Composition and medicine for relieving drug-induced liver injury as well as preparation method and application of composition and medicine

Abstract

The invention relates to the field of biological medicines, and discloses a composition and a medicine for relieving drug-induced liver injury as well as a preparation method and application of the composition and the medicine. The composition for relieving the drug-induced liver injury is prepared from clostridium praeparatum, mannan oligosaccharide, PQQ, IP6 and isogenistein. The inner layer structure of the medicine for relieving the drug-induced liver injury is used for embedding clostridium praeparatum and mannan oligosaccharide, and the outer layer structure of the medicine comprises PQQ, IP6 and isogenistein. The inner layer structure is formed by coating sodium alginate gel microspheres with chitosan; the outer layer structure is composed of hydroxypropyl methyl cellulose and sodium alginate. The survival rate of the composition for APAP-induced human hepatocyte injury reaches 85.5%, and hepatocyte injury and inflammatory response can be effectively relieved. The microcapsule structure enables the release rate of PQQ in gastric juice to be lower than 20%, the release rate of probiotics in gastric juice to be lower than 10%, and the total release rate of probiotics in intestinal segments to be higher than 90%, so that accurate delivery and stable protection of medicinal components are realized.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine, and more specifically, to a composition for alleviating drug-induced liver injury, a drug, and a preparation method and application thereof. Background Art

[0002] Drug-induced liver injury (DILI) is a common clinical condition, yet prevention and treatment options remain limited. Its occurrence is associated with a variety of medications, including commonly used antipyretics and analgesics (such as acetaminophen), antibiotics, anti-tumor drugs, immunosuppressants, traditional Chinese medicines, and health supplements. In my country, DILI has become one of the most common types of unexplained liver injury. The clinical manifestations of DILI range from mild transaminase elevations to severe acute liver failure, and are characterized by significant individual variability, difficult diagnosis, and a lack of specific therapeutic agents.

[0003] Current treatment strategies for DILI primarily include drug discontinuation, supportive care, and intervention with antioxidants (such as N-acetylcysteine). However, these approaches are primarily symptomatic and lack effective intervention mechanisms specific to the pathogenesis of DILI, limiting their effectiveness in clinical practice. Some natural products have been explored as adjunctive therapies for DILI. For example, CN108653267A discloses the use of dihydroquercetin for ameliorating drug-induced liver injury, primarily through regulating glutathione, ALT / AST, and other indicators to achieve liver protection. CN112773813A discloses the use of sola gum in DILI, aiming to alleviate acute liver tissue damage by reducing inflammatory cytokines.

[0004] On the other hand, the application of intestinal microecological regulation in the treatment of liver disease has gradually become a research hotspot. After the intestinal-liver axis theory was proposed, the role of probiotics and their metabolites in liver health has attracted attention. CN113993529A discloses a butyrate-producing bacterial composition, including Bifidobacterium, Lactobacillus, etc., which aims to improve liver metabolism and inflammatory status and has a certain positive effect on liver function. However, most current probiotic combinations are still focused on the superposition of traditional strains, with similar mechanism pathways, and the effects are mainly reflected in chronic metabolic liver damage, which is difficult to cope with the rapid cytotoxic process of DILI.

[0005] In addition, in terms of the delivery method of functional ingredients, CN109621852A discloses a natural polymer double-layer structure carrier, while CN115154425A and CN109125726A respectively proposed pH-responsive nanoparticles and targeted controlled-release nanoparticles. Although they have certain structural hierarchies and controlled-release properties, they mostly use a single film-forming material such as sodium carboxymethyl cellulose or polyacrylamide. The structural and functional versatility is strong but the specificity is weak. They are mostly oriented to non-liver injury indications such as tumors, and it is difficult to meet the comprehensive needs of synergistic delivery of probiotics and small molecules, precise treatment of drug-induced liver injury and segmented intestinal release.

