Water-soluble fullerene material and its application in preventing and treating liver injury / liver failure

Abstract

The present disclosure relates to a new water-soluble fullerene derivative and a preparation method thereof, and application of the water-soluble fullerene material in preventing and treating liver damage / liver failure. The fullerene material of the present disclosure can efficiently scavenge active free radicals, reduce liver oxidative stress, restore normal liver function indicators, reduce liver necrosis, and prevent and treat drug-induced liver damage / liver failure.

Description

Technical Field

[0001] This disclosure pertains to the fields of chemistry and biomedicine, and relates to a water-soluble fullerene material, its preparation method, and its application in the prevention and treatment of liver injury / liver failure. Background Technology

[0002] Drug-induced liver injury / failure is the most common condition, particularly induced by acetylphenol (APAP). APAP is one of the most commonly used medications worldwide for treating pain and fever. Overdose of APAP can lead to acute liver injury (ALI) and eventually acute liver failure (ALF). However, treatment options for APAP-induced hepatotoxicity are relatively limited. Clinically available drugs are limited, and the therapeutic window is narrow. Liver transplantation is an option to improve survival rates. However, due to the lack of suitable donor tissue and the lifelong immunosuppression associated with it, liver transplantation is not an ideal treatment intervention. Therefore, new treatment methods to protect the liver are urgently needed.

[0003] Fullerenes are allotropes of carbon. These substances refer to cage-like structures composed of carbon atoms, with C being the dominant element. 60 C 70 These materials, represented by [examples of materials mentioned earlier], possess a large conjugated electronic structure, exhibiting excellent reactive oxygen species (ROS) scavenging ability. They can also regulate mitochondrial function, reverse non-alcoholic fatty liver disease, and protect the liver, among other therapeutic effects. Previous studies have also found that fullerene tricarboxylic acids can act like SOD enzymes to combat oxidative stress, protecting cells from superoxide anion damage. Summary of the Invention

[0004] The purpose of this disclosure is to provide a novel water-soluble fullerene derivative, its preparation method, and its application.

[0005] Another object of this disclosure is to provide a pharmaceutical composition and method for preventing and / or treating liver injury / liver failure caused by a drug.

[0006] Specifically, this disclosure provides the following technical solutions:

[0007] The purpose of this disclosure is to provide a water-soluble fullerene derivative, wherein the water-soluble fullerene derivative is a fullerene isoperidinic acid derivative.

[0008] In one aspect of this disclosure, the ratio of fullerene molecules to isoperidine acid molecules in each molecule of fullerene isoperidine acid derivative is 1:(1-6), preferably 1:(1-4).

[0009] In one aspect of this disclosure, a method for preparing a water-soluble fullerene derivative is provided, the method comprising the following steps:

[0010] (1) Dissolve fullerene or endogenous metal-embedded fullerene in a solvent;

[0011] (2) Add isopiridine ester, and after purification, acid hydrolysis and drying, water-soluble carboxylated fullerene material is obtained.

[0012] In one aspect of this disclosure, the isoperidol ester is methyl isoperidol formate or tert-butyl isoperidol formate.

[0013] In one aspect of this disclosure, a method for preparing a water-soluble fullerene derivative is provided, wherein the preparation method includes the following steps:

[0014] (1) C 60 Dissolved in o-dichlorobenzene, dimethyl sulfoxide was added under inert gas protection, and the mixture was bubbled with oxygen to replace it.

[0015] (2) Add isoperidine ester, stir with oxygen, and concentrate to obtain a reddish-brown solid;

[0016] (3) The reddish-brown solid was separated by column chromatography to obtain the elution product;

[0017] (4) The elution product is hydrolyzed with hydrochloric acid-acetic acid or trifluoroacetic acid, centrifuged, the precipitate is collected, washed with dilute acid, neutralized, and finally dialyzed and freeze-dried to obtain the water-soluble fullerene derivative.

[0018] This disclosure provides a pharmaceutical composition, characterized in that the pharmaceutical composition comprises the above-mentioned water-soluble fullerene derivative.

[0019] In one aspect of this disclosure, the use of water-soluble fullerene derivatives and pharmaceutical compositions in the preparation of medicaments for the prevention and / or treatment of liver injury / liver failure is provided.

[0020] In one aspect of this disclosure, liver injury / liver failure is caused by any drug that can cause liver injury / liver failure, preferably, said drug includes one or more of acetylphenol, isoniazid, cyclosporine A, tetracycline, tripterygium wilfordii, concanavalin A, and BCG.

[0021] In one aspect of this disclosure, the prevention and / or treatment of liver injury / liver failure includes improving liver function, reducing the area of ​​liver necrosis, reducing the degree of liver necrosis, and improving one or more of the following liver biochemical indicators: preferably, improving liver function to normalize liver function indicator values; preferably, the liver function indicator values ​​are selected from one or more of the following indicators: alanine aminotransferase (ALT) and aspartate aminotransferase (AST).

