A traditional Chinese medicine composition for treating non-alcoholic fatty liver and a preparation method thereof

Abstract

The present application relates to a kind of traditional Chinese medicine composition for treating non-alcoholic fatty liver and its preparation method, the traditional Chinese medicine composition in the present application is by ginseng 5~10 parts, red yeast 3~6 parts, daide flower 3~6 parts, dried tangerine or orange peel 6~12 parts, Herba Lycopi 6~12 parts, sea buckthorn 6~12 parts.The traditional Chinese medicine composition provided by the present application has the effect of resolving phlegm, promoting blood circulation to remove blood stasis, benefiting qi and invigorating the spleen, and cuts the pathogenesis of non-alcoholic fatty liver, and is suitable for liver stagnation and spleen deficiency type non-alcoholic fatty liver patients.

Description

TECHNICAL FIELD

[0001] The application relates to a traditional Chinese medicine composition for treating non-alcoholic fatty liver and a preparation method thereof, and belongs to the field of traditional Chinese medicine. BACKGROUND

[0002] Non-alcoholic fatty liver disease (NAFLD) is a chronic metabolic disease characterized by hepatocyte steatosis and dyslipidemia. In recent years, the number of NAFLD patients has been rising, and the adult prevalence rate has reached as high as 25%. The disease has become the world's first chronic liver disease. NAFLD can develop into cirrhosis and even liver cancer, posing a great threat to human life and health, and patients often bear a huge economic burden. So far, there is no specific treatment drug for NAFLD, and currently, lipid-lowering drugs, diet therapy and exercise therapy are generally used for treatment in the clinic. Although the above drug treatment has a certain effect, long-term use will have many side effects. The diet therapy and exercise therapy have no obvious effect, and patients are difficult to adhere to for a long time. Some lipid-lowering drugs also have potential hepatotoxicity, and some long-term use may aggravate liver damage.

[0003] There is no clear disease name record of 'fatty liver' in ancient Chinese medical books, but there are many records similar to its symptoms. It is recorded in 'Zhuozhengyuanhoulun' that 'pi is a disease in which the side is on both sides of the chest, and sometimes it is painful'. In addition, 'Nanjing' also records that 'the accumulation of liver is called fat'. The National Administration of Traditional Chinese Medicine defines the Chinese medicine name of NAFLD as 'liver pi', which is helpful for its research. In terms of etiology and pathogenesis, ancient physicians believe that fatty liver patients often eat fatty food, overeat and lack exercise, emotional disorders, causing liver to lose dredging, spleen to lose transportation, internal generation of dampness, internal retention of phlegm and turbidity, kidney essence deficiency, and phlegm and turbidity not being transformed, leading to dysfunction of liver, spleen and kidney, and mutual knotting of dampness, heat and phlegm in the liver.

[0004] The present application systematically sorts out the prior art documents, and it can be found that the prior art patent document CN101204573A discloses a traditional Chinese medicine composition. The composition is made of the following raw materials by weight: bitter gourd slices 30g, sophora 9g, rhizoma coptidis 30g, anemarrhena 30g, semen ziziphi spinosae 15g, prepared ginger 6g, Monascus 3g, dried tangerine or orange peel 9g, Rheum officinale 2g, persicae semen 6g. The heat-clearing and turbidity-reducing prescription of the application can treat type 2 diabetes, and can reduce blood glucose, blood lipid and blood pressure, and improve insulin resistance and blood stasis. InsulinThe resistance and improvement of islet cell function significantly improve the abnormal glucose metabolism of type 2 diabetes model rats, and reduce fatty liver and maintain normal blood lipids. CN101574476A discloses a medicinal composition for auxiliary lipid-lowering and liver protection, comprising the following medicinal materials: jujube fruit, alisma, turtle shell, papaya, dried tangerine or orange peel, atractylodes, salvia miltiorrhiza, polygonatum, hawthorn and lotus leaf. Through animal experiment verification, the drug composition has the function of auxiliary lipid-lowering, and plays a protective role on the liver.

[0005] At present, there is no effective treatment drug and clear treatment scheme for the disease, and some drugs are still in the preclinical research or clinical trial stage, and so far no drug has been approved in the United States and the European Union and used for treating the disease. Therefore, developing safe and effective traditional Chinese medicine compound preparations will be more suitable for the treatment of NAFLD, and has wide application prospect and practical significance. SUMMARY

[0006] The present application relates to a kind of traditional Chinese medicine composition for treating nonalcoholic fatty liver and preparation method thereof, the traditional Chinese medicine composition provided by the present application has the effect of eliminating phlegm, promoting blood circulation and removing blood stasis, benefiting qi and invigorating the spleen, and it is suitable for liver stagnation and spleen deficiency type nonalcoholic fatty liver patients.The present application is simple, and raw materials are easy to obtain.The preparation process is scientific and reasonable, simple and feasible, and has the prospect of industrial application.

[0007] The etiology, pathogenesis and treatment principle of the present application are as follows: the nonalcoholic fatty liver treated by the present application is a chronic metabolic disease characterized by hepatocyte steatosis and dyslipidemia. According to its clinical symptoms, nonalcoholic fatty liver belongs to the category of liver distension, hypochondriac pain, fat and accumulation in traditional Chinese medicine. The earliest record of this disease is in the Difficult Classic, which mentions "liver accumulation, called fat". It is believed that the formation of "fat" is caused by overeating fatty and sweet foods, and excessive accumulation in the liver. Fatty liver patients eat fatty food, overeat and have little exercise, emotional disorder, causing liver to lose its function of dredging, spleen to lose its function of transportation, internal generation of dampness, internal retention of phlegm and turbidity, kidney essence deficiency, phlegm and turbidity not transformed, etc., leading to dysfunction of liver, spleen and kidney, and mutual retention of dampness, heat and phlegm in the liver. The main syndrome types are liver stagnation and spleen deficiency, internal retention of phlegm and turbidity, internal retention of dampness and heat, and mutual retention of phlegm and blood stasis. The most common one in clinic is liver stagnation and spleen deficiency. Therefore, the cause of the disease is summarized as improper diet or overeating fatty food, leading to spleen deficiency, phlegm and dampness not transformed, obstruction of qi, liver losing its function of dredging, phlegm and dampness staying in the liver, liver qi stagnation for a long time, blood stasis staying, and mutual retention of blood stasis and phlegm and dampness. The core pathogenesis is stagnation of earth and stagnation of wood, mutual retention of phlegm and blood stasis. The treatment is to remove blood stasis and phlegm for the symptoms, and to regulate liver and spleen qi for the root.

