Development method of soft pear beverage capable of dispelling effects of alcohol

Abstract

The development method comprises the following steps: (1) extraction of soft pear juice: after ripening soft pears, peeling, naturally unfreezing and taking juice, and controlling the content of total phenols in the pear juice to be not less than 120mg / kg and the content of total flavonoids to be not less than 40mg / kg; (2) preparing a prepared liquid: mixing chrysanthemum, fructus lycii and pitted jujube, then adding drinking water to soak for 30 minutes, decocting for 20 minutes, and filtering to respectively obtain a liquid medicine A and a medicine residue A; adding drinking water into the medicine residue A, continuously decocting for 10 minutes, and filtering to respectively obtain medicine liquid B and medicine residue B; fully mixing the liquid medicine A and the liquid medicine B, and concentrating to obtain a prepared solution; and (3) cooling the prepared liquid to room temperature, uniformly mixing the cooled liquid with the soft pear juice, filling the soft pear juice according to 400ml / can, and sterilizing the soft pear juice at normal temperature. The method is simple, industrial production is easy, and the obtained hangover-alleviating drink not only has the hangover-alleviating effect, but also can enhance the immunity of the organism and strengthen the body.

Description

Technical Field

[0001] The invention relates to the technical field of hangover-relief beverages, in particular to a method for preparing a soft pear hangover-relief beverage. Background Art

[0002] Alcoholic liver injury, also known as alcoholic liver disease (ALD), refers to liver damage caused by long-term heavy drinking. In the early stage, it usually manifests as fatty liver, which can then develop into alcoholic hepatitis, liver fibrosis and cirrhosis. Modern research shows that alcoholic liver injury is the result of the interaction of multiple factors such as inflammatory response, oxidative stress, enterogenic endotoxins, inflammatory mediators and nutritional imbalance (especially protein) induced directly or indirectly by alcohol and its metabolites and derivatives in the metabolic process. It is a complex pathological process with multiple factors, multiple links and multiple pathways. With the increase of modern social activities, wine culture occupies an important position in people's daily life, alcohol consumption continues to increase, and the incidence of alcoholic liver disease is on the rise. According to the statistical analysis of hospitalized patients in hospitals, the proportion of alcoholic liver disease has continued to rise from 1.7% in 2002 to 10.5% in 2023. During the same period, the number of patients with severe alcoholic hepatitis has also increased by 3.35 times. The proportion of alcoholic liver disease in all hospitalized patients with liver disease and the proportion of alcoholic cirrhosis in all cirrhosis are constantly increasing. How to prevent and treat alcoholic liver damage is increasingly becoming the focus of attention.

[0003] Since hangover-detoxifying drinks can effectively relieve discomfort after drinking, protect the liver, and reduce the harm of alcohol to the body, the market demand is very strong. With the popularization of globalization and the Internet, the demand for hangover-detoxifying drinks in overseas markets is also increasing. However, due to the low threshold of the hangover-detoxifying drink industry, the product quality varies greatly, and there is a lack of high-quality high-end products.

[0004] TCM believes that: ① alcohol is a damp-heat toxic evil, which can easily turn into fire and produce phlegm, disturbing the mind; ② alcohol is a yang evil, which is ascending and dispersing, and can easily disturb the mind; ③ alcohol is pungent and dispersing, and can easily move to the internal organs, causing many symptoms. TCM syndrome differentiation and treatment of drunkenness is based on the basic principle of detoxification, clearing heat and removing dampness, among which the elimination of alcohol toxicity runs through the entire treatment. Specific methods mainly include sobering up and detoxifying, clearing heat and detoxifying, harmonizing the stomach and digesting food, and tonifying the spleen and replenishing qi. Summary of the invention

[0005] The technical problem to be solved by the invention is to provide a preparation method of a soft pear alcohol-relieving drink which is simple in method and easy in industrial production.

[0006] In order to solve the above problems, the preparation method of a soft pear alcohol-relieving drink of the present invention comprises the following steps:

[0007] ⑴ Extraction of soft pear juice:

[0008] After ripening, peel off the skin of soft pears, thaw naturally, and extract the juice. The total phenol content in the pear juice should be no less than 120 mg / kg, and the total flavonoid content should be no less than 40 mg / kg.

