Composition, combined preparation and use with the effect of dispelling alcohol and protecting liver

Abstract

This invention relates to a composition, combined formulation, and application with hangover-relieving and liver-protecting effects, belonging to the field of biopharmaceutical technology. The composition comprises a probiotic compound powder and a kudzu root and Japanese raisin tree fruit compound liquid. The probiotic compound powder includes: *Lactobacillus reuteri* ZKAW05, *Lactobacillus acidophilus* NCFM, *Lactobacillus plantarum* ZKAW02, *Lactobacillus paracasei* Lpc-37, *Lactobacillus casei* LC15, *Bifidobacterium breve* M-16V, fructooligosaccharides, isomaltooligosaccharides, xylooligosaccharides, konjac glucomannan, L-arabinose, sorbitol, and magnesium stearate. The kudzu root and Japanese raisin tree fruit compound liquid includes: kudzu root powder, prickly pear powder, Japanese raisin tree fruit powder, *Ficus hirta* powder, corn oligopeptide powder, fructooligosaccharides, flavoring fruit juice powder, citric acid monohydrate, and water. The composition of this invention repairs alcohol-induced structural and functional damage to the liver from multiple dimensions, achieving a level of liver protection comparable to existing chemical drugs.

Description

Technical Field

[0001] This invention relates to a composition, combination formulation, and application that has hangover-relieving and liver-protecting effects, belonging to the field of biopharmaceutical technology. Background Technology

[0002] Alcoholic liver disease (ALD) is one of the most prevalent liver diseases worldwide. Its pathogenesis is complex, involving multiple pathological processes such as alcohol metabolism disorders, oxidative stress, inflammatory responses, and gut microbiota imbalance. After alcohol enters the body, approximately 90% is metabolized and broken down by the liver. Its metabolic pathways are mainly divided into two major pathways: oxidative metabolism and non-oxidative metabolism. In the oxidative pathway, the alcohol dehydrogenase (ADH)-acetaldehyde dehydrogenase (ALDH) system is the core metabolic pathway for low-concentration alcohol intake, while long-term heavy drinking abnormally activates the cytochrome P450 2E1 (CYP2E1) enzyme system. Overactivation of the CYP2E1 pathway continuously produces large amounts of reactive oxygen species (ROS), inducing oxidative stress damage and disrupting the normal physiological function of hepatocytes. Simultaneously, acetaldehyde, an intermediate metabolite of alcohol, is a core toxic substance mediating liver damage. It can form stable adducts with proteins and DNA, triggering lipid peroxidation, depleting glutathione (GSH), inducing mitochondrial toxicity, and directly inducing hepatocyte apoptosis and systemic inflammatory responses. In addition, toxic metabolites such as fatty acid ethyl esters (FAEEs) and ethyl glucuronide (EtG) generated by non-oxidative metabolic pathways can also directly damage the mitochondrial structure and biomembrane integrity of hepatocytes, further accelerating the liver damage process.

[0003] The vicious cycle of oxidative stress and inflammation is the core pathological mechanism driving the continuous progression of alcoholic liver injury. Excessive ROS can activate key inflammatory signaling pathways such as NF-κB, prompting the massive release of pro-inflammatory factors such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), forming a cascade amplification effect of "oxidative stress-inflammatory response." Simultaneously, long-term alcohol consumption directly damages the integrity of the intestinal mucosal barrier, leading to intestinal flora imbalance, excessive proliferation of Gram-negative bacteria, and ectopic lipopolysaccharide (LPS). Ectopic endotoxins enter the liver via the portal vein circulation, continuously activating hepatic Kupffer cells and further amplifying the local inflammatory response in the liver. This disruption of the "gut-hepatic axis" regulatory pathway has been proven to be a crucial pathogenic mechanism in the development and progression of alcoholic liver injury. Gut flora imbalance not only exacerbates intestinal mucosal barrier damage but also interferes with fatty acid metabolism, bile acid homeostasis, and immune regulation, becoming a significant driver of the progression of alcoholic liver disease.