[0008] The prescription of the present application is as follows:

[0009] The traditional Chinese medicine composition for treating non-alcoholic fatty liver of the present application is Shenqu Jiangzhi Decoction, which is composed of six medicinal ingredients of ginseng rootlet, red yeast, dried tangerine or orange peel, Lantana camara, citrus aurantium and Hippophae rhamnoides, and is mainly used for treating non-alcoholic fatty liver of the type of liver stagnation and spleen deficiency and phlegm and blood stasis. The selected medicinal ingredients are all traditional Chinese medicines with homology of food and medicine, high safety, remarkable curative effect, and dual effects of prevention and treatment.

[0010] There is no disease name of fatty liver in traditional Chinese medicine, and it is classified into the categories of hypochondriac pain, distension, fat, and phlegm syndrome according to clinical symptoms and syndrome of traditional Chinese medicine. The Difficult Classic·Fifty-sixth Difficulty accurately describes the characteristics of the disease, that is, liver accumulation is called fat, and emphasizes that the lesion site is in the liver, which is caused by the accumulation of fat. Numerous clinical data show that liver stagnation and spleen deficiency are the key to the disease, and phlegm and blood stasis are important pathological factors, and liver stagnation and spleen deficiency are the premise and basis of the disease.

[0011] The Shenqu Jiangzhi Decoction is the effective experience prescription of Professor Ding Hui of Shaanxi Institute of Traditional Chinese Medicine for many years. The selected six medicinal ingredients mainly enter the liver, spleen and stomach meridians, wherein the ginseng rootlet and red yeast are the monarch drugs, which have the effects of invigorating the spleen, promoting digestion, promoting blood circulation and removing blood stasis. The ginseng rootlet is the fine branch root of Panax ginseng of the Araliaceae family, which not only plays the role of tonifying qi and invigorating the spleen of the ginseng medicine, but also strengthens the function of dredging collaterals and promoting qi on the basis of tonifying qi. The red yeast is the most wonderful one, which enters the liver and spleen meridians, and plays the multiple roles of invigorating the spleen and stomach, resolving phlegm and turbidity, and removing blood stasis; the dried tangerine or orange peel and Lantana camara are the ministerial drugs, which are used with the red yeast to strengthen the effects of resolving phlegm and turbidity and removing blood stasis; the citrus aurantium is the flower of Citrus aurantium, and the fruit of Citrus aurantium has the effect of regulating the qi of liver and spleen, and the citrus aurantium is mild in nature and pungent and sweet in taste, which is good at regulating the qi of liver and spleen without damaging the qi, and is more suitable for long-term taking, and is the auxiliary drug of the other drugs; the Hippophae rhamnoides is sour, astringent and sweet in taste, and enters the liver meridian, which plays the role of the messenger of the drugs, and also has the effects of resolving phlegm, removing blood stasis and dispersing stagnation, which is closely related to the pathogenesis of fatty liver, and modern medical research finds that it can effectively reduce cholesterol and blood lipids.

[0012] Clinical observation shows that the Shenqu Jiangzhi Decoction can improve fat deposition, reduce blood lipid level, restore transaminase abnormality, and has good therapeutic effect on non-alcoholic fatty liver, and this effect is also verified in animal experiments.

[0013] The messenger of the prescription of the present application can also be the seed of Astragalus complanatus, which is warm in nature and sweet in taste, and has the effects of warming and tonifying liver and kidney and brightening eyes. Combined with other drugs, it can play the effects of protecting liver and reducing blood lipids.

[0014] The traditional Chinese medicine composition Shenqu Jiangzhi Decoction provided by the present application hits the pathogenesis of non-alcoholic fatty liver, and is suitable for patients with non-alcoholic fatty liver of the type of liver stagnation and spleen deficiency and phlegm and blood stasis.

[0015] The technical scheme of the present application is as follows:

[0016] The dosage and ratio of the components of the traditional Chinese medicine composition in the application are obtained by the inventors of the application through a large amount of clinical practice and summarization, and the components in the above weight range have good clinical effects.

[0017] A traditional Chinese medicine composition for treating alcoholic fatty liver, which comprises the following raw medicinal materials in the weight ratio: ginseng stem 5-10 parts, red yeast 3-6 parts, Daidai flower 3-6 parts, dried tangerine or orange peel 6-12 parts, and Herba Lycopi 6-12 parts.

[0018] Preferably, the traditional Chinese medicine composition comprises the following raw medicinal materials in the weight ratio: ginseng stem 5-10 parts, red yeast 3-6 parts, Daidai flower 3-6 parts, dried tangerine or orange peel 6-12 parts, Herba Lycopi 6-12 parts, and sea buckthorn 6-12 parts.

[0019] Preferably, the traditional Chinese medicine composition comprises the following raw medicinal materials in the weight ratio: ginseng stem 5-10 parts, red yeast 3-6 parts, Daidai flower 3-6 parts, dried tangerine or orange peel 6-12 parts, Herba Lycopi 6-12 parts, and sea buckthorn 6-12 parts.

[0020] As a further preferred embodiment of the application, the traditional Chinese medicine composition comprises the following raw medicinal materials in the weight ratio: ginseng stem 6 parts, red yeast 3 parts, Daidai flower 3 parts, dried tangerine or orange peel 10 parts, Herba Lycopi 10 parts, and sea buckthorn 6 parts.

