A traditional Chinese medicine composition for treating liver fibrosis and its preparation method

Abstract

This invention relates to a traditional Chinese medicine composition for treating liver fibrosis and its preparation method. The components or raw materials of this traditional Chinese medicine composition include: 20-30 parts of *Lysimachia christinae*, 8-15 parts of *Trionyx sinensis* shell, 10-16 parts of *Litchi chinensis* seed, 5-15 parts of *Panax notoginseng*, 5-10 parts of *Curcuma zedoaria*, and 8-12 parts of *Poria cocos*. This traditional Chinese medicine composition can be prepared into various pharmaceutical preparations using conventional processes. The components of the above-mentioned traditional Chinese medicine composition are rationally combined, making it a safe and effective treatment for liver fibrosis.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine technology, specifically to a traditional Chinese medicine composition for treating liver fibrosis and its preparation method. Background Technology

[0002] The liver is a vital organ in the human body. Liver fibrosis is a common pathological stage in the development of many chronic liver diseases. It is a pathological process caused by the abnormal growth of connective tissue in the liver and excessive accumulation of diffuse extracellular matrix (ECM). If timely and effective treatment is not taken, it will lead to cirrhosis and liver cancer.

[0003] In the prior art, there are some traditional Chinese medicine compositions used to treat liver fibrosis. For example, patent document CN112043758A provides a traditional Chinese medicine composition for treating cirrhosis and liver fibrosis, which includes formula 1 and formula 2. Formula 1 includes bezoar, artificial bezoar, safflower, safflower root, madder root, costus root, melon seeds, rock essence, blue-flowered meconopsis, dayflower, swert, and sea buckthorn paste. Formula 2 includes pomegranate, agarwood, cinnamon, cardamom, long pepper, dried ginger, nutmeg, jujube, frankincense, costus root, chebula, kapok flower, bamboo shavings, caragana, and bergamot. For example, patent document CN103330882A provides a traditional Chinese medicine tablet for treating chronic hepatitis and liver fibrosis and its preparation method. The traditional Chinese medicine tablet is composed of Oldenlandia diffusa, Acorus tatarinowii, Paeonia lactiflora, Panax notoginseng, Polygonum hydropiper, Angelica sinensis, Vespa nidus, Sophora flavescens, Hirudo medicinalis, Sparganium stoloniferum, Corydalis yanhusuo, carbonized human hair, Vaccaria segetalis, Coix lacryma-jobi, Coptis chinensis, Dioscorea opposita, Polygonatum sibiricum, Paeonia lactiflora and Spatholobus suberectus.

[0004] There are many traditional Chinese medicine (TCM) compositions for treating liver fibrosis, but they all have drawbacks, such as irrational treatment principles and compatibility, insignificant efficacy, and poor treatment results. Therefore, there is an urgent need for a TCM composition that is effective, practical, has high compliance, and is safe and effective in treating liver fibrosis. Summary of the Invention

[0005] The purpose of this invention is to provide a safe and effective traditional Chinese medicine composition for treating liver fibrosis and its preparation method. To this end, the present invention adopts the following technical solution:

[0006] In a first aspect, the present invention provides a traditional Chinese medicine composition for treating liver fibrosis, the raw materials for preparing it comprising the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of Trionyx sinensis, 10-16 parts of Litchi chinensis seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, and 8-12 parts of Poria cocos.

[0007] In one or more embodiments, the raw materials comprise the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of Trionyx sinensis, 10-16 parts of Litchi chinensis seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, 8-12 parts of Poria cocos, and 10-15 parts of Cyperus rotundus.

[0008] In one or more embodiments, the raw materials comprise the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of Trionyx sinensis, 10-16 parts of Litchi chinensis seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, 8-12 parts of Poria cocos, 10-15 parts of Cyperus rotundus, and 5-10 parts of Atractylodes macrocephala.

[0009] In one or more embodiments, the raw materials comprise the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of Trionyx sinensis, 10-16 parts of Litchi chinensis seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, 8-12 parts of Poria cocos, 10-15 parts of Cyperus rotundus, 5-10 parts of Atractylodes macrocephala, and 2-6 parts of Glycyrrhiza uralensis.

[0010] Secondly, the present invention provides a method for preparing the above-mentioned traditional Chinese medicine composition, which involves mixing and pulverizing the raw materials to obtain the traditional Chinese medicine composition.

[0011] In one or more embodiments, the raw materials are mixed and pulverized to 10-200 mesh.

[0012] Thirdly, the present invention provides another method for preparing the above-mentioned traditional Chinese medicine composition, which involves extracting the raw materials with water to obtain an aqueous extract, using the aqueous extract as a traditional Chinese medicine preparation, or further preparing the aqueous extract with pharmaceutically acceptable excipients into other traditional Chinese medicine dosage forms, and the resulting traditional Chinese medicine preparation is the traditional Chinese medicine composition.

[0013] In one or more embodiments, the dosage form of the traditional Chinese medicine preparation is a decoction, oral liquid, drops, ointment, pill, powder, tablet or capsule.