[0009] ⑵Preparation of solution:

[0010] Mix chrysanthemum, wolfberry and pitted jujube in a mass ratio of 1:1:1, then add 10 times the mass of drinking water of the mixture to soak for 30 minutes, and then boil for 20 minutes, and filter to obtain liquid medicine A and residue A respectively; add 5 times the mass of drinking water to the residue A and continue to boil for 10 minutes, and filter to obtain liquid medicine B and residue B respectively; the liquid medicine A and the liquid medicine B are fully mixed and concentrated to obtain a prepared solution;

[0011] ⑶ After the prepared liquid is cooled to room temperature, it is mixed with the soft pear juice at a volume ratio of 3:1, and filled into a can at 400 ml / can, and sterilized at room temperature.

[0012] Compared with the prior art, the present invention has the following advantages:

[0013] 1. The present invention uses Lanzhou specialty fruit soft pear as the main raw material, supplemented with appropriate Chinese medicinal materials, and has the characteristics of easy availability of raw materials and stable efficacy.

[0014] 2. The method of the present invention is simple and easy to industrialize. The obtained hangover-relieving drink not only has the effect of relieving hangover, but also can increase the body's immunity and strengthen the body.

[0015] 3. The present invention makes full use of the local characteristic resource of Lanzhou, soft pear, not only enhancing the development and utilization of the resource, but also expanding the utilization scope of the resource, further mobilizing the enthusiasm of fruit farmers, and promoting agricultural production and income increase. At the same time, it can also provide reference and reference for the utilization of other characteristic resources in Lanzhou. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] The specific implementation modes of the present invention are further described in detail below in conjunction with the accompanying drawings.

[0017] Figure 1 This is a graph showing the effect of soft pears in the present invention on liver function indicators in rats with alcoholic liver damage.

[0018] Figure 2 This is a graph showing the effect of soft pears in the present invention on blood lipid indexes in rats with alcoholic liver damage.

[0019] Figure 3 This is a graph showing the effect of soft pear of the present invention on SOD and CAT activities (left figure) and MDA, LPO contents and GSH-Px activity (right figure) in liver tissue of rats with alcoholic liver damage.

[0020] Figure 4This is a diagram showing the effect of soft pear in the present invention on pathological changes in the liver of rats with alcoholic liver injury (HE staining, ×200).

[0021] Figure 5 The left figure is the PLS-DA result and the right figure is the PLS-DA permutation test result.

[0022] Figure 6 The OPLS-DA and permutation test of the samples in the present invention. Among them: the upper left figure is the OPLS-DA results of the blank group and the model group; the upper right figure is the OPLS-DA permutation test results of the blank group and the model group; the lower left figure is the OPLS-DA results of the model group and the Ruan'erli group; the lower right figure is the OPLS-DA permutation test results of the model group and the Ruan'erli group.

[0023] Figure 7 The present invention is used to screen and identify differential metabolites. Among them: a is the effect of Ruan'er pear on the content of differential metabolites in the serum of rats with alcoholic liver injury; b is the correlation diagram of differential metabolites in the serum of rats with alcoholic liver injury; c is the clustering heat map of differential metabolites in the serum of rats with alcoholic liver injury.

[0024] Figure 8 This is a diagram of enrichment analysis of differential metabolite pathways in the present invention. DETAILED DESCRIPTION

[0025] A method for preparing a soft pear alcohol-relief drink comprises the following steps:

[0026] ⑴ Extraction of soft pear juice:

[0027] After the soft pears are ripened, the skin is peeled off, and the pears are thawed naturally to extract the juice. The total phenol content in the pear juice is controlled to be not less than 120 mg / kg, and the total flavonoid content is not less than 40 mg / kg.

[0028] Every 1000mL of pear juice is equivalent to 1460.7g of soft pear frozen pears, and its component content can be controlled by high performance liquid chromatography.

[0029] ⑵Preparation of solution:

[0030] Mix chrysanthemum, wolfberry and pitted jujube in a mass ratio of 1:1:1 (g / g), then add drinking water 10 times the mass of the mixture and soak for 30 minutes, then boil for 20 minutes, and filter to obtain medicinal solution A and medicinal residue A respectively; add drinking water 5 times the mass of the medicinal residue A and continue to boil for 10 minutes, and filter to obtain medicinal solution B and medicinal residue B respectively; after fully mixing and concentrating, the medicinal solution A and the medicinal solution B are obtained.