[0021] As a further preferred embodiment of the application, the traditional Chinese medicine composition comprises the following raw medicinal materials in the weight ratio: ginseng stem 6 parts, red yeast 3 parts, Daidai flower 3 parts, dried tangerine or orange peel 10 parts, Herba Lycopi 10 parts, and sea buckthorn 6 parts.

[0022] The preparation method of the traditional Chinese medicine composition comprises the following steps:

[0023] The medicinal materials in the above weight ratio are added with water in an amount of 6-13 times the weight of the medicinal materials, soaked, decocted for 1-3 times, and the decoction time is 0.5-1.5 hours, filtered, and the water decoction extraction filtrates are combined, dried by vacuum concentration to obtain a dry paste powder intermediate, and then added with pharmaceutical adjuvants to prepare a traditional Chinese medicine oral preparation.

[0024] As a preferred embodiment of the application, in the preparation method, water in an amount of 8 times the weight of the medicinal materials is added, and the decoction is performed for 3 times, and the decoction time is 1 hour each time.

[0025] The traditional Chinese medicine composition can be prepared into granules and oral liquid.

[0026] The traditional Chinese medicine composition of the application is used for preparing a medicine for treating alcoholic fatty liver, viral hepatitis, and chronic hepatitis B.

[0027] In order to enable the related technical personnel to better understand the application, the traditional Chinese medicinal materials used in the present application are raw medicinal materials, and the sources are as follows.

[0028] Ginseng rootlets: These are the fine roots of Panax ginseng CAMey., a plant belonging to the Araliaceae family.

[0029] Red yeast rice: It is made from indica rice as raw material, using modern bioengineering technology to isolate high-quality red yeast mold (monascus purpureus) and refine it through deep liquid fermentation.

[0030] Citrus aurantium L. (Bitter Orange Flower): The source is the flower bud of Citrus aurantium L., a plant in the Rutaceae family.

[0031] Dried tangerine peel: It is derived from the mature peel of Citrus reticulata Blanco., a plant of the Rutaceae family, and its cultivated varieties.

[0032] Eupatorium: The source is the aerial parts of Lycopus lucidus Turcz. var. hirtus Regel, a plant in the Lamiaceae family.

[0033] Sea buckthorn: The source is the mature seeds of Astragalus complanatus R.Br., a legume.

[0034] Sha Yuan Zi: Sourced from the legume *Astragalus complanatus*. Astragalus membranaceus Astragalus complanatus R.BrI ex Bunge dried mature seeds.

[0035] In order to obtain a scientific and reasonable preparation process route for the traditional Chinese medicine composition of this invention, a large amount of preliminary research was conducted, and the specific scheme is as follows:

[0036] I. Screening of Process Conditions

[0037] 1. Investigation on the preparation process of the sea buckthorn formula of the present invention

[0038] 1.1 Water Absorption Rate Test

[0039] The composition of the traditional Chinese medicine composition consists of the following raw materials: ginseng rootlets 5-10g, red yeast rice 3-6g, bitter orange blossom 3-6g, dried tangerine peel 6-12g, eupatorium fortunei 6-12g, and sea buckthorn 6-12g. Weigh each herb in the prescription, add 12 times the amount of water, and soak until all herbs are completely wet. Calculate the water absorption rate using the following formula: Water absorption rate (%) = (wet weight of herbs - dry weight of herbs) / dry weight of herbs × 100%. The calculated water absorption rate is 166.32%.

[0040] 1.2 Determination of hesperidin content

[0041] 1.2.1 Chromatographic conditions Chromatographic column: Kromasil 100-5-C18 chromatographic column (4.6 mm x 250 mm, 2.5 μm); mobile phase: acetonitrile-water (19:81); flow rate: 1.0 mL min -1 ; column temperature: 30℃; detection wavelength: 283 nm; injection volume: 10 μL.

[0042] 1.2.2 Preparation of solutions

[0043] (1) Preparation of the control solution

[0044] Accurately weigh 4.0 mg of hesperidin control substance into a 10 mL volumetric flask, add methanol to the mark, and obtain a 0.4 mg mL -1 hesperidin control substance solution.

[0045] (2) Preparation of the test solution

[0046] Take the water decoction of the Hippophae rhamnoides L. formula (Example 1 group formula) and filter it through a micropore filter. Inject the control solution (5 μL) and the test solution (10 μL) into the liquid chromatograph for determination, and calculate the content of hesperidin in the test solution.

[0047] 1.2.3 Content determination

[0048] Weigh each raw material of the traditional Chinese medicine composition, prepare the test solution according to the method in item (2) under "1.2.2", inject 10 μL, and analyze the sample injection according to the chromatographic conditions in item (1) under "1.2.1", determine the chromatographic peak area, and calculate the content of hesperidin. The chromatograms of the control solution and the test solution are shown in Figure 1 .

[0049] 1.3 Calculation of extract yield

[0050] Accurately take 25 mL of the water extract, place it in an evaporating dish that has been dried to a constant weight, evaporate it in a water bath, dry it to a constant weight at 105℃ under normal pressure, take it out, cool it in a desiccator for 0.5 h, quickly and accurately weigh it, and calculate the dry extract yield.

[0051] Extract yield (%) = (m1V / mV1) x 100%, where m1 is the extract weight, V is the total volume of the extract, m is the weight of the prescription medicinal material, and V1 is the amount of test solution taken for measurement.

[0052] 1.4 Single factor investigation

[0053] 1.4.1 Investigation of extraction time

[0054] Take 1 of each prescription amount of medicinal materials, a total of 5, each add 10 times the amount of water, respectively in 15, 30, 60, 90, 120 min conditions extraction 2 times, filter, combined extract, determine the volume, with hesperidin content as evaluation index, to investigate the influence of different extraction time on the sample. In the water multiple 10 times, extraction times 2 times, respectively in 15, 30, 60, 90, 120 min extraction, analysis of extraction time on the content of hesperidin and dry extract yield, the results are shown in table 1-1.