[0014] In one or more embodiments, the water extraction method includes: boiling the raw material with water several times, and finally combining the decoctions obtained from each boiling.

[0015] In one or more embodiments, the decoction is performed 1-3 times, with the amount of water added each time being 5-15 times the total weight of the raw materials, and the decoction time being 30-120 minutes each time.

[0016] This invention achieves the following beneficial effects: Through a specific ratio of nine herbs including Lysimachia christinae, turtle shell, and Panax notoginseng, it achieves a synergistic effect of "softening and dispersing nodules" and "strengthening the spleen and soothing the liver," overcoming the limitations of existing technologies where single-drug targets are limited or traditional compound formulas suffer from imbalances in efficacy. This significantly improves the overall efficacy in reversing liver fibrosis and alleviating clinical symptoms. Furthermore, the adjuvant effect of tonifying herbs (Atractylodes macrocephala, Poria cocos, and Glycyrrhiza uralensis) effectively reduces the potential gastrointestinal irritation risk of blood-activating and stasis-removing herbs (Curcuma zedoaria and Panax notoginseng). Compared to existing anti-fibrotic Western medicines, this product has fewer toxic side effects, making it more suitable for long-term use by liver disease patients to consolidate therapeutic effects, and resulting in high patient compliance. Attached Figure Description

[0017] Figure 1 Inhibition rate of each group in the HSC-T6 cell proliferation inhibition experiment.

[0018] Figure 2 : Pathological morphological changes in liver tissue of rats in different groups during animal experiments; from left to right, they are the normal control group, the model group, and the freeze-dried Chinese medicine powder group.

[0019] Figure 3 Comparative experiment: HSC-T6 cell proliferation inhibition rate in each group. Detailed Implementation

[0020] The present invention will now be described in detail with reference to exemplary embodiments. However, it should be understood that the present invention may be implemented in many different forms and should not be construed as limited to the embodiments set forth herein. These embodiments are provided herein to make the disclosure of the present invention more complete and to fully convey the inventive concept to those skilled in the art.

[0021] First, this invention provides a traditional Chinese medicine composition for treating liver fibrosis, comprising, by weight, at least the following components: 20-30 parts of *Gynostemma pentaphyllum*, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of *Panax notoginseng*, 5-10 parts of *Curcuma zedoaria*, and 8-12 parts of *Poria cocos*. The raw materials may also include *Cyperus rotundus*, meaning the raw materials comprise the following components by weight: 20-30 parts of *Gynostemma pentaphyllum*, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of *Panax notoginseng*, 5-10 parts of *Curcuma zedoaria*, 8-12 parts of *Poria cocos*, and 10-15 parts of *Cyperus rotundus*. Further, the raw material may also include the following components in parts by weight: 20-30 parts of *Lysimachia christinae*, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of *Panax notoginseng*, 5-10 parts of *Curcuma zedoaria*, 8-12 parts of *Poria cocos*, 10-15 parts of *Cyperus rotundus*, and 5-10 parts of *Atractylodes macrocephala*. Even further, the raw material may also include licorice, in which case the raw material comprises the following components in parts by weight: 20-30 parts of *Lysimachia christinae*, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of *Panax notoginseng*, 5-10 parts of *Curcuma zedoaria*, 8-12 parts of *Poria cocos*, 10-15 parts of *Cyperus rotundus*, 5-10 parts of *Atractylodes macrocephala*, and 2-6 parts of licorice.

[0022] The above raw material components are common Chinese medicinal materials in the field, and their general sources are as follows: Phyllodium pulchellum is the dried whole herb of Phyllodium pulchellum (L.) Desv. of the Leguminosae family. Litchi seed is the dried mature seed of Litchi chinensis Sonn. of the Sapindaceae family. Panax notoginseng is the dried root and rhizome of Panax notoginseng (Burkill) F.H.Chen of the Araliaceae family. Cyperus rotundus is the dried rhizome of Cyperus rotundus L. of the Cyperaceae family. Atractylodes macrocephala is the dried rhizome of Atractylodes macrocephala Koidz. of the Asteraceae family. Rhizoma curcumae is the dried rhizome of Curcuma phaeocaulis Val., Curcuma kwangsiensis S.G.Lee et C.F.Liang or Curcuma wenyujin Y.H.Chen et C.Ling of the Zingiberaceae family. Turtle shell is the dried carapace of Trionyx sinensis Wiegmann of the Trionychidae family. Poria cocos is the dried sclerotium of Poria cocos (Schw.) Wolf of the Polyporaceae family. Licorice is the dried root and rhizome of Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Bat. or Glycyrrhiza glabra L. of the Leguminosae family.