[0031] ⑶ After the prepared liquid is cooled to room temperature, mix it with soft pear juice in a volume ratio of 3:1 (mL / mL), fill it in 400 ml / can, and sterilize it at room temperature.

[0032] Soft pear, also known as frozen pear and perfume pear, is a specialty fruit of Gansu. It is not only sweet like honey, but also has the effects of promoting fluid and resolving phlegm, moistening dryness and relieving cough, clearing away heat and detoxifying, nourishing the body and curing diseases. It is a rare fruit that can be used as both medicine and food. Studies have shown that soft pear can not only remove 1,1-diphenyl-2-picrylhydrazyl radical (DPPH), hydroxyl radical (OH-) and superoxide anion free radical (O2 - ), can also enhance the activity of antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px), improve oxidative stress, eliminate inflammation, etc. "Compendium of Materia Medica" records that soft pear has the effects of cooling the heart, relieving inflammation, reducing fire and relieving alcohol.

[0033] The alcohol-detoxifying drink of the present invention uses Lanzhou's special fruit soft pear as the main raw material, supplemented with Chinese medicine for clearing away heat and detoxification, and soothing alcohol and invigorating the spleen, which conforms to the dialectical treatment rules of Chinese medicine for drunkenness. The medicinal properties of the ingredients of the detoxifying drink are as follows:

[0034] Soft pear: cool in nature, sweet in taste. It has many functions such as moistening the lungs, relieving cough, cooling the heart and reducing inflammation, reducing fire, relieving sores and toxins, and sobering up.

[0035] Chrysanthemum: slightly cold in nature, bitter and sweet in taste. Enters the lung and liver meridians. Has the effects of dispersing wind and clearing heat, calming the liver and improving eyesight, clearing away heat and detoxifying. Chrysanthemum tea can accelerate alcohol metabolism and relieve physical discomfort such as sore throat, mouth sores, etc. caused by excessive internal heat after drinking.

[0036] Lycium barbarum: neutral in nature, sweet in taste, and enters the liver and kidney meridians. Lycium barbarum has the effect of nourishing the liver and kidneys, improving intelligence and improving eyesight. It can relieve headaches and dizziness after drinking. By nourishing the kidneys, it can enhance the liver's detoxification and metabolic functions, reduce the time alcohol stays in the body, and relieve the degree of drunkenness.

[0037] Example 1

[0038] A method for preparing a soft pear alcohol-relief drink comprises the following steps:

[0039] ⑴ Extraction of soft pear juice:

[0040] After ripening, peel the soft pears, thaw naturally, and take 300 mL of juice. Test the pear juice to ensure that the total phenol content is not less than 120 mg / kg and the total flavonoid content is not less than 40 mg / kg.

[0041] ⑵Preparation of solution:

[0042] According to the preparation of 1000mL of the finished alcohol-relieving drink, 20g of chrysanthemum, 20g of wolfberry and 20g of pitted jujube are mixed, then 600g of drinking water is added to soak for 30 minutes, and then decocted for 20 minutes, and the liquid medicine A and the residue A are obtained by filtration; 300g of drinking water is added to the residue A and the decocted for 10 minutes, and the liquid medicine B and the residue medicine B are obtained by filtration; the liquid medicine A and the liquid medicine B are fully mixed and concentrated to 250mL to obtain the prepared solution.

[0043] (3) After 250 mL of the prepared liquid is cooled to room temperature, mix it with 750 mL of soft pear juice and fill it into a can at a rate of 400 mL / can. Sterilize it at room temperature to obtain the hangover drink.

[0044] The efficacy experiment of the obtained hangover-relieving drink was carried out:

[0045] Metabolomics is a new method for exploring the mechanism of drug action by studying the changes in small molecule metabolites. This method reveals the complex relationships within and between metabolic networks in organisms, as well as their response mechanisms to internal and external environments, by analyzing information such as the types, contents, and temporal changes of metabolites. Therefore, the present invention uses a non-targeted metabolomics method to explore the intervention effect of soft pear on alcoholic liver damage, providing new ideas for the further development and utilization of this resource.