[0004] Currently, clinical treatment for alcoholic liver injury primarily relies on hepatoprotective and anti-inflammatory chemotherapeutic drugs (such as silymarin and polyene phosphatidylcholine). While these drugs can lower transaminase levels and alleviate hepatocellular damage in the short term, long-term use has significant drawbacks, including marked toxic side effects, poor patient tolerance, and a high risk of relapse, making it difficult to meet the clinical needs for long-term prevention and radical treatment of alcoholic liver injury. Therefore, developing safe, efficient, natural, non-toxic, and long-term intervention methods has become a core focus in the current research field of alcoholic liver injury.

[0005] Probiotics, as a class of active microbial preparations that colonize the host's gut and effectively regulate the balance of the gut microbiota, have shown great application potential in the intervention and treatment of alcoholic liver injury due to their multiple biological functions, including regulating gut microbiota structure, repairing the intestinal mucosal barrier, anti-oxidation, and anti-inflammation. In recent years, numerous basic studies have confirmed the significant protective effect of probiotics against alcoholic liver injury. For example, *Lactobacillus plantarum* can exert liver-protective effects by activating the Nrf2 antioxidant signaling pathway and increasing the expression of intestinal tight junction proteins; *Bifidobacterium* can effectively regulate gut microbiota structure and improve the body's immune homeostasis; single strains such as *Lactobacillus acidophilus*, as well as multi-strain compound probiotic preparations, can alleviate hepatocyte damage through pathways such as repairing the intestinal mucosal barrier, reducing endotoxin ectopication, lowering the body's oxidative stress level, and inhibiting liver inflammatory responses. However, the following shortcomings still exist in the existing technology: (1) The target of a single strain is limited, making it difficult to comprehensively combat the multi-pathway pathogenic mechanism of alcoholic liver injury; (2) The strain compatibility of existing compound probiotic preparations lacks systematic optimization for alcohol metabolism pathways, and the synergistic effect on acetaldehyde toxicity clearance and CYP2E1 pathway inhibition is insufficient; (3) The strategy of combining probiotics with specific prebiotics, plant extracts, etc. is not yet clear, and the synergistic potential of multiple components in the regulation of the "gut-liver axis" has not been fully utilized. Summary of the Invention

[0006] To address the shortcomings of the existing technologies, this invention provides a composition, combined formulation, and application with hangover-relieving and liver-protecting effects. By scientifically combining probiotic strains with specific alcohol metabolism regulation functions with functional prebiotics and natural plant extracts, it aims to exert synergistic liver-protecting effects from multiple dimensions, such as accelerating alcohol metabolism, eliminating acetaldehyde toxicity, inhibiting oxidative stress and inflammatory responses, and repairing the intestinal barrier, providing a safe and efficient solution for long-term intervention of alcoholic liver damage.

[0007] The technical solution provided by this invention is as follows: One objective of this invention is to provide a composition with hangover-relieving and liver-protecting effects, comprising a probiotic compound powder and a kudzu root and Japanese raisin tree fruit compound liquid. The probiotic compound powder comprises: *Lactobacillus reuteri* ZKAW05, *Lactobacillus acidophilus* NCFM, *Lactobacillus plantarum* ZKAW02, *Lactobacillus paracasei* Lpc-37, *Lactobacillus casei* LC15, *Bifidobacterium breve* M-16V, fructooligosaccharides, isomaltooligosaccharides, xylooligosaccharides, konjac glucomannan, L-arabinose, sorbitol, and magnesium stearate. The kudzu root and Japanese raisin tree fruit compound liquid comprises: kudzu root powder, prickly pear powder, Japanese raisin tree fruit powder, *Ficus hirta* powder, corn oligopeptide powder, fructooligosaccharides, flavoring fruit juice powder, citric acid monohydrate, and water.

[0008] Lactobacillus reuteri ZKAW05 was deposited on December 26, 2024, at the Guangdong Provincial Center for Microbial Culture Collection (GDMCC), Guangzhou, China, with accession number GDMCC NO: 65682, and classified as follows: Limosilactobacillus reuteri.

[0009] Lactobacillus plantarum ZKAW02 was deposited on September 27, 2024, at the Guangdong Provincial Center for Microbial Culture Collection, Guangzhou, China, with accession number GDMCC NO: 65219, and classified as follows: Lactiplantibacillus paraplantarums.