[0055] Table 1-1 extraction time investigation

[0056]

[0057] From the results, with the increase of extraction time, the content of hesperidin first increased and then basically stable, in the extraction time of 60 min, the content of hesperidin and dry extract yield was the largest, 7.2117 mg / g and 27.51%, respectively, in 60~120 min, the content of hesperidin and dry extract yield change was not obvious, it was proved that the optimal extraction time was 60 min. Therefore, 30, 60, 90 min were selected for orthogonal test.

[0058] 1.4.2 extraction times investigation

[0059] Take 1 of each prescription amount of medicinal materials, a total of 3, each add 10 times the amount of water, respectively extraction 1, 2, 3 times, 60 min each time, filter, combined extract, determine the volume, with hesperidin content as evaluation index, to investigate the influence of different extraction times on the sample. In the water multiple 10 times, extraction time for 60 min, respectively extraction 1, 2, 3 times, analysis of extraction times on the content of hesperidin and dry extract yield, the results are shown in table 1-2.

[0060] Table 1-2 extraction times investigation

[0061] The test results found that, with the increase of extraction times, the content of hesperidin and dry extract yield also increased, the content of hesperidin was 9.0859 mg / g, dry extract yield was 28.4215 % when extracted 3 times, so the optimal extraction times was 3 times. Therefore, 1, 2, 3 times were selected for orthogonal test.

[0062] 1.4.3 water multiple investigation

[0063] Take 1 prescription amount of each medicinal material, 4 parts, each add 6, 8, 10, 12 times the amount of water, extract 2 times, each for 60 min, filter, combine the extract, determine the volume, take the content of hesperidin as the evaluation index, and investigate the influence of different water multiples on the sample.

[0064] Under the conditions of 2 extraction times and 60 min extraction time, each add 6, 8, 10, 12 times the amount of water for extraction, analyze the influence of water multiple on the content of hesperidin and the yield of dry extract, and the results are shown in Tables 1-3, Figures 1-4

[0065] Table 1-3 Water multiple investigation

[0066]

[0067] According to the results of water multiple investigation, the content of hesperidin and the yield of dry extract showed an upward trend when the water multiple was 6-8 times, and the content of hesperidin and the yield of dry extract reached the maximum when the water multiple was 8 times, which were 7.0693 mg / g and 24.0278 %, respectively. From 8 times, it showed a downward trend, and then basically tended to be balanced, so 8 times water was selected as the optimal water multiple. Therefore, 6, 8 and 10 times water were selected for orthogonal test.

[0068] 1.5 Orthogonal test design

[0069] The prescription was extracted by water decoction method, the yield of extract (%) and the content of hesperidin were used as evaluation indexes, the water amount, extraction time and extraction times were used as investigation factors to carry out L9(34) 4 ) orthogonal test, and the best extraction process conditions were investigated. The orthogonal test factor level is shown in Table 1-4. Comprehensive score = (y i - y min ) / (y max - y min ) × 40% + (c i -c min ) / (c max - c min ) × 60% Note: y i is the yield of dry extract, y max is the maximum value of the yield of dry extract, y min is the minimum value of the yield of dry extract; c i is the content of hesperidin, c max is the maximum value of the content of hesperidin; c min is the minimum value of the content of hesperidin.

[0070] Table 1-4 Orthogonal test factor level table ​

[0071] The optimal extraction time, extraction times and water amount obtained by single factor test are used as the conditions for extraction, and the content of hesperidin and the yield of dry extract are used as indexes, 3 factors and 3 levels are designed, and 9 tests are carried out, and the results are shown in Tables 1-5. The variance analysis results are shown in Table 1-6.

[0072] Table 1-5 Orthogonal test results

[0073] Table 1-6 Variance analysis table

[0074] It can be known from the results in Table 1-5 that the primary and secondary order of the three investigated factors is extraction times (C), water amount (A) and extraction time (B), and the optimal combination is A3B3C3. It can be known from the variance analysis results in Table 1-6 that the extraction times and the water amount have significant differences (P<0.05, P<0.01), and the extraction time has no obvious difference (P>0.1), so the optimal combination finally determined is A3B2C3, that is, 8 times of water is added, and decoction is carried out for 3 times, and each time is 60 min.

[0075] Three batches of prescriptions are extracted by using the optimal process parameters in the orthogonal test, the yield of dry extract and the content of hesperidin are determined, the optimal extraction process is verified, and the preparation method of the prescription is determined.

[0076] Three verification tests are carried out according to the optimized optimal method, and the results of the content of hesperidin and the yield of dry extract in the three tests are shown in Table 1-7, which are higher than those of each test number in the orthogonal test, so the optimized extraction method is feasible, and is suitable for the extraction process of the traditional Chinese medicine compound.

[0077] Table 1-7 Verification results of the optimal extraction process

[0078] 2 The preparation process of the semen astragali prescription is investigated

[0079] 2.1 Water absorption rate investigation

[0080] The composition of the traditional Chinese medicine composition is: ginseng 5-10 g, red yeast 3-6 g, daye flower 3-6 g, dried tangerine peel 6-12 g, aeonides 6-12 g, semen astragali 9-15 g. The water absorption rate of the medicine is calculated according to the method.

[0081] The water absorption rate of the medicine is calculated according to the method. The calculated water absorption rate of the medicine is 177 %.

[0082] 2.2 Determination of Hesperidin Content

[0083] 2.2.1 Chromatographic Conditions Column: Kromasil 100-5-C18 column (4.6 mm x 250 mm, 2.5 μm); mobile phase: acetonitrile-water (19:81); flow rate: 1.0 mL / min; column temperature: 30 °C; detection wavelength: 283 nm; injection volume: 10 μL. -1

[0084] 2.2.2 Preparation of Solutions

[0085] (1) Preparation of Reference Solution

[0086] Accurately weigh 4.25 mg of hesperidin reference substance into a 10 mL volumetric flask, add methanol to the mark, and obtain a 0.425 mg / mL hesperidin reference solution. -1

[0087] (2) Preparation of Test Solution

[0088] Take the supernatant of the water extract of the traditional Chinese medicine composition after centrifugation and filter through a microporous filter membrane. Inject the reference solution (5 μL) and the test solution (10 μL) into the liquid chromatograph for determination, and calculate the content of hesperidin in the test solution.

