Secondary ginsenoside composition and application thereof in prevention and treatment of pulmonary fibrosis and acute lung injury

Abstract

The invention belongs to the technical field of medicines, and particularly relates to a secondary ginsenoside composition and application thereof in prevention and treatment of pulmonary fibrosis and acute lung injury. The secondary ginsenoside composition is prepared from any two or more components selected from the following components: (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3 and Rh4; (20S)-Rg3, (20R)-Rg3, Rk1, Rg5, (20S)-Rh2, (20R)-Rh2, Rk2 and Rh3 are selected from the group consisting of (20S)-Rg3, (20R)-Rg3, Rk1 and Rg5. The medicine effect of the secondary ginsenoside composition provided by the invention on the aspects of preventing and treating pulmonary fibrosis and acute lung injury is obviously superior to that of a primary ginsenoside composition and secondary saponin Rg5 and Rg3 monomer components. Meanwhile, the secondary ginsenoside composition is more easily absorbed by intestinal tracts than a primary ginsenoside composition, and is slightly influenced by individual differences of intestinal flora. Compared with a secondary saponin monomer component, the compound is easier to prepare, lower in production cost and safer and more efficient compared with existing commercially available drugs.

Description

[0001] Cross-reference information

[0002] This application claims priority to Chinese patent application No. 2024109099904 filed on July 8, 2024, the entire contents of which are incorporated herein by reference. Technical Field

[0003] The present invention belongs to the field of medical technology, and specifically relates to a secondary ginsenoside composition and an application thereof in preventing and treating pulmonary fibrosis and acute lung injury. Background Art

[0004] Pulmonary fibrosis (PF) is a serious respiratory disease with a high mortality rate. Pathogenic microbial infection, inducing drugs, and aging are all high-risk factors for PF. Among them, idiopathic pulmonary fibrosis (IPF) has no clear cause, with a median survival of only 2 to 3 years and a five-year survival rate of about 20%. It is also called a tumor-like disease, and there is no specific treatment. With the invasion of pathogens such as environmental pollution, respiratory viruses (such as SARS-CoV-2) and bacteria, post-inflammatory pulmonary fibrosis has also become a problem that cannot be ignored.

[0005] At present, two anti-pulmonary fibrosis drugs have been launched on the market - pirfenidone and nintedanib, both of which are suitable for PF patients with mild to moderate pulmonary dysfunction. Among them, pirfenidone has side effects such as photosensitivity, fatigue, rash, stomach discomfort and anorexia. Nintedanib is a multi-target tyrosine kinase inhibitor that can inhibit platelet-derived growth factor receptor, vascular endothelial growth factor receptor and fibroblast growth factor receptor, but it can only alleviate the disease process to a certain extent, and there are adverse reactions of liver damage and kidney damage in clinical applications.

[0006] In order to solve the adverse reactions of existing drugs, searching for functional ingredients or drugs for preventing / treating pulmonary fibrosis and lung injury from traditional Chinese medicines that are both medicinal and edible has always been a hot topic in the research and development of drugs against pulmonary fibrosis and lung injury. Currently, some patent documents have been published on the application of ginsenosides in the treatment of pulmonary fibrosis.

[0007] It is proposed in CN116869977 A that total ginsenosides can be used to prepare aerosol inhalation preparations for the treatment of pulmonary fibrosis or acute lung injury, and it is found that its effective therapeutic effect is achieved by activating the smad2 signaling pathway.

[0008] CN112587539 B provides the use of ginsenoside Rg5 alone or in combination with other drugs in the preparation of drugs for the prevention and treatment of pulmonary fibrosis. In this technical solution, the animal model of pulmonary fibrosis induced by bleomycin and the in vitro model of pulmonary fibrosis induced by TGF-β are used respectively, and it is proved that ginsenoside Rg5 has pharmacological activity against pulmonary fibrosis, and can inhibit the occurrence of inflammation and collagen fiber formation, and promote the degradation of collagen fibers and the transformation of lung fibroblasts into myofibroblasts, which hinders the development of pulmonary fibrosis through multiple mechanisms and exerts a pharmacological effect against pulmonary fibrosis.

[0009] CN101757009 B provides the use of notoginsenoside Rg1 (i.e. ginsenoside Rg1, which belongs to the PPT type native ginsenoside) in the preparation of drugs for treating pulmonary interstitial fibrosis. The technical solution has been verified by pharmacological experiments, and notoginsenoside Rg1 can inhibit the activity of human embryonic lung fibroblasts in a dose-dependent manner in vitro; it can reduce the lung coefficient and the content of hydroxyproline in lung tissue of mice with pulmonary interstitial fibrosis in vivo; it can also reduce the degree of alveolitis and pulmonary interstitial fibrosis in mice with pulmonary interstitial fibrosis.

[0010] CN116459267 A discloses a new application of ginsenoside Rd (PPD-type native ginsenoside) in the preparation of drugs for relieving or / and treating inflammation and health foods / foods with medicinal and edible properties / functional foods / dietary supplements. Experiments have shown that ginsenoside Rd has significant and rapid effects in treating inflammation, and is better than dexamethasone; it is suitable for treating acute inflammation such as influenza virus pneumonia, new coronavirus infection, acute lung injury inflammation, etc.; it is also suitable for treating chronic inflammation such as chronic obstructive pneumonia, chronic asthmatic airway inflammation, etc., and is a new source of drugs for treating acute and chronic inflammation and its complications.

[0011] In addition, CN107635564 B also mentions the new use of ginsenoside M1 for the treatment or prevention of silicosis. Among them, ginsenoside M1 cannot be extracted from ginseng medicinal materials (ginseng, American ginseng, Panax notoginseng, etc.). It is actually one of the metabolites of PPD-type native ginsenosides produced by human intestinal bacteria metabolism. Its chemical structure has a glucose at the C-20 position of protopanaxadiol PPD, and its full name is 20-β-D-pyranose glucopyranosyl protopanaxadiol (20-O-β-D-glucopyranosyl-20(S)-protopanaxadiol).

[0012] In addition to the above-mentioned patent documents, some non-patent documents also mention the use of ginsenosides in the prevention and treatment of lung diseases.

[0013] For example, Professor Deng Jiang of Zunyi Medical College mentioned in his research paper "Inhibitory Effects of total ginsenoside on bleomycin-induced pulmonary fibrosis in mice" published in Biomedicine & Pharmacotherapy, 2019, 114: 108851 that total ginsenosides (TG), as the active ingredient of ginseng, can inhibit pulmonary fibrosis. The protective effect of TG on bleomycin-induced pulmonary fibrosis is related to the regulation of TGF-β1 / Smad signaling pathway and MMP system.

[0014] The research paper "Ginsenoside Rg3 inhibits pulmonary fibrosis by preventing HIF-1αnuclear localization" published in BMC Pulmonary Medicine, 2021, 21:70 mentioned that bleomycin induces activation of the HIF-1α / TGFβ1 signaling pathway and further enhances the migration and proliferation of fibroblasts through epithelial-mesenchymal transition (EMT). Ginsenoside Rg3 can bind to HIF-1α to inhibit its nuclear localization, thereby slowing the progression of pulmonary fibrosis. This finding suggests that early targeted treatment of hypoxia may have potential value in the treatment of pulmonary fibrosis.

[0015] The research paper "Ginsenoside Rg3 attenuates pulmonary fibrosis by inhibiting endothelial tomesenchymal transition" published in Animal Cells and Systems, 2023, 27(1): 159-170 mentioned that endothelial-mesenchymal transition (EndMT), as a source of fibroblasts, contributes to the progression of PF, and Rg3 inhibits EndMT by regulating the Smad2 / 3 signaling pathway, thereby exerting an anti-PF effect. Therefore, Rg3 can be identified as a potential therapeutic agent for the treatment of PF using an EndMT inhibition strategy. In addition, Rg3 can also be considered as a candidate drug for the treatment of various EndMT-related vascular diseases.

[0016] Professor Dongxian Jin of Kyung Hee University in South Korea published a research paper titled "Ginsenoside Rg5 ameliorates lung inflammation in mice by inhibiting the binding of LPS to toll-like receptor-4 on macrophages" in International Immunopharmacology, 2012, 12:110-116. He mentioned that the main component of ginsenoside Rg5 was isolated from steamed ginseng, and the anti-inflammatory effect of Rg5 on lipopolysaccharide (LPS)-stimulated macrophages and LPS-induced lung inflammation was investigated by intraperitoneal injection (ip). It was found that Rg5 may improve lung inflammation by inhibiting the binding of LPS to Toll-like receptor (TLR)-4 on macrophages.

[0017] In summary, the prior art has reported that the ginsenoside composition mainly composed of primary ginsenosides, the primary ginsenoside monomer components Rg1 and Rd, and the secondary ginsenoside monomers Rg5 and Rg3 have different degrees of efficacy in preventing bleomycin (BLM)-induced pulmonary fibrosis (PF) in mice or rats. The evaluation method for preventing PF efficacy is to administer the drug immediately after BLM modeling. This stage is considered to be the inflammatory stage of the lungs, and it is impossible to know whether the ginsenoside composition or monomer component has an anti-pulmonary inflammation or anti-pulmonary fibrosis effect. The secondary ginsenoside Rg5 can improve lipopolysaccharide (LPS)-induced lung inflammation by intraperitoneal injection (ip).

[0018] However, in vivo metabolism studies in recent years have shown that native ginsenosides have a large molecular weight and high molecular polarity, and their absorption rate in the human body is extremely low through oral administration. The active ingredients that are truly absorbed by the human body are secondary ginsenosides or a small amount of metabolites such as aglycones produced by metabolism in the stomach (acidic environment) and intestinal bacteria. These metabolites include secondary ginsenoside types that only hydrolyze C-20 sugars, secondary metabolites produced by hydrolyzing C-6 (PPT type) sugars and C-3 (PPD type) sugars while retaining C-20 sugars (such as PPT type ginsenoside F1 and PPD type ginsenoside M1) and a variety of other types of metabolites. Due to individual differences in the intestinal flora of different people, oral administration of native ginsenosides will lead to different metabolic characteristics, which in turn cause differences in drug efficacy.

[0019] However, the preparation of secondary ginsenoside monomer components is difficult, the production cost is high, and it is difficult to promote them in practical applications.

[0020] At the same time, there are differences in the induction models used in existing patents or studies, such as rats or mice, animal strains, the source and dosage of the inducer bleomycin (BLM), whether BLM is modeled by tracheal instillation or surgical tracheotomy (which may cause animal infection), etc.; there are differences in the PF prevention mode of administering the drug immediately after modeling or the PF treatment mode of administering the drug 7 days after BLM modeling (pulmonary fibrosis has been induced by lung inflammation) (the PF disease peaks 10 days after BLM modeling, and the treatment mode of starting to administer the drug at the peak of the disease can better reflect the drug's anti-fibrosis treatment effect on PF disease); marketed drugs (pirfenidone or nintedanib) were not used as positive controls, and it is impossible to obtain information on the comparison of their effects with PF treatment drugs; there are also differences in the administration of ginsenosides, such as oral gavage, intraperitoneal injection or pulmonary atomization. Therefore, based on the disclosure of the above-mentioned prior art, it is difficult to see the substantial difference in the efficacy of the primary ginsenoside composition, the secondary ginsenoside composition, and the secondary ginsenoside Rg3 and Rg5 monomer components in preventing or treating pulmonary fibrosis, which also brings certain technical obstacles to further research. Summary of the invention

[0021] The present invention first provides a secondary ginsenoside composition, which contains any two or more components selected from the components shown in the following 1) to 2):

[0022] 1) PPT type secondary ginsenosides: (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3, Rh4;

[0023] 2) PPD type secondary ginsenosides: (20S)-Rg3, (20R)-Rg3, Rk1, Rg5, (20S)-Rh2, (20R)-Rh2, Rk2, Rh3.

[0024] The present invention further provides a method for preparing a secondary ginsenoside composition, which comprises: using at least one of a primary ginsenoside composition, a PPT-type primary ginsenoside composition, and a PPD-type primary ginsenoside composition as a raw material, hydrolyzing only the C-20 sugar in the raw material and the C-20 alcohol hydroxyl group of the hydrolysis product, and further dehydrating the resulting product to obtain a secondary ginsenoside composition.

[0025] The present invention also provides a secondary ginsenoside composition, which is prepared by the preparation method.

[0026] The present invention further provides the use of the secondary ginsenoside composition in the preparation of medicines or foods, wherein the medicines or foods are used to prevent, treat or alleviate at least one disease selected from pulmonary fibrosis and acute lung injury.

