complications, and its safety and effectiveness are supported by a large amount of data.

[0038] As an optional embodiment, the number of viable Lactobacillus plantarum LP-Onlly in the product of the present invention is ≥1×10 9 CFU / mL, or 1×10 9 ~1×10 10 CFU / mL.

[0039] As an optional embodiment, the relief described in the present invention can be to relieve the symptoms of alcoholic liver disease, improve alcoholic liver disease, prevent alcoholic liver disease or treat alcoholic liver disease. The plant lactobacillus LP-Onlly is used for any one or more of the following (1) to (9): (1) reducing liver index; (2) reducing triglyceride content in the liver; (3) reducing cholesterol content in the liver; (4) reducing the activity of liver alanine aminotransferase and / or aspartate aminotransferase, and increasing the activity of alcohol dehydrogenase and / or acetaldehyde dehydrogenase in the liver; (5) increasing the activity of superoxide dismutase and / or catalase in the liver; (6) increasing liver glutathione content; (7) reducing liver malondialdehyde level; (8) reducing liver inflammatory response; (9) increasing the mRNA expression level of liver lipid metabolism-related genes Lipin-1α, PPAR-α and PGC-1α. As an optional embodiment, the relief of liver inflammatory response described in the present invention is to significantly reduce the expression level of TNF-α and / or IL-6 in the liver.

[0040] The verification experiment was carried out with mice as the research object. The results in the embodiment proved that the plant lactobacillus LP-Onlly of the present invention can significantly reduce the liver index and the triglyceride and cholesterol content in the liver of alcohol-fed mice (P<0.05); can significantly reduce the activity of liver transaminase, and increase the activity of alcohol dehydrogenase and acetaldehyde dehydrogenase in the liver (P<0.05); can significantly increase the activity of superoxide dismutase and catalase in the liver, increase the glutathione content in the liver, and reduce the level of malondialdehyde (P<0.05); can significantly reduce the body's inflammatory response, reduce the expression level of TNF-α and / or IL-6, and restore the serum endotoxin content to the same level as the normal group (P<0.05); and can up-regulate the mRNA expression levels of liver lipid metabolism-related genes Lipin-1α, PPAR-α, and PGC-1α, and has the effect of improving alcoholic fatty liver.

[0041] As an optional embodiment, the alcoholic liver disease of the present invention includes alcoholic fatty liver. The examples show that the Lactobacillus plantarum LP-Onlly of the present invention has the effect of improving alcoholic fatty liver in mice.

[0042] As an optional embodiment, the product of the present invention includes a medicine.

[0043] The invention provides application of Lactobacillus plantarum LP-Onlly in preparing functional food and / or health care product for protecting or assisting in protecting the liver.

[0044] As an optional embodiment, the protection of the present invention is to alleviate the symptoms of alcoholic liver disease and improve alcoholic liver disease. The plant lactobacillus LP-Onlly of the present invention is used for any one or more of the following functions (1) to (9): (1) reducing liver index; (2) reducing triglyceride content in the liver; (3) reducing cholesterol content in the liver; (4) reducing the activity of liver alanine aminotransferase and / or aspartate aminotransferase, and increasing the activity of alcohol dehydrogenase and / or acetaldehyde dehydrogenase in the liver; (5) increasing the activity of superoxide dismutase and / or catalase in the liver; (6) increasing liver glutathione content; (7) reducing liver malondialdehyde level; (8) reducing liver inflammatory response; (9) increasing the mRNA expression level of liver lipid metabolism-related genes Lipin-1α, PPAR-α and PGC-1α. As an optional embodiment, the reduction of liver inflammatory response of the present invention is to significantly reduce the expression level of TNF-α and / or IL-6 in the liver.

[0045] As an optional embodiment, the health care product of the present invention includes probiotic powder. The number of viable Lactobacillus plantarum LP-Onlly in the probiotic powder prepared by the present invention is not less than 1.0×10 11 CFU / g.

[0046] As an optional embodiment, the preparation method of the probiotic powder of the present invention comprises: mixing the Lactobacillus plantarum LP-Onlly and a protective agent, and then freeze-drying to obtain the probiotic powder; as another optional embodiment, the preparation method of the probiotic powder of the present invention comprises: mixing the bacterial bodies of the Lactobacillus plantarum LP-Onlly and a protective agent, and then freeze-drying to obtain the probiotic powder.