[0046] 1 Experimental Materials and Methods

[0047] (1) Animals, reagents, and instruments

[0048] Forty 6-week-old SPF Wistar rats, half male and half female, weighing 180-200 g, were provided by the Animal Center of Gansu University of Chinese Medicine, with animal production license number SCXK (Gan) 2015-0002.

[0049] The soft pear frozen pears were provided by Bailutong Fruit Professional Cooperative in Gaolan, Gansu. After the frozen pears were naturally thawed, the skin was peeled off and 1000mL of original juice (equivalent to 1460.70g of soft pear frozen pears) was taken. After determination by high performance liquid chromatography, the total phenol content was 149.14mg / kg and the total flavonoid content was 58.23mg / kg.

[0050] Physiological saline, paraffin, hematoxylin, eosin, etc. were provided by the laboratory of Gansu University of Chinese Medicine. Red Star Erguotou liquor with an alcohol volume fraction of 0.52 was produced by Beijing Brewing Factory.

[0051] All biochemical detection kits used were purchased from Nanjing Jiancheng Bioengineering Institute, with batch numbers: MDA (20230812), LPO (20231015), SOD (20231106), GSH-Px (20230914), and CAT (20231104).

[0052] SFY3-400 fully automatic biochemical detector (Tianjin Jinbeier Technology Co., Ltd.), 721 visible light spectrophotometer (Shanghai Tingzheng Instrument Technology Co., Ltd.), electronic balance (Sartorius Scientific Instrument Co., Ltd.), ultra-low temperature electric centrifuge (XYJ 80-2, Jintan Hengfeng Instrument Factory), AB Triple TOF 6600 mass spectrometer (AB SCIEX, USA); Agilent 1290 Infinity LC ultra-high pressure liquid chromatograph (Agilent, USA).

[0053] (2) Animal grouping and drug administration

[0054] Rats were randomly divided into 4 groups according to body weight and sex: blank group (equal amount of NS + equal amount of NS), model group (equal amount of NS + Red Star Erguotou 10 mL / kg), low-dose Ruaner Pear group (Ruaner Pear juice 10 mL / kg + Red Star Erguotou 10 mL / kg), and high-dose Ruaner Pear group (Ruaner Pear juice 20 mL / kg + Red Star Erguotou 10 mL / kg). Rats were given drugs by gavage once a day for 4 consecutive weeks.

[0055] (3) Collection of biological tissue samples

[0056] 4 hours after the last administration, the rats were weighed, blood was collected from the femoral artery, and the whole blood was allowed to stand at room temperature for 45 minutes, then centrifuged at 3000r / min for 10 minutes, and the supernatant was taken. A portion of the serum samples was quickly frozen in liquid nitrogen for 5 minutes and stored at -80°C for metabolomics detection (only the high-dose group was taken for metabolomics detection in the soft pear administration group); the other part of the serum samples was used for automatic biochemical analyzer to detect liver function and blood lipid indicators. The animals were killed, the liver was removed and weighed, and the liver index was calculated according to the formula. An appropriate amount of liver tissue was cut and made into a homogenate, centrifuged at 4°C and 2500r / min for 10 minutes, and the supernatant was taken. The MDA and LPO content, as well as the SOD, GSH-Px and CAT activities were detected by colorimetry. The remaining liver was fixed with 10% formaldehyde solution, stained with HE, dehydrated with ethanol gradient, transparent, and sealed, and pathological changes were observed under an optical microscope.

[0057]

[0058] (4) Non-targeted metabolomics detection in rat serum

[0059] ① Serum metabolomics sample processing

[0060] Take 50uL of serum sample, add 250uL of acetonitrile-formic acid (1:4) solution, vortex for 2min, centrifuge at 4℃12000r / min for 10min, take 200uL of supernatant, let stand at -20℃ for 30min, centrifuge again at 4℃12000r / min for 3min, take 150uL of supernatant and put it in the injection bottle for analysis. Take equal amount of all serum samples to make quality control samples.

[0061] ②Chromatographic conditions

[0062] A Waters Acquity UPLC HSS T3 chromatographic column (2.1 mm×100 mm, 1.8 μm) was used; the mobile phase was 0.1% formic acid aqueous solution (A)-0.1% formic acid-acetonitrile (B), gradient elution (0-3 min, 5%-40% B; 3-6 min, 40%-60% B; 6-8 min, 60%-80% B; 8-15 min, 80% B; 15-20 min, 80%-95% B; 20-28 min, 95%-5% B); flow rate was 0.5 mL / min, injection volume was 2 μL; column temperature was 45°C.