[0089] 2.2.3 Content Determination

[0090] Weigh each raw material of the traditional Chinese medicine composition, prepare the test solution according to the method in item "1.2.2 (2)", inject 10 μL, and analyze the sample under the chromatographic conditions in item "1.2.1" of this chapter, determine the chromatographic peak area, and calculate the content of hesperidin. The chromatograms of the reference solution and the test solution are shown in Figure 3 .

[0091] 2.3 Calculation of Extract Yield

[0092] After measuring the total volume of the filtrate, take an appropriate amount of the water extract, centrifuge (3000 rpm), accurately measure 25 mL of the supernatant, place it in an evaporating dish that has been dried to a constant weight, evaporate it in a water bath, dry it at 105 °C for 3 h, cool it in a desiccator for 0.5 h, quickly weigh it, and calculate the dry extract yield (%).

[0093] Extract yield (%) = (m1·V / m·V1) x 100%, where m1 is the weight of the extract, V is the total volume of the extract, m is the weight of the prescription medicinal material, and V1 is the amount of test solution measured.

[0094] 2.4 Orthogonal Test Design

[0095] ​​The six herbs in the prescription were extracted by decoction with water. The yield (%) of the extract and the content of hesperidin were used as evaluation indicators, and the amount of water added, extraction time and number of extractions were used as factors to investigate L9 (3) 4 An orthogonal experiment was conducted to investigate the optimal extraction process conditions. The factor levels for the orthogonal experiment are shown in Table 1-4. The overall score = (y...) i - y min ) / (y max - y min ) × 40% + (c i -c min ) / (c max - c min ) × 60%, Note: y i For the yield of dry extract, y max y represents the maximum yield of the dry extract. min c is the minimum yield of dry extract; i For hesperidin content, c max This represents the maximum content of hesperidin; c min This represents the minimum hesperidin content.

[0096] Table 2-1 Factor Level Table for Orthogonal Experiments

[0097] Extraction was performed using the optimal extraction time, number of extractions, and water addition ratio obtained from single-factor experiments. Nine experiments with three factors and three levels were conducted, using hesperidin content and dry extract yield as indicators. The results are shown in Tables 1-5. Analysis of variance results are shown in Tables 2-6.

[0098] Table 2-2 Results of Orthogonal Experiments

[0099] Table 2-6 Analysis of Variance Table

[0100] The results show that the order of importance of the three factors is: number of extractions (C), amount of water added (A), and extraction time (B). The optimal combination is A3B3C3. According to the analysis of variance results in Table 2-6, there are significant differences in the number of extractions and amount of water added (P<0.05, P<0.01), while there is no significant difference in extraction time (P>0.1). Therefore, considering the actual situation, the optimal combination finally determined is A3B2C3, that is, add 8 times the amount of water, decoct 3 times, and each time for 60 minutes.

[0101] Extract 3 batches of prescriptions with the optimal process parameters selected in the orthogonal test, determine the dry extract yield and hesperidin content, verify the optimal extraction process, and determine the preparation method of the prescription. According to the optimized optimal method, 3 verification tests are carried out, and the results of the hesperidin content and dry extract yield of the 3 tests are shown in Tables 2-7, which are higher than the results of each test number in the orthogonal test, indicating that the optimized extraction method is feasible and suitable for the extraction process of the compound medicine.

[0102] Table 2-7 Verification results of the optimal extraction process

[0103]

[0104] Advantages of the traditional Chinese medicine composition in the application

[0105] (1) The prescription is formed by repeated optimization, summary and combination of the clinical practice experience of the inventors and the clinical efficacy feedback. The prescription is prepared from ginseng, red yeast rice, daye flower, dried tangerine or orange peel, aeonidium, and hippophae rhamnoides (sandy yam). The whole prescription is reasonably matched, has the effects of reducing phlegm and dampness, promoting blood circulation to remove blood stasis, and tonifying qi and invigorating the spleen. It is suitable for patients with non-alcoholic fatty liver with liver stagnation and spleen deficiency and phlegm and blood stasis.

[0106] (2) The preparation process has the following advantages: In order to obtain complete extraction of the effective components, the preparation process of the prescription (the corresponding clinical prescription is: Shenqu Jiangzhi prescription) is optimized according to the physicochemical properties of the effective components in the above-mentioned medicinal materials. The dry extract yield and the content of hesperidin are used as the index components for screening. The optimal preparation process parameters of the hippophae rhamnoides prescription are as follows: 8 times the amount of water is added, and decocting is carried out for 3 times, each time for 60 min. According to the optimal process, the dry extract yield of the hippophae rhamnoides prescription is 29.39-29.73%, and the content of hesperidin is 10.18-10.92 (mg·g -1 ). The optimal preparation process parameters of the sandy yam prescription are as follows: 8 times the amount of water is added, and decocting is carried out for 3 times, each time for 60 min. According to the optimal process, the dry extract yield of the sandy yam prescription is 28.09-28.37%, and the content of hesperidin is 10.18-10.41 (mg·g -1 ). The whole prescription is simple and the raw materials are easy to obtain. The preparation process is scientific, reasonable, simple and feasible, and has the prospect of industrial application.

[0107] (3) The pharmacological test results show that the water decoction prepared according to the extraction process of the traditional Chinese medicine composition (hippophae rhamnoides prescription) and the traditional Chinese medicine composition (sandy yam prescription) has a reducing effect on the liver function, blood lipid level and antioxidant index of mice, has a reducing effect on the liver coefficient, and can significantly improve the accumulation of liver fat. This indicates that the traditional Chinese medicine composition has a significant therapeutic effect on non-alcoholic fatty liver.