[0047] As an optional embodiment, the mixing method includes resuspension; as an optional embodiment, the mass ratio of the bacteria and the protective agent is 1:1. The present invention does not specifically limit the preparation method of the bacteria, and a conventional method can be used. As an optional embodiment, the preparation method of the bacteria of the present invention includes: inoculating the plant lactobacillus LP-Onlly in MRS culture medium for culturing, and centrifuging the obtained culture solution to obtain bacteria.

[0048] As an optional embodiment, the freeze drying includes vacuum freeze drying. The present invention has no special limitation on the freeze drying parameters, and conventional methods can be used.

[0049] As an optional embodiment, the protective agent of the present invention includes one or more of skim milk, gelatin, maltodextrin, modified starch, gum arabic, sucrose, lactose, trehalose, sorbitol and xylitol. In a specific embodiment of the present invention, skim milk is used as an example for verification. The present invention resuspends Lactobacillus plantarum LP-Onlly cell bodies in a skim milk solution, the mass concentration of the skim milk solution is 10%, and the mass ratio of the Lactobacillus plantarum LP-Onlly to the skim milk solution is 1:1.

[0050] As an optional embodiment, the functional food of the present invention includes yogurt and / or fermented soy milk beverage.

[0051] As an optional embodiment, the preparation method of yogurt of the present invention comprises: inoculating the plant lactobacillus LP-Onlly into a dairy product for fermentation to obtain yogurt. As an optional embodiment, the dairy product comprises skim milk; before the skim milk is fermented, it also comprises sterilization, and the sterilization method is to maintain it at 90°C for 10 minutes. The fermentation temperature of the present invention is 40°C, and the fermentation is carried out until the pH of the fermentation product is 4.5±0.05. After the fermentation, the present invention also comprises post-ripening the fermentation product obtained by the fermentation to obtain yogurt. As an optional embodiment, the post-ripening temperature is 4°C, and the post-ripening time is 44h. The yogurt prepared by the present invention has the advantages of high public acceptance and easy daily availability; at the same time, the lactose in the skim milk is degraded, avoiding and reducing the occurrence of lactose intolerance, and also has the health care function of preventing or treating alcoholic liver disease.

[0052] As an optional embodiment, the method for preparing the fermented soymilk beverage of the present invention comprises: inoculating the Lactobacillus plantarum LP-Onlly into a base material for fermentation to obtain the fermented soymilk beverage.

[0053] As an optional embodiment, the base material includes soy milk, a sweetener and a stabilizer. As an optional embodiment, the sweetener includes glucose and / or white sugar. The fermentation is performed until the pH value of the soy milk beverage is 3.9 to 4.1. As an optional embodiment, the stabilizer of the present invention includes one or more of monoglyceride, sucrose ester, carrageenan, CMC (sodium carboxymethyl cellulose) and sodium alginate. The fermented soy milk beverage prepared by the present invention can relieve alcoholic fatty liver. The advantage of the fermented soy milk beverage prepared by the present invention is that the number of live bacteria of Lactobacillus plantarum LP-Onlly in the fermented soy milk beverage is not less than 1.0×10 9 CFU / mL.

[0054] The present invention provides a product for alleviating alcoholic liver disease, wherein the active ingredient comprises the Lactobacillus plantarum LP-Onlly described in the above technical solution. The present invention has no special limitation on the auxiliary material components of the product, and conventional components can be used.

[0055] In order to further illustrate the present invention, the technical solution provided by the present invention is described in detail below in conjunction with the accompanying drawings and embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0056] (1) The culture medium involved in the following examples is composed as follows:

[0057] The composition of MRS solid medium is: 10g / L peptone, 10g / L beef extract, 20g / L glucose, 2g / L sodium acetate, 5g / L yeast powder, 2g / L diammonium hydrogen citrate, 2.6g / L K 2 PO 4 ·3H 2 O, 0.1g / LMgSO 4 7H 2 O, 0.05g / LMnSO 4 , 1mL / L Tween 80, 0.5g / L cysteine ​​amino acid salt and 20g / L agar, pH 6.8.