[0063] ③Mass spectrometry conditions

[0064] Electrospray ionization (ESI) was used for collection in positive and negative ion modes, respectively. ESI+ spray voltage was 2500 V, and ESI- spray voltage was -1500 V. The nebulization temperature was 300 °C, the nebulization gas flow rate was 10 L / min, the nebulization gas voltage was 40 V, and the mass spectrometry collection range was m / z 60-1 200.

[0065] ④Data processing and multivariate statistical analysis

[0066] After alignment, filtering and normalization, the original data from mass spectrometry were subjected to multivariate statistical analysis, unsupervised principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA) and orthogonal partial least squares discriminant analysis (OPLS-DA) using SIMCA14.0.2; differential metabolites with variable importance in projection (VIP)>1 and P<0.05 in OPLS-DA analysis were screened out, and the differential metabolites were identified with reference to the HMDB database (http: / / www.hmdb.ca / ); Oringin 2022 software was used to draw a clustering heat map of differential metabolites based on the relative peak areas of the differential metabolites; the differential metabolites were imported into Metabo Analyst 5.0 platform (https: / / www.metaboanalyst.ca / ), pathway enrichment analysis was performed on the differential metabolites to obtain the metabolic pathways that may be involved in the intervention of soft pear on alcoholic liver injury.

[0067] (5) Statistical processing

[0068] All experimental data were processed using SPSS 24.0 software and expressed as mean ± standard deviation. The t-test was used for comparison between two groups, and one-way analysis of variance was used for comparison among multiple groups. P < 0.05 indicated that the difference was statistically significant.

[0069] 2 Experimental results and analysis

[0070] (1) Effects of Ruaner Pear on Liver Index in Rats with Alcoholic Liver Injury

[0071] The organ index, also known as the organ-body ratio, is the ratio of the weight of a certain organ of an experimental animal to its body weight. Under normal circumstances, the ratio of each organ of an animal to its body weight is relatively constant. When an animal is poisoned and its organs are damaged, the weight of the damaged organs may change, so the organ index also changes accordingly. An increase in the organ index indicates that the organ may have congestion, edema, or hyperplasia; a decrease in the organ index indicates that the organ may have atrophy and other degenerative changes. The liver index is the ratio of the liver weight to the animal's body weight. As shown in Table 1, compared with the blank group, the liver index of rats increased significantly after 4 weeks of alcohol damage (P<0.05), indicating that the liver may have congestion, edema, and other phenomena. After being treated with soft pears, the liver index decreased, indicating that soft pears can reduce the phenomenon of liver congestion and edema.

[0072] Table 1 Effects of Ruan'er pear on liver indexes in rats with alcoholic liver injury

[0073]

[0074] Note: Compared with the blank group, P*<0.05; compared with the model group, P # <0.05.

[0075] (2) Effects of Ruaner Pear on Liver Function Indexes in Rats with Alcoholic Liver Injury

[0076] The liver is the largest solid gland in the human body, and it undertakes complex and important metabolic functions. Liver function indicators can reflect the impact of different pathogenic factors on liver function. Commonly used liver function indicators include: alanine aminotransferase (glutamic-pyruvictransaminase, ALT), aspartate aminotransferase (glutamic oxalacetic transaminase, AST) and alkaline phosphatase (alkaline phosphatase, ALP). Under normal circumstances, the concentrations of ALT and AST in the blood are very low. When the liver is damaged, the permeability of the liver cell membrane increases. At this time, the ALT and AST originally present in the cells will overflow from the cells and transfer to the blood, causing the ALT and AST levels in the blood to increase. At the same time, when the liver is damaged, liver cells will synthesize a large amount of ALP and release it into the blood, causing a significant increase in serum ALP. Therefore, the detection of ALT, AST and ALP can directly reflect the degree of liver damage. By Figure 1 It can be seen that after rats were damaged by alcohol, the levels of ALT, AST and ALP in the serum were significantly increased (P<0.05), indicating that liver cells were damaged. The relevant indicators of the soft pear treatment group were significantly decreased (P<0.05), among which the effect of the high-dose group was more obvious (P<0.05), indicating that soft pear can alleviate the increase of liver function indicators caused by alcohol.