[0010] Based on the above technical solution, the present invention can be further improved as follows: Further, by weight percentage, the probiotic compound powder comprises: Lactobacillus reuteri ZKAW05 0.5~1.5%, Lactobacillus acidophilus NCFM 0.5~1.5%, Lactobacillus plantarum ZKAW02 0.5~1.5%, Lactobacillus paracasei Lpc-37 0.05~0.15%, Lactobacillus casei LC15 0.05~0.15%, Bifidobacterium breve M-16V 0.05~0.15%, fructooligosaccharides 8~12%, isomaltooligosaccharides 8~12%, xylooligosaccharides 8~12%, konjac glucomannan 3~7%, L-arabinose 3~7%, sorbitol 50~60%, and magnesium stearate 1~3%.

[0011] Further, by weight percentage, the kudzu root and Japanese raisin tree compound liquid comprises: 0.6-0.8% kudzu root powder, 0.6-0.8% prickly pear powder, 0.6-0.8% Japanese raisin tree powder, 0.6-0.8% five-finger peach powder, 0.3-0.4% corn oligopeptide powder, 6-8% fructooligosaccharides, 9-12% flavoring fruit juice powder, 0.06-0.08% citric acid monohydrate, and 75-85% water. Furthermore, in each gram of the probiotic compound powder, the viable count of *Lactobacillus reuteri* ZKAW05 is 2-5 billion CFU, the viable count of *Lactobacillus acidophilus* NCFM is 1-5 billion CFU, the viable count of *Lactobacillus plantarum* ZKAW02 is 5-10 billion CFU, the viable count of *Lactobacillus paracasei* Lpc-37 is 1-5 billion CFU, the viable count of *Lactobacillus casei* LC15 is 250-250 million CFU, and the viable count of *Bifidobacterium breve* M-16V is 1-5 billion CFU.

[0012] Furthermore, the ratio of probiotic compound powder to kudzu root and Japanese raisin tree fruit compound liquid is 0.5~3g: 25~50mL.

[0013] Furthermore, the ratio of probiotic compound powder to kudzu root and Japanese raisin tree fruit compound liquid is 1~3g: 25~35mL.

[0014] The second objective of this invention is to provide a combined formulation comprising an upper chamber and a lower chamber, wherein the upper chamber is a probiotic compound tablet made from the probiotic compound powder in the composition described above, and the lower chamber is a kudzu root and Japanese raisin tree compound liquid in the composition described above.

[0015] A third objective of this invention is to provide the application of the composition described above, which has the effect of sobering up and protecting the liver, in the preparation of products that have an auxiliary protective effect against chemically induced liver damage.

[0016] The technical solution provided by this invention has the following advantages compared with the prior art: 1. This invention, through acute alcohol poisoning experiments in mice, confirms that the probiotic group, the compound liquid group, and the combination of both can significantly shorten the sleep time and prolong the latency period of alcohol intoxication in mice. Among them, the combined intervention group of probiotics and compound liquid components showed the best effect, which could quickly alleviate the intoxication state in mice, indicating that the composition of this invention can effectively accelerate alcohol metabolism and has good prospects for acute alcohol detoxification. Experiments using a mouse alcoholic liver injury model showed that, compared with the model group, the serum liver function indicators (ALT, AST) and blood lipid indicators (TG, TC) levels in each intervention group were significantly reduced, and the liver index decreased significantly, indicating that the composition of this invention can repair alcohol-induced liver structural and functional damage from multiple dimensions.

[0017] 2. The combined intervention group of the probiotics and compound liquid components of this invention showed significantly better liver damage relief than any single intervention group, confirming a clear synergistic effect between the two. Simultaneously, the improvement effect of the combined group on key biochemical indicators was similar to that of the positive control group of silymarin, a commonly used hepatoprotective drug, indicating that the composition of this invention can achieve the level of existing chemical drugs in liver protection.

[0018] 3. The combination of *Lactobacillus acidophilus* NCFM and *Lactobacillus paracasei* Lpc-37 in the probiotic compound powder of this invention, the combination of *Lactobacillus paracasei* and *Lactobacillus reuteri*, and the compatibility of kudzu root powder, *Hovenia dulcis* powder, corn oligopeptide powder, and *Prickly pear* powder in the kudzu root and *Hovenia dulcis* compound liquid all exhibit clear synergistic effects. Konjac glucomannan and L-arabinose exert their synergistic liver-protective potential through complementary mechanisms of physical barrier and metabolic regulation. These synergistic effects together constitute a systematic intervention system from intestinal microecological regulation to liver function protection.