[0006] At present, no combination solution has been found for DILI that uses probiotics, small molecule inducers, anti-inflammatory / antioxidant ingredients in combination with a multi-layered controlled-release delivery system. Summary of the Invention

[0007] The present invention overcomes the above-mentioned lack of prior art solutions for DILI by using a combination of probiotics, small molecule inducers, and anti-inflammatory / antioxidant ingredients in combination with a multi-layered controlled-release delivery system to provide a composition for alleviating drug-induced liver injury. Another object of the present invention is to provide an application of a composition for alleviating drug-induced liver injury; Another object of the present invention is to provide a drug for alleviating drug-induced liver injury; Another object of the present invention is to provide a method for preparing a drug for alleviating drug-induced liver injury.

[0008] In order to solve the above technical problems, the technical solutions of the present invention are as follows: A composition for treating drug-induced liver injury, comprising the following components in parts by weight: Faecalibacterium prausnitzii ( Faecalibacterium prausnitzii ) 1-5 parts, mannooligosaccharide (MOS) 8-15 parts, pyrroloquinoline quinone (PQQ) 0.1-0.72 parts, inositol hexaphosphate (IP6) 1.8-5 parts and isoscoparin 0.5-1.8 parts.

[0009] Furthermore, the mass ratio of Faecalibacterium prausnitzii to mannooligosaccharide is 1-5:8-15, and the mass ratio of pyrroloquinoline quinone, phytic acid and isogenysin is 0.2-2:5-10:1-5.

[0010] Preferably, the mass ratio of Faecalibacterium prausnitzii to mannooligosaccharide is 1-3:10-15.

[0011] Furthermore, the viable count of Faecalibacterium prausnitzii is not less than 1×10 7 CFU / g.

[0012] An application of the composition for treating drug-induced liver injury is used for preparing a drug for treating drug-induced liver injury.

[0013] A drug for treating drug-induced liver injury, comprising the composition for treating drug-induced liver injury and a double-layer carrier; the inner layer structure of the double-layer carrier embeds Faecalibacterium prausnitzii and mannooligosaccharide, and the outer layer structure contains pyrroloquinoline quinone, phytic acid and isogenycin.

[0014] Furthermore, the inner layer structure is formed by cross-linking sodium alginate and calcium ions, and its surface is treated with chitosan to form a coating layer; the outer layer structure is composed of a membrane-forming liquid composed of hydroxypropyl methylcellulose and sodium alginate.

[0015] A method for preparing the drug for treating drug-induced liver injury comprises the following steps: S1, adding Faecalibacterium prausnitzii and mannooligosaccharide into sodium alginate solution to form a mixed suspension; S2, adding the mixed suspension dropwise to a calcium chloride solution for cross-linking to form gel microspheres; S3, transferring a number of gel microspheres into a chitosan solution and stirring to connect the gel microspheres and form a coating layer to obtain core particles; S4, dissolving pyrroloquinoline quinone, phytic acid and isogenin in hydroxypropyl methylcellulose and sodium alginate to obtain a film-forming solution; S5. After forming an outer coating on the surface of the inner core particles with a film-forming liquid, a drug for alleviating drug-induced liver injury is obtained.

[0016] Furthermore, the total concentration of Faecalibacterium prausnitzii and mannooligosaccharide in the mixed suspension is 5.5%~8% (w / v), and the total mass concentration of pyrroloquinoline quinone, phytic acid and isogenistein in the film-forming solution is 4.3%~7.2% (w / v).

[0017] Furthermore, the chitosan solution has a concentration of 0.8% to 1% (w / v), and is taken out after soaking for 1 to 3 hours.

[0018] Preferably, the concentration of the chitosan solution is 1% (w / v) Furthermore, the mass ratio of the coating layer to the core particles is 0.1~0.15:1.

[0019] Preferably, the mass ratio of the coating layer to the core particles is 0.15:1.

[0020] This invention is the first to combine Faecalibacterium prausnitzii and mannan oligosaccharides as core prebiotic factors, utilizing their high colonization in the ileum and butyrate production potential to synergistically regulate the gut-liver axis, combined with small molecules PQQ, IP6 and isogenycin to target liver damage from the aspects of antioxidant and anti-inflammatory, respectively, to construct a systematic, multi-pathway synergistic treatment plan.