[0058] The composition of MRS liquid medium is: 10g / L peptone, 10g / L beef extract, 20g / L glucose, 2g / L sodium acetate, 5g / L yeast powder, 2g / L diammonium hydrogen citrate, 2.6g / L K 2 PO4·3H 2 O, 0.1g / LMgSO 4 7H 2 O, 0.05g / L MnSO 4 , 1 mL / L Tween 80 and 0.5 g / L cysteine ​​salt, pH 6.8.

[0059] (2) The bacterial suspension preparation method involved in the following examples

[0060] The activated liquid of LP-Onlly was inoculated into MRS liquid culture medium at a volume ratio of 2% (v / v), and cultured at a constant temperature of 37°C. The obtained fermentation liquid was centrifuged at 8000r / min for 5min at 4°C, and the supernatant was discarded. The bacteria were washed twice with sterile PBS and resuspended in sterile PBS to obtain a bacterial suspension. The number of viable bacteria was determined using MRS solid culture medium, and the bacterial suspension was mixed with 10% sterilized skim milk at a mass ratio of 1:1 and frozen, and the number of viable bacteria in the bacterial suspension was adjusted to 1.0×10 9 CFU / mL and 1.0×10 10CFU / mL, the bacterial suspension was packaged according to the daily requirement, stored in an ultra-low temperature refrigerator for use in the following examples.

[0061] Example 1: Improvement of alcoholic fatty liver disease in mice by Lactobacillus plantarum LP-Onlly

[0062] 1. Animal grouping and establishment of alcoholic fatty liver model

[0063] Thirty-five C57BL / 6J mice were adaptively fed for 7 days and then randomly divided into 5 groups, with 7 mice in each group (n=7), namely, blank control group (Con group), alcohol model group (Mod group), low-dose Lactobacillus plantarum LP-Onlly group (low-dose LP-Onlly group), high-dose Lactobacillus plantarum LP-Onlly group (high-dose LP-Onlly group), and positive drug tiopronin group (LP group).

[0064] All the mice in the above groups were fed with basic feed, and the mice were free to eat and drink water.

[0065] Mod group: mice were gavaged with alcohol every morning. The alcohol concentration was 20% (v / v) on the 1st to 3rd day of gavage, and 25% (v / v) on the 4th to 6th day of gavage. The alcohol concentration of gavage was increased by 5% every three days until the final concentration of alcohol was 40% (v / v). It was no longer increased. The alcohol concentration was maintained at 40% for gavage until the end of the experiment. The time of gavage with alcohol was from the 1st week to the 10th week. The gavage volume of alcohol was 10mL / kg based on the weight of the mouse. The mice were intervened from the 11th week, and the gavage mass concentration of skim milk powder solution was 10mL / kg. The gavage volume of skim milk powder solution was 10mL / kg based on the weight of the mouse. The mice were killed at the end of the 14th week of gavage. The mice were fasted for 12 hours before being killed, and water was freely available. Blood was collected from the eyeball vein of the mice.

[0066] Low-dose LP-Onlly group: mice were gavaged with alcohol every morning. The alcohol concentration was 20% (v / v) on the 1st to 3rd day of gavage, and 25% (v / v) on the 4th to 6th day of gavage. The alcohol concentration was increased by 5% every three days until the final concentration of alcohol was 40% (v / v). It was no longer increased and the final concentration of alcohol was maintained at 40% for gavage until the end of the experiment. The duration of alcohol gavage was from the 1st week to the 10th week. The amount of alcohol gavage was 10mL / kg based on the weight of the mice. The mice were treated starting from the 11th week. The bacterial concentration in the low-dose group was approximately 10 9 The LP-Onlly suspension with CFU / mL was administered intragastrically at a volume of 10 mL / kg based on the mouse body weight. The mice were killed at the end of the 14th week of intragastrically administration. The mice were fasted for 12 hours before being killed, and were allowed to drink water freely. Blood was collected from the eyeball vein of the mice.