[0077] (3) Effects of Ruaner Pear on Blood Lipid Indexes in Rats with Alcoholic Liver Injury

[0078] Blood lipid tests can detect abnormalities in blood lipid metabolism. Commonly used blood lipid indicators include: total triglycerides (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C), commonly known as the four blood lipids. TG contains saturated fatty acids and unsaturated fatty acids. Elevated TG is often associated with cardiovascular events such as thrombosis and atherosclerosis. TC refers to the sum of cholesterol loaded by all lipoproteins in the blood, including free cholesterol (10%) and cholesterol esters (90%). Among them, low-density lipoprotein-cholesterol (LDL-C) is the cholesterol component in low-density lipoprotein particles (LDL), which can transport cholesterol from the liver to tissues throughout the body, and can also be deposited on the blood vessel walls to form atherosclerotic plaques, playing a core role in the development of cardiovascular diseases. High-density lipoprotein particles (HDL) send cholesterol from various tissues back to the liver for metabolism, which is conducive to the stability of cholesterol substances. Studies have shown that acetaldehyde, a metabolite of alcohol, can not only reduce the liver's ability to oxidize fatty acids, but also damage liver cell mitochondria, inhibit the tricarboxylic acid cycle, affect the liver cell microtubule system, reduce the secretion of microparticle protein, and cause lipid and protein deposition in liver cells; at the same time, it can mobilize subcutaneous fatty acids to transfer to the liver, causing an increase in TG content in the liver and gradually forming fatty liver. In this study, the TG, TC, LDL-C and HDL-Cj in the blood of rats after alcohol damage were significantly increased (P<0.05)( Figure 2 ), indicating that the blood lipid level was increased overall. The four blood lipid indicators of the Ruan'er pear treatment group were significantly lower than those of the model group (P<0.05), and the effect of the high-dose group was more obvious (P<0.05), indicating that Ruan'er pear can reduce the blood lipid level of rats with alcoholic liver injury.

[0079] (4) Effects of Ruan'er Pear on MDA and LPO Contents and SOD, GSH-Px, and CAT Activities in Liver Tissue of Rats with Alcoholic Liver Injury

[0080] Studies have shown that when alcohol is metabolized in the liver, it can convert the oxidized coenzyme nicotinamide adenine dinucleotide (NAD+) into the reduced form of the coenzyme nicotinamide adenine dinucleotide (NADH), thereby changing the NADH / DAD+ ratio and the intracellular redox state, thus affecting the metabolism of the three major substances. The oxidative stress and lipid peroxidation induced during alcohol metabolism can also produce a large number of oxygen stress products, such as OH-, O2-, H2O2 and other free radicals. These free radicals can directly destroy the integrity and fluidity of the liver cell membrane structure, thereby changing the microenvironment of membrane-bound enzymes, receptors and ion channels, affecting their functions; at the same time, a large number of free radicals can also destroy mitochondria and lysosomes, causing liver cells to apoptosis. Alcohol metabolite acetaldehyde can participate in the synthesis and metabolic reactions of antioxidant substances such as cysteine, glutathione and vitamin E, leading to a decrease in cysteine ​​and glutathione in the liver and a decrease in antioxidant capacity. At the same time, a large number of free radicals and alcohol metabolites can also cause abnormal function of liver Kupffer cells, thereby stimulating immune activation, inducing inflammatory cells to release various inflammatory mediators, triggering inflammatory reactions and further damaging liver cells. LPO can reflect the rate and degree of lipid peroxidation in the body. MDA is a metabolite of lipid peroxidation, so the determination of LPO and MDA content can often reflect the degree of lipid peroxidation in the body. GSH-Px, SOD and CAT are important enzymes for the body to scavenge free radicals, and are also key enzymes in the biological defense system. They play an important role in combating alcohol liver damage. Figure 3 It can be seen that after alcohol injury in rats, the content of oxidative stress products MDA and LPO increased significantly (P<0.05), indicating that the oxidative stress response was activated; the activities of SOD, GSH-Px, and CAT decreased significantly (P<0.05), indicating that the oxidation-antioxidation system in the liver environment was unbalanced. The content of MDA and LPO in the Ruan'er Pear treatment group decreased, and the activities of SOD, GSH-Px, and CAT increased to varying degrees (P<0.05), indicating that Ruan'er Pear can enhance the activity of antioxidant enzymes and reduce lipid peroxidation damage.