[0019] 4. The body weight of mice in the alcoholic liver injury model group decreased significantly, while the body weight of mice in the probiotic intervention group, compound solution intervention group, and combined group of this invention all recovered significantly. However, no body weight recovery was observed in the silymarin-positive control group. These results indicate that the composition of this invention can effectively improve the overall health of the body and promote weight recovery after injury while exerting liver-protective effects. This is an additional advantage that existing chemical hepatoprotective drugs do not possess.

[0020] 5. The raw materials used in this invention are all probiotics and natural plant-based ingredients. Animal experiments have verified that no adverse reactions were observed, demonstrating good safety. This invention overcomes the shortcomings of traditional chemical liver-protecting drugs, such as significant toxic side effects, poor tolerance, and easy relapse after long-term use. It is suitable for long-term prevention and daily health maintenance of alcoholic liver injury. Based on the "gut-liver axis" regulation theory, this invention repairs the intestinal mucosal barrier and regulates the intestinal flora structure through probiotics. Combined with the antioxidant, anti-inflammatory, and alcohol metabolism-accelerating effects of the compound liquid components, it achieves multi-pathway and multi-target systemic intervention for alcoholic liver injury. The mechanism of action is clear, providing a new approach for a natural, safe, and long-term prevention and treatment strategy for alcoholic liver injury.

[0021] 6. The compositions and combined formulations of the present invention have controllable costs and stable quality, and are easy to achieve large-scale industrial production. They have good social and economic benefits and provide experimental evidence and industrialization basis for promoting the clinical translation and application of probiotics in the prevention and treatment of liver diseases. Attached Figure Description

[0022] Figure 1 The changes in body weight of mice in different groups (****P<0.0001); Figure 2 The changes in liver index in mice of different groups (*P<0.05, **P<0.01, ***P<0.001, ****P<0.0001). Figure 3 The changes in AST and ALT activity in mice from different groups are shown in Figure A; Figure B shows the changes in AST activity and Figure C shows the changes in ALT activity (*P<0.05, **P<0.01, ns indicates no statistical difference). Figure 4The changes in TG and TC in mice from different groups are shown; A is the graph showing the changes in TG, and B is the graph showing the changes in TC activity (*P<0.05, ***P<0.001, ****P<0.0001). Figure 5 The changes in MDA and SOD in mice from different groups are shown in Figure A; Figure B shows the changes in MDA activity (**P<0.01, ***P<0.001, ****P<0.0001, ns indicates no statistical difference). Detailed Implementation

[0023] The principles and features of the present invention are described below with reference to examples. The examples are only used to explain the present invention and are not intended to limit the scope of the present invention.

[0024] Lactobacillus acidophilus NCFM (Beijing Taipingsha Biotechnology Co., Ltd.) Lactobacillus paracasei Lpc-37 (Beijing Taipingsha Biotechnology Co., Ltd.) Lactobacillus casei LC15 (WeCon Probiotics Co., Ltd.) Bifidobacterium breve M-16V (Hebei Hanlan Biotechnology Co., Ltd.) Lactobacillus reuteri ZKAW05 was deposited on December 26, 2024, at the Guangdong Provincial Center for Microbial Culture Collection (GDMCC), Guangzhou, China, with accession number GDMCC NO: 65682, and classified as follows: Limosilactobacillus reuteri.

[0025] Lactobacillus plantarum ZKAW02 was deposited on September 27, 2024, at the Guangdong Provincial Center for Microbial Culture Collection, Guangzhou, China, with accession number GDMCC NO: 65219, and classified as follows: Lactiplantibacillus paraplantarums.