[0023] In traditional Chinese medicine, liver fibrosis is mostly caused by prolonged illness invading the collaterals, resulting in the liver losing its function of dredging and dispersing, qi stagnation and blood stasis; the spleen losing its function of transportation and transformation, and dampness accumulating to form phlegm; phlegm and blood stasis intermingling and blocking the liver collaterals, and over time, a mass (fibrosis) is formed. Its characteristic is deficiency of healthy qi (deficiency of the spleen and kidney). The formula of the present invention closely adheres to the pathogenesis characteristics of liver fibrosis, namely "deficiency in origin and excess in superficiality, blood stasis blocking collaterals, and phlegm-dampness intermingling", and adopts the treatment methods of strengthening healthy qi and eliminating pathogenic factors, softening hardness and dissipating nodules, and promoting blood circulation to remove blood stasis. Thus, it supplements qi and strengthens the spleen to strengthen healthy qi, promotes blood circulation to remove blood stasis, softens hardness and dissipates nodules to eliminate pathogenic factors, and promotes qi movement and diuresis to dredge collaterals. The analysis of the compatibility of monarch, minister, assistant and envoy is as follows:

[0024] The herb *Lysimachia christinae* is used in the largest quantity, making it the king of the formula. It possesses extremely strong effects in promoting blood circulation, removing blood stasis, softening and dispersing masses, and promoting diuresis and reducing swelling. Modern pharmacological research shows that it has a significant anti-liver fibrosis effect, inhibiting the activation of hepatic stellate cells and reducing collagen deposition. *Turtle shell* is a key ingredient; its salty and cold properties enter the liver, excelling at nourishing yin and suppressing yang, softening and dispersing masses. It is a crucial medicine for treating hypochondriac masses (hepatosplenomegaly, fibrosis) in liver disease, and can assist *Lysimachia christinae* in softening already formed hard fibrous tissue, hence the saying "saltyness can soften hardness." Panax notoginseng and Curcuma zedoaria serve as assistant herbs to the principal herb. Panax notoginseng promotes blood circulation, stops bleeding, and relieves pain; it stops bleeding without leaving blood stasis and promotes blood circulation without harming the body's vital energy, improving liver microcirculation and promoting the repair of damaged liver cells. Curcuma zedoaria breaks up blood stasis, promotes qi circulation, eliminates stagnation, and relieves pain; its blood-breaking power is strong, specifically targeting stagnation of qi and blood. Combined with Panax notoginseng, one breaks up blood stasis while the other stops, combining movement and stillness to powerfully clear blood stasis in the liver meridians. Together, they assist the principal herb in strengthening its ability to promote blood circulation, eliminate blood stasis, and resolve masses, addressing the key pathogenic factor of "blood stasis." Litchi seed is also an assistant herb; it promotes qi circulation, disperses stagnation, dispels cold, and relieves pain, specifically entering the liver meridian. It is used for hypochondriac pain due to liver qi stagnation. By soothing the liver and regulating qi, it assists the blood-activating herbs in better exerting their blood-stasis-removing effects, preventing qi stagnation from worsening blood stasis. Atractylodes macrocephala, Poria cocos, and Cyperus rotundus are adjuvant herbs. Atractylodes macrocephala and Poria cocos are both essential herbs for strengthening the spleen and replenishing qi, as well as drying / draining dampness. Liver disease inevitably affects the spleen; spleen deficiency leads to dampness, and dampness accumulation promotes phlegm and blood stasis. Atractylodes macrocephala and Poria cocos strengthen the spleen and eliminate dampness, cutting off the source of phlegm and dampness and preventing the formation of fibrosis. Furthermore, the formula contains many blood-breaking herbs (Curcuma zedoaria and Panax notoginseng), which can easily damage the body's vital energy. Atractylodes macrocephala and Poria cocos can protect the spleen and stomach qi, allowing the attack on pathogenic factors to proceed without harming the body's vital energy. Cyperus rotundus soothes the liver, relieves depression, regulates qi, relieves chest congestion, regulates menstruation, and alleviates pain. It can assist Litchi chinensis in strengthening the liver-soothing and qi-regulating effects, resolving the fundamental problem of liver qi stagnation. When qi flows smoothly, blood stasis naturally dissipates. Licorice is the guiding herb, harmonizing the other herbs and guiding them to their respective meridians. Licorice itself has liver-protecting and detoxifying effects, providing direct protection for the liver. In summary, this formula uses Lysimachia christinae and turtle shell to soften and disperse nodules as the principal herbs; Panax notoginseng, Curcuma zedoaria, and litchi seed to invigorate blood and regulate qi as the assistant herbs; Atractylodes macrocephala, Poria cocos, and Cyperus rotundus to strengthen the spleen and soothe the liver as the adjuvant herbs; and Glycyrrhiza uralensis to harmonize the formula as the guiding herb. The entire formula works synergistically to soothe the liver and strengthen the spleen, invigorate blood and remove blood stasis, soften and disperse nodules, effectively addressing the core pathogenesis of liver fibrosis: "qi deficiency and blood stasis, phlegm and blood stasis intertwined."