[0027] The present invention also provides a medicine comprising the secondary ginsenoside composition.

[0028] The present invention also provides a food, which comprises the secondary ginsenoside composition.

[0029] The secondary ginsenoside composition provided by the present invention is significantly superior to the primary ginsenoside composition, and the secondary saponin Rg5 and Rg3 monomer components in terms of the efficacy in preventing and treating pulmonary fibrosis and acute lung injury. At the same time, compared with the primary ginsenoside composition, the secondary ginsenoside composition has a smaller molecular weight and smaller polarity, and is more easily absorbed by the intestine through oral administration, and is less affected by individual differences in intestinal flora. Compared with the secondary saponin Rg5 and Rg3 monomer components, the secondary ginsenoside composition of the present invention is not only easier to prepare, but also has a lower production cost. Compared with the clinical effective dose (200 mg / kg) of the positive drug pirfenidone, the dosage (20 mg / kg, 50 mg / kg and 100 mg / kg) of the secondary ginsenoside composition of the present invention is small, and the efficacy and biosafety of treating pulmonary fibrosis are significantly better than pirfenidone. Compared with the clinical effective dose (50 mg / kg) of the positive drug nintedanib, the secondary ginsenoside composition of the present invention significantly improves the survival rate of PF mice, significantly reduces the lung index of PF mice, and significantly improves various indicators of lung function of model animals under the same dosage (50 mg / kg) or lower dosage (20 mg / kg) conditions, and has a better efficacy in treating pulmonary fibrosis. In addition, the secondary ginsenoside composition of the present invention can also effectively protect the lungs, promote the barrier function of alveolar capillary endothelial cells, and prevent post-inflammatory fibrosis of the lungs and related lung diseases caused by drugs such as bleomycin or bacteria and other inducing factors. The safety evaluation results of the total secondary ginsenoside composition of the present invention for 14 consecutive days of oral administration under the condition of gavage dose (100 mg / kg) show that it has good safety in animals. Compared with existing marketed drugs for the treatment of pulmonary fibrosis, the secondary ginsenoside composition is safer and more efficient, and can not only be used as a drug for the treatment of pulmonary fibrosis and related lung injury diseases, but also as the main functional ingredient in health foods for the prevention of pulmonary fibrosis and lung injury. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] In order to more clearly illustrate the specific implementation methods of the present invention or the technical solutions in the prior art, the drawings required for use in the specific implementation methods or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some implementation methods of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0031] Figure 1This is the UHPLC-MS test result of the total native saponin composition in Example 1 of the present invention.

[0032] Figure 2 This is the UHPLC-MS test result of the total secondary ginsenoside composition in Example 1 of the present invention.

[0033] Figure 3 This is the UHPLC-MS test result of the PPT type secondary ginsenoside composition in Example 1 of the present invention.

[0034] Figure 4 This is the UHPLC-MS test result of the PPD type secondary ginsenoside composition in Example 1 of the present invention.

[0035] Figure 5 Schematic diagram of the experimental process of using a secondary ginsenoside composition to prevent pulmonary fibrosis in Example 2 of the present invention.

[0036] Figure 6 These are Micro-CT scan images of the lung tissues of mice in each experimental group and photographs of the lungs of mice in Example 2 of the present invention.

[0037] Figure 7 It is the H&E staining image of the lung tissue of mice in each experimental group in Example 2 of the present invention; in this figure, A is the sham operation group, B is the model group, C is the nintedanib group, D is the Rg3 group, E is the Rg5 group, F is the total saponin (control) group, G is the total primary glycoside group, H is the total secondary glycoside group, I is the PPT type secondary glycoside group, and J is the PPD type secondary glycoside; in the figure, the scale bar is 50 microns.

[0038] Figure 8 The statistical results of the total leukocytes, lymphocytes and macrophages in the bronchoalveolar lavage fluid of mice in each experimental group in Example 2 of the present invention; in this figure, ****p<0.0001vs sham operation group; # p<0.05, ## p<0.01, ### p<0.001vs model group; & p<0.05, && p<0.01vs total protoglycoside group; ％ p<0.05, ％％ p<0.01, ％％％ p<0.001vs total secondary glycosides group; $ p<0.05, $$ p<0.01, $$$ p<0.001vs PPD type secondary glycoside group.

[0039] Fig. 9The Masson staining images of the lung tissues of mice in each experimental group in Example 2 of the present invention and the statistical results of the collagen fiber positive area ratio; in the figure, A to J are Masson staining images of the lung tissues of mice, and the scale is 50 microns; A is the sham operation group, B is the model group, C is nintedanib, D is the Rg3 group, E is the Rg5 group, F is the total saponin (control) group, G is the total primary glycoside group, H is the total secondary glycoside group, I is the PPT type secondary glycoside group, and J is the PPD type secondary glycoside; Figure K is the statistical results of the percentage of collagen positive area, and in Figure K, ****p<0.0001vs the sham operation group; # p<0.05, ### p<0.001, #### p<0.0001vs model group; & p<0.05, && p<0.01vs total protoglycosides; ％％ p<0.01, ％％％％ p<0.0001vs total secondary glycosides; $$ p<0.01vs PPD-type secondary glycosides.

[0040] Fig.10 The results of the detection of hydroxyproline content in the lung tissue of mice in each experimental group in Example 2 of the present invention; in this figure, ****p<0.0001vs sham operation group; # p<0.05, ## p<0.01, ### p<0.001, #### p<0.0001vs model group; &&&& p<0.0001vs total protoglycosides; ％％％ p<0.001, ％％％％ p<0.0001vs total secondary glycosides; $$$$ p<0.0001vs PPD-type secondary glycosides.

[0041] Fig.11 This is a schematic diagram of the experimental process of using a secondary ginsenoside composition to treat pulmonary fibrosis in Example 3 of the present invention.

[0042] Fig.12 The statistical results of the survival period of mice in each experimental group in Example 3 of the present invention; in this figure, *p<0.05 model group vs sham operation group; # p<0.05 each secondary glycoside group vs model group.

[0043] Fig.13 These are Micro-CT scan images of the lung tissues of mice in each experimental group and photographs of the lungs of mice in Example 3 of the present invention.

[0044] Fig.14This is the H&E staining image of the lung tissue of mice in each experimental group in Example 3 of the present invention; in the figure, the scale bar is 50 microns.

[0045] Fig.15 The statistical results of the number of total leukocytes, lymphocytes and macrophages in the bronchoalveolar lavage fluid of mice in each experimental group in Example 3 of the present invention; in this figure, ***p<0.001, ****p<0.0001vs sham operation group; # p<0.05, ## p<0.01, ### p<0.001, #### p<0.0001vs model group.

[0046] Fig.16 The Masson staining diagram of the lung tissue of each experimental group of mice in Example 3 of the present invention and the statistical results of the collagen fiber positive area ratio; in the Masson staining diagram, the scale bar is 50 microns. Among them, in the statistical results of the collagen positive area ratio, ****p<0.0001vs sham operation group; #### p<0.0001vs model group; & p<0.05vs pirfenidone.

[0047] Fig.17 The results of the detection of hydroxyproline content in the lung tissue of mice in each experimental group in Example 3 of the present invention; in this figure, ****p<0.0001vs sham operation group; # p<0.05, #### p<0.0001vs model group; & p<0.05, && p<0.01, &&& p<0.001, &&&& p<0.0001vs pirfenidone.

[0048] Fig.18 The results of protein immunoblotting of fibronectin, collagen COL1A1, and smooth muscle actin α-SMA expressed in the lung tissue of mice in the PPT-type secondary glycoside group in Example 3 of the present invention are shown in the figure. * p<0.05, *** p<0.001, **** p<0.0001vs sham operation group; # p<0.05, ## p<0.01, ### p<0.001, #### p<0.0001vs model group; &&& p<0.001vs pirfenidone; $$p<0.01vs PPT type secondary glycoside-L.

[0049] Fig.19 The results of protein immunoblotting of fibronectin, collagen COL1A1, and smooth muscle actin α-SMA expressed in the lung tissue of mice in the PPD-type secondary glycoside group in Example 3 of the present invention are shown in the figure. * p<0.05, **** p<0.0001vs sham operation group; ### p<0.001, #### p<0.0001vs model group.

[0050] Fig. 20 The immunohistochemical detection results (scale bar: 100 micrometers) and the statistical results of the protein positive area ratio of fibronectin, collagen I, and smooth muscle actin α-SMA expressed in the lung tissues of mice in each experimental group in Example 3 of the present invention; in the statistical results, ****p<0.0001vs sham operation group; #### p<0.0001vs model group; & p<0.05, && p<0.01, &&&& p<0.0001vs pirfenidone.

[0051] Fig.21 These are H&E staining images of the heart, liver, spleen and kidney of mice in each experimental group in Example 3 of the present invention; in this figure, the scale bar is 100 microns.

[0052] Fig. 22 This is a schematic diagram of the experimental process of using the total secondary ginsenoside composition to treat bleomycin (2.5 mg / kg)-induced pulmonary fibrosis in Example 4 of the present invention.

[0053] Fig.23 The therapeutic effect of total secondary ginsenosides in Example 4 of the present invention on bleomycin-induced pulmonary fibrosis in mice; in this figure, A is the statistical result of the survival period of mice, **p<0.01 model group vs sham operation group; # p<0.05 total secondary glycosides group vs model group; B is the effect on the lung index of model animals, **p<0.01 vs sham operation group; # p<0.05vs model group.

[0054] Fig.24 These are Micro-CT scan images of the lung tissues of mice in each experimental group and photographs of the lungs of mice in Example 4 of the present invention.

[0055] Fig.25Schematic diagram of the experimental process of using the total secondary ginsenoside composition to prevent and treat bleomycin (1.8 mg / kg)-induced pulmonary fibrosis in Example 5 of the present invention.

[0056] Fig.26 The protective / preventive and therapeutic effects of total secondary ginsenosides on bleomycin-induced pulmonary fibrosis in mice in Example 5 of the present invention; in this figure, A is the statistical result of the survival period of the prevention model mice, **p<0.01 model group vs sham operation group; # p<0.05 total secondary glycosides group vs model group; B is the effect of total secondary glycosides in the preventive mode on the lung index of model animals, ****p<0.0001 vs sham operation group; ### p<0.001 vs model group; C is the statistical result of survival of mice in treatment model, **p<0.01 model group vs sham operation group; # p<0.05, ## p<0.01 vs model group; D is the effect of total secondary glycosides in treatment mode on the lung index of model animals, ***p<0.001 vs sham operation group; # p<0.05, ## p<0.01vs model group.

[0057] Fig. 27 The results of the test of total secondary glycosides on the lung function of model animals in Example 5 of the present invention; in the figure, A is the prevention mode, B is the treatment mode, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 vs. sham operation group; # p<0.05, ## p<0.01vs model group; $ p<0.05, $$ p<0.01, $$$ p<0.001vs nintedanib group.

[0058] Fig.28 1 is the effect of total secondary glycosides on the curve of exhaled gas flow rate (Flow) and exhaled gas volume (Volume) of model animals in Example 5 of the present invention; in this figure, A is the prevention mode and B is the treatment mode.

[0059] Fig.29 These are Micro-CT scan images of the total secondary glycosides on the lung tissue of model animals and photographs of mouse lungs in Example 5 of the present invention.

[0060] Fig.30 The effect of total secondary glycosides on pathological changes in lung tissue of model animals in Example 5 of the present invention (H&E staining); in the figure, the scale bar is 100 microns. In the statistical analysis of inflammation scores, ****p<0.0001vs sham operation group; #p<0.05, ### p<0.001, #### p<0.0001vs model group.

[0061] Fig.31 This is the effect of total secondary glycosides in Example 5 of the present invention on the number of inflammatory cells in bronchoalveolar lavage fluid of model animals; in this figure, A is the prevention mode, B is the treatment mode, **p<0.01, ****p<0.0001vs sham operation group; # p<0.05, ## p<0.01vs model group.

[0062] Fig.32 The effect of total secondary glycosides on pathological changes in lung tissue of model animals in Example 5 of the present invention (Masson staining); in the figure, the scale bar is 100 microns. In the statistical analysis of fibrosis scores, ****p<0.0001vs sham operation group; # p<0.05, ## p<0.01, #### p<0.0001vs model group.

[0063] Fig.33 The immunohistochemical detection results (scale bar: 100 micrometers) and the statistical results of the protein positive area ratio of the total secondary glycosides in Example 5 of the present invention on fibronectin, collagen I, and smooth muscle actin α-SMA expressed in the lung tissue of the model animal; in the statistical results, ***p<0.001, ****p<0.0001vs sham operation group; # p<0.05, ## p<0.01, ### p<0.001, #### p<0.0001vs model group.