[0026] Silymarin (MADAUS GmbH, Germany) ALT Detection Kit (Chongqing Bainian Chuangxing Biotechnology Co., Ltd.) AST detection kit (Chongqing Bainian Chuangxing Biotechnology Co., Ltd.) TC Detection Kit (Chongqing Bainian Chuangxing Biotechnology Co., Ltd.) TG Detection Kit (Chongqing Bainian Chuangxing Biotechnology Co., Ltd.) SOD Detection Kit (Chongqing Bainian Chuangxing Biotechnology Co., Ltd.) MDA Detection Kit (Chongqing Bainian Chuangxing Biotechnology Co., Ltd.) Example 1 A composition with hangover-relieving and liver-protecting effects includes a probiotic compound powder and a kudzu root and Japanese raisin tree fruit compound liquid; The probiotic compound powder, by weight percentage, comprises: *Lactobacillus reuteri* ZKAW05 1%, *Lactobacillus acidophilus* NCFM 1%, *Lactobacillus plantarum* ZKAW02 1%, *Lactobacillus paracasei* Lpc-37 0.1%, *Lactobacillus casei* LC15 0.1%, and *Bifidobacterium breve* M-16V. 0.1%, fructooligosaccharides 10%, isomaltooligosaccharides 10%, xylooligosaccharides 10%, konjac glucomannan 5%, L-arabinose 5%, sorbitol 54.7%, and magnesium stearate 2%; in each gram of the probiotic compound powder, the viable count of *Lactobacillus reuteri* ZKAW05 is 2-5 billion CFU, the viable count of *Lactobacillus acidophilus* NCFM is 1-5 billion CFU, the viable count of *Lactobacillus plantarum* ZKAW02 is 5-10 billion CFU, the viable count of *Lactobacillus paracasei* Lpc-37 is 1-5 billion CFU, the viable count of *Lactobacillus casei* LC15 is 250-250 million CFU, and the viable count of *Bifidobacterium breve* M-16V is 1-5 billion CFU.

[0027] The kudzu root and Japanese raisin tree fruit compound liquid comprises: 0.69% kudzu root powder, 0.69% prickly pear powder, 0.69% Japanese raisin tree fruit powder, 0.69% five-finger peach powder, 0.35% corn oligopeptide powder, 6.9% fructooligosaccharides, 10.4% flavoring fruit juice powder (3.5% passion fruit juice powder and 6.9% concentrated apple juice), 0.069% citric acid monohydrate, and 79.521% water.

[0028] The ratio of the probiotic compound powder to the kudzu root and Japanese raisin tree fruit compound liquid is 2g:30mL.

[0029] A combined formulation comprising an upper chamber and a lower chamber, wherein the upper chamber is a probiotic compound tablet made from the probiotic compound powder in the composition as described above, and the lower chamber is a kudzu root and Japanese raisin tree compound liquid in the composition as described above.

[0030] test: I. Sobering-up test: Twenty-four Kunming mice (6 weeks old, weighing approximately 28g) were divided into four groups: a model group (MC group), a probiotic group (PG group), a compound liquid group (MG group), and a probiotic + compound liquid combined group (CG group), with six mice in each group. The animals in each group were fasted for 12 hours the night before the experiment, but had free access to water. Each group was administered 52° Erguotou liquor by gavage at a dose of 17 mL / kg. After administration, the mice were placed back-down, and their position was adjusted every 5 seconds until they could no longer immediately turn back down. When a mouse could not roll over on its own for 30 seconds, the reflex was considered to have disappeared (intoxication); when the mouse could roll over on its own, the reflex was considered to have recovered (awake state). After administering the liquor, the latency period of intoxication (time from intoxication to the disappearance of the reflex) and the sleep time (time from the disappearance of the reflex to its recovery) were recorded sequentially. The gavage dose was the dose with the highest intoxication rate and the lowest mortality rate. The time of liquor administration and the time of intoxication were recorded.

[0031] Thirty minutes later, the MC group was administered physiological saline by gavage, the PG group was administered a probiotic compound solution (2g of the probiotic compound powder from Example 1 dissolved in 30mL of physiological saline by gavage), the MG group was administered a kudzu root and Japanese raisin tree fruit compound solution from Example 1 by gavage, and the CG group was administered the composition prepared in Example 1 by gavage. Each group was administered the solution at a volume of 0.4 mL. The gavage time, intoxication time, and sobering time were recorded for each group of intoxicated mice, and the sleep time was calculated. The results are shown in Table 1. Table 1. Intoxication time, sobering time, and sleep time in mice.

[0032] Note: Compared with the MC group, **P<0.01, ****P<0.0001.