[0021] The present invention combines mannooligosaccharide with Faecalibacterium prausnitzii. On the one hand, MOS can be F. prausnitzii Specific utilization, promoting its colonization and butyrate production, on the other hand F. prausnitzii The butyrate produced can effectively inhibit inflammatory factors (such as TNF-α) and EMT process related to drug-induced liver injury. F. prausnitzii Substituting other probiotics will not form the above metabolic interactions and pathway synergy, and the effect will be significantly reduced. This proves that the composition of the present invention has clear non-obviousness.

[0022] The present invention innovatively constructs a double-layer microcapsule delivery system. The inner layer is a gel structure formed by cross-linking sodium alginate and chitosan, which is used to encapsulate probiotics and mannan oligosaccharides. It can effectively protect the activity of probiotics in the gastric acid environment and delay their release, ensuring their directional release and colonization in the distal small intestine, such as the ileum. The outer coating film is composed of a composite of HPMC and sodium alginate, forming a stable controlled-release structure. In the stomach, HPMC is insoluble, and sodium alginate undergoes acidic contraction, together forming a dense protective film that inhibits the premature release of small molecule active substances; after entering the jejunum, the increase in pH causes HPMC to swell, sodium alginate to gradually degelatinize, the membrane layer disintegrates, and the small molecules are quickly released and absorbed. This structural design successfully achieves the functional zoning control of "small molecule release in the jejunum + probiotic release in the ileum", significantly improving the bioavailability of various components and the synergistic intervention effect.

[0023] Compared with the prior art, the beneficial effects of the technical solution of the present invention are: 1. Effectively alleviate drug-induced liver injury. In an acetaminophen (APAP)-induced human hepatocyte HepG2 injury model, the present invention significantly increased cell viability at three concentration gradients, with a survival rate of 85.5% at a 200-fold dilution, demonstrating that the composition of the present invention can effectively alleviate drug-induced liver injury.

[0024] 2. Significant improvement in liver function indicators. In the mouse DILI model, the present invention significantly reduced ALT and AST levels, with a 24-hour average ALT level of 108.7 U / L. TNF-α concentration was also kept at a minimum level (84.3 pg / mg at 24 hours), demonstrating that the present invention can effectively alleviate liver cell damage and inflammatory responses.

[0025] 3. Excellent controlled-release performance. Simulated gastrointestinal release experiments showed that the release rate of PQQ in simulated gastric fluid (pH 1.5) was less than 20%, and the release rate of probiotics was less than 10%. Small molecules such as PQQ were rapidly released in simulated jejunum fluid (pH 4.5), while the release of probiotics was highest in simulated ileum fluid (pH 7.0). The total release rate exceeded 90%, achieving precise delivery and stable protection of the active ingredients. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 This is a comparison of the cell activities of different compositions on APAP-induced HepG2 cell damage; Figure 2 The effects of the examples and comparative examples on ALT levels in mouse liver; Figure 3 The effects of the examples and comparative examples on AST levels in mouse liver; Figure 4 The following are the effects of the examples and comparative examples on the TNF-α level in mouse liver tissue. DETAILED DESCRIPTION

[0027] The present invention will be further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0028] Unless otherwise specified, the reagents and materials used in the following examples were commercially available.

[0029] In the following embodiments, the Faecalibacterium prausnitzii was purchased from Zhili Zhongte (Wuhan) Biotechnology Co., Ltd. Example 1

[0030] Prepare the components of the composition in the following proportions: 3 parts of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1 × 10 7 CFU / g), 8 parts of mannooligosaccharide, 0.6 parts of pyrroloquinoline quinone, 4.8 parts of phytic acid, and 1.8 parts of isocyanin.