[0064] Fig.34 The results of the determination of total leukocytes and neutrophils in the bronchoalveolar lavage fluid of mice in each experimental group in Example 6 of the present invention; in the figure, A is the flow cytometry analysis of BALF neutrophils, B is the total leukocyte count in BALF, and C is the neutrophil count in BALF; **p<0.01, ***p<0.001vs control group; # p<0.05, ## p<0.01vs model group; & p<0.05vs total protoglycosides; $ p<0.05vs Rg5.

[0065] Fig.35The results of the determination of the contents of MPO and proinflammatory factors TNF-α, IL-6, and IL-1β in the bronchoalveolar lavage fluid of mice in each experimental group in Example 6 of the present invention; in this figure, ***p<0.001, ****p<0.0001vs the control group; # p<0.05vs model group; & p<0.05vs total protoglycosides; $ p<0.05vs Rg5.

[0066] Fig.36 The results of the determination of protein concentration in the bronchoalveolar lavage fluid of mice in each experimental group in Example 6 of the present invention; in this figure, ****p<0.0001vs control group; # p<0.05vs model group; $ p<0.05vs Rg5.

[0067] Fig.37 The results of Western blotting of tight junction protein ZO-1, cyclooxygenase protein COX-2, phosphorylated nuclear transcription factor NF-κB p-p65 and total p65 protein and tumor necrosis factor TNF-α expressed in the lung tissues of mice in each experimental group in Example 6 of the present invention; in the figure, **p<0.01, ***p<0.001, ****p<0.0001vs control group; # p<0.05, ## p<0.01, ### p<0.001, #### p<0.0001vs model group; && p<0.01,; &&& p<0.001vs total protoglycosides; $$ p<0.01, $$$ p<0.001, $$$$ p<0.0001vs Rg5.

[0068] Fig.38 The safety assessment results of mice of Application Example 7 of the present invention include (A) changes in mouse body weight, (B) mouse organ indexes, (C) H&E staining of mouse heart, liver, spleen, lung, and kidney, and the levels of (D) aspartate aminotransferase AST, (E) alanine aminotransferase ALT, and (F) blood creatinine CRE in mouse serum. DETAILED DESCRIPTION

[0069] The specific embodiments of the present invention are described in detail below. It should be understood that the specific embodiments described herein are only used to illustrate and explain the present invention, and are not used to limit the present invention. Those skilled in the art may make various modifications and changes to the present invention without departing from the scope or spirit of the present invention. For example, a feature described or illustrated as part of one embodiment may be used in another embodiment to produce a further embodiment.

[0070] Explanation of terms

[0071] Unless otherwise indicated, the meaning of all terms (including technical and scientific terms) used to disclose the present invention is the same as that commonly understood by those of ordinary skill in the art to which the present invention belongs. By way of further guidance, the following definitions are used to better understand the teachings of the present invention. The terms used herein in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention.

[0072] The terms "and / or", "or / and", and "and / or" used in this article include any one of two or more related listed items, and also include any and all combinations of related listed items, and the arbitrary and all combinations include any two related listed items, any more related listed items, or a combination of all related listed items. It should be noted that when at least three items are connected by at least two conjunctions selected from "and / or", "or / and", and "and / or", it should be understood that in this application, the technical solution undoubtedly includes technical solutions that are all connected by "logical and", and undoubtedly includes technical solutions that are all connected by "logical or". For example, "A and / or B" includes three parallel solutions of A, B and A+B. For example, the technical solution of "A, and / or, B, and / or, C, and / or, D" includes any one of A, B, C, and D (that is, the technical solution that is all connected by "logical OR"), and also includes any and all combinations of A, B, C, and D, that is, the combination of any two or any three of A, B, C, and D, and also includes the combination of four of A, B, C, and D (that is, the technical solution that is all connected by "logical AND").

[0073] As used herein, the terms "comprising", "including" and "comprising" are synonymous and are inclusive or open-ended and do not exclude additional, unrecited members, elements or method steps.

[0074] The term "above" used in the present invention includes the number itself. For example, "two or more" includes two, three, four, etc.; "70% or more" includes 70%, 71%, 72%, etc.

[0075] Numerical ranges expressed as endpoints herein include all numbers and fractions subsumed within the range, as well as the recited endpoints.

[0076] The present invention relates to concentration values, and its meaning includes fluctuations within a certain range. For example, it can fluctuate within the corresponding accuracy range. For example, 2% can allow fluctuations within the range of ±0.1%. For values ​​that are large or do not require too fine control, its meaning is also allowed to include greater fluctuations. For example, 100mM can allow fluctuations within the range of ±1%, ±2%, ±5%, etc. Involving molecular weight, its meaning is allowed to include fluctuations of ±10%.

[0077] In the present invention, descriptions such as "plurality" and "multiple" refer to quantities greater than or equal to 2 unless otherwise specified.

[0078] In the present invention, the technical features described in an open manner include closed technical solutions composed of the listed features, and also include open technical solutions containing the listed features.

[0079] In the present invention, “preferred”, “better”, “more preferred” and “suitable” are only used to describe implementation methods or examples with better effects. It should be understood that they do not constitute limitations on the scope of protection of the present invention.

[0080] In the present invention, "optionally", "optional", "optionally", "optionally", "optional", "optional", "optional" means optional, that is, it means to be selected from any one of the two parallel schemes of "yes" or "no". If multiple "optional" or "optional" appear in a technical solution, unless otherwise specified and there is no contradiction or mutual restriction, each "optional" or "optional" is independent.

[0081] In the present invention, the term "primary ginsenoside" refers to a triterpene glycoside compound that exists in large quantities in plants of the genus Panax in the Araliaceae family. Primary ginsenosides have similar basic triterpene aglycone structures, all of which contain a dammarane-type tetracyclic triterpene mother nucleus composed of 30 carbon atoms. Primary ginsenosides can be divided into protopanaxatriol (PPT) primary ginsenosides and protopanaxadiol (PPD) primary ginsenosides, and both PPT-type primary ginsenosides and PPD-type primary ginsenosides contain the following 20 (S) configuration, i.e., sugar or sugar chain at C-20, and in addition, PPT-type primary ginsenosides also contain sugar or sugar chain at C-6, while PPD-type primary ginsenosides also contain sugar or sugar chain at C-3.

[0082]

[0083] In the present invention, the term "secondary ginsenoside" refers to the secondary metabolites of primary ginsenosides, which are less or non-existent in plants of the genus Panax in the Araliaceae family. Generally, secondary metabolites include secondary ginsenoside types that only hydrolyze sugars at the C-20 position, secondary metabolites produced by hydrolyzing sugars at the C-6 position (PPT type) and sugars at the C-3 position (PPD type) while retaining sugars at the C-20 position (such as PPT-type ginsenoside F1 and PPD-type ginsenoside M1), and various other types of metabolites. In the present invention, secondary ginsenosides mainly refer to secondary ginsenoside types that only hydrolyze sugars at the C-20 position, and secondary ginsenoside compositions refer to compositions with secondary ginsenosides of this type as the main component (mass content is 50% or more, more preferably 70% or more). PPT-type secondary ginsenosides refer to products of PPT-type primary ginsenosides that only hydrolyze sugars at the C-20 position. PPD-type secondary ginsenosides refer to the products of hydrolysis of only the C-20 sugar of PPD-type primary ginsenosides.

[0084] The structures of several specific PPT-type secondary ginsenosides mentioned in the present invention are as follows:

[0085]

[0086] The structures of several specific PPD-type secondary ginsenosides mentioned in the present invention are as follows:

[0087] In the present invention, the term "pulmonary fibrosis" refers to pulmonary fibrosis, abbreviated as PF, which is a terminal manifestation of interstitial lung disease. This disease is characterized by fibroblast proliferation and a large amount of extracellular matrix deposition, leading to the destruction of lung tissue structure.

[0088] In the present invention, the term "acute lung injury" refers to acute lung injury, abbreviated as ALI, which is caused by various direct and indirect injury factors that damage alveolar epithelial cells and capillary endothelial cells, resulting in diffuse pulmonary interstitial and alveolar edema, leading to acute hypoxic respiratory insufficiency. This injury usually occurs due to a systemic inflammatory response caused by severe diseases or injuries, such as infection, trauma, burns, drowning, etc. The pathophysiological characteristics are reduced lung volume, reduced lung compliance, and ventilation / perfusion imbalance. Clinically, it manifests as progressive hypoxemia and respiratory distress, and lung imaging shows non-uniform exudative lesions. Acute lung injury develops to a severe stage, namely Acute Respiratory Distress Syndrome (ARDS), which is characterized by extensive damage to the alveoli and microvessels, resulting in severe lung function impairment, and usually requires important supportive treatments such as mechanical ventilation.

[0089] In the present invention, the term "oral dosage form" refers to a drug dosage form taken orally. Drugs in this dosage form are usually designed to be absorbed by the gastrointestinal tract to achieve a therapeutic effect. Oral dosage forms include tablets, capsules, granules, liquids (including oral solutions, suspensions and syrups, etc.), oral dissolving tablets, oral suspensions, oral emulsions and other forms, and these drug dosage forms can be selected according to the properties of the drug and treatment needs. Oral dosage forms usually have the advantages of being easy to use and rapidly absorbed.

[0090] Secondary ginsenoside composition

[0091] The present invention relates to a secondary ginsenoside composition, which contains any two or more components selected from the components shown in the following 1) to 2):

[0092] 1) PPT type secondary ginsenosides: (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3, Rh4;

[0093] 2) PPD type secondary ginsenosides: (20S)-Rg3, (20R)-Rg3, Rk1, Rg5, (20S)-Rh2, (20R)-Rh2, Rk2, Rh3.

[0094] The present invention found that the above secondary ginsenoside composition is significantly superior to the primary ginsenoside composition in terms of the efficacy in preventing and treating BLM-induced pulmonary fibrosis and LPS-induced acute lung injury. Moreover, the secondary ginsenoside composition can not only prevent pulmonary fibrosis (the prior art adopts a dosing mode for preventing pulmonary fibrosis, that is, dosing starts the day after modeling), but also treat pulmonary fibrosis (modeling for 7 days, from lung inflammation to pulmonary fibrosis, dosing starts after pulmonary fibrosis, and the therapeutic efficacy is investigated; modeling for 10 days, dosing starts at the peak of pulmonary fibrosis, and the therapeutic efficacy is investigated). At the same time, compared with the secondary saponin monomers Rg5 and Rg3, the above secondary ginsenoside composition has a more excellent efficacy in preventing pulmonary fibrosis. In addition, the secondary ginsenoside composition is also superior to the secondary saponin monomer Rg5 in terms of the efficacy of protecting LPS-induced acute lung injury in mice. It can be seen that the above secondary ginsenoside composition has a synergistic effect.

[0095] In some embodiments, the secondary ginsenoside composition comprises (20S)-Rg3, (20R)-Rg3, Rg5, and at least one selected from (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3, Rh4, Rk1, (20S)-Rh2, (20R)-Rh2, Rk2, Rh3.

[0096] In some embodiments, the secondary ginsenoside composition is a PPT-type secondary ginsenoside composition, comprising all the components shown in 1), namely (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3 and Rh4.

[0097] In some embodiments, the secondary ginsenoside composition is a PPD-type secondary ginsenoside composition, comprising all components shown in 2), namely (20S)-Rg3, (20R)-Rg3, Rk1, Rg5, (20S)-Rh2, (20R)-Rh2, Rk2 and Rh3.

[0098] In some embodiments, the secondary ginsenoside composition is a total secondary ginsenoside composition, comprising all the components shown in 1) and all the components shown in 2), namely, (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3, Rh4, (20S)-Rg3, (20R)-Rg3, Rk1, Rg5, (20S)-Rh2, (20R)-Rh2, Rk2 and Rh3.

[0099] The present invention finds that secondary ginsenoside compositions (including PPT-type secondary ginsenoside compositions, PPD-type secondary ginsenoside compositions, and total secondary ginsenoside compositions) can prevent and treat bleomycin (BLM)-induced mouse pulmonary fibrosis (PF), which is specifically manifested in improving the lung imaging changes of (PF) mice, reducing the pathological damage of lung tissue, reducing the degree of lung inflammation and fibrosis, improving the survival rate of PF mice, and reducing the lung index of PF mice. At the same time, the secondary ginsenoside composition can also reduce the infiltration of inflammatory cells in the lungs of mice with acute lung injury (ALI) induced by lipopolysaccharide (LPS), reduce the permeability of the lungs and the expression of proinflammatory factors in the bronchoalveolar lavage fluid (BALF), and provide alveolar vascular endothelial barrier protection. In terms of the efficacy of treating pulmonary fibrosis, the secondary ginsenoside composition has better efficacy and in vivo safety than the pulmonary fibrosis drug pirfenidone; the secondary ginsenoside composition has better efficacy than the pulmonary fibrosis drug nintedanib.