[0108] (4) The traditional Chinese medicine composition (sea buckthorn formula) and the traditional Chinese medicine composition (astragalus complanatus formula) have been clinically verified to have significant clinical efficacy in treating non-alcoholic fatty liver disease. The total effective rate of clinical treatment is over 90%. After taking the traditional Chinese medicine of this invention for 3 courses of treatment, the symptoms of most patients are significantly improved and their transaminase levels return to normal. Moreover, no adverse reactions have been found during the use of the above-mentioned drugs, which has the advantage of high safety.

[0109] Instruction manual illustrations

[0110] The accompanying drawings are provided to further explain the preparation, detection methods, and efficacy of the compositions of the present invention and constitute a part of the present invention. The illustrative embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute an improper limitation of the present invention.

[0111] Figure 1 -HPLC chromatograms of the reference standard and the test sample of the traditional Chinese medicine composition (sea buckthorn formula) of the present invention, wherein A: reference standard (hesperidin) and B: test sample solution (1-hesperidin).

[0112] Figure 2 - The number of extractions of the traditional Chinese medicine composition (sea buckthorn formula) of this invention was investigated, including the content of hesperidin and the content of dry extract;

[0113] Figure 3 -HPLC chromatograms of the reference standard and the test sample of the traditional Chinese medicine composition (Shayuanzi formula) of the present invention, wherein A: reference standard (1-hesperidin) and B: test sample solution (1-hesperidin).

[0114] Figure 4 - Changes in body weight, liver coefficient, liver function, blood lipid levels, and oxidative and antioxidant indices in the liver of mice in each group of the traditional Chinese medicine composition (sea buckthorn formula) of this invention; Note: Compared with the blank group. # P<0.05 ## P<0.01 ### P<0.001; compared with the model group, *P<0.05, **P<0.01.

[0115] Figure 5 - HE staining (×200) and Oil Red O staining (×100) of mouse liver tissue using the traditional Chinese medicine composition (sea buckthorn formula) of this invention;

[0116] Figure 6 - Changes in body weight, liver coefficient, liver function, blood lipid levels, and oxidative and antioxidant indices in the liver of mice in each group of the traditional Chinese medicine composition (Sha Yuan Zi Fang) of this invention; Note: Compared with the blank group. # P<0.05 ## P<0.01 ###P<0.001; compared with the model group, *P<0.05, **P<0.01.

[0117] Figure 7 - The HE staining and oil red O staining of the liver tissue of the mouse treated with the traditional Chinese medicine composition (Sahuyanzi recipe) in the application (×200). DETAILED DESCRIPTION

[0118] In order to more fully understand the implementation of the application, one or more experimental examples are listed below, and the application is further described below through typical examples.

[0119] Test Example 1

[0120] 1 The intervention effect of the traditional Chinese medicine composition (Sahujia recipe) in the application on non-alcoholic fatty liver mice induced by high-fat diet

[0121] 1.1 Establishment of non-alcoholic fatty liver mouse model

[0122] Test materials

[0123] Male C57BL6 / J mice, 7 weeks old, weighing 20±2 g, SPF level. The mice were allowed to drink water and eat freely, and the bedding was changed regularly. The mice were raised in a quiet environment with a temperature of 23-25℃, a relative humidity of 40%-60%, and a 12 h day-night alternation. After adaptive feeding for 7 days, the mice were randomly divided into a control group (10 mice) and a modeling group (30 mice). The modeling mice were fed with high-fat feed for 12 weeks. The sample of the Sahujia recipe in the application was prepared by the method of Example 1 of the application.

[0124] 1.2 Grouping and administration

[0125] At the end of the 8th week of modeling, the modeling mice were randomly divided into a model group, a high-dose group, and a low-dose group, 10 mice in each group. From the 9th week of modeling, the mice were administered once a day by gavage, with a dosage of 0.2 mL·10g-1, and the administration was continued for 8 weeks. The control group and the model group were administered with the same volume of normal saline by gavage.

[0126] 1.3 Sample preparation

[0127] After the last administration, the mice were fasted for 12 h without water, and the next day, the body weight of each group of mice was measured, the eyeball was taken for blood sampling, and the serum was prepared by centrifugation and stored in a-80℃ refrigerator for testing. The liver tissue was taken, weighed, and the organ coefficient was calculated; two pieces of liver tissue were taken from the left lobe of the liver, fixed with 4% paraformaldehyde, and the rest was stored at-80℃ for testing. Organ coefficient = organ weight (g) / mouse body weight (g) × 100%.

[0128] 1.4 Detection of blood lipids, serum transaminase levels, and antioxidant indicators in liver tissue

[0129] The levels of triglyceride (TG), total cholesterol (TC), alanine aminotransferase (ALT), aspartate aminotransferase (ALT) in mouse serum, and the levels of catalase (CAT), reduced glutathione (GSH), superoxide dismutase (SOD) in liver tissue were determined according to the kit instructions. The results are shown in Figure 2 .

[0130] 1.5 HE staining and oil red O staining to observe the pathological changes and lipid deposition in liver tissue

[0131] About 0.2 g of left lobe tissue of each group of mice was taken, 4% paraformaldehyde was added for fixation for 24 h, dehydration, wax immersion, and embedded in paraffin. The trimmed wax block was placed in a paraffin sectioning machine to cut into 4 μm thick sections. Hematoxylin-eosin (HE) staining was performed, then the sections were dehydrated and immediately mounted on glass slides with neutral gum. The sections were observed and photographed under a white light microscope, and pathological analysis was performed.