[0067] High-dose LP-Onlly group: mice were gavaged with alcohol every morning. The alcohol concentration was 20% (v / v) on the 1st to 3rd day of gavage, and 25% (v / v) on the 4th to 6th day of gavage. The alcohol concentration was increased by 5% every three days until the final concentration of alcohol was 40% (v / v). It was no longer increased and the final concentration of alcohol was maintained at 40% for gavage until the end of the experiment. The duration of alcohol gavage was from the 1st week to the 10th week. The amount of alcohol gavage was 10mL / kg based on the weight of the mice. The mice were treated starting from the 11th week. The bacterial concentration in the high-dose group was about 10 10 The LP-Onlly suspension with CFU / mL was administered intragastrically at a volume of 10 mL / kg based on the mouse body weight. The mice were killed at the end of the 14th week of intragastrically administration. The mice were fasted for 12 hours before being killed, and were allowed to drink water freely. Blood was collected from the eyeball vein of the mice.

[0068] LP group: mice were gavaged with alcohol every morning. The alcohol concentration was 20% (v / v) on the 1st to 3rd day of gavage, and 25% (v / v) on the 4th to 6th day of gavage. According to the above method, the concentration of alcohol for gavage was increased by 5% every three days until the final concentration of alcohol was 40% (v / v), and it was no longer increased. The final concentration of alcohol was maintained at 40% for gavage until the end of the experiment. The time of gavage with alcohol was from the 1st week to the 10th week. The amount of alcohol gavage was 10mL / kg. The mice were treated and intervened from the 11th week, and the positive drug intervention dose was 65mg / kg / d. The mice were killed at the end of the 14th week of gavage. The mice were fasted for 12 hours before being killed, and water was freely available. Blood was collected from the eye vein of the mice.

[0069] Con group: mice were gavaged with an equal volume of sterile distilled water every morning, with a gavage dose of 10 mL / kg, and the time of gavage of distilled water was from week 1 to week 10. The mice were intervened from week 11, and the gavage mass concentration of skim milk powder solution was 10 mL / kg based on the weight of the mice. The mice were killed at the end of the 14th week after gavage. The mice were fasted for 12 hours before being killed, and water was freely available. Blood was collected from the eyeball vein of the mice.

[0070] Six mice were sacrificed in each group for measurement. The results are as follows:

[0071] 1. Mouse liver histopathological results

[0072] The tissue at the same position of the left lobe of the mouse liver was fixed with a 4% paraformaldehyde solution for 5 to 6 hours, then embedded in paraffin, sectioned, stained with Oil Red and H&E, and observed and photographed under a microscope. The results are as follows: Figure 1 and Figure 2As shown. In the Con group, the hepatocyte cord structure was intact and radial, the hepatic lobule structure was clear, the hepatocytes were clearly demarcated and the cytoplasm was uniform, there was no fat cavitation and inflammatory infiltration, and only a small amount of lipid accumulation; in the Mod group, there was obvious fatty degeneration, the hepatocytes were swollen and deformed, the cytoplasm was turbid with a large number of fat cavities, a large amount of lipid accumulation in the hepatocytes, and inflammatory cell infiltration appeared. It can be seen that compared with the Con group mice, the liver of the alcohol modeling group mice had severe pathological damage; compared with the Mod group mice, the pathological damage of the liver tissue of the high and low doses of LP-Onlly and LP group mice was significantly reduced, mainly manifested in the reduction of fat vacuoles, reduced fatty degeneration, and reduced lipid accumulation. Among them, the high dose of LP-Onlly group had the best improvement effect.

[0073] 2. Effects of Lactobacillus plantarum LP-Onlly on body weight and liver index of mice

[0074] The liver index is the ratio of liver to body weight, which can reflect the nutritional status of experimental animals and the health status of the liver to a certain extent. The body weight, liver weight and liver index of each group of mice are shown in Table 1. As can be seen from Table 1, there was no significant difference in the initial body weight of each group of mice. The body weight of each group of mice increased after 14 weeks of gavage, but there was no significant difference in the body weight of mice between the groups. After the experiment, the liver index of the Mod group mice was significantly increased compared with the Con group (P<0.05); compared with the Mod group mice, the liver index of the high and low doses of LP-Onlly and LP groups was significantly reduced (P<0.05), but did not return to the normal level of the Con group.