[0025] Using the above-mentioned components as raw materials, two main types of traditional Chinese medicine (TCM) products can be prepared: one is by directly pulverizing the mixed raw materials, generally to a particle size of 10-200 mesh; the other is by extracting the raw materials with water to obtain an aqueous extract, which is then used as a TCM preparation, or the aqueous extract can be further processed with pharmaceutically acceptable excipients to prepare other TCM dosage forms. The TCM preparations include liquid, semi-solid, or solid preparations. For example, they can be decoctions, oral liquids, drops, ointments, pills, powders, tablets, capsules, or any other usable dosage form. The aqueous extraction involves decocting the raw materials several times and finally combining the decoctions obtained from each decoction. The number of decoctions is 1-3 times, the amount of water added each time is 5-15 times the total weight of the raw materials, and the decoction time is 30-120 minutes each time.

[0026] Furthermore, the present invention provides several specific embodiments to further illustrate the composition and preparation method of the above-mentioned drug for treating liver fibrosis, specifically as follows:

[0027] Example 1

[0028] 1) Weigh the raw materials according to the following weight parts: 25 parts of Lysimachia christinae, 12 parts of turtle shell, 12 parts of litchi seed, 10 parts of Panax notoginseng, 8 parts of Curcuma zedoaria, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, 8 parts of Atractylodes macrocephala, and 5 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct twice, each time adding water 10 times the total weight of the raw materials, and decoct for 30 minutes. Filter, concentrate the filtrate, and freeze-dry to obtain freeze-dried Chinese medicine powder. The freeze-dried powder is used as a powder.

[0029] Example 2

[0030] 1) Weigh the raw materials according to the following weight proportions: 20 parts of Lysimachia christinae, 8 parts of turtle shell, 10 parts of litchi seed, 5 parts of Panax notoginseng, 5 parts of Curcuma zedoaria, and 8 parts of Poria cocos; 2) Add water to the raw materials and decoct once, with the amount of water being 5 times the total weight of the raw materials. The decoction time is 30 minutes. Filter the solution, and the resulting filtrate is the traditional Chinese medicine decoction for treating liver fibrosis.

[0031] Example 3

[0032] 1) Weigh the raw materials according to the following weight proportions: 30 parts of Lysimachia christinae, 15 parts of turtle shell, 16 parts of litchi seed, 15 parts of Panax notoginseng, 10 parts of Curcuma zedoaria, 12 parts of Poria cocos, and 15 parts of Cyperus rotundus; 2) Add water to the raw materials and decoct three times, each time adding water at a rate of 15 times the total weight of the raw materials, and each decoction time being 120 minutes. Combine the three decoctions, filter, and concentrate the filtrate into an extract; 3) Dry and pulverize the extract, add an appropriate amount of honey, and refine it into pills. Concentrate the pills according to conventional processes in this field.

[0033] Example 4

[0034] 1) Weigh the raw materials according to the following weight parts: 25 parts of Lysimachia christinae, 12 parts of turtle shell, 12 parts of litchi seed, 10 parts of Panax notoginseng, 8 parts of Curcuma zedoaria, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, and 8 parts of Atractylodes macrocephala; 2) Add water to the raw materials and decoct twice, each time adding water at a rate of 8 times the total weight of the raw materials, and each decoction time is 60 minutes. Combine the two decoctions, filter, and concentrate the filtrate into an extract; 3) Dry and granulate the extract, add starch and magnesium stearate, and compress it into tablets according to conventional processes in the art.

[0035] Example 5

[0036] 1) Weigh the raw materials according to the following weight parts: 28 parts of Lysimachia christinae, 14 parts of turtle shell, 15 parts of litchi seed, 12 parts of Panax notoginseng, 9 parts of Curcuma zedoaria, 11 parts of Poria cocos, 14 parts of Cyperus rotundus, 9 parts of Atractylodes macrocephala, and 5 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct once, with the amount of water being 12 times the total weight of the raw materials, for a decoction time of 90 minutes, filter, concentrate and dry the filtrate into a fine powder; 3) Fill the fine powder into capsule shells and prepare capsules according to conventional processes in the art.

[0037] Example 6

[0038] 1) Weigh the raw materials according to the following weight proportions: 22 parts of Lysimachia christinae, 9 parts of turtle shell, 11 parts of litchi seed, 6 parts of Panax notoginseng, 6 parts of Curcuma zedoaria, 9 parts of Poria cocos, 11 parts of Cyperus rotundus, 6 parts of Atractylodes macrocephala, and 3 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct three times, each time adding water at a rate of 6 times the total weight of the raw materials, and each decoction time being 40 minutes. Combine the three decoctions, filter, and concentrate the filtrate to an appropriate amount; 3) Add sucrose and preservatives to the filtrate, adjust the pH value, fill and sterilize, and prepare an oral liquid according to conventional processes in this field.

[0039] Example 7

[0040] 1) Weigh the raw materials according to the following weight parts: 24 parts of Lysimachia christinae, 10 parts of turtle shell, 13 parts of litchi seed, 8 parts of Panax notoginseng, 7 parts of Curcuma zedoaria, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, 7 parts of Atractylodes macrocephala, and 4 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct twice, each time adding water at a rate of 10 times the total weight of the raw materials, and each decoction time is 80 minutes. Combine the decoctions, filter, and concentrate the filtrate into a clear extract; 3) Add dextrin and stevia to the clear extract, granulate and dry, and prepare granules according to conventional processes in the field.