[0081] (5) Effects of Ruaner Pear on Pathological Changes of Liver in Rats with Alcoholic Liver Injury

[0082] The development of liver disease often goes through multiple stages, each with its own characteristics. The pathological changes in each stage are closely related to liver disease. Liver pathology is a discipline that studies the development mechanism and treatment methods of liver disease. It studies the morphology and structural changes of liver tissue to infer the type and development process of liver disease, determine the diagnosis, staging and treatment plan of liver disease, and provide reference for clinical practice. Figure 4 It can be seen that the hepatocytes of normal rats are complete in morphology, uniform in size, with clear boundaries and neat and orderly arrangement. After 28 days of alcohol damage, the hepatocytes are no longer complete in morphology and are not uniform in size. A large number of irregularly shaped eosinophilic glassy bodies (Mallory bodies) formed by the denaturation of the prokeratin of the intermediate filaments appear in the cytoplasm, the number of hepatic lobules increases, blood vessels proliferate, and obvious fragmentary necrosis and nuclear condensation can be seen. These pathological changes have improved in the Ruan'er Pear treatment group, as shown by the reduction of inflammatory infiltration and lobular hyperplasia, the alleviation of fragmentary necrosis, the clear cell boundaries, and the relatively complete morphology. The improvement effect of the Ruan'er Pear high-dose group is more obvious.

[0083] (6) Results of serum non-targeted metabolomics analysis

[0084] ①Multivariate statistical analysis results

[0085] The results of PLS-DA showed that the metabolic profiles of the blank group, model group and Ruan'erli drug-treated group were significantly separated, indicating that there were significant differences in the endogenous metabolites in the blood of the three groups of rats ( Figure 5 ). In this model, R2X is 0.949, indicating that the variable has a good explanatory power for the model, R2Y is 0.845, indicating that the model has good stability, and Q2 is 0.861, indicating that the model has good predictive power. After 200 permutation tests, it was found that all simulated values ​​were lower than the true value on the far right, and the intercept of the Q2 regression line on the Y axis was less than -0.05 ( Figure 5 ), indicating that the model has good fit and prediction ability, and there is no overfitting phenomenon. Further OPLS-DA results show that ( Figure 6 ), in the blank group and the model group, R2X was 0.974, R2Y was 0.912, and Q2 was 0.893. In the Ruan'erli drug group and the model group, R2X was 0.883, R2Y was 0.907, and Q2 was 0.875, all meeting the model requirements. The two OPLS-DA models were verified by 200 permutation tests, indicating that both models had good fit and prediction ability, and there was no overfitting phenomenon.

[0086] ② Screening and identification results of differential metabolites

[0087] According to the results of OPLS-DA, small molecule compounds with VIP>1 and P<0.05 were selected as potential differential metabolites of alcoholic liver injury through S-plot analysis. Based on the metabolite mass spectrometry data of the blank group, model group and Ruanerli drug administration group, combined with literature information and HMND and other databases, 20 differential metabolites with significant differences compared with the model group were finally screened and identified, as shown in Table 2.

[0088] Table 2 List of differential metabolites

[0089]

[0090] Note: Compared with the model group, P * <0.05, P ** <0.01.

[0091] The relative peak areas of these 20 differential metabolites were significantly different among the blank group, model group and Ruan'erli treatment group ( Figure 7 a)(P<0.05), and there was a strong correlation between the 20 differential metabolites ( Figure 7 b). Cluster analysis results showed that the blank group, model group and Ruan'erli treatment group had good clustering within the three groups and obvious separation between the groups ( Figure 7 c), suggesting that the intervention effect of soft pear on alcoholic liver injury may be related to the regulation of differential metabolite levels.

[0092] (3) Differential metabolite pathway enrichment analysis

[0093] The 20 differential metabolites were imported into the MetaboAnalyst platform and screened with impact value>0.1, P<0.05 as the standard, and a total of 4 metabolic pathways related to the intervention of Ruaner pear on alcoholic liver injury were obtained ( Figure 8 ), namely aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine and tryptophan biosynthesis, pyrimidine metabolism, and linoleic acid metabolism, suggesting that soft pear may intervene in alcoholic liver injury by regulating these signaling pathways and affecting nucleotide synthesis and protein metabolism.