[0100] In the present invention, whether it is a PPT-type secondary ginsenoside composition or a PPD-type secondary ginsenoside composition or a total secondary ginsenoside composition composed of them, it has the same efficacy in preventing pulmonary fibrosis (administered the day after modeling). The PPT-type secondary ginsenoside composition and the PPD-type secondary ginsenoside composition also have the same efficacy in treating pulmonary fibrosis in mice (administered one week after modeling). Therefore, it is reasonable to believe that the total secondary ginsenoside composition composed of PPT-type secondary ginsenoside and PPD-type secondary ginsenoside in different proportions should have the same efficacy in preventing pulmonary fibrosis.

[0101] In industrial production, the preparation of total secondary ginsenoside composition is simpler, easier and less costly than that of PPT-type secondary ginsenoside composition and PPD-type secondary ginsenoside composition. And at the same oral administration dose (greater than or equal to 100 mg / kg), the solubility of total secondary ginsenoside composition is better than that of PPT-type secondary ginsenoside composition and PPD-type secondary ginsenoside composition. As an example, in 0.5% sodium carboxymethyl cellulose solution, the maximum solubility of each secondary ginsenoside is as follows: total secondary ginsenoside composition: 250 mg / kg; PPT-type secondary saponin composition: 100 mg / kg; PPD-type secondary saponin composition: 100 mg / kg. Therefore, in the application of drugs and health foods for the prevention and treatment of pulmonary fibrosis and acute lung injury, total secondary ginsenoside composition can be given priority.

[0102] In some embodiments, the mass content of the components shown in 1) to 2) in the secondary ginsenoside composition is 50% or more, preferably 60% or more, and more preferably 70% or more.

[0103] In some specific embodiments, the secondary ginsenoside composition (i.e., PPT-type secondary ginsenoside composition) contains, by mass percentage, 8-13% (20S)-Rg2, 15-25% (20S)-Rh1, 5-10% (20R)-Rg2, 10-15% (20R)-Rh1, 2-5% Rg6, 5-10% F4, 5-10% Rk3, and 10-15% Rh4.

[0104] In some specific embodiments, the secondary ginsenoside composition (i.e., PPD-type secondary ginsenoside composition) contains, by mass percentage, 30-40% (20S)-Rg3, 4-8% (20R)-Rg3, 3-5% Rk1, 8-13% Rg5, 8-13% (20S)-Rh2, 2-4% (20R)-Rh2, 1-2% Rk2, and 2-4% Rh3.

[0105] In some specific embodiments, the secondary ginsenoside composition (i.e., the total secondary ginsenoside composition) contains, by mass percentage, 1-3% (20S)-Rg2, 5-7% (20S)-Rh1, 1-3% (20R)-Rg2, 5-8% (20R)-Rh1, 0.5-0.8% Rg6, 2-4% F4, 2-4% Rk3, 7-10% Rh4, 10-20% (20S)-Rg3, 5-7% (20R)-Rg3, 5-7% Rk1, 10-20% Rg5, 0.5-0.7% (20S)-Rh2, 0.1-0.3% (20R)-Rh2, 0.1-0.3% Rk2, and 0.5-0.8% Rh3.

[0106] In some embodiments, the secondary ginsenoside composition further contains a small amount of other types of secondary ginsenosides.

[0107] Preparation method

[0108] The present invention also provides a method for preparing a secondary ginsenoside composition, which comprises: using at least one of a primary ginsenoside composition, a PPT-type primary ginsenoside composition, and a PPD-type primary ginsenoside composition as a raw material, hydrolyzing only the C-20 sugar in the raw material and the C-20 alcohol hydroxyl group of the hydrolysis product, and further dehydrating the resulting product to obtain a secondary ginsenoside composition.

[0109] By hydrolyzing only the C-20 sugar in the above raw materials and further dehydrating the C-20 alcoholic hydroxyl group of the hydrolysis product, a secondary ginsenoside composition with better effect in preventing and treating pulmonary fibrosis and acute lung injury can be obtained.

[0110] The sugar at the C-20 position of the primary ginsenoside is unstable and is easily hydrolyzed under heating or acidic conditions to produce various secondary ginsenosides, such as PPT-type secondary ginsenosides Rg2 and Rh1, PPD-type secondary ginsenosides Rg3 and Rh2, etc. The sugar at the C-6 and C-3 positions of PPT and PPD-type ginsenosides is very stable and is difficult to hydrolyze under heating and acidic conditions. By utilizing the above characteristics of the primary ginsenosides, it is easy to prepare a secondary ginsenoside composition that only hydrolyzes the sugar at the C-20 position under heated acidic conditions.

[0111] In some embodiments, the hydrolysis conditions are: acidic conditions at 70-90°C.

[0112] In some embodiments, the acidic condition is provided by a mixed solution of methanol and 0.2±0.02N hydrochloric acid solution in a volume ratio of 1:0.8-1.2.

[0113] In some embodiments, the preparation method further comprises: after the hydrolysis reaction is completed, using n-butanol to extract the secondary ginsenoside composition.

[0114] In some specific embodiments, the preparation method comprises: dissolving the raw material in a mixed solution of methanol and 0.2±0.02N hydrochloric acid solution in a volume ratio of 1:0.8-1.2, hydrolyzing at 70-90°C for 3-4h, and then neutralizing with 8-12N sodium hydroxide, followed by removal of methanol by rotary evaporation. After the solution is cooled at room temperature, pure water is added, followed by three extractions with n-butanol, the n-butanol layers are combined and dried. The dried product obtained is dissolved in methanol again, and activated carbon is added for decolorization and stirring. After completing these steps, the secondary ginsenoside composition is finally dried.

[0115] In some embodiments, the main components of the native ginsenoside composition include Rg1, Re, Rf, Rb1, Rc, Rb2, Rb3 and Rd, and the content of the main components in the native ginsenoside composition is greater than 70%.

[0116] In some embodiments, the PPT-type native ginsenoside composition and the PPD-type native ginsenoside composition are separated from the native ginsenoside composition by medium pressure liquid chromatography (MPLC); in the medium pressure liquid chromatography, water is used as mobile phase A, methanol is used as mobile phase B, the flow rate is 50±5 mL / min, and a linear gradient elution method is adopted: 0-5 min, 10% mobile phase B; 5-45 min, 10%-100% mobile phase B; 45-55 min, 100% mobile phase B.

[0117] In some specific embodiments, the medium pressure liquid chromatography technology uses a Büchi brand medium pressure liquid chromatography (MPLC) preparative system and an ODS medium pressure preparative glass chromatography column (with a size of 26×460 mm) produced by Büchi.

[0118] In some specific embodiments, the detection wavelength in the medium pressure liquid chromatography technique is 210±5 nm.

[0119] In some specific embodiments, the primary ginsenoside composition can be obtained by extracting ginseng roots, stems, leaves, and / or fruits as raw materials. However, other Ginseng plants containing primary ginsenosides (such as roots, stems, leaves, flowers and fruits of American ginseng and Panax notoginseng, etc.) or commercial products such as Korean ginseng, red ginseng and black ginseng can all be prepared by the same or similar methods of the present invention to obtain the secondary ginsenoside composition.

[0120] In some specific embodiments, when performing the extraction, the extracting solution is a 70-90% methanol aqueous solution.

[0121] In some specific embodiments, when performing the extraction, ultrasound is used to assist the extraction, which can improve the extraction efficiency.

[0122] In some specific embodiments, after the extraction is completed, the methanol is evaporated and removed, the obtained aqueous solution is separated using a macroporous adsorption resin, and the saponin-rich eluates are combined and concentrated and dried to obtain the native ginsenoside composition.

[0123] The above-mentioned embodiments can be combined with common sense in this field to obtain a preferred embodiment of the preparation method of the present invention.

[0124] The present invention also provides a secondary ginsenoside composition, which is prepared by the preparation method described in any of the aforementioned embodiments.

[0125] application

[0126] The present invention further provides use of the secondary ginsenoside composition described in any of the aforementioned embodiments in the preparation of medicines or foods, wherein the medicines or foods are used to prevent, treat or alleviate at least one disease selected from pulmonary fibrosis and acute lung injury.

[0127] In some embodiments, the drug or food is used for at least one of the following aspects: i) improving the survival rate of an individual and prolonging the survival period; ii) reducing the lung index; iii) improving lung function; iv) improving lung imaging; v) reducing the degree of alveolar structural lesions; vi) reducing the degree of lung inflammatory cell infiltration; vii) reducing the degree of lung fibrosis; viiii) reducing collagen deposition in lung tissue; ix) reducing the hydroxyproline content in lung tissue; x) reducing the expression of fibrosis-related proteins in lung tissue; xi) reducing the total number of white blood cells and neutrophils in bronchoalveolar lavage fluid; xii) reducing the expression of proinflammatory factors in bronchoalveolar lavage fluid; xiii) reducing the myeloperoxidase content in bronchoalveolar lavage fluid; xiv) reducing the total protein concentration in bronchoalveolar lavage fluid, increasing the expression of tight junction protein ZO-1 in lung tissue, and providing alveolar vascular endothelial barrier protection; xv) inhibiting the activation of the nuclear transcription factor NF-κB signaling pathway in the lungs, and reducing the expression of downstream inflammation-related proteins COX-2 and proinflammatory factors TNF-α regulated by the NF-κB signaling pathway.

[0128] In some embodiments, the inflammatory cells include at least one of total leukocytes, lymphocytes, neutrophils, eosinophils, monocytes, and macrophages.

[0129] In some embodiments, the fibrosis-related protein includes at least one of fibronectin, collagen I, and smooth muscle actin α-SMA.

[0130] In some embodiments, the pro-inflammatory factor includes at least one of TNF-α, IL-1β, and IL-6.

[0131] In some embodiments, the dosage form of the drug is an oral dosage form.

[0132] In some embodiments, the drug further comprises at least one of the following components: (1) excipients: ingredients used to fix, disperse, improve the taste or increase the stability of the drug, such as fillers, adhesives, dispersants, etc.; (2) flavoring agents: ingredients used to improve the taste of the drug or mask the unpleasant taste of the drug, such as sweeteners, flavors, etc.; (3) coating agents: ingredients used to encapsulate the drug, protect the drug from damage by gastric acid, etc., delay the release rate of the drug or control the release rate, such as capsule shells, tablet coatings, etc.; (4) dissolution aids: ingredients used to enhance the solubility of the drug in the gastrointestinal tract and improve the bioavailability of the drug, such as surfactants, solubilizers, etc.; (5) fillers: ingredients used to adjust the drug dosage and improve the physical properties or volume of the oral dosage form, such as cellulose, starch, etc.

[0133] In some embodiments, the drug or food is administered to a mammal.

[0134] In some embodiments, the subjects of administration of the drug or food include but are not limited to: humans, primates (such as apes, apes and gorillas), canines (such as dogs, wolves, etc.), felines (such as cats, leopards, lions, etc.), rodents (such as mice, hamsters, etc.), even-toed ungulates (such as cattle, sheep, pigs, etc.), and odd-toed ungulates (such as horses, etc.).

[0135] The present invention also provides a medicine comprising the secondary ginsenoside composition described in any of the aforementioned embodiments.

[0136] In some embodiments, the dosage form of the drug is an oral dosage form.

[0137] The present invention also provides a food, comprising the secondary ginsenoside composition described in any of the aforementioned embodiments.

[0138] The present invention also provides a method for preventing, treating or alleviating a disease, wherein the disease includes at least one selected from pulmonary fibrosis and acute lung injury, and the method comprises: using the secondary ginsenoside composition, medicine and / or food described in any of the aforementioned embodiments.

[0139] The dosage of the secondary ginsenoside composition of the present invention can vary widely according to the nature and severity of the disease to be prevented or treated and the individual situation. Generally speaking, the suitable daily dosage range of the secondary ginsenoside composition of the present invention is 0.01-200 mg / kg body weight, preferably 0.1-100 mg / kg body weight, more preferably 0.1-50 mg / kg body weight, and most preferably 0.2-50 mg / kg body weight. The above dosage can be administered in one dosage unit or divided into several dosage units, depending on the doctor's clinical experience and the dosage regimen including the use of other treatment means.