[0022] In one aspect of this disclosure, liver biochemical parameters are improved to normalize one or more activity levels of GSH, MDA, and SOD.

[0023] In one aspect of this disclosure, the prevention and / or treatment of liver injury / liver failure includes one or more of the following: significantly reducing drug-induced hepatocyte death and significantly reducing intracellular oxidative stress levels.

[0024] In one aspect of this disclosure, water-soluble fullerenes may also include (1) fullerenes with hydrophilic groups modified on their surface; (2) fullerenes encapsulated by hydrophilic biomolecules; (3) fullerenes loaded with a biocompatible carrier material; and (4) water-soluble supramolecular fullerenes formed by self-assembly.

[0025] In one aspect of this disclosure, water-soluble fullerenes may also include water-soluble fullerene derivatives, water-soluble endogenous metal-encapsulated fullerene derivatives, compositions of water-soluble fullerene derivatives and water-soluble endogenous metal-encapsulated fullerene derivatives, etc.

[0026] Compared with the prior art, the beneficial effects of this disclosure are as follows:

[0027] 1) The purpose of making the raw material fullerene and the raw material endogenous metal fullerene water-soluble is to enable them to be rapidly transported to the liver in the body and exert their effects in the liver, thereby reducing the degree and area of ​​liver necrosis, normalizing liver function indicators (alanine aminotransferase ALT and aspartate aminotransferase AST), normalizing GSH, MDA and SOD activity indicators, thereby reducing the toxicity caused by drug-induced liver injury / liver failure.

[0028] 2) After the water-soluble fullerene structure of the active ingredient in this disclosure enters the liver in vivo, it retains its conjugated structure and thus has a good effect on scavenging free radicals. It can regulate the liver microenvironment and improve the level of oxidative stress in the liver.

[0029] 3) The water-soluble fullerene structure of the active ingredient in this disclosure has no obvious biological toxicity, can be rapidly metabolized in the body, and has no toxicity to the organs in the body. Attached Figure Description

[0030] Figure 1 Synthetic route for the water-soluble carboxylated fullerene material (TIFE) prepared in Example 1.

[0031] Figure 2 MALDI-TOF-MS image of the water-soluble carboxylated fullerene material (TIFE) prepared in Example 1.

[0032] Figure 3 The water-soluble carboxylated fullerene material (TIFE) prepared in Example 1 1 H-NMR spectrum.

[0033] Figure 4The FTIR image is of the water-soluble carboxylated fullerene material (TIFE) prepared in Example 1.

[0034] Figure 5 The image shows the electron spin nuclear magnetic resonance (ESR) pattern of the water-soluble carboxylated fullerene material (TIFE) in Example 2.

[0035] Figure 6 The cell viability of HepaRG hepatocytes in Example 3 after co-incubation with different concentrations of water-soluble carboxy-fullerene material (TIFE) for 24 hours.

[0036] Figure 7 The cell viability of HepaRG hepatocytes after oxidative stress induced by 500 μM hydrogen peroxide in Example 3 was obtained by co-incubation with different concentrations of water-soluble carboxylated fullerene material (TIFE) for 3 hours.

[0037] Figure 8 The cell viability of HepaRG hepatocytes in Example 3 after co-incubation with 5mM p-acetylphenol (APAP) and different concentrations of water-soluble carboxy-fullerene material (TIFE) for 24 hours.

[0038] Figure 9 Optical photographs of the livers of different groups of C57BL / 6J mice in Example 4.

[0039] Figure 10 The H&E test was performed on the livers of different groups of C57BL / 6J mice in Example 4.

[0040] Figure 11 The values ​​of ALT and AST in the serum of different groups of C57BL / 6J mice in Example 4 are shown.

[0041] Figure 12 Biochemical indicators of livers from different groups of C57BL / 6J mice in Example 4.

[0042] Figure 13 The changes in p-JNK protein expression in the livers of different groups of C57BL / 6J mice in Example 4 are shown. Detailed Implementation

[0043] I. Definition

[0044] As used in this disclosure, the term "treatment" includes its generally accepted meaning, which includes preventing, inhibiting, improving, and slowing down, stopping, or reversing the development of resulting symptoms or anticipated lesions. Accordingly, this disclosure covers both therapeutic and preventative applications.

[0045] As used in this disclosure, the terms "active ingredient," "active ingredient water-soluble fullerene structure," or "water-soluble fullerene structure" refer to at least one of a water-soluble fullerene, a water-soluble endogenous metal-encapsulated fullerene, a combination of a water-soluble fullerene and a water-soluble endogenous metal-encapsulated fullerene