[0094] 3 Experimental Discussion and Conclusion

[0095] Studies have shown that the pathogenesis of alcoholic liver injury or alcoholic liver disease includes not only the direct toxicity of ethanol to the liver and the metabolic disorders caused by the toxicity of its metabolites, but also inflammatory mediators, cytokines, cell apoptosis mechanisms and free radical damage, as well as malnutrition and choline deficiency, which are considered to be associated factors in the onset of the disease.

[0096] The present invention establishes an animal model of alcoholic liver injury and finds that soft pear can not only improve the liver function of rats with alcoholic liver injury, reduce blood lipid indexes, but also enhance the activity of antioxidant enzymes and reduce lipid peroxidation damage. Further metabolomics studies have found that the intervention effect of soft pear on alcoholic liver injury may be related to the effect on aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine and tryptophan biosynthesis, pyrimidine metabolism, linoleic acid metabolism and other pathways, thereby affecting nucleotide synthesis and protein metabolism. Aminoacyl-tRNA is a transfer RNA (also known as transfer RNA) that can bind to the corresponding amino acid. It can transport the corresponding amino acid to a specific site on the ribosome, merge it with the extending polypeptide chain, and then form a protein. Phenylalanine, tyrosine and tryptophan are all essential amino acids for the human body, and play an important role in the body's metabolism, growth and development. Phenylalanine can be oxidized to tyrosine by phenylalanine hydroxylase catalysis. Tyrosine is not only a substrate for the synthesis of neurotransmitters adrenaline and norepinephrine, but also a raw material for the synthesis of thyroid hormones, and participates in the body's sugar metabolism and fat metabolism. Tryptophan is also a precursor of serotonin, an important neurotransmitter in the human body. Pyrimidine is a nitrogen-containing compound and an important base that constitutes nucleic acids. The pyrimidine metabolic pathway includes pyrimidine synthesis and catabolism. In pyrimidine catabolism, pyrimidine is broken down into uric acid and other metabolites. If pyrimidine metabolism is abnormal, it will lead to increased uric acid levels and cause diseases such as gout. In addition, abnormal pyrimidine metabolism may also lead to abnormal neurotransmitter synthesis and metabolism. Linoleic acid metabolism is one of the important metabolic pathways in the human body. By oxidizing fatty acids in the body into linoleic acid, it can not only degrade and consume lipids and provide energy for cells, but the sugar ketokinase produced during the oxidation process can also promote blood sugar metabolism and maintain blood sugar stability. Abnormal linoleic acid metabolism can induce fat and sugar metabolism disorders.

[0097] The experimental results of the present invention show that the soft pear may affect nucleotide synthesis and protein metabolism by regulating metabolic pathways such as aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine and tryptophan biosynthesis, pyrimidine metabolism and linoleic acid metabolism, thereby playing an intervention role in alcoholic liver injury. Therefore, the results of the present invention will provide new ideas for the further development and utilization of Lanzhou soft pear, a special resource.

Claims

1. A method for preparing a soft pear alcohol-relief drink, comprising the following steps: ⑴ Extraction of soft pear juice: After ripening, peel off the skin of soft pears, thaw naturally, and extract the juice. The total phenol content in the pear juice should be no less than 120 mg / kg, and the total flavonoid content should be no less than 40 mg / kg. ⑵Preparation of solution: Mix chrysanthemum, wolfberry and pitted jujube in a mass ratio of 1:1:1, then add 10 times the mass of drinking water of the mixture to soak for 30 minutes, and then boil for 20 minutes, and filter to obtain liquid medicine A and residue A respectively; add 5 times the mass of drinking water to the residue A and continue to boil for 10 minutes, and filter to obtain liquid medicine B and residue B respectively; the liquid medicine A and the liquid medicine B are fully mixed and concentrated to obtain a prepared solution; ⑶ After the prepared liquid is cooled to room temperature, it is mixed with the soft pear juice at a volume ratio of 3:1, and filled into a can at 400 ml / can, and sterilized at room temperature.

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