[0033] The results showed that the sobering-up time and sleep time of mice in the probiotic (PG), compound liquid (MG), and combined (CG) groups were significantly shorter than those in the model (MC) group, demonstrating good hangover relief effects, with the CG group showing the most significant effect.

[0034] II. Mouse Liver Injury Modeling and Intervention Experiment Forty-eight 6-8 week old C57 / 6J mice (weighing approximately 22g) were randomly divided into six groups of eight each: normal (NC), model (MC), probiotic (PG), compound solution (MG), probiotic + compound solution combination (CG), and positive control (PC). Every day at 9:00 AM, the PG group was administered the probiotic compound solution (2g of the probiotic compound powder from Example 1 dissolved in 30 mL of physiological saline), the MG group was administered the kudzu root and Japanese raisin fruit compound solution from Example 1, the CG group was administered the composition prepared in Example 1, the PC group was administered silymarin (10.8 mg of silymarin dissolved in 4 mL of physiological saline), and the NC group was administered physiological saline; the gavage volume for all groups was 0.4 mL. At 4:00 PM, except for the NC group, all other groups were administered 52° Erguotou liquor by gavage at a volume of 13 mL / kg for 14 consecutive days. After the intervention, mouse blood was collected, and after being placed at room temperature for 30 min, the serum was separated by centrifugation at 4℃ and 3000×g for 20 min and stored at -80℃. Serum ALT, AST, TC, TG levels, SOD activity and MDA content were detected, and liver wet weight was measured to calculate liver index.

[0035] 1. Changes in mouse body weight To investigate the effects of probiotics, kudzu root and Japanese raisin tree fruit compound solution, and the combined preparation on mouse body weight, the mice were weighed daily during the intervention period, and changes in body weight were observed. The results are as follows: Figure 1 As shown.

[0036] The results showed that compared with the normal (NC) group, the model (MC) group mice had a significant decrease in body weight, indicating that long-term heavy drinking can cause weight loss. However, the probiotic (PG) group, the compound liquid (MG) group, and the probiotic + compound liquid combination (CG) group all showed a significant increase in body weight, indicating that taking probiotics and the components of the kudzu root and Japanese raisin tree compound liquid after drinking alcohol had significant protective effects. However, the positive control (PC) group mice experienced the same degree of weight loss as the model (MC) group, indicating that silymarin has no protective effect against weight loss caused by long-term excessive alcohol consumption.

[0037] 2. Comparison of liver indices The formula for calculating the liver index is: Liver Index (%) = (Wet weight of liver / Body weight before sacrifice) × 100%.

[0038] The results are as follows Figure 2 As shown in the figure. The results showed that compared with the NC group, the liver index of mice in the MC group increased, indicating liver edema. Compared with the MC group, the liver index of the PG, MG, and CG groups was significantly reduced and returned to the normal level, indicating that the probiotics and the components of the Pueraria lobata and Hovenia dulcis compound liquid both have the effect of relieving liver edema.

[0039] 3. Measurement of alanine aminotransferase (ALT) and aspartate aminotransferase (AST): (1) Take a sterile 96-well plate and set up test wells and control wells. Add 10 μL of serum sample to be tested to the test well, and leave the control wells empty. Add 20 μL of reagent one (matrix solution) to each of the test wells and control wells, and gently pipette to mix, avoiding the formation of air bubbles. Incubate the 96-well plate at 37°C for 30 min to start the enzymatic reaction.

[0040] (2) After incubation, add 20 μL of reagent 2 (2,4-dinitrophenylhydrazine solution) to each of the assay and control wells and mix gently. Add 10 μL of the serum sample to be tested to the control well and mix gently by pipetting to ensure no air bubbles are generated. Do not add additional sample to the assay wells. Incubate the 96-well plate at 37°C for 10 min to terminate the enzymatic reaction and initiate the colorimetric reaction.

[0041] (3) Add 200 μL of reagent three (sodium hydroxide solution) to each well and mix thoroughly. Incubate the 96-well plate at 25℃ (room temperature) for 10 min to allow the colorimetric reaction to proceed fully. After incubation, immediately measure the absorbance values ​​of the test wells and control wells at a wavelength of 520 nm using a microplate reader. Calculate the corresponding enzyme activity units.