[0041] Example 8

[0042] 1) Weigh the raw materials according to the following weight proportions: 26 parts of Lysimachia christinae, 13 parts of turtle shell, 14 parts of litchi seed, 11 parts of Panax notoginseng, 8 parts of Curcuma zedoaria, 11 parts of Poria cocos, 13 parts of Cyperus rotundus, 8 parts of Atractylodes macrocephala, and 5 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct once, with the amount of water being 15 times the total weight of the raw materials, for a decoction time of 100 minutes, filter, and spray-dry the filtrate into a fine powder; 3) Directly package the fine powder to make a powder.

[0043] Example 9

[0044] 1) Weigh the raw materials according to the following weight proportions: 23 parts of Lysimachia christinae, 11 parts of turtle shell, 12 parts of litchi seed, 9 parts of Panax notoginseng, 7 parts of Curcuma zedoaria, 9 parts of Poria cocos, 11 parts of Cyperus rotundus, 7 parts of Atractylodes macrocephala, and 4 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct 3 times, each time adding water at a rate of 5 times the total weight of the raw materials, and each decoction time being 50 minutes. Combine the decoctions, filter, and concentrate the filtrate to a suitable density; 3) Mix the concentrated liquid with the matrix, drip it into the cooling liquid, and make drop pills according to the conventional process in this field.

[0045] Example 10

[0046] 1) Weigh the raw materials according to the following weight proportions: 27 parts of Lysimachia christinae, 12 parts of turtle shell, 13 parts of litchi seed, 10 parts of Panax notoginseng, 8 parts of Curcuma zedoaria, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, 8 parts of Atractylodes macrocephala, and 4 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct twice. The first time, add water equal to 8 times the total weight of the raw materials and decoct for 120 minutes; the second time, add water equal to 8 times the total weight of the raw materials and decoct for 30 minutes. Combine the decoctions, filter, and concentrate the filtrate; 3) Add refined honey to the concentrated liquid to make a paste, thus preparing a decoction paste.

[0047] Comparative Example 1

[0048] 1) Weigh the raw materials according to the following weight parts: 12 parts turtle shell, 12 parts litchi seed, 10 parts Panax notoginseng, 8 parts Curcuma zedoaria, 10 parts Poria cocos, 12 parts Cyperus rotundus, 8 parts Atractylodes macrocephala, and 5 parts licorice; 2) Add water to the raw materials and decoct twice, each time adding water 10 times the total weight of the raw materials, and decoct for 30 minutes. Filter, concentrate the filtrate, and freeze-dry to obtain freeze-dried Chinese medicine powder.

[0049] Comparative Example 2

[0050] 1) Weigh the raw materials according to the following weight parts: 25 parts of Lysimachia christinae, 12 parts of Litchi chinensis seed, 10 parts of Panax notoginseng, 8 parts of Curcuma zedoaria, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, 8 parts of Atractylodes macrocephala, and 5 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct twice, each time adding water 10 times the total weight of the raw materials, and decoct for 30 minutes. Filter, concentrate the filtrate, and freeze-dry to obtain freeze-dried Chinese medicine powder.

[0051] Comparative Example 3

[0052] 1) Weigh the raw materials according to the following weight parts: 25 parts of Lysimachia christinae, 12 parts of turtle shell, 10 parts of Panax notoginseng, 8 parts of Curcuma zedoaria, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, 8 parts of Atractylodes macrocephala, and 5 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct twice, each time adding water at a rate of 10 times the total weight of the raw materials, for a decoction time of 30 minutes. Filter, concentrate the filtrate, and freeze-dry to obtain freeze-dried Chinese medicine powder.

[0053] Comparative Example 4

[0054] 1) Weigh the raw materials according to the following weight parts: 25 parts of Lysimachia christinae, 12 parts of turtle shell, 12 parts of litchi seed, 10 parts of Poria cocos, 12 parts of Cyperus rotundus, 8 parts of Atractylodes macrocephala, and 5 parts of Glycyrrhiza uralensis; 2) Add water to the raw materials and decoct twice, each time adding water 10 times the total weight of the raw materials, and decoct for 30 minutes. Filter, concentrate the filtrate, and freeze-dry to obtain freeze-dried Chinese medicine powder.

[0055] Experimental Example 1: Inhibitory effect on HSC-T6 cell proliferation

[0056] 1. Experimental Materials and Reagents

[0057] Cell line: Rat hepatic stellate cell line HSC-T6. Main reagents: DMEM high-glucose medium (containing 10% fetal bovine serum FBS, 1% penicillin-dextrose antibody); CCK-8 assay kit (Dojin Chemical, Japan); Samples to be tested: Lyophilized powder prepared in Example 1 (reconstituted with DMSO to a 100mM stock solution and stored at -20℃); Positive control: Colchicine (a known anti-fibrotic drug); 0.25% trypsin-EDTA digestion solution; PBS buffer (calcium and magnesium-free). Main instrument: CO2 incubator (37℃, 5%). ); ELISA reader (detection wavelength 450nm); 96-well cell culture plate (flat bottom, sterile).