[0132] The results showed that the liver lobule structure of normal mice was complete, the hepatocyte morphology was complete, the arrangement was compact, the cytoplasm was rich, there was no fatty degeneration and inflammatory cell infiltration in the lobule. Obvious fatty degeneration, fat vacuoles and inflammatory cell infiltration were observed in the model mice, the liver cells were enlarged and rounded, the cytoplasm was loose, containing large fat droplets, the nucleus was pushed to one side of the cell, showing different degrees of edema degeneration. The liver fatty degeneration and inflammatory reaction of the treatment group were significantly reduced compared with the model group, the liver cell volume was smaller, the liver lobule outline was clear, and the number of fat droplets was reduced.

[0133] Fresh liver left lobe tissue of each group was taken for frozen section, immersed in formaldehyde solution, and stained with oil red O according to the requirements and steps of the saturated oil red O kit, and finally mounted with glycerol gelatin. The distribution of red lipid droplets in liver tissue was observed under a microscope. The results showed that the liver of normal mice had almost no lipid droplets, the liver of model mice had the largest area of fat droplets, and the liver of treatment mice had different degrees of reduction in texture. The results are shown in the description Figure 5 .

[0134] In summary, the water decoction prepared by the extraction process of the traditional Chinese medicine composition (sea buckthorn recipe) has a reducing effect on liver function, blood lipid level and liver antioxidant index of mice, has a reducing effect on liver coefficient, and can significantly improve liver fat accumulation, indicating that the present application has a certain therapeutic effect on non-alcoholic fatty liver.

[0135] Test Example 2

[0136] Intervention effect of traditional Chinese medicine composition (Hawthorn recipe) on high-fat diet-induced non-alcoholic fatty liver mice

[0137] 2.1 Establishment of non-alcoholic fatty liver mouse model

[0138] Experimental materials

[0139] Male C57BL6 / J mice, 7 weeks old, weighing 20±2 g, SPF level. The mice were free to drink water and eat, and the bedding was changed regularly, and were raised in a quiet environment, temperature 23-25 ℃, relative humidity 40%-60%, and 12 h day-night alternation. After adaptive feeding for 7 days, the mice were randomly divided into a control group (10) and a modeling group (30), and the modeling mice were fed with high-fat feed for 12 weeks. The preparation of the Hippophae rhamnoides L. formula sample was prepared by the method of Example 4 of the present application.

[0140] 2.2 Grouping and administration

[0141] At the end of the 8th week of modeling, the modeling mice were randomly divided into a model group, a high-dose group, and a low-dose group, 10 in each group; from the 9th week of modeling, the mice were administered once a day by gavage, with a dosage of 0.2 mL·10g-1, for 8 consecutive weeks; the control group and the model group were given the same volume of normal saline by gavage.

[0142] 2.3 Sample preparation

[0143] After the last administration, the mice were fasted for 12 h without water, and the next day, the body weight of the mice in each group was measured, the mice were exophthalmos, and the blood was centrifuged to prepare serum, which was stored in a-80 ℃ refrigerator for testing. The liver tissue was taken out, weighed, and the organ coefficient was calculated; two pieces of liver tissue were cut from the left lobe of the liver, fixed with 4 % paraformaldehyde, and the rest was stored at-80 ℃ for testing. Organ coefficient = organ weight (g) / mouse body weight (g) x 100%

[0144] 2.4 Detection of blood lipids, serum transaminase levels, and antioxidant indexes in liver tissue

[0145] The levels of triglyceride (TG), total cholesterol (TC), alanine aminotransferase (ALT), and aspartate aminotransferase (ALT) in mouse serum, and the levels of catalase (CAT), reduced glutathione (GSH), and superoxide dismutase (SOD) in liver tissue were determined according to the instructions of the kit. The results are shown in the instructions Figure 6 .

[0146] 2.5 HE staining and oil red O staining to observe the pathological changes and lipid deposition in liver tissue

[0147] About 0.2 g of liver tissue from the left lobe of the mice in each group was taken, and 4 % paraformaldehyde was added for fixation for 24 h, followed by dehydration and wax immersion, and embedding in paraffin. The trimmed wax block was cut into a 4 μm thick section on a paraffin sectioning machine. Hematoxylin-eosin (HE) staining was performed, and then the section was dehydrated and immediately sealed with neutral gum on a glass slide. The section was observed and photographed under a white light microscope, and pathological analysis was performed.

[0148] The results show that the liver lobule structure of the normal group mice is complete, the hepatocyte morphology is complete, the arrangement is compact, the cytoplasm is rich, there is no fatty degeneration and inflammatory cell infiltration in the lobule. The model group mice can see obvious fatty degeneration, fat vacuoles and inflammatory cell infiltration, the hepatocyte is enlarged and rounded, the cytoplasm is loose, contains large fat droplets, the nucleus is pushed to one side of the cell, presents different degrees of edema degeneration, the liver fatty degeneration and inflammatory reaction of the treatment group mice are obviously reduced compared with the model group, the liver cell volume is smaller, the liver lobule contour is clear, and the number of fat droplets is reduced.

[0149] The left lobe of the liver of each group is taken to make frozen sections, is soaked in formaldehyde solution, is dyed by oil red O according to the requirements and steps of the saturated oil red O kit, and finally is sealed with glycerol gelatin. The distribution of red lipid droplets in the liver tissue is observed under a microscope. The results show that the liver of the normal group mice has almost no lipid droplets, the liver fat lipid droplet area of the model group mice is the largest, and the liver texture of the treatment group mice is reduced to different degrees. The results are shown in Figure 7 .

[0150] In summary, the water decoction prepared by the extraction process of the traditional Chinese medicine composition (Sahyunzi recipe) has a reducing effect on the liver function, blood lipid level, liver oxidation and antioxidant index of mice, has a reducing effect on the liver coefficient, and can significantly improve the liver fat accumulation, which indicates that the traditional Chinese medicine composition has a certain therapeutic effect on non-alcoholic fatty liver.