[0031] 1. Preparation of Core Particles Add 3 g of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1 × 10 7 CFU / g) and 8 g of mannooligosaccharide were stirred evenly to form a mixed suspension. The mixed suspension was slowly dripped into 200 mL of 2% CaCl2 solution and allowed to stand at room temperature for 30 minutes to form preliminary cross-linked gel microspheres. Several gel microspheres were filtered out and immersed in 0.8% chitosan solution at 15 mg per unit. The solution was soaked at room temperature for 1 hour to form a coating layer. After removal, the core particles were lightly washed with sterile water.

[0032] 2. Preparation and coating of outer film-forming liquid To 50 mL of 4% (w / v) hydroxypropyl methylcellulose, add 50 mL of 1% sodium alginate solution and mix. Then, add 0.6 g of pyrroloquinoline quinone, 4.8 g of phytic acid, and 1.8 g of isogenin, stirring until completely dissolved, to prepare a film-forming solution. Place the core particles in a petri dish and drip the film-forming solution onto their surface. Dry at 35°C for 30 minutes, repeating three cycles. After coating, microcapsules of the drug were obtained (mass ratio of coating layer to core particles was approximately 0.15:1). Example 2

[0033] Prepare the components of the composition in the following proportions: 1 part of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1 × 10 7 CFU / g), 15 parts of mannooligosaccharide, 0.1 parts of pyrroloquinoline quinone, 5 parts of phytic acid, and 0.5 parts of isocyanin.

[0034] 1. Preparation of Core Particles Add 1 g of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1 × 10 7 CFU / g) and 15 g of mannooligosaccharide were stirred evenly to form a mixed suspension. The mixed suspension was slowly dripped into 200 mL of 2% CaCl2 solution and allowed to stand at room temperature for 30 minutes to form preliminary cross-linked gel microspheres. Several gel microspheres were filtered out and immersed in 1% chitosan solution at a rate of 15 mg each. The solution was then soaked at room temperature for 2 hours to form a coating. After removal, the core particles were gently washed with sterile water to obtain the core particles.

[0035] 2. Preparation and coating of outer film-forming liquid To 50 mL of 4% (w / v) hydroxypropyl methylcellulose, add 50 mL of 1% sodium alginate solution and mix. Then, add 0.1 g of pyrroloquinoline quinone, 5 g of phytic acid, and 0.5 g of isogenin, stirring until completely dissolved, to prepare a film-forming solution. Place the core particles in a petri dish and drip the film-forming solution onto their surface. Dry at 35°C for 30 minutes, repeating three cycles. After coating, microcapsules of the drug were obtained (mass ratio of coating layer to core particles was approximately 0.12:1). Example 3

[0036] Prepare the components of the composition in the following proportions: 5 parts of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1×10 7 CFU / g), 10 parts of mannooligosaccharide, 0.72 parts of pyrroloquinoline quinone, 1.8 parts of phytic acid, and 1.8 parts of isocyanin.

[0037] 1. Preparation of Core Particles Add 5 g of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1 × 10 7 CFU / g) and 10 g of mannooligosaccharide were stirred evenly to form a mixed suspension. The mixed suspension was slowly dripped into 200 mL of 2% CaCl2 solution and allowed to stand at room temperature for 30 minutes to form preliminary cross-linked gel microspheres. Several gel microspheres were filtered out and immersed in 1% chitosan solution at a rate of 15 mg each. The solution was then soaked at room temperature for 3 hours to form a coating. After removal, the core particles were gently washed with sterile water to obtain the core particles.

[0038] 2. Preparation and coating of outer film-forming liquid To 50 mL of 4% (w / v) hydroxypropyl methylcellulose, add 50 mL of 1% sodium alginate solution and mix. Then, add 0.72 g of pyrroloquinoline quinone, 1.8 g of phytic acid, and 1.8 g of isogenin, stirring until completely dissolved, to prepare a film-forming solution. Place the core particles in a petri dish and drip the film-forming solution onto their surface. Dry at 35°C for 30 minutes, repeating three cycles. After coating, microcapsules of the drug are obtained (mass ratio of coating layer to core particles is approximately 0.1:1). Comparative Example 1

[0039] A composition was prepared according to the proportions of the components in Example 1, except that Faecalibacterium prausnitzii was replaced by Bifidobacterium.