[0140] The secondary ginsenoside composition of the present invention or the pharmaceutical composition containing the same can be taken alone or used in combination with other therapeutic drugs or symptomatic drugs. When the secondary ginsenoside composition of the present invention has a synergistic effect with other therapeutic drugs, its dosage should be adjusted according to the actual situation.

[0141] The secondary ginsenoside composition of the present invention has significant therapeutic effects on the bleomycin-induced mouse pulmonary fibrosis model, can significantly prolong the survival of the model animals, improve various lung functions, alleviate lung inflammation, reduce pathological damage and fibrosis of the lungs, is safe and non-toxic, and provides a new drug option for the clinical treatment of pulmonary fibrosis.

[0142] Example

[0143] Embodiments of the present invention will be described in detail below in conjunction with examples. It should be understood that these examples are only used to illustrate the present invention and are not intended to limit the scope of the present invention. The experimental methods in the following examples that do not specify specific conditions are preferably referred to the guidance provided in the present invention, and can also be based on the experimental manual or normal conditions in this area, and can also be based on other experimental methods known in the art, or according to the conditions recommended by the manufacturer.

[0144] In the following specific embodiments, the measured parameters of raw material components may have slight deviations within the range of weighing accuracy unless otherwise specified. For temperature and time parameters, acceptable deviations caused by instrument test accuracy or operation accuracy are allowed.

[0145] Example 1 Preparation and content determination results of secondary ginsenoside composition

[0146] First, the ginseng root medicinal material was ground into powder and extracted with 10, 8 and 6 times the amount of 80% methanol for 1 hour by ultrasonic assisted extraction. After the extracts were combined, the methanol was removed by rotary evaporation, and the resulting aqueous solution was then separated using a macroporous adsorption resin, and the saponin-rich eluents were combined. Finally, the total primary ginsenoside composition of ginseng root was successfully obtained through concentration and drying. The total primary ginsenoside composition from ginseng root was used to prepare the total secondary ginsenoside composition.

[0147] The ginseng stem medicinal material was ground into powder, and 10, 8 and 6 times the amount of 80% methanol were used in sequence, and ultrasonic-assisted extraction was performed for 1 hour. After the extracts were combined, the methanol was removed by rotary evaporation, and the obtained aqueous solution was then separated using a macroporous adsorption resin, and the eluents rich in saponins were combined. Finally, by concentration and drying, the total native ginsenoside composition of ginseng stem was successfully obtained. Then, the total native ginsenosides from ginseng stem were separated by ODS medium-pressure preparative chromatography columns to obtain PPT-type native ginsenosides and PPD-type native ginsenosides. In order to achieve the separation of PPT-type and PPD-type native ginsenoside compositions, this embodiment uses a medium-pressure liquid chromatography (MPLC) preparative system of the Büchi brand and an ODS medium-pressure preparative glass chromatographic column (size 26×460mm) of Büchi. During the separation process, water was used as mobile phase A, methanol was used as mobile phase B, the flow rate was set to 50mL / min, and the detection wavelength was set to 210nm. The linear gradient elution method was used: 0-5min, 10% B; 5-45min, 10%-100% B; 45-55min, 100% B. Each 200mL sample was collected as a group. The analysis results showed that the 9th component mainly contained the PPT-type primary ginsenoside composition, while the 12th component was mainly the PPD-type primary ginsenoside composition. The PPT-type primary ginsenoside composition and the PPD-type primary ginsenoside composition derived from ginseng stems were used to prepare the PPT-type secondary ginsenoside composition and the PPD-type secondary ginsenoside composition.

[0148] Take 40g of the total primary ginsenoside composition from ginseng root, the PPT type primary ginsenoside composition from ginseng stem, and the PPD type primary ginsenoside composition from ginseng stem respectively, and dissolve them in a mixed solution of 200mL methanol and 0.2N hydrochloric acid (volume ratio of 50 / 50). Stir the reaction at a temperature of 80°C for 3 to 4 hours. After the reaction, 10N sodium hydroxide was used for neutralization treatment, and then methanol was removed by rotary evaporation. After the solution was cooled at room temperature, pure water was added to 300mL, and then 150mL of n-butanol was used for three extractions, the n-butanol layers were combined, and dried. The dried product obtained was dissolved in 200mL of methanol again, and 5g of activated carbon was added for decolorization and stirring. After completing these steps, the total secondary ginsenoside composition, the PPT type secondary ginsenoside composition, and the PPD type secondary ginsenoside composition were finally dried.

[0149] The composition and content of the total primary ginsenoside composition, the total secondary ginsenoside composition, the PPT type secondary ginsenoside composition and the PPD type secondary ginsenoside composition of ginseng root were detected by UHPLC-MS. The UHPLC-MS conditions are as follows:

[0150] Instrument: Agilent UHPLC-TOF-MS (6230);

[0151] Chromatographic column: Agilent Zorbax Extend C18 column (2.1×100mm, 1.8μm);

[0152] Mobile phase system: water containing 0.1% formic acid as the aqueous phase (A), acetonitrile containing 0.1% formic acid as the organic phase (B);

[0153] Flow rate: 0.35 mL / min;

[0154] Gradient elution program: 0-5 minutes, 5-20% B; 5-20 minutes, 20-45% B; 20-23 minutes, 45-80% B; 23-25 ​​minutes, 80-100% B; 25-27 minutes, 100-100% B; 27.1-32 minutes, 5-5% B;

[0155] Mass spectrometry conditions: ion source: electrospray ion source (ESI); ionization mode: negative ion mode; capillary voltage: 3500 V; nebulizer gas pressure: 35 psi; dry nitrogen temperature: 300 ° C; dry nitrogen flow rate: 11 L / min; sheath gas temperature: 325 ° C; sheath gas flow rate: 11 L / min; scanning range: m / z 200-1700;

[0156] Four different types of standard samples Rg1, Rb1, 20(S)-Rg2, and 20(S)-Rg3 were used to prepare standard curves for quantitative analysis of each monomer ginsenoside component. Rg1 was used to quantify each monomer component of PPT-type primary ginsenosides; Rb1 was used to quantify each monomer component of PPD-type primary ginsenosides; 20(S)-Rg2 was used to quantify each monomer component of PPT-type secondary ginsenosides; and 20(S)-Rg3 was used to quantify each monomer component of PPD-type secondary ginsenosides.

[0157] The test results of total primary ginsenoside composition, total secondary ginsenoside composition, PPT type secondary ginsenoside composition and PPD type secondary ginsenoside composition are shown in Table 1. Figures 1 to 4 And Tables 1 to 4.

[0158] Table 1 Main components and contents of total native ginsenoside composition

[0159]

[0160] Table 2 Main components and contents of total secondary ginsenoside composition

[0161]

[0162] Table 3 Main components and contents of PPT-type secondary ginsenoside compositions

[0163]

[0164] Table 4 Main components and contents of PPD-type secondary ginsenoside compositions

[0165]

[0166] Example 2 Secondary ginsenoside composition prevents and protects against bleomycin (BLM)-induced pulmonary fibrosis in mice

[0167] 82 male C57BL / 6J mice (aged 8-10 weeks) were selected and randomly divided into 10 groups (n=8-10), respectively named as sham operation group, model group (bleomycin), nintedanib group (bleomycin + 30 mg / kg nintedanib), total primary glycoside group (bleomycin + 50 mg / kg total primary ginsenoside composition), total secondary glycoside group (bleomycin + 50 mg / kg total secondary ginsenoside composition), PPT type secondary glycoside group (bleomycin + 50 mg / kg PPT type secondary ginsenoside composition), PPD type secondary glycoside group (bleomycin + 50 mg / kg PPD type secondary ginsenoside composition), Rg3 group (bleomycin + 50 mg / kg Rg3), Rg5 group (bleomycin + 50 mg / kg Rg5) and total saponin (control) group (bleomycin + 50 mg / kg total saponins), as shown in Table 5.

[0168] Table 5

[0169]

[0170] Among them, the total saponin (control) group was blended with monomer saponin components according to the mass percentage according to the ginsenoside ratio in the reference "Inhibitory effects of totalginsenoside on bleomycin-induced pulmonary fibrosis in mice. Biomedicine & Pharmacotherapy, 2019, 114: 108851. Deng Jiang, Zunyi Medical College". The Rg3 group (50 mg / kg) was blended with monomer components according to the ratio of 20(S)-Rg3 and 20(R)-Rg3 in the total secondary ginsenoside composition.

[0171] like Figure 5As shown, during the experiment, according to the dosage requirements of each group, the corresponding compound was dissolved in 0.5% sodium carboxymethyl cellulose (0.5% CMC-Na) aqueous solution, and each group of mice was administered by gavage. On day 0, the sham operation group was instilled with an equal amount of 0.9% sterile saline once through the trachea, and the model group and each drug group were instilled with 2.5 mg / kg bleomycin (Nippon Kayaku Co., Ltd., production batch number 600700) once through the trachea for modeling. Starting from the first day of modeling, the sham operation group and the model group were gavaged with an equal amount of 0.5% CMC-Na aqueous solution every day, the positive drug control group was gavaged with 30 mg / kg nintedanib (McLean) every day, and each ginsenoside administration group was gavaged according to the set dose every day, once a day, for 20 consecutive days. And on the 21st day of modeling, after anesthetizing the mice in each group, the mouse lung imaging was performed using a Micro-CT scanner. Finally, the mice were sampled and the degree of pulmonary fibrosis in the model animals was evaluated according to the experimental methods known in the art, including: H&E and Masson staining of mouse lung tissue, determination of hydroxyproline (HYP) content in mouse lung tissue, determination of total leukocytes, lymphocytes and macrophages in mouse bronchoalveolar lavage fluid (BALF) (Luna TM Automatic cell counter) etc.

[0172] Micro-CT scanning image results of mouse lung tissue ( Figure 6 ) showed that there were a large number of dense fibrous foci in the lungs of mice in the model group, indicating that bleomycin can induce radiological changes of pulmonary fibrosis in mice. Total primary glycosides and total saponins (control) can reduce the radiological changes of pulmonary fibrosis in mice. Total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides significantly reduced the radiological changes of pulmonary fibrosis in mice, and the lung images of mice in the total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides groups were closer to those of mice in the sham operation group. Ginsenosides Rg3 and Rg5 had a certain degree of improvement on the lung images of PF mice, but not as good as the imaging manifestations of other drug-treated groups.

[0173] Based on macroscopic observation of lung tissue ( Figure 6), the lung tissue of mice in the sham operation group was pink, soft, smooth and full. The lungs of mice in the model group were dark red, hard, and had poor elasticity, with visible bleeding spots and scar-like textures. The lung tissue of mice in the total primary glycosides and total saponins (control) groups was dark red in color, with many bleeding spots and obvious fibrous texture. Compared with the model group, total primary glycosides and total saponins (control) alleviated pulmonary fibrosis in mice. The lung tissue of mice in the total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides groups was light red, with few bleeding spots and no obvious fibrous texture, which was close to that of mice in the sham operation group. It shows that total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides have significant preventive and protective effects on pulmonary fibrosis in mice. Ginsenosides Rg3 and Rg5 have a slight improvement effect on pulmonary fibrosis in PF mice.

[0174] H&E staining of mouse lung tissues showed Figure 7 As shown in the figure, the alveolar structure of the lung tissue of mice in the sham operation group was normal, without inflammatory cell infiltration; the local alveolar septa of the lung tissue of mice in the model group were significantly thickened, a large number of alveolar cavities were significantly reduced or disappeared, and a large number of inflammatory cells infiltrated the lung tissue interstitium. Compared with the lung tissue of mice in the model group, the vacuolar degeneration of the lung tissue of mice in the total primary glycosides, total saponins (control), total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides groups was significantly reduced, the thickening of the alveolar wall was significantly improved, and the inflammatory infiltration was significantly improved. Moreover, the effects of total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides were significantly better than those of total primary glycosides, and closer to the performance of the sham operation group. Ginsenosides Rg3 and Rg5 also had a certain degree of improvement on the lung pathology of PF mice, but their effects were far less than those of total secondary glycosides and PPD-type secondary glycosides.

[0175] Results of total leukocytes, lymphocytes and macrophages in mouse bronchoalveolar lavage fluid ( Figure 8 ) showed that compared with the sham operation group, the number of inflammatory cells in the model group mice increased significantly (p<0.0001). Compared with the model group, the number of total leukocytes, lymphocytes and macrophages in the bronchoalveolar lavage fluid of mice treated with total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides was significantly reduced (p<0.05 or p<0.01), indicating that secondary glycosides can significantly reduce the number of inflammatory cells in the bronchoalveolar lavage fluid of mice. Compared with total native glycosides, the number of total leukocytes and lymphocytes in the bronchoalveolar lavage of mice treated with total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides was significantly reduced (p<0.05 or p<0.01), indicating that the effect of total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides in reducing inflammatory cells in the alveoli of mice was significantly better than that of total native glycosides. In addition, the effect of total secondary glycosides and PPD-type secondary glycosides in reducing inflammatory cells in the alveoli of mice was also significantly better than that of Rg3 and Rg5 (p<0.05).