[0058] 2. Experimental Procedure

[0059] (1) HSC-T6 cells were placed in DMEM complete medium containing 10% FBS and incubated at 37°C and 5% FBS. Incubate in a saturated humidity incubator.

[0060] (2) Take HSC-T6 cells in the logarithmic growth phase, digest them with 0.25% trypsin, centrifuge after digestion (1000 rpm, 5 min), and discard the supernatant.

[0061] (3) Resuspend the cells in complete culture medium, count them, and adjust the cell density to the specified values. - cells / mL.

[0062] (4) Inoculate the cell suspension into a 96-well plate, 100 μL per well; add 100 μL of sterile PBS to the outermost wells of the 96-well plate to prevent liquid evaporation during culture from affecting the experimental results (edge ​​effect). Experimental data are only read from the inner 60 wells.

[0063] (5) Place the culture plate in the incubator for 12-24 hours to allow the cells to adhere fully.

[0064] (6) After the cells adhere to the wall, remove the old culture medium and add fresh complete culture medium containing different concentrations of drugs (100 μL per well, 6 replicates) according to the following groups: blank control group (complete culture medium containing 0.1% DMSO), positive control group (colchicine, 0.20 μg / mL), low-dose group (lyophilized powder of Example 1, 10 μg / mL), medium-dose group (lyophilized powder of Example 1, 50 μg / mL), high-dose group (lyophilized powder of Example 1, 100 μg / mL); blank well, 100 μL culture medium + 10 μL CCK-8 (cell-free).

[0065] (7) After administration, culture for 48 hours; after reaching the specified time point, add 10 μL of CCK-8 solution directly to each well; gently shake the culture plate to mix, put it back into the incubator and continue incubation for 1 hour; take out the culture plate and use an ELISA reader to measure the absorbance (OD value) of each well at a wavelength of 450 nm.

[0066] 3. Data Processing and Computation

[0067] Record the OD value of each well, remove outliers (such as excessive differences between replicates), and calculate the average value of each group. ) and standard deviation ( The cell proliferation inhibition rate (IR) is calculated using the following formula:

[0068]

[0069] 4. Experimental Results

[0070] See results Figure 1 The inhibition rates of the low, medium, and high dose groups of the traditional Chinese medicine were 26.5%, 50.7%, and 62.3%, respectively, while the inhibition rate of the positive control group was 48.9%. Compared with the blank control group, the traditional Chinese medicine inhibited the proliferation of HSC-T6 cells in a dose-dependent manner within the concentration range of 10-100 μg / mL. Moreover, at medium and high doses, the inhibition rates showed the expected strong inhibitory effect compared with the control group, and the differences were statistically significant (P<0.01).

[0071] Experiment Example 2: Animal Experiment

[0072] 1. Materials and Methods

[0073] 1.1 Laboratory Animals and Grouping

[0074] Eighty SPF-grade Wistar rats were selected, half male and half female, with a body weight of 160-200g.

[0075] Modeling and grouping strategy: 16 mice were randomly selected as the normal control group. The remaining 64 mice were treated with carbon tetrachloride (…). A liver fibrosis model was constructed using a composite factor method. At the end of the fourth week of modeling, eight model rats were randomly sacrificed for pre-assessment. After confirming fibrosis formation, the remaining 56 successfully modeled rats were randomly divided into two groups (n=28 in each group, with 16 rats in each group ultimately included in the statistical analysis). Model group: administered an equal volume of physiological saline by gavage. Traditional Chinese medicine lyophilized powder group (Example 1): administered a suspension of the traditional Chinese medicine lyophilized powder prepared in Example 1 by gavage.

[0076] 1.2 Dosing regimen

[0077] Model maintenance: Except for the normal control group, the other groups continued to receive intraperitoneal injections of 40% weekly during the drug administration period. Oil (0.3 mL / kg) is used to maintain the fibrotic state.

[0078] Intervention measures: The normal control group and the model group were administered physiological saline by gavage at a dose of 0.1 mL / 100g body weight. In Example 1, the lyophilized traditional Chinese medicine powder group was administered a drug-containing suspension by gavage at a dose of 0.1 mL / 100g body weight, with each milliliter of suspension containing 0.2g of lyophilized powder.

[0079] Treatment course: Once daily for 12 consecutive weeks.

[0080] 1.3 Detection Indicators and Methods

[0081] Hepatic microcirculation contrast ultrasound: After anesthetizing rats, ultrasound microbubble contrast agent was injected via the femoral vein. The time to onset of hepatic artery (AT), time to onset of hepatic vein (VT), and hepatic arteriovenous transit time (TT = VT - AT) were recorded. A shortened TT value indicated increased intrahepatic vascular shunts and exacerbated microcirculatory disturbances.

[0082] Serum vasoactive substance assay: Blood was collected from the abdominal aorta and centrifuged to obtain serum. The levels of endothelin-1 (ET-1) and nitric oxide (NO) were measured using ELISA.