[0040] The technical solution for preparing the microcapsule drug is similar to that in Example 1, except that Faecalibacterium prausnitzii is replaced by Bifidobacterium. Comparative Example 2

[0041] A composition was prepared according to the proportions of the components in Example 1, except that mannooligosaccharide was replaced by chitosan oligosaccharide. Comparative Example 3

[0042] A composition was prepared according to the proportions of the components in Example 1, except that isozygotone was not added. Comparative Example 4

[0043] Prepare the composition according to the following proportions: 0.5 parts of Faecalibacterium prausnitzii freeze-dried powder (viable cell count ≥ 1 × 10 7 CFU / g), 18 parts of mannooligosaccharide, 0.1 parts of pyrroloquinoline quinone, 3 parts of phytic acid, and 0.5 parts of isocyanin. Comparative Example 5

[0044] The technical solution for preparing the microcapsule drug is similar to that of Example 1, except that the freeze-dried powder of Faecalibacterium prausnitzii, mannooligosaccharide, pyrroloquinoline quinone, phytic acid, and isogenycin are mixed simultaneously to prepare the core particles, and no outer coating treatment is performed. Comparative Example 6

[0045] The technical solution for preparing microcapsule drugs is similar to that of Example 1, except that 15 mg of gel microspheres are immersed in 0.6% chitosan solution and soaked at room temperature for 3.5 hours. Comparative Example 7

[0046] The technical solution for preparing the microcapsule drug is similar to that of Example 1, except that the mass ratio of the coating layer to the core particles is approximately 0.05:1. Detection method

[0047] 1. APAP-induced liver injury model An APAP-induced drug-induced liver injury model was established using the human hepatocyte cell line HepG2. Specifically, HepG2 cells were seeded in a 96-well plate during the logarithmic growth phase. After cell attachment, acetaminophen (APAP) was added at a final concentration of 15 mM for 24 hours to induce injury.

[0048] The compositions of Example 1 and Comparative Examples 1-4 were prepared as stock solutions at a concentration of 100 mg / mL and diluted at 1:100, 1:200, and 1:400, respectively, to treat an APAP-induced HepG2 liver injury model. Cell viability in each group was assessed using the CCK-8 assay, with the absorbance (OD) value reflecting the change in viability.

[0049] 2. Mouse Acute DILI Model SPF-grade C57BL / 6 female mice (weighing 25-35 g) were selected and grouped to adapt to feeding. Acetaminophen (APAP) was injected intraperitoneally (300 mg / kg) to establish an acute liver injury model.

[0050] Example 1 and Comparative Examples 1 and 5 served as intervention groups. They were administered 100 mg / kg via gavage for 5 consecutive days before modeling and again on the 6th day after modeling. Blood was collected from the eyeballs 12 and 24 hours after modeling. Serum samples were used to measure alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels; liver tissue homogenates were assayed for TNF-α levels using ELISA to assess the extent to which the drug improves hepatocyte damage and systemic inflammatory responses.

[0051] 3. Release experiment at different pH 200 mg of microcapsules were added to 10 mL of pH 1.5 simulated solution and incubated in a 37°C water bath with gentle shaking for 2 h. Filtered through a sterile filter membrane, the filtrate was collected for PQQ and probiotic release. The filter residue (containing residual microcapsules) was transferred to 10 mL of pH 4.5 simulated solution and incubated in a 37°C shaker for another 2 h. Filtered again, the filtrate was collected for PQQ and probiotic release. The filter residue was again transferred to 10 mL of pH 7.0 simulated solution and incubated in a 37°C shaker for another 2 h. Filtered again, the filtrate was collected for PQQ and probiotic release. 200 mg of microcapsules were added to 10 mL of sterile PBS and thoroughly disrupted by ultrasonication or mechanical homogenization. The filtrate was collected for PQQ and probiotic release. Release rate = (released amount / total microcapsule loading) × 100%.