[0075] Table 1 Effects of Lactobacillus plantarum LP-Onlly on body weight and liver index of mice

[0076] Group Initial body weight (g) Final body weight (g) Liver weight (g) Liver index (%) Con <![CDATA[21.11±1.14 a ]]> <![CDATA[25.40±3.30 a ]]> <![CDATA[0.85±0.07 a ]]> <![CDATA[3.35±0.16 c ]]> Mod <![CDATA[20.94±0.85 a ]]> <![CDATA[22.65±1.63 a ]]> <![CDATA[1.01±0.12 a ]]> <![CDATA[4.47±0.15 a ]]> Low-dose LP-Onlly <![CDATA[20.85±1.02 a ]]> <![CDATA[24.74±1.62 a ]]> <![CDATA[0.99±0.09 a ]]> <![CDATA[4.01±0.15 b ]]> High Dose LP-Onlly <![CDATA[21.32±0.70 a ]]> <![CDATA[24.14±0.63 a ]]> <![CDATA[0.93±0.09 a ]]> <![CDATA[3.80±0.37 b ]]> LP <![CDATA[21.29±1.17 a ]]> <![CDATA[22.03±1.66 a ]]> <![CDATA[0.85±0.14 a ]]> <![CDATA[3.80±0.40 b ]]>

[0077] Note: Data are expressed as Mean±SD (n=5-6, different letters represent significant differences among different groups, P<0.05).

[0078] 3. Effects of Lactobacillus plantarum LP-Onlly on TG and TC in mouse liver

[0079] In order to evaluate the effect of chronic alcohol intake on lipid accumulation in the liver of mice, the contents of triglyceride (TG) and cholesterol (TC) in the liver of each group of mice were measured. Figure 3 As shown in Table 2. Figure 3As shown in Table 2, after alcohol intervention, the TG and TC contents in the liver of the Mod group mice were significantly increased compared with those in the Con group (P<0.05). Compared with the Mod group, the TG and TC contents in the liver of the mice were significantly decreased after intervention with high-dose LP-Onlly, low-dose LP-Onlly and LP groups, among which high-dose LP-Onlly and LP intervention restored the TC content in the liver of the mice to the normal level of the Con group, but there was no significant difference among the three intervention methods in improving the liver TG level.

[0080] Table 2 Data on TG and TC content in mouse liver

[0081]

[0082] Note: “-” means no data, the same below.

[0083] 4. Effects of Lactobacillus plantarum LP-Onlly on liver damage-related indicators in mice

[0084] Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH) were measured to measure liver damage, and the results are shown in Table 3 .

[0085] Table 3 Indicators related to liver damage in mice

[0086]

[0087] Note: Data are expressed as Mean±SD (n=5-6), different letters represent significant differences among different groups, P<0.05.

[0088] ALT is mainly distributed in the cytoplasm of liver cells. Elevated serum ALT reflects damage to the liver cell membrane. AST is mainly distributed in the cytoplasm of liver cells and mitochondria of liver cells. Elevated serum AST indicates that liver cell damage has reached the organelle level. According to Table 2, liver damage biomarkers are all increased due to long-term chronic drinking, but are reduced by high LP-Onlly, low-dose LP-Onlly and LP intervention, which indicates that all three intervention methods can play a role in protecting liver damage. Overall, high-dose LP-Onlly has the best effect.

[0089] 5. Effects of Lactobacillus plantarum LP-Onlly on the activities of enzymes related to alcohol metabolism in mice

[0090] Normal liver has two ethanol oxidation metabolism systems, one is composed of alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH), and the other is the microsomal ethanol oxidation system (MEOS) composed of cytochrome enzyme CYP2E1. Figure 4As shown in Table 4, drinking significantly reduced the activities of ADH and ALDH in the liver. After high-dose and low-dose LP-Onlly interventions and low-dose LP-Onlly interventions, the ADH activity in the liver of mice increased by 48.05% and 55.96%, respectively, and the ALDH activity increased by 175.32% and 99.59%, respectively, compared with the Mod group. Therefore, LP-Onlly can significantly reverse the decrease in alcohol metabolism enzyme activity caused by alcohol. CYP2E1 is considered an important factor in liver steatosis and increased oxidative stress. Excessive alcohol intake can lead to a significant increase in CYP2E1 activity (P<0.05), but all three intervention methods can significantly reduce its activity, but there is no significant difference.