[0042] The results are as follows Figure 3 As shown in the figure. The results showed that the serum ALT and AST activities in the MC group were significantly higher than those in the normal group, indicating successful model establishment. Compared with the MC group, the AST activities in the MG and CG groups were significantly lower, and better than those in the PC group, indicating that the MG and CG groups can effectively alleviate alcoholic liver injury, with the CG group showing the best effect. In terms of ALT activity, the PG, MG, CG, and PC groups were all significantly lower than the MC group, with the CG group showing the most significant decrease, indicating a better effect in alleviating alcoholic liver injury.

[0043] 4. Determination of triglycerides (TG) and total cholesterol (TC) (1) Take a sterile 96-well plate and set up test wells, standard wells (for one-time testing only), and blank wells (for one-time testing only). Add 5 μL of serum sample to the test well, add 5 μL of standard to the standard well, and add no sample or standard to the blank well. (The amount of TG and TC added may differ; refer to the instructions for use.) (2) Add 100 μL of reagent one to each of the test wells, standard wells and blank wells, and then add 100 μL of reagent two to each well. Gently blow and mix to avoid generating air bubbles. Place the 96-well plate at room temperature (25°C) and incubate in the dark for 10 min to allow the colorimetric reaction to proceed fully.

[0044] (3) After incubation, immediately measure the absorbance of each well at a wavelength of 510 nm using an ELISA reader and record it as A assay, A standard and A blank. Calculate the TG and TC contents.

[0045] The results are as follows Figure 4 As shown in the figure. The results showed that compared with the NC group, the serum TG and TC levels of mice in the MC group were significantly increased, indicating disordered hepatic lipid metabolism. Compared with the MC group, the serum TG and TC levels of mice in the PG, MG, and CG groups were significantly decreased, with the CG group showing the most significant decrease. The effect was comparable to that of the positive control group, indicating that the composition of the present invention has a certain effect on promoting lipid metabolism.

[0046] 5. Determination of malondialdehyde (MDA) and superoxide dismutase (SOD) (1) Take a sterile EP tube and add 300 μL of working solution and 25 μL of serum sample to be tested (and bring the total volume to 200 μL with physiological saline or distilled water), and mix gently. Subsequent calculations need to be corrected according to the actual sample volume. Place the EP tube in a 90-95℃ water bath for 30 min. After the reaction is complete, remove it quickly and place it in an ice bath to cool. Centrifuge the cooled sample at 4℃ and 12000 r / min for 10 min. Transfer 200 μL of supernatant to a 96-well plate and measure the absorbance at wavelengths of 532 nm and 600 nm, respectively. Calculate the MDA content in the serum.

[0047] (2) Set up sample tubes, sample control tubes, blank tube 1 (only once), and blank tube 2 (only once) in a 96-well plate. Add 70 μL of reagent 1 to all wells. Add samples and reagents according to the instructions. After adding samples, place the 96-well plate at room temperature (25℃) and in the dark for 30 min (the reaction time must be strictly controlled). After incubation, immediately measure the absorbance of each well at a wavelength of 450 nm using a microplate reader. Calculate the SOD activity in liver tissue.

[0048] The results are as follows Figure 5 As shown. By Figure 5 As shown in Figure (A), compared with the NC group, the MDA activity in the serum of mice in the MC group was significantly increased, indicating that liver oxidative stress was aggravated. Compared with the MC group, the MDA levels in the serum of mice in the PG, MG, and CG groups were significantly decreased, indicating that taking probiotics and the compound liquid of Pueraria lobata and Hovenia dulcis during alcohol consumption has significant antioxidant and liver-damage-reducing protective effects, with the CG group showing better results, comparable to the positive control group.

[0049] Depend on Figure 5 As shown in Figure (B), the SOD activity in the liver tissue of the MC group mice was slightly lower than that in the normal NC group. The SOD activity in the PC group, PG group, MG group, and CG group mice all showed varying degrees of improvement, but there was no significant difference.

[0050] III. Volunteer Experience Verification Twenty volunteers, aged 35-55, were selected for the trial. Volunteers drank alcohol until they were intoxicated. After drinking, each volunteer took this product (the composition prepared in Example 1) once, went to sleep, and upon waking the next day, compared their subjective feelings with those experienced without taking the product at the same level of alcohol consumption. The comparison results are shown in Table 1 below.