[0052] 4. Synergy effect assessment To validate the synergistic effect of the prebiotic factor combination (Faecalibacterium prausnitzii and mannooligosaccharides) and the small molecule combination (PQQ, IP6, and isogenistein) in alleviating drug-induced liver injury, the Chou-Talalay assay was used to evaluate the cytoprotective effect of their combined intervention. A human hepatocyte HepG2 cell model was constructed with APAP-induced DILI. Prebiotic factor and small molecule groups were treated individually and in combination, and cell viability recovery was measured at different concentrations in each group (CCK-8 assay).

[0053] Based on the distribution ratio of each group in Example 1, the IC values ​​of the prebiotic factor group and the small molecule combination were determined. 50 , and press IC 50 The concentration was set to 1 / 4, 1 / 2, 1 times and other gradient concentrations, and then mixed at a 1:1 ratio to set the combined treatment group. The cell survival rate of each group was recorded, and the Combination Index (CI) was calculated by the following formula: CI = (D1) / (D x1 ) + (D2) / (D x2 ). D1 and D2 are the doses used in each group of combined medication, D x1 、D x2 The dose required to achieve the same effect as a single drug. CI < 1 indicates synergy, CI = 1 indicates addition, and CI > 1 indicates antagonism. Analysis

[0054] 1. Cell survival rate is significantly improved according to Figure 1The results show that Example 1 of the present invention exhibited significant cytoprotective effects in the acetaminophen (APAP)-induced HepG2 cell injury model, superior to the various comparative examples. Specifically, at three dilution concentrations, the cell viability of Example 1 ranged from 74.2% to 85.5%, significantly higher than that of the model group (approximately 28%) and also higher than the survival rates of Comparative Examples 1 to 4 at the corresponding concentrations. While Comparative Examples 1 and 2 showed some improvement, they were significantly lower than those of Example 1 at all three concentration gradients, indicating that the synergistic pairing of Faecalibacterium prausnitzii and mannooligosaccharide is key to maintaining the efficacy of the present composition. Substituting Bifidobacterium for the probiotic in Comparative Example 1 resulted in butyrate deficiency, deregulation of EMT, and inflammatory pathways, significantly reducing the efficacy. Because the anti-inflammatory and antioxidant activities of chitosan oligosaccharides rely on their inherent structure, they are unable to modulate the intestinal microbiome, provide specific support for Faecalibacterium prausnitzii, or establish a liver-protective pathway centered on butyrate. Therefore, the substitution of mannosan oligosaccharide for chitosan oligosaccharide in Comparative Example 2 failed to achieve the same cytoprotective effect. Comparative Example 3, in which no isogenisin was added, also showed a significant decrease in cell activity, suggesting that it plays an important role in antioxidant or anti-inflammatory pathways. Comparative Example 4 showed a severe imbalance in the ratio of active ingredients, resulting in poor cell protection. Overall, the results demonstrate that the composition of the present invention exhibits significant synergistic advantages in terms of ingredient selection and ratio, significantly improving the survival rate of APAP-induced hepatocytes and demonstrating its significant advantages in alleviating drug-induced liver injury.

[0055] 2. Biochemical indicators and inflammatory responses of drug-induced liver injury were significantly improved from Figure 2 As can be seen, ALT levels in the model group increased significantly at 12 and 24 hours, confirming the successful establishment of the model. ALT levels in the Example 1 group decreased significantly, reaching approximately 178.3 U / L and 108.7 U / L at 12 and 24 hours, respectively, demonstrating the composition's significant protective effect against hepatocellular damage. Although the formulation structure of Comparative Example 1 is similar to that of Example 1, the substitution of Faecalibacterium prausnitzii with Bifidobacterium prausnitzii results in a lack of a stable synergistic mechanism with mannooligosaccharides, preventing sustained efficacy. ALT levels were only 19% lower after 24 hours (255.8 U / L) compared to 12 hours (207.1 U / L), a significantly lower decrease than in Example 1. Comparative Example 5, lacking the protective outer film-forming coating and staged release mechanism, resulted in partial disintegration of the microcapsules in the stomach, leading to more pronounced issues such as interference between components, premature release of small molecules, and decreased probiotic activity. ALT values ​​were approximately 290.7 U / L and 238.7 U / L, respectively.