[0083] Liver histopathological observation: The right lobe of the liver was fixed, embedded, sectioned, and stained with hematoxylin and eosin (HE). Following the modified Scheuer scoring system and literature methods, the degree of liver fibrosis was classified into grades 0-6, and the average pathological grade for each group was calculated.

[0084] 1.4 Statistical Analysis

[0085] Data with The analysis was performed using SPSS 22.0 software, and the LSD-t test was used for comparisons between groups. The difference is considered statistically significant.

[0086] 2. Results

[0087] 2.1 Pathological morphological changes in liver tissue of rats in each group

[0088] See results Figure 2 As shown in Table 1, in the normal control group: the liver lobule structure was clear and intact, the hepatocyte cords were neatly arranged, and they were radially distributed with the central vein as the center. No fibrous tissue hyperplasia or inflammatory cell infiltration was observed (pathological grade 0 for all groups). In the model group: the liver lobule structure was severely damaged, with a large number of coarse collagen fiber septa forming, dividing the liver tissue into pseudolobules of varying sizes; hepatocytes showed extensive fatty degeneration and ballooning degeneration, and there was significant inflammatory cell infiltration in the portal areas and septa. The pathological grade was mainly concentrated in grades 4-6, with the average grade significantly higher than that of the normal group. Example 1: The freeze-dried herbal powder group showed significantly reduced liver tissue structural damage compared to the model group. Collagen fiber deposition decreased, the number of pseudolobules significantly decreased, and the structure of some liver lobules tended to recover; hepatocyte degeneration and necrosis decreased, and inflammatory infiltration lessened. Pathological grading was mainly distributed in grades 2-3, with no cases of severe fibrosis in grades 5-6 observed.

[0089] Table 1. Comparison of pathological grading of liver fibrosis in rats of different groups (n=16)

[0090]

[0091] Note: Compared with the normal control group, Compared with the model group, .

[0092] 2.2 Effects of freeze-dried traditional Chinese medicine powder on serum ET-1 and NO levels in Example 1

[0093] The results are shown in Table 2. Serum ET-1 and NO levels in the model group rats were significantly higher than those in the normal control group. This indicates a severe imbalance in microcirculatory regulation. After 12 weeks of intervention with the lyophilized herbal powder from Example 1, the serum ET-1 and NO levels in rats significantly decreased, with statistically significant differences compared to the model group. This indicates that the freeze-dried powder of this traditional Chinese medicine can effectively inhibit the overexpression of vasoconstrictor factors and improve vasomotor function.

[0094] Table 2. Comparison of serum ET-1 and NO levels in rats of different groups ( (n=16)

[0095]

[0096] Note: Compared with the normal control group, Compared with the model group, .

[0097] 2.3 Effects of freeze-dried traditional Chinese medicine powder on hepatic microcirculation hemodynamics in Example 1

[0098] The results are shown in Table 3. Ultrasound imaging revealed that the hepatic artery initiation time was slightly shortened and the hepatic vein initiation time was significantly advanced in the model group rats, resulting in a significantly shorter hepatic arteriovenous transit time (TT) compared to the normal control group. This reflects the presence of numerous arteriovenous shunts within the liver and abnormal microcirculation perfusion. In Example 1, the time of hepatic venous initiation in rats in the lyophilized herbal powder group was significantly delayed compared to the model group, and the TT value was significantly prolonged. The levels were close to normal. This indicates that the freeze-dried powder of this traditional Chinese medicine can effectively reduce abnormal intrahepatic shunting and prolong the residence time of blood in the hepatic sinusoids, thereby improving hepatic microcirculation perfusion.

[0099] Table 3. Comparison of time parameters of liver ultrasound contrast imaging in rats of different groups ( (n=16)

[0100]

[0101] Note: Compared with the normal control group, Compared with the model group, .

[0102] 3. Discussion

[0103] This experiment was conducted through... A rat model of induced liver fibrosis was established to evaluate the intervention effect of lyophilized traditional Chinese medicine (TCM) powder on hepatic microcirculatory disturbances. The results showed that the rats in the liver fibrosis model exhibited significant microcirculatory dysfunction, manifested by abnormally elevated serum ET-1 and NO levels, and a significantly shortened hepatic arteriovenous transit time as observed by contrast-enhanced ultrasound. This microcirculatory disturbance, leading to hepatic sinusoidal capillaryization and blood flow shunting, is a key factor exacerbating hepatocyte ischemia and hypoxia and promoting fibrosis progression. Experimental data showed that the TCM lyophilized powder group exhibited significant therapeutic effects in multiple dimensions:

[0104] 1. Improvement of pathological structure: The average grade of liver fibrosis in this group of rats decreased from 4.81 in the model group to 2.69, and the formation of pseudolobules decreased, suggesting that the freeze-dried powder of this traditional Chinese medicine has a clear anti-fibrotic effect and can reverse pathological changes.