[0051] Table 1. Volunteer Overall Subjective Feelings Survey

[0052] In Table 1, "significantly effective" means waking up the next day without the smell of alcohol, dizziness, headache, nausea, gastrointestinal discomfort, or other symptoms, and with a return to normal energy and spirit; "effective" means that the above symptoms have changed to some extent; and "ineffective" means that the above symptoms have not changed significantly. As can be seen from Table 1, the composition prepared in this invention has the effect of sobering up and relieving hangovers.

[0053] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A composition with hangover-relieving and liver-protecting effects, characterized in that, The product comprises a probiotic compound powder and a kudzu root and Japanese raisin tree fruit compound liquid. The probiotic compound powder includes: *Lactobacillus reuteri* ZKAW05, *Lactobacillus acidophilus* NCFM, *Lactobacillus plantarum* ZKAW02, *Lactobacillus paracasei* Lpc-37, *Lactobacillus casei* LC15, *Bifidobacterium breve* M-16V, fructooligosaccharides, isomaltooligosaccharides, xylooligosaccharides, konjac glucomannan, L-arabinose, sorbitol, and magnesium stearate. The kudzu root and Japanese raisin tree fruit compound liquid includes: kudzu root powder, prickly pear powder, Japanese raisin tree fruit powder, *Ficus hirta* powder, corn oligopeptide powder, fructooligosaccharides, flavoring fruit juice powder, citric acid monohydrate, and water.

2. The composition with hangover-relieving and liver-protecting effects according to claim 1, characterized in that, By weight percentage, the probiotic compound powder comprises: Lactobacillus reuteri ZKAW05 0.5-1.5%, Lactobacillus acidophilus NCFM 0.5-1.5%, Lactobacillus plantarum ZKAW02 0.5-1.5%, Lactobacillus paracasei Lpc-37 0.05-0.15%, Lactobacillus casei LC15 0.05-0.15%, Bifidobacterium breve M-16V 0.05-0.15%, fructooligosaccharides 8-12%, isomaltooligosaccharides 8-12%, xylooligosaccharides 8-12%, konjac glucomannan 3-7%, L-arabinose 3-7%, sorbitol 50-60%, and magnesium stearate 1-3%.

3. The composition with hangover-relieving and liver-protecting effects according to claim 1, characterized in that, The kudzu root and Japanese raisin tree fruit compound liquid comprises, by weight percentage: 0.6-0.8% kudzu root powder, 0.6-0.8% prickly pear powder, 0.6-0.8% Japanese raisin tree fruit powder, 0.6-0.8% five-finger peach powder, 0.3-0.4% corn oligopeptide powder, 6-8% fructooligosaccharides, 9-12% flavoring fruit juice powder, 0.06-0.08% citric acid monohydrate, and 75-85% water.

4. The composition with hangover-relieving and liver-protecting effects according to claim 1 or 2, characterized in that, In each gram of the probiotic compound powder, the viable count of *Lactobacillus reuteri* ZKAW05 is 2-5 billion CFU, the viable count of *Lactobacillus acidophilus* NCFM is 1-5 billion CFU, the viable count of *Lactobacillus plantarum* ZKAW02 is 5-10 billion CFU, the viable count of *Lactobacillus paracasei* Lpc-37 is 1-5 billion CFU, the viable count of *Lactobacillus casei* LC15 is 250-250 million CFU, and the viable count of *Bifidobacterium breve* M-16V is 1-5 billion CFU.

5. The composition with hangover-relieving and liver-protecting effects according to claim 1, characterized in that, The ratio of probiotic compound powder to kudzu root and Japanese raisin tree fruit compound liquid is 0.5~3g: 25~50mL.

6. A combination formulation, characterized in that, It includes an upper compartment and a lower compartment, wherein the upper compartment is a probiotic compound tablet made from the probiotic compound powder in the composition according to any one of claims 1 to 5, and the lower compartment is a kudzu root and Japanese raisin tree compound liquid in the composition according to any one of claims 1 to 5.

7. The use of the composition with hangover-relieving and liver-protecting effects as described in any one of claims 1 to 5 in the preparation of products with auxiliary protective effects against chemically induced liver damage.

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