[0056] In the examples, the outer coating plays a role in layered release and protection of the active ingredients, allowing the timely release of small molecule antioxidant factors in the stomach and effective colonization of probiotics in the small intestine, while reducing mutual interference and early inactivation between components, thereby maximizing the synergistic effect of the drugs. AST testing further confirmed this conclusion ( Figure 3 The AST value of Example 1 was significantly lower than that of the two control groups at 12 hours and remained at the lowest level at 24 hours, indicating that it has a stronger inhibitory effect on liver cell necrosis and inflammatory factor release. This also confirms the targeting advantage of the double-layer microcapsules in the delivery route and the synergistic rationality of the component design.

[0057] from Figure 4 It can be seen that the level of inflammatory factors in the model group increased significantly, while that in Example 1 dropped to 84.3 pg / mg after 24 hours, which was significantly better than that in Comparative Examples 1 and 5 (both above 110 pg / mg). This shows that the combination of Faecalibacterium prausnitzii and MOS in the example, coupled with the layered delivery structure, can effectively reduce the release of pro-inflammatory factors and improve the inflammatory response associated with liver injury, reflecting the synergistic advantage of the present invention in the anti-inflammatory mechanism.

[0058] 3. Targeting and controlled release advantages of the delivery system In simulated gastrointestinal release experiments, Examples 1-3 all demonstrated clear segmented release characteristics and excellent controlled-release effects. The release of small molecules, such as PQQ, was strictly controlled within 20% at pH 1.5, indicating that the composite membrane composed of the outer HPMC and sodium alginate layer remained stable and did not disintegrate in the stomach. As the pH increased to 4.5 and 7.0, the outer membrane gradually swelled, and PQQ release increased significantly, demonstrating typical pH-responsive release behavior, meeting the "intestinal delivery" requirements for small molecule absorption (Table 1). Regarding probiotic release, the core structure of the examples released less than 10% of the probiotic in the stomach, effectively preventing acidic damage and ensuring that active bacteria reached the intestine before release, resulting in an overall release rate exceeding 90% (Table 2).

[0059] Table 1 Average release rate of PQQ (%)

[0060] Table 2 Average release rate of Faecalibacterium prausnitzii (%)

[0061] Comparative Examples 5-7, however, exhibit significant deficiencies. Although chitosan treatment was employed in Comparative Example 5, the outer coating was omitted. Both the small molecules and probiotics were encapsulated within the inner core, allowing gastric fluid to penetrate directly into the core structure. At pH 1.5, a significant amount of PQQ (up to 43%) was released, leading to premature leakage and the risk of acidic inactivation. While the probiotic release rate reached a high 31.5%, this was largely passive, and the CFU activity was far lower than that of the Examples, resulting in poor intestinal delivery. Comparative Example 6 employed a lower concentration of chitosan cross-linking, resulting in a less dense shell and weaker gastric barrier. While the release rate was slightly better than that of Comparative Example 5, premature PQQ release and bacterial leakage persisted. The extremely thin coating in Comparative Example 7, while providing some sustained-release capability, struggled to establish a pH-responsive barrier. Consequently, gastric PQQ and probiotic release remained higher than in the Examples, indicating a failure in controlled-release.

[0062] 4. Synergy According to the results in Table 3, the cell survival rates of the combined groups were significantly higher than those of the single-use groups, reaching 91.2%, 83.6% and 73.1% respectively, with corresponding CI values ​​of 0.46, 0.60 and 0.69, all less than 1, clearly showing a synergistic effect. In contrast, the cell survival rates of the probiotic factor group and the small molecule group were generally low. The prebiotic factor combination and the small molecule combination in the present invention have a good synergistic effect, and a strong protective effect can be achieved at a lower dose, which helps to reduce the dosage, improve biosafety and drug efficacy persistence, and highlight the structural advantages and functional synergy of the combination in the intervention of liver injury.