[0105] 2. Regulation of vasoactive substances: This freeze-dried Chinese medicine powder significantly reduced serum ET-1 (a potent vasoconstrictor) and excess NO levels. The reduction in ET-1 helps relieve the contraction of hepatic stellate cells and expand the diameter of hepatic sinusoids; the return of NO levels to normal reduces the toxic damage of free radicals to hepatocytes and restores normal vasomotor regulation mechanisms.

[0106] 3. Optimization of microcirculatory blood flow: The most crucial finding is that the lyophilized herbal powder in Example 1 significantly prolonged the hepatic arteriovenous transit time (from 2.10s in the model group to 4.64s). This improvement directly demonstrates that the drug can reduce pathological arteriovenous shunts in the liver, increase effective hepatic blood flow, and improve blood perfusion of liver tissue.

[0107] In conclusion, even after removing the comparison of individual formulations, the lyophilized powder of the traditional Chinese medicine in Example 1 still exhibits strong comprehensive therapeutic effects when used alone. Its anti-liver fibrosis mechanism is likely closely related to its multi-target regulation of vasoactive factors, correction of hepatic microcirculatory hemodynamic disturbances, and thus blocking the vicious cycle of "ischemia-hypoxia-fibrosis." These experimental results provide a solid experimental basis for the clinical development and application of the traditional Chinese medicine composition of this invention.

[0108] Experiment Example 3: Comparative Experiment

[0109] 1. Experimental Design

[0110] To demonstrate the synergistic effect of each herb in the prescription of this invention, comparative groups were set up to identify the missing herbs: the group lacking Lysimachia christinae (i.e., Comparative Example 1), the group lacking Trionyx sinensis (i.e., Comparative Example 2), the group lacking Litchi chinensis seed (i.e., Comparative Example 3), and the group lacking Panax notoginseng and Curcuma zedoaria (i.e., Comparative Example 4). Extracts were prepared using the same method, and the inhibition rate of each group was compared with that of HSC-T6 cell proliferation inhibition assay (50 μg / mL) as the indicator, referring to Experiment 1.

[0111] 2. Experimental Results

[0112] See results Figure 3 The inhibition rate of the group without Lysimachia christinae (Comparative Example 1) was 50.7%, while the inhibition rate of the group lacking Lysimachia christinae (Comparative Example 1) decreased to 18.3%, the group lacking Trionyx sinensis (Comparative Example 2) was 25.6%, the group lacking Litchi chinensis seed (Comparative Example 3) was 28.1%, and the group lacking Panax notoginseng and Curcuma zedoaria (Comparative Example 4) was 31.5%. The results indicate that the inhibition rate of each herb in combination was significantly higher than that of the groups lacking any of the herbs, suggesting a synergistic effect among the components of the composition of this invention.

Claims

1. A traditional Chinese medicine composition for treating liver fibrosis, characterized in that, The raw materials for its preparation include the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, and 8-12 parts of Poria cocos.

2. The traditional Chinese medicine composition according to claim 1, characterized in that, The raw materials include the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, 8-12 parts of Poria cocos, and 10-15 parts of Cyperus rotundus.

3. The traditional Chinese medicine composition according to claim 1, characterized in that, The raw materials include the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, 8-12 parts of Poria cocos, 10-15 parts of Cyperus rotundus, and 5-10 parts of Atractylodes macrocephala.

4. The traditional Chinese medicine composition according to claim 1, characterized in that, The raw materials include the following components in parts by weight: 20-30 parts of Lysimachia christinae, 8-15 parts of turtle shell, 10-16 parts of litchi seed, 5-15 parts of Panax notoginseng, 5-10 parts of Curcuma zedoaria, 8-12 parts of Poria cocos, 10-15 parts of Cyperus rotundus, 5-10 parts of Atractylodes macrocephala, and 2-6 parts of Glycyrrhiza uralensis.

5. A method for preparing a traditional Chinese medicine composition according to any one of claims 1-4, characterized in that, The raw materials are mixed and pulverized to obtain the traditional Chinese medicine composition.

6. The preparation method according to claim 5, characterized in that, The raw materials are mixed and pulverized to 10-200 mesh.

7. A method for preparing a traditional Chinese medicine composition according to any one of claims 1-4, characterized in that, The raw material is extracted with water to obtain an aqueous extract, which is then used as a traditional Chinese medicine preparation. Alternatively, the aqueous extract can be further processed with pharmaceutically acceptable excipients to form other traditional Chinese medicine dosage forms, and the resulting traditional Chinese medicine preparation is the traditional Chinese medicine composition.

8. The preparation method according to claim 7, characterized in that, The dosage forms of the traditional Chinese medicine preparations are decoctions, oral liquids, drops, ointments, pills, powders, tablets, or capsules.

9. The preparation method according to claim 7, characterized in that, The water extraction method includes: boiling the raw material with water several times, and finally combining the decoctions obtained from each boiling.

10. The preparation method according to claim 9, characterized in that: The decoction is performed 1-3 times, with the amount of water added each time being 5-15 times the total weight of the raw materials, and the decoction time being 30-120 minutes each time.

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