[0091] Table 4 Activity data of enzymes related to alcohol metabolism in mouse liver

[0092]

[0093] 6. Effects of Lactobacillus plantarum LP-Onlly on oxidative stress in mouse liver

[0094] Oxidative stress is a recognized factor in the progression of liver damage. The oxidative and antioxidant systems in a normal body always maintain a balanced state, but alcohol can break this balance, reduce the activity of related antioxidant enzymes and reduce antioxidant substances, which is consistent with our research results. Figure 5 As shown in Table 5, long-term alcohol intake reduces superoxide dismutase (SOD), catalase (CAT) activity and glutathione (GSH) content, produces a large amount of ROS, and thus increases liver MDA content. Compared with the Con group, the SOD, CAT activity and GSH content in the liver of the Mod group mice decreased by 40.34%, 38.65% and 50.75%, respectively. The three intervention methods effectively reversed this decrease and inhibited the increase in oxidative stress caused by drinking, but there was no significant difference in their effects. Drinking also caused a significant increase in the MDA content in the liver of the Mod group mice compared with the Con group mice (P<0.05). After the three intervention methods, the MDA levels were reduced by 25.83%, 29.06% and 24.00%, respectively, compared with the Mod group mice, but did not return to normal values.

[0095] Table 5 Data on oxidative stress levels in mouse liver

[0096]

[0097] 7. Effects of Lactobacillus plantarum LP-Onlly on inflammatory factors in mouse liver and serum LPS content

[0098] TNF-α is a mediator of mammalian inflammatory response. As the main proinflammatory cytokine in alcoholic liver disease, it participates in inflammatory response, leading to fatty degeneration and cell death. IL-6 can come from lymphocytes or be synthesized by non-lymphocytes, such as hepatocytes and Kupffer cells. The secreted IL-6 can in turn stimulate the synthesis of more other cytokines, thereby accelerating the progression of liver disease and aggravating liver damage. Figure 6 As shown in Table 6, after alcohol intervention, the levels of TNF-α and IL-6 in the Mod group mice increased by 25.53% and 54.21% respectively compared with the Con group mice. After the three interventions, the inflammatory level decreased. The high-dose LP-Onlly group restored TNF-α to 581.18±31.38g / mL, which was not significantly different from the Con group. This indicates that the three interventions can alleviate the inflammatory response caused by drinking to a certain extent, and the high-dose LP-Onlly group has the best effect in reducing TNF-α. Figure 6 As shown in Figure c, after 14 weeks of alcohol intervention, the serum LPS level of mice in the Mod group increased significantly compared with that in the Con group (P<0.05). After the three intervention methods, the serum LPS levels decreased. Among them, the serum LPS content of mice in the high LP-Onlly group and the low LP-Onlly group recovered to a level that was not significantly different from that in the Con group.

[0099] Table 6 Data on mouse liver inflammatory factors and serum LPS content

[0100]

[0101] 8. Effects of Lactobacillus plantarum LP-Onlly on lipid oxidation-related genes in mice

[0102] Lipin-1α, PPAR-α, and PGC-1α are important genes regulating liver lipid oxidation. Figure 7 As shown in Table 7, after 14 weeks of alcohol intervention, the expression of the three genes in the liver of the Mod group mice was downregulated by 64.8%, 71.39% and 61.5% respectively compared with the Con group. After the three interventions, the expression of the three genes was significantly increased (P<0.05), among which the high-dose LP-Onlly group and the low-dose LP-Onlly group had better effects. Compared with the Mod group, the low-dose LP-Onlly group upregulated the three genes of Lipin-1α, PPAR-α and PGC-1α by 210%, 337% and 285%, respectively, and the high-dose LP-Onlly group upregulated them by 224%, 376% and 281%, respectively. There was no significant difference between the high-dose and low-dose LP-Onlly groups in regulating the expression of the three genes.

[0103] Table 7 Expression levels of lipid oxidation-related genes in mouse liver

[0104]

[0105]

[0106] Example 2 Preparation of probiotic freeze-dried powder for alleviating alcoholic fatty liver by Lactobacillus plantarum LP-Onlly

[0107] The plant lactobacillus LP-Onlly was inoculated into MRS culture medium, fermented and cultured at 37°C to obtain bacterial culture solution, centrifuged at 8000r / min for 5 minutes, the supernatant was removed to obtain bacterial cells, the bacterial cells were resuspended in skim milk with a mass concentration of 10% to obtain a resuspension, and the resuspension was freeze-dried to obtain probiotic freeze-dried bacterial powder.