[0063] Table 3 Synergistic effect

[0064] Obviously, the above embodiments of the present invention are merely examples for the purpose of clearly illustrating the present invention, and are not intended to limit the embodiments of the present invention. Those skilled in the art will appreciate that other variations or modifications can be made based on the above description. It is not necessary and impossible to enumerate all embodiments here. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention shall be included within the scope of protection of the claims of the present invention.

Claims

1. A composition for alleviating drug-induced liver injury, characterized in that: The invention comprises the following components in parts by weight: 1-5 parts of Faecalibacterium prausnitzii, 8-15 parts of mannooligosaccharide, 0.1-0.72 parts of pyrroloquinoline quinone, 1.8-5 parts of phytic acid and 0.5-1.8 parts of isocyanidin.

2. The composition for alleviating drug-induced liver injury according to claim 1, characterized in that: The mass ratio of Faecalibacterium prausnitzii to mannooligosaccharide is 1-5:8-15, and the mass ratio of pyrroloquinoline quinone, phytic acid and isogenysin is 0.2-2:5-10:1-5.

3. The composition for alleviating drug-induced liver injury according to claim 1, characterized in that: The viable count of Faecalibacterium prausnitzii is not less than 1×10 7 CFU / g.

4. Use of the composition for alleviating drug-induced liver injury according to any one of claims 1 to 3, characterized in that: Used to prepare drugs for alleviating drug-induced liver injury.

5. A drug for alleviating drug-induced liver injury, characterized in that: It comprises the composition for alleviating acute drug-induced liver injury as described in any one of claims 1 to 3 and a double-layer carrier; the inner layer structure of the double-layer carrier encapsulates Faecalibacterium prausnitzii and mannooligosaccharide, and the outer layer structure contains pyrroloquinoline quinone, phytic acid and isogenycin.

6. The drug for alleviating drug-induced liver injury according to claim 5, characterized in that: The inner layer structure is formed by cross-linking sodium alginate and calcium ions, and its surface is treated with chitosan to form a coating layer; the outer layer structure is composed of a membrane-forming liquid composed of hydroxypropyl methylcellulose and sodium alginate.

7. A method for preparing the drug for alleviating drug-induced liver injury according to any one of claims 5 to 6, characterized in that: The following steps are involved: S1, adding Faecalibacterium prausnitzii and mannooligosaccharide into sodium alginate solution to form a mixed suspension; S2, adding the mixed suspension dropwise to a calcium chloride solution for cross-linking to form gel microspheres; S3, transferring a number of gel microspheres into a chitosan solution and stirring to connect the gel microspheres and form a coating layer to obtain core particles; S4, dissolving pyrroloquinoline quinone, phytic acid and isogenin in hydroxypropyl methylcellulose and sodium alginate to obtain a film-forming solution; S5. After forming an outer coating on the surface of the inner core particles with a film-forming liquid, a drug for alleviating drug-induced liver injury is obtained.

8. The method for preparing the drug for alleviating drug-induced liver injury according to claim 7, characterized in that: The total concentration of Faecalibacterium prausnitzii and mannooligosaccharide in the mixed suspension is 5.5%-8% (w / v), and the total mass concentration of pyrroloquinoline quinone, phytic acid and isogenistein in the film-forming solution is 4.3%-7.2% (w / v).

9. The method for preparing the drug for alleviating drug-induced liver injury according to claim 7, characterized in that: The chitosan solution has a concentration of 0.8% to 1% (w / v), and is taken out after soaking for 1 to 3 hours.

10. The method for preparing the drug for alleviating drug-induced liver injury according to claim 7, characterized in that: The mass ratio of the coating layer to the core particles is 0.1~0.15:1.

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