[0151] Example 1 Preparation of the traditional Chinese medicine composition (Sahyunzi recipe) in the present application

[0152] Take 6g of ginseng, 3g of red yeast, 3g of daida flower, 10g of dried tangerine or orange peel, 10g of Herba Lycopi, and 6g of Hippophae, immerse the above six medicinal materials in water with a weight of 8 times that of the medicinal materials, decoct for 3 times, decoct for 1 hour, filter, combine the above water decoction filtrates, reduce pressure to concentrate the liquid, then add 50g of sucrose and 0.3g of sodium benzoate, mix uniformly, and adjust with water to prepare an oral solution.

[0153] Example 2 Preparation of the traditional Chinese medicine composition (Sahyunzi recipe) in the present application

[0154] Take 10g of ginseng, 3g of red yeast, 6g of daida flower, 12g of dried tangerine or orange peel, 6g of Herba Lycopi, and 12g of Hippophae, immerse the above six medicinal materials in water with a weight of 13 times that of the medicinal materials, decoct for 1 time, decoct for 1.5 hours, filter, combine the above water decoction filtrates, reduce pressure to concentrate the liquid, then add 60g of aspartame and 0.4g of sorbic acid, mix uniformly, and adjust with water to prepare an oral solution.

[0155] Example 3 Preparation of the traditional Chinese medicine composition (Sahyunzi recipe) in the present application

[0156] Take 5g of ginseng rootlets, 6g of red yeast rice, 3g of bitter orange blossom, 6g of dried tangerine peel, 12g of eupatorium, and 6g of sea buckthorn. Add 6 times the weight of the above six medicinal materials to water, soak them, decoct 3 times, and decoct for 0.5 hours each time. Filter, combine the above decoction extracts, concentrate under reduced pressure and dry into a dry powder. Then add 40g of dextrin, 20g of starch, and appropriate amount of water, mix well, granulate, dry, and make into granules.

[0157] Example 4: Preparation of the traditional Chinese medicine composition (Shayuanzi formula) of the present invention

[0158] Take 6g of ginseng rootlets, 3g of red yeast rice, 3g of bitter orange blossom, 10g of dried tangerine peel, 10g of Lycopus lucidus, and 15g of Astragalus complanatus. Add 8 times the weight of the above six medicinal materials to water, soak them, and decoct twice for 1 hour each time. Filter the decoction, combine the above decoction filtrates, concentrate them under reduced pressure, add 50g of white sugar and 0.5g of sodium benzoate, mix well, and add water to make an oral liquid.

[0159] Example 5: Preparation of the traditional Chinese medicine composition (Sha Yuan Zi Fang) of the present invention

[0160] Take 10g of ginseng rootlets, 3g of red yeast rice, 3g of bitter orange blossom, 12g of dried tangerine peel, 6g of Lycopus lucidus, and 15g of Astragalus complanatus. Add 10 times the weight of the above six medicinal materials to water and decoct once, each time for 1 hour. Filter the decoction, combine the above decoction filtrates, concentrate under reduced pressure, add 50g of white sugar and 0.5g of sorbic acid, mix well, and add water to make an oral liquid.

[0161] Example 6: Preparation of the traditional Chinese medicine composition (Shayuanzi formula) of the present invention

[0162] Take 5g of ginseng rootlets, 6g of red yeast rice, 6g of bitter orange blossom, 6g of dried tangerine peel, 12g of Lycopus lucidus, and 9g of Astragalus complanatus. Add 11 times the weight of the above six medicinal materials to water, soak them, decoct twice for 1.5 hours each time, filter, combine the above decoction extracts, concentrate under reduced pressure and dry into a dry powder, then add 50g of lactose, 30g of starch, and appropriate amount of water, mix well, granulate, dry, and make into granules.

Claims

1. A traditional Chinese medicine composition for treating non-alcoholic fatty liver, characterized in that, The traditional Chinese medicine composition is prepared from raw medicinal materials in the following proportions by weight: ginseng stem 5-10 parts, red yeast 3-6 parts, day day flower 3-6 parts, dried tangerine or orange peel 6-12 parts, and herba lycopi 6-12 parts, and sea buckthorn 6-12 parts.

2. The traditional Chinese medicine composition of claim 1, wherein, The traditional Chinese medicine composition is prepared from raw medicinal materials in the following proportions by weight: ginseng stem 6 parts, red yeast 3 parts, day day flower 3 parts, dried tangerine or orange peel 10 parts, and herba lycopi 10 parts, and sea buckthorn 6 parts.

3. A traditional Chinese medicine composition for treating non-alcoholic fatty liver, characterized in that, The traditional Chinese medicine composition is prepared from raw medicinal materials in the following proportions by weight: ginseng stem 5-10 parts, red yeast 3-6 parts, day day flower 3-6 parts, dried tangerine or orange peel 6-12 parts, and herba lycopi 6-12 parts, and sea buckthorn 9-15 parts.

4. The traditional Chinese medicine composition of claim 3, wherein, The traditional Chinese medicine composition is prepared from raw medicinal materials in the following proportions by weight: ginseng stem 6 parts, red yeast 3 parts, day day flower 3 parts, dried tangerine or orange peel 10 parts, and herba lycopi 10 parts, and sea buckthorn 15 parts.

5. The preparation method of the traditional Chinese medicine composition according to any one of claims 1-4, characterized in that, The preparation method comprises the following steps: taking the medicinal materials in the above proportions by weight, adding 6-13 times the weight of water, soaking, decocting 1-3 times, each time for 0.5-1.5 hours, filtering, combining the water decoction extraction filtrates, and drying the filtrates under reduced pressure to obtain a dry paste powder intermediate, and adding pharmaceutical adjuvants to prepare a traditional Chinese medicine oral preparation.

6. The preparation method of the traditional Chinese medicine composition according to claim 5, characterized in that, In the preparation method, 8 times the weight of water is added, and the decoction is performed 3 times, each time for 1 hour.

7. The preparation method of the traditional Chinese medicine composition according to claim 5, characterized in that, The traditional Chinese medicine oral preparation is an oral liquid or granules.

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