[0176] Masson staining of mouse lung tissues was performed. Fig. 9As shown in the figure, there was no collagen deposition in the lung tissue of mice in the sham operation group; a large amount of blue collagen deposition appeared in the lung tissue of mice in the model group. Compared with the lung tissue of mice in the model group, the collagen deposition in the lung tissue of mice in the total primary glycosides (p<0.001), total secondary glycosides (p<0.0001), PPT-type secondary glycosides (p<0.0001), PPD-type secondary glycosides (p<0.0001), total saponin (control) group (p<0.0001), Rg3 (p<0.05) and Rg5 (p<0.05) was significantly reduced. Moreover, the effects of total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides were significantly better than those of total primary glycosides (p<0.05); the performance of the total secondary glycosides and PPD-type secondary glycosides groups was closer to that of the positive control drug nintedanib group. The effects of total secondary glycosides and PPD-type secondary glycosides in slowing down the collagen deposition in the lung tissue of PF mice were also significantly better than those of Rg3 and Rg5 (p<0.01).

[0177] Results of hydroxyproline content detection in mouse lung tissue ( Fig.10 ) showed that compared with the sham operation group, the hydroxyproline content in the lung tissue of the mice in the model group was significantly increased (p<0.0001). Compared with the model group, the hydroxyproline content in the lung tissue of the mice in the total native glycosides, total secondary glycosides, PPT-type secondary glycosides, PPD-type secondary glycosides and total saponins (control) groups was significantly decreased (p<0.01). It shows that total native glycosides, total secondary glycosides, PPT-type secondary glycosides, PPD-type secondary glycosides and total saponins (control) can significantly reduce the hydroxyproline content in the lung tissue of PF mice. Compared with total native glycosides, total secondary glycosides, PPT-type secondary glycosides and PPD-type secondary glycosides are significantly better than total native glycosides in reducing the hydroxyproline content in the lung tissue of PF mice (p<0.0001). In addition, although the decrease in hydroxyproline content in the lung tissue of mice in the Rg3 and Rg5 groups was statistically significant compared with the model group (p<0.05), the efficacy of Rg3 and Rg5 in reducing the hydroxyproline content in the lung tissue of PF mice was far less than that of total secondary glycosides and PPD-type secondary glycosides (p<0.0001).

[0178] Example 3 Treatment of BLM-induced pulmonary fibrosis in mice with secondary ginsenoside composition

[0179] Male C57BL / 6J mice (8-10 weeks old) were selected and fed for one week before modeling. On day 0, 2.5 mg / kg bleomycin (Nippon Kayaku Co., Ltd., production batch number 600700) was instilled into the trachea of ​​mice once to establish a bleomycin-induced pulmonary fibrosis animal model; mice in the sham operation group were instilled once through the trachea with an equal amount of 0.9% sterile saline (NS). The subjects were divided into sham operation group (Sham), model group (bleomycin), pirfenidone group (bleomycin + 200 mg / kg pirfenidone), low-dose PPT-type secondary glycoside group (bleomycin + 50 mg / kg PPT-type secondary glycoside), high-dose PPT-type secondary glycoside group (bleomycin + 100 mg / kg PPT-type secondary glycoside), low-dose PPD-type secondary glycoside group (bleomycin + 50 mg / kg PPD-type secondary glycoside) and high-dose PPD-type secondary glycoside group (bleomycin + 100 mg / kg PPD-type secondary glycoside), as shown in Table 6.

[0180] Table 6

[0181]

[0182] like Fig.11 As shown, according to the dosage requirements of each group, the corresponding drugs were dissolved in 0.5% sodium carboxymethyl cellulose (0.5% CMC-Na) aqueous solution. Starting from the 7th day after modeling, the sham operation group and the model group were gavaged with an equal amount of 0.5% CMC-Na aqueous solution every day, the positive drug control group was gavaged with 200 mg / kg pirfenidone (McLean) every day, and the secondary ginsenosides were gavaged at the set dose every day, once a day, for 14 consecutive days. The survival of the mice was observed and recorded every day during the experiment. On the 21st day of modeling, after anesthetizing the mice in each group, a Micro-CT scanner was used to image the mouse lungs. Finally, the mice were sampled and the degree of pulmonary fibrosis in the model animals was evaluated according to experimental methods known in the art, including: pathological examination of mouse lung tissue (H&E staining and Masson staining), immunohistochemical staining of mouse lung tissue to detect the expression levels of pulmonary fibrosis-related proteins Fibronectin, Collagen I, and α-SMA, determination of hydroxyproline (HYP) content in mouse lung tissue, determination of the number of total leukocytes, lymphocytes, and macrophages in mouse bronchoalveolar lavage fluid (BALF), Western blot analysis to detect pulmonary fibrosis-related proteins in mouse lung tissue, etc. At the same time, the pathological conditions of the heart, liver, spleen, and kidney of each group of mice were evaluated (H&E staining).

[0183] During the experiment, the survival time of each group of mice was recorded and counted. Fig.12The survival rate of mice in the model group was 58.3%, the survival rate of the positive drug pirfenidone group was 66.7%, and the survival rate of mice in each experimental group of PPT-type secondary glycosides (including low-dose group and high-dose group) and PPD-type secondary glycosides (including low-dose group and high-dose group) was 100% (completely overlapped with the sham operation group in the figure). Compared with the PF model group, each dose of PPT-type secondary glycosides and PPD-type secondary glycosides significantly increased the survival rate of PF mice (p<0.05).

[0184] Micro-CT scanning image results of mouse lung tissue ( Fig.13 ) showed that there were a large number of dense fibrous foci (high-density images) in the lungs of mice in the model group. Compared with the model group, the areas of lung fibrosis in the two dose groups of PPT-type secondary glycosides and PPD-type secondary glycosides were significantly reduced, indicating that PPT-type secondary glycosides and PPD-type secondary glycosides can significantly improve the imaging changes of lung tissue in PF mice. The lung images of mice in the high-dose (100 mg / kg) group of PPT-type secondary glycosides and PPD-type secondary glycosides were close to those of mice in the sham operation group, which was significantly better than the positive drug pirfenidone.

[0185] According to the lung tissue ( Fig.13 ) were observed with the naked eye, and the lungs of the mice in the model group were dark red, with visible scar-like textures. Compared with the model group, the lung tissues of mice in the low-dose (50 mg / kg) group of PPT-type secondary glycosides and PPD-type secondary glycosides had fewer bleeding spots and no obvious fibrous lines; the lung tissues of mice in the high-dose (100 mg / kg) group of PPT-type secondary glycosides and PPD-type secondary glycosides were light pink, with no bleeding spots and fibrous lines, which was close to that of mice in the sham operation group. This shows that PPT-type secondary glycosides and PPD-type secondary glycosides have significant therapeutic effects on mouse pulmonary fibrosis, and are superior to the positive drug pirfenidone.

[0186] H&E staining of mouse lung tissues showed Fig.14 As shown in the results, the alveolar structure of the lung tissue of mice in the sham operation group was normal, without inflammatory cell infiltration; the alveolar septa of the lung tissue of mice in the model group were significantly thickened, a large number of alveolar cavities were significantly reduced or disappeared, and a large number of inflammatory cells infiltrated the lung tissue interstitium; compared with the model group, both doses of PPT-type secondary glycosides and PPD-type secondary glycosides significantly reduced the degree of inflammation in the lung tissue of BLM-induced pulmonary fibrosis mice, and significantly improved the alveolar wall thickness, alveolar cavity volume and inflammatory cell infiltration. The therapeutic effect of high doses (100 mg / kg) of PPT-type secondary glycosides and PPD-type secondary glycosides was close to that of the sham operation group.

[0187] Results of total leukocytes, lymphocytes and macrophages in mouse bronchoalveolar lavage fluid ( Fig.15) showed that compared with the model group, both doses of PPT-type secondary glycosides and PPD-type secondary glycosides could significantly reduce the number of total leukocytes (p<0.001), lymphocytes (p<0.05 or p<0.01) and macrophages (p<0.05 or p<0.001) in the bronchoalveolar lavage fluid of PF mice, indicating that PPT-type secondary glycosides and PPD-type secondary glycosides can significantly improve the lung inflammation of PF mice.

[0188] Masson staining of mouse lung tissues was performed. Fig.16 As shown in the figure, the lung tissue structure of mice in the sham operation group was normal, and the fiber staining was fine and light; the alveolar wall structure of the lung tissue of mice in the model group was significantly destroyed, and a large amount of blue collagen deposition was observed (p<0.0001). Compared with the lung tissue of mice in the model group, the collagen deposition in the lung tissue of mice in the low-dose group (50 mg / kg) of PPT-type secondary glycosides and PPD-type secondary glycosides was significantly reduced, and the degree of pulmonary fibrosis was significantly reduced (p<0.0001), which was equivalent to the positive control drug pirfenidone group. The high-dose group (100 mg / kg) of PPT-type secondary glycosides and PPD-type secondary glycosides was closer to the performance of the sham operation group, and was significantly better than the positive drug pirfenidone (p<0.05).

[0189] Results of hydroxyproline content detection in mouse lung tissue ( Fig.17 ) showed that compared with the sham operation group, the hydroxyproline content in the lung tissue of the model group mice increased significantly (p<0.0001). Compared with the model group, both doses of PPT-type secondary glycosides and PPD-type secondary glycosides can significantly reduce the hydroxyproline content in the lung tissue of mice (p<0.0001), that is, improve the excessive deposition of collagen in the lung tissue of mice. Moreover, the efficacy of both doses of PPT-type secondary glycosides and PPD-type secondary glycosides in reducing the hydroxyproline content in the lung tissue of mice is significantly better than that of the positive control drug pirfenidone (p<0.05 or p<0.01 or p<0.001 or p<0.0001).

[0190] Western blot detection results of fibronectin, collagen COL1A1 and smooth muscle actin α-SMA in lung tissue of mice in PPT-type secondary glycoside group ( Fig.18 ) showed that compared with the sham operation group, the expression levels of fibronectin, collagen COL1A1 and smooth muscle actin α-SMA in the lung tissue of the model group mice were significantly increased. Compared with the model group, PPT-type secondary glycosides significantly reduced the expression of fibronectin, collagen COL1A1 and smooth muscle actin α-SMA in the lung tissue of PF mice. And the high dose (100 mg / kg) of PPT-type secondary glycosides was significantly better than pirfenidone in reducing smooth muscle actin α-SMA in the lung tissue of PF mice (p<0.001).

[0191] Western blot detection results of fibronectin, collagen COL1A1 and smooth muscle actin α-SMA in lung tissue of mice in PPD-type secondary glycoside group ( Fig.19 ) showed that compared with the sham operation group, the expression levels of fibronectin, collagen COL1A1 and smooth muscle actin α-SMA in the lung tissue of the model group mice were significantly increased. Compared with the model group, PPD-type secondary glycosides significantly reduced the expression of fibronectin and collagen COL1A1 in the lung tissue of PF mice (p<0.001).

[0192] Immunohistochemistry was used to detect the expression of fibrosis-related target proteins in lung tissue. Fig. 20 As shown. Compared with the model group, the two doses of PPT-type secondary glycosides and PPD-type secondary glycosides significantly reduced the expression levels of fibronectin, collagen I and smooth muscle actin α-SMA in the lung tissue of PF mice (p<0.0001). Moreover, the efficacy of PPT-type secondary glycosides and PPD-type secondary glycosides in reducing fibronectin, collagen COL1A1 and smooth muscle actin α-SMA in the lung tissue of PF mice was significantly better than that of the positive drug pirfenidone (p<0.05).

[0193] Pathological examination results of the heart, liver, spleen and kidney of mice ( Fig.21 ) showed that compared with the sham-operated mice, high doses of PPT-type secondary glycosides (100 mg / kg) did not cause significant pathological changes in the heart, liver, spleen, and kidney of mice, indicating that PPT-type secondary glycosides and PPD-type secondary glycosides were biosafe under this experimental condition. However, pirfenidone (200 mg / kg) caused significant pathological changes in the spleen of mice, suggesting that pirfenidone has certain toxicity to mice.