[0108] Example 3 Preparation of hangover-relief and liver-protecting yogurt products by Lactobacillus plantarum LP-Onlly

[0109] Heat 100 kg of skim milk to 90°C and maintain for 10 min, immediately cool to 40°C, inoculate with 0.1 kg of freeze-dried powder of Lactobacillus plantarum LP-Onlly, ferment at 40°C until the pH reaches 4.5±0.05, then terminate the fermentation, and place at 4°C for post-ripening for 44 h.

[0110] Example 4 Preparation of fermented soybean milk beverage by Lactobacillus plantarum LP-Onlly

[0111] A fermented soy milk beverage for relieving alcoholic fatty liver is prepared. 0.01 kg of freeze-dried powder of Lactobacillus plantarum LP-Onlly is added into a base material, which is composed of 100 kg of soybean milk, 3 kg of glucose, 35 kg of white sugar and 1 kg of stabilizer. The base material is fermented at 37°C to a pH value of 3.9 to 4.1, and 0.03 kg of edible flavor is added to obtain the fermented soy milk beverage. The number of live bacteria of Lactobacillus plantarum LP-Onlly in the obtained fermented soy milk beverage is not less than 1.0×10 9 CFU / mL.

[0112] In summary, the Lactobacillus plantarum LP-Onlly of the present invention can alleviate alcoholic fatty liver caused by chronic alcohol intake, reduce liver lipid accumulation caused by long-term alcohol intake, increase liver lipid consumption, and alleviate liver inflammatory response.

[0113] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.

Claims

1. Application of Lactobacillus plantarum LP-Only in the preparation of products for alleviating alcoholic liver disease.

2. The use according to claim 1, characterized in that: The number of viable Lactobacillus plantarum LP-Onlly in the product is ≥ 1×10 9 CFU / mL.

3. The use according to claim 1, characterized in that: The Lactobacillus plantarum LP-Only is used for any one or two or more of the following (1) to (9): (1) Reduce liver index; (2) Reduce triglyceride levels in the liver; (3) Reduce cholesterol levels in the liver; (4) Reduce the activity of alanine aminotransferase and / or aspartate aminotransferase in the liver, and increase the activity of alcohol dehydrogenase and / or acetaldehyde dehydrogenase in the liver; (5) increasing superoxide dismutase and / or catalase activity in the liver; (6) Increase liver glutathione content; (7) Reduce liver malondialdehyde levels; (8) Reduce liver inflammation; (9) Increase the mRNA expression levels of liver lipid metabolism-related genes Lipin-1α, PPAR-α and PGC-1α.

4. The use according to claim 1, characterized in that: The alcoholic liver disease includes alcoholic fatty liver.

5. The use according to claim 1 or 2, characterized in that: The products include pharmaceutical products.

6. Use of Lactobacillus plantarum LP-Only in the preparation of functional foods and / or health products for protecting or assisting in protecting the liver.

7. The use according to claim 6, characterized in that: The functional food includes yogurt and / or fermented soybean milk beverage, and the health care product includes probiotic powder.

8. The use according to claim 7, characterized in that: The preparation method of the probiotic powder comprises: mixing the plant lactobacillus LP-Onlly and a protective agent, and then freeze-drying to obtain the probiotic powder; the protective agent comprises one or more of skim milk, gelatin, maltodextrin, modified starch, gum arabic, sucrose, lactose, trehalose, sorbitol and xylitol.

9. The use according to claim 7, characterized in that: The method for preparing yogurt comprises: inoculating the Lactobacillus plantarum LP-Onlly into dairy products for fermentation to obtain yogurt; The method for preparing the fermented soybean milk beverage comprises: inoculating the lactobacillus plantarum LP-Onlly into a base material for fermentation to obtain the fermented soybean milk beverage.

10. A product for alleviating alcoholic liver disease, characterized in that: Active ingredients include Lactobacillus plantarum LP-Onlly.

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