[0194] Example 4 Total secondary ginsenoside composition treats bleomycin (2.5 mg / kg)-induced pulmonary fibrosis in mice

[0195] Male C57BL / 6J mice (8-10 weeks old) were selected and bred for one week before modeling. On day 0, 2.5 mg / kg bleomycin (Nippon Kayaku Co., Ltd., production batch number 330810) was instilled into the trachea of ​​mice once. In this experiment, 104 mice were instilled with bleomycin. On the 10th day after modeling, 43 mice died. At this time, the survival rate of PF model animals was 58%, and most of the surviving PF model animals had significantly reduced weight; mice in the sham operation group were instilled once with an equal amount of 0.9% sterile saline (NS) through the trachea. On the 10th day after bleomycin modeling (peak of the disease), the model animals were divided into groups and drug administration, and were divided into sham operation group (Sham), model group (bleomycin), nintedanib group (bleomycin + 50 mg / kg nintedanib), total primary glycoside group (bleomycin + 50 mg / kg total primary ginsenoside composition) and total secondary glycoside group (bleomycin + 50 mg / kg total secondary ginsenoside composition), as shown in Table 7.

[0196] Table 7

[0197]

[0198] like Fig. 22 As shown, according to the dosage requirements of each group, the corresponding drugs were dissolved in 0.5% sodium carboxymethyl cellulose (0.5% CMC-Na) aqueous solution. Starting from the 10th day after modeling, the sham operation group and the model group were gavaged with an equal amount of 0.5% CMC-Na aqueous solution every day, the positive drug control group was gavaged with 50 mg / kg nintedanib (McLean) every day, and each ginsenoside administration group was gavaged according to the set dose every day, once a day, for 11 consecutive days. During the experiment, the survival of the mice was observed and recorded every day. On the 21st day of modeling, the mice in each group were anesthetized and the mouse lungs were imaged using a Micro-CT scanner. Finally, the mice were sampled, the lung weight was weighed, and the lung index was calculated.

[0199] The statistical results of the survival time of mice in each group are shown in Fig.23 A in the figure. The survival rate of mice in the model group was 41.2%, the survival rate of the positive drug nintedanib group was 58.3%, the survival rate of mice in the total secondary glycoside group was 75.0%, and the survival rate of mice in the total primary glycoside group was 62.5%. Compared with the PF model group, the total secondary glycosides significantly improved the survival rate of PF mice (p<0.05). The lung weight index can reflect the degree of fibrosis in the animal's lungs to a certain extent, such as Fig.23 As shown in B, only total secondary glycosides significantly reduced the lung index of model animals (p<0.05), and total native glycosides and nintedanib had a tendency to reduce the lung index of model animals, but there was no statistical difference. Both mouse survival rate and lung index showed that total secondary glycosides were more effective than total native glycosides and positive drug nintedanib in treating pulmonary fibrosis.

[0200] Micro-CT scanning image results of mouse lung tissue ( Fig.24 ) showed that there were large areas of dense fibrous foci in the lungs of mice in the model group (high-density images). Compared with the model group, the area of ​​fibrous foci in the lungs of mice in the total secondary glycosides group was significantly reduced, and the area of ​​fibrous foci in the lungs of mice in the total primary glycosides and nintedanib groups showed an improvement trend, but far less than that in the total secondary glycosides group.

[0201] Example 5 Total secondary ginsenoside composition prevents and treats bleomycin (1.8 mg / kg)-induced pulmonary fibrosis in mice

[0202] The present invention uses the same modeling dose of bleomycin used in Example 2, Example 3 (bleomycin, Nippon Kayaku Co., Ltd., production batch number 600700) and Example 4 (bleomycin, Nippon Kayaku Co., Ltd., production batch number 330810), which is 2.5 mg / kg. The present invention further found that the bleomycin in Example 4 caused serious deaths in the model animals, and most of the model animals were slow in action, their hair lost its luster, and the samples were taken on the 21st day after modeling. The reason for the analysis may be that the different production batches of bleomycin may cause differences in its potency. Therefore, in Example 5, the present invention slightly adjusted the induction dose of bleomycin, from 2.5 mg / kg in Example 4 to 1.8 mg / kg in Example 5, and measured the lung function (the gold standard for evaluating pulmonary fibrosis) and other indicators of the model animals on the 28th day after modeling. The specific operation is as follows:

[0203] Male C57BL / 6J mice (8-10 weeks old) were selected and fed for one week before modeling. On day 0, 1.8 mg / kg bleomycin (Nippon Kayaku Co., Ltd., production batch number 330810) was instilled into the trachea of ​​mice once to establish a bleomycin-induced pulmonary fibrosis animal model; mice in the sham operation group were instilled once through the trachea with an equal amount of 0.9% sterile saline (NS).

[0204] Preventive administration mode: Administration began on the first day after bleomycin modeling, and the groups were divided into sham operation group (Sham), model group (bleomycin), and total secondary glycoside group (bleomycin + 20 mg / kg total secondary ginsenoside composition), as shown in Table 8.

[0205] Treatment administration mode: Group administration began on the 10th day after bleomycin modeling (the peak of the disease, when the survival rate of PF model animals was 94%), and was divided into a model group (bleomycin), a nintedanib group (bleomycin + 50 mg / kg nintedanib), a low-dose group of total secondary glycosides (bleomycin + 20 mg / kg total secondary ginsenoside composition) and a high-dose group of total secondary glycosides (bleomycin + 50 mg / kg total secondary ginsenoside composition), as shown in Table 8.

[0206] Table 8

[0207]

[0208] like Fig.25 As shown, according to the dosage requirements of each group, the corresponding drugs were dissolved in 0.5% sodium carboxymethyl cellulose (0.5% CMC-Na) aqueous solution. In the preventive administration mode, oral administration began on the first day after BLM modeling. The sham operation group and the model group were gavaged with an equal amount of 0.5% CMC-Na aqueous solution every day. The total secondary glycosides preventive administration group was gavaged with 20 mg / kg of total secondary ginsenoside composition every day, once a day, for 27 consecutive days. In the therapeutic administration mode, group administration began on the 10th day after BLM modeling. The nintedanib treatment group was gavaged with 50 mg / kg nintedanib (McLean) every day, and each treatment group of total secondary ginsenosides was gavaged according to the set dose every day, once a day, for 18 consecutive days. During the experiment, the survival of the mice was observed and recorded every day. On the 28th day of modeling, after anesthesia of each group of mice, the pulmonary function test system (DSI PFT, USA) was used to measure the lung function of mice (6-12 mice per group); Micro-CT scanner was used to image the lungs of mice (5-17 mice per group). Finally, the mice were sampled, and the survival rate and lung weight index (lung weight mg / body weight g) were evaluated and statistically analyzed according to experimental methods known in the art; H&E staining and Masson staining were used to perform pathological evaluation of lung inflammation and fibrosis, and an automatic blood cell analyzer (Mindary BC-5000Vet) was used to determine the content of inflammatory cells in bronchoalveolar lavage fluid; immunohistochemistry was used to analyze fibrosis-related proteins in lung tissue, etc.

[0209] The statistical results of the survival time of mice in each group are shown in Fig.26 A (prevention mode) and C (treatment mode) in the experiment. The results show that in the prevention mode, the survival rate of mice in the model group was 69.0%, and the survival rate of mice in the total secondary glycosides (20 mg / kg) preventive administration group was 92.6%, which significantly prolonged the survival of the model animals (p<0.05). In the treatment mode, the survival rate of mice in the model group was 65.5%, the survival rate of the positive drug nintedanib group was 87.5%, the survival rate of mice in the total secondary glycosides (20 mg / kg) group was 96.3%, and the survival rate of mice in the total secondary glycosides (50 mg / kg) group was 88.9%. Compared with the PF model group, both doses of total secondary glycosides significantly increased the survival rate of PF mice (p<0.01 or p<0.05). The lung weight index can reflect the degree of fibrosis in the animal's lungs to a certain extent, such as Fig.26 As shown in B (preventive mode) and D (treatment mode), total secondary glycosides significantly reduced the lung weight index of model animals (p<0.001 or p<0.05).

[0210] The results of mouse lung function test are shown in Fig. 27 A (prevention mode) and B (treatment mode) in the figure. Regardless of the prevention mode or the treatment mode, total secondary glycosides have a significant improvement effect on various lung function indicators of mice, and many indicators can be restored to the level of normal animals. Moreover, the improvement effect of total secondary glycosides on the FRC (functional residual capacity), PEF (maximum expiratory flow rate), RI (airway resistance) and Cdyn (dynamic lung compliance) of model animals is significantly better than that of nintedanib (p<0.05). The curve diagram between the expiratory flow rate and the expiratory volume of PF mice, such as Fig.28 A (prevention mode) and B (treatment mode) in the figure intuitively present the improvement effect of total secondary glycosides on the lung function of model animals.

[0211] Micro-CT scanning image results of mouse lung tissue ( Fig.29 ) showed that there were large areas of dense fibrous lesions (high-density images) in the lungs of mice in the model group. In both the prevention and treatment modes, total secondary glycosides significantly reduced the fibrotic lesions in the lung tissues of PF mice compared with the model group.

[0212] H&E staining of mouse lung tissues showed Fig.30 As shown, in both the prevention and treatment modes, total secondary glycosides significantly reduced inflammatory damage in the lungs of model animals compared with the model group.

[0213] The number of inflammatory cells in the bronchoalveolar lavage fluid of mice ( Fig.31 ) showed that in both the prevention and treatment modes, compared with the model group, total secondary glycosides significantly reduced the content of inflammatory cells such as lymphocytes, neutrophils, eosinophils and monocytes infiltrating the lungs of model animals (p<0.05).

[0214] Masson staining of mouse lung tissues was performed. Fig.32 As shown, total secondary glycosides significantly reduced the degree of lung fibrosis in model animals in both prevention and treatment modes (p<0.05).

[0215] Immunohistochemistry was used to detect the expression of fibrosis-related proteins in lung tissue. Fig.33 As shown in the figure, in both the prevention and treatment modes, total secondary glycosides significantly reduced the expression levels of fibronectin, collagen I and smooth muscle actin α-SMA in the lung tissue of PF mice (p<0.05 or p<0.01).

[0216] It can be seen from the results of the above Examples 2, 3, 4 and 5 that under the two dosage modes of preventing and treating pulmonary fibrosis, the total secondary ginsenoside composition, the PPT-type secondary ginsenoside composition and the PPD-type secondary ginsenoside composition can effectively prevent and treat pulmonary fibrosis, which is manifested by significantly prolonging the survival time of model animals, reducing the lung index of model animals, significantly improving the lung function of model animals, alleviating lung inflammation of model animals, and reducing the pathological damage and fibrosis degree of the lungs of mice.

[0217] The efficacy of PPT-type secondary ginsenoside composition and PPD-type secondary ginsenoside composition in treating pulmonary fibrosis is better than that of the listed drug pirfenidone, and at the effective dose for treating pulmonary fibrosis, the in vivo safety of PPT-type secondary ginsenoside composition and PPD-type secondary ginsenoside composition (100 mg / kg) is also better than that of pirfenidone (200 mg / kg). Moreover, the efficacy of total secondary ginsenoside composition in treating pulmonary fibrosis is better than that of the listed drug nintedanib.

[0218] Example 6 Secondary ginsenoside composition alleviates lipopolysaccharide (LPS)-induced acute lung injury

[0219] Lipopolysaccharide (LPS)-induced acute lung injury is a model that replicates acute lung injury caused by sepsis. It can destroy the vascular endothelial barrier and cause edema in the lung tissue space and alveoli, and is widely used by researchers. LPS is the main component of endotoxin and the main substance of acute lung injury induced by sepsis caused by Gram-negative bacteria. The present invention establishes an animal model of acute lung injury induced by a one-time tracheal instillation of LPS. The model is established by instilling LPS (5 mg / kg, Sigma) through the mouse trachea 2 hours after the last administration, and further verifies the protective effect of the secondary ginsenoside composition on lung injury.

[0220] Thirty C57BL / 6J male mice (6-8 weeks old) were randomly divided into a control group, a model group (LPS group), a total primary glycoside group (LPS + 50 mg / kg total primary ginsenoside composition), a total secondary glycoside group (LPS + 50 mg / kg total secondary ginsenoside composition) and an Rg5 group (LPS + 50 mg / kg Rg5) after one week of adaptive feeding, with 6 mice in each group, as shown in Table 9.

[0221] Table 9

[0222]

[0223] According to the dosage requirements of each group, the corresponding secondary ginsenoside drugs were dissolved in 0.5% sodium carboxymethyl cellulose (0.5% CMC-Na) aqueous solution. Starting from the 0th day of the experiment, the control group and the model group were gavaged with an equal amount of 0.5% CMC-Na aqueous solution every day, and each medication group was gavaged every day according to the set dose for 3 consecutive days. 2 hours after the last administration, after anesthetizing the mice, LPS saline solution was instilled through the trachea at a dose of 5 mg / kg to model acute lung injury (ALI) in mice, and after 6 hours of LPS tracheal instillation, the mice were anesthetized and the samples were taken; the control group was only anesthetized and the mice were taken. The detection indicators include: taking the mouse bronchoalveolar lavage fluid (BALF), centrifuging it, determining the total white blood cell count, using a cell flow cytometer to analyze the neutrophil ratio, and calculating the number of neutrophils; using the enzyme-linked immunosorbent assay (ELISA) method to detect the levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6) and myeloperoxidase (MPO) in the BALF supernatant to evaluate inflammatory mediators; and determining the total protein concentration in the BALF supernatant. The Western blot method was used to analyze the tight junction protein ZO-1 in the mouse lung tissue and the inflammatory-related protein cyclooxygenase COX-2 and TNF-α protein in the nuclear transcription factor NF-κB signaling pathway, and to investigate the degree of damage to the mouse alveolar capillary endothelial barrier and the molecular mechanism of anti-inflammatory effects. The experimental results are as follows:

[0224] The total number of leukocytes and neutrophils in the bronchoalveolar lavage fluid (BALF) of mice Fig.34 ) The results showed that compared with the control group, the number of total white blood cells and neutrophils in the BALF of the model group mice was significantly increased (p<0.01). After intervention with the total secondary ginsenoside composition, the number of white blood cells (p<0.01) and neutrophils (p<0.05) in the BALF of ALI mice were significantly reduced; while the total primary ginsenoside composition could only reduce the number of white blood cells in the BALF of ALI mice (p<0.05). This shows that the total secondary ginsenoside composition is better than the total primary ginsenoside composition in intervening in the neutrophils and total white blood cells in the bronchoalveolar lavage fluid of ALI mice.

[0225] TNF-α, IL-1β, and IL-6 are typical pro-inflammatory factors in various inflammatory diseases. In order to further evaluate the anti-inflammatory activity of total secondary ginsenosides, the contents of TNF-α, IL-1β, and IL-6 in mouse bronchoalveolar lavage fluid were detected according to the ELISA kit operating instructions. The results are as follows Fig.35As shown: After LPS tracheal instillation modeling, the levels of TNF-α, IL-6 and IL-1β in the bronchoalveolar lavage fluid of mice were significantly increased (p<0.0001). After intervention with total secondary ginsenosides, the levels of TNF-α, IL-6 and IL-1β in the bronchoalveolar lavage fluid of mice were significantly reduced (p<0.05). In particular, myeloperoxidase (MPO) is mainly present in neutrophils and is released into the extracellular fluid during inflammatory reactions. The large-scale aggregation of neutrophils can lead to changes in tissue and cell permeability and cause tissue damage. The results of this experiment ( Fig.35 ) showed that compared with the control group, the MPO content in the bronchoalveolar lavage fluid of the model group mice was significantly increased (p<0.001). After intervention with total secondary ginsenosides, the MPO content in the bronchoalveolar lavage fluid of mice was significantly decreased (p<0.05).

[0226] According to the total protein concentration in mouse bronchoalveolar lavage fluid ( Fig.36 ) The results showed that compared with the control group, the total protein concentration of bronchoalveolar lavage fluid in the model group was significantly increased (p<0.0001). After intervention with total secondary ginsenosides, the total protein concentration of bronchoalveolar lavage fluid in mice was significantly decreased (p<0.05).

[0227] Based on the expression of tight junction protein ZO-1, cyclooxygenase protein COX-2, phosphorylated nuclear transcription factor NF-κB p-p65 and total p65 protein and tumor necrosis factor TNF-α in mouse lung tissue by western blot ( Fig.37 ) The results showed that compared with the control group, the tight junction protein ZO-1 in the lung tissue of the model group mice was significantly decreased (p<0.01), indicating that the tight junction of the alveolar vascular endothelial barrier of the model group mice was severely damaged. Compared with the model group, after intervention with total secondary ginsenosides, the expression level of tight junction protein ZO-1 in the lung tissue of mice was significantly upregulated (p<0.001), indicating that total secondary ginsenosides can promote the tight junction of the alveolar vascular endothelial barrier and provide barrier protection. Compared with the control group, the nuclear transcription factor NF-κBp-p65 protein (p<0.001) and its regulated downstream inflammatory-related proteins cyclooxygenase COX-2 (p<0.0001) and tumor necrosis factor TNF-α (p<0.0001) proteins in the model group mice were significantly increased, indicating that the NF-κB signaling pathway in the lung tissue of the model group mice was activated. Both total primary ginsenosides and total secondary ginsenosides significantly inhibited the expression levels of NF-κB p-p65, cyclooxygenase COX-2 and tumor necrosis factor TNF-α protein in lung tissue; and the effect of total secondary ginsenosides in inhibiting TNF-α protein was significantly better than that of total primary ginsenosides (p<0.001). This indicates that total primary ginsenosides and total secondary ginsenosides exert anti-inflammatory effects by regulating the NF-κB signaling pathway.

[0228] Example 7 Safety Assessment of Secondary Ginsenoside Compositions

[0229] Safety assessment: 16 male C57BL / 6J mice (8-10 weeks old) were randomly divided into 2 groups (n=8), named blank control group and total secondary ginsenoside composition (100 mg / kg) administration group. The experiment used 0.5% sodium carboxymethyl cellulose (0.5% CMC-Na) aqueous solution as solvent, and the experiment was conducted by continuous gavage for 14 days (i.e., gavage once a day from the 1st day to the 14th day). The blank control group was gavaged with an equal amount of blank solvent. During the experiment, the weight of mice was measured every day, and samples were collected after anesthesia on the second day after the last administration, including: mouse serum, heart, liver, spleen, lungs and kidneys. The changes in mouse body weight during the administration period were plotted, and the indexes of various mouse organs were calculated. The organ tissues of the two groups of mice were fixed with 0.4% paraformaldehyde and further stained with H&E. At the same time, the levels of aspartate aminotransferase AST, alanine aminotransferase ALT and blood creatinine CRE in mouse serum were measured with the help of a kit to evaluate the effects of the total secondary ginsenoside composition on the liver and kidney functions of mice.

[0230] The experimental results are as follows Fig.38 As shown, the weight changes of mice ( Fig.38 A) in the figure shows that there was no significant difference in the body weight change between the blank control group and the total secondary ginsenoside composition (100 mg / kg) group during the entire administration period; the organ indexes of the two groups of mice ( Fig.38 B) in the figure shows that there is no significant difference in the organ indexes of mice in the blank control group and the total secondary ginsenoside composition (100 mg / kg) group, among which the heart index p = 0.5321, liver index p = 0.7409, spleen index p = 0.9910, lung index p = 0.9999, and kidney index p = 0.765; the H&E staining results of the organs of the two groups of mice ( Fig.38 C) shows that the morphology of various organs of mice in the total secondary ginsenoside composition (100 mg / kg) group did not show any difference compared with the blank control group; the results of the serum aspartate aminotransferase AST, alanine aminotransferase ALT and blood creatinine CRE in the two groups of mice ( Fig.38 DF in the data showed that there was no significant difference in AST, ALT and CRE in the serum of mice between the blank control group and the total secondary ginsenoside composition (100 mg / kg) group, among which AST (p=0.7333), ALT (p=0.9838) and CRE (p=0.9951). The safety evaluation results showed that the total secondary ginsenoside composition had good safety in animals.

[0231] The above-mentioned embodiments only express several implementation methods of the present invention, and the description is relatively specific and detailed, but it cannot be understood as limiting the scope of the invention patent. It should be pointed out that for ordinary technicians in this field, several modifications and improvements can be made without departing from the concept of the present invention, which all belong to the protection scope of the present invention.

Claims

1. A secondary ginsenoside composition comprising any two or more components selected from the following components 1) to 2): 1) PPT type secondary ginsenosides: (20S)-Rg2, (20S)-Rh1, (20R)-Rg2, (20R)-Rh1, Rg6, F4, Rk3, Rh4; 2) PPD type secondary ginsenosides: (20S)-Rg3, (20R)-Rg3, Rk1, Rg5, (20S)-Rh2, (20R)-Rh2, Rk2, Rh3.

2. The secondary ginsenoside composition according to claim 1, comprising: all the components shown in 1); and / or all the components shown in 2).

3. The secondary ginsenoside composition according to claim 1 or 2, wherein: The components listed in 1) to 2) account for 50% or more by mass in the secondary ginsenoside composition, and more preferably 70% or more by mass.

4. A method for preparing a secondary ginsenoside composition, comprising: Using at least one of the native ginsenoside composition, the PPT-type native ginsenoside composition, and the PPD-type native ginsenoside composition as raw materials, only the C-20 sugar in the raw materials and the C-20 alcoholic hydroxyl group of the hydrolysis product are hydrolyzed and further dehydrated to obtain a product to obtain a secondary ginsenoside composition.

5. The method for preparing the secondary ginsenoside composition according to claim 4, wherein: The hydrolysis conditions are: acidic conditions at 70-90° C.; preferably, the acidic conditions are provided by a mixed solution of methanol and 0.2±0.02N hydrochloric acid solution in a volume ratio of 1:0.8-1.

2.

6. The method for preparing the secondary ginsenoside composition according to claim 4 or 5, wherein: The method further comprises: after the hydrolysis reaction is completed, using n-butanol to extract the secondary ginsenoside composition.

7. The method for preparing the secondary ginsenoside composition according to claim 4, wherein: The main components of the native ginsenoside composition include Rg1, Re, Rf, Rb1, Rc, Rb2, Rb3 and Rd, and the content of the main components in the native ginsenoside composition is more than 70%.

8. The method for preparing the secondary ginsenoside composition according to claim 4 or 7, wherein: The PPT-type native ginsenoside composition and the PPD-type native ginsenoside composition are separated from the native ginsenoside composition by medium pressure liquid chromatography; In the medium pressure liquid chromatography technology, water is used as mobile phase A, methanol is used as mobile phase B, the flow rate is 50±5mL / min, and a linear gradient elution method is adopted: 0-5min, 10% mobile phase B; 5-45min, 10%-100% mobile phase B; 45-55min, 100% mobile phase B.

9. A secondary ginsenoside composition, prepared by the preparation method according to any one of claims 4 to 8.

10. Use of the secondary ginsenoside composition according to any one of claims 1 to 3 and 9 in the preparation of medicines or foods, wherein: The medicine or food is used to prevent, treat or alleviate at least one disease selected from pulmonary fibrosis and acute lung injury.

11. The use according to claim 10, wherein: The medicine or food is used for at least one of the following aspects: i) improving the survival rate of an individual and prolonging the survival period; ii) reducing the lung index; iii) improving lung function; iv) improving lung imaging conditions; v) reducing the degree of alveolar structure lesions; vi) reduce the degree of inflammatory cell infiltration in the lungs; preferably, the inflammatory cells include at least one of total leukocytes, lymphocytes, neutrophils, eosinophils, monocytes and macrophages; vii) reduce the degree of pulmonary fibrosis; viii) reduce collagen deposition in lung tissue; ix) reduce the hydroxyproline content in lung tissue; x) reduce the expression of fibrosis-related proteins in lung tissue; preferably, the fibrosis-related proteins include fibronectin, collagen I and smooth muscle actin α-SMA; xi) reducing the total white blood cell count and neutrophil count in alveolar lavage fluid; xii) reducing the expression of proinflammatory factors in alveolar lavage fluid; preferably, the proinflammatory factors include at least one of TNF-α, IL-1β, and IL-6; xiii) reducing the myeloperoxidase content in alveolar lavage fluid; xiv) reducing the total protein concentration in alveolar lavage fluid, increasing the expression of tight junction protein ZO-1 in lung tissue, and providing alveolar vascular endothelial barrier protection; xv) inhibiting the activation of the nuclear transcription factor NF-κB signaling pathway in the lung, and reducing the expression of downstream inflammatory-related protein COX-2 and proinflammatory factor TNF-α regulated by the NF-KB signaling pathway.

12. The use according to claim 10 or 11, wherein: The dosage form of the drug is an oral dosage form.

13. A medicine comprising the secondary ginsenoside composition according to any one of claims 1 to 3 and 9.

14. The drug according to claim 13, wherein The dosage form of the drug is an oral dosage form.

15. A food comprising the secondary ginsenoside composition according to any one of claims 1 to 3 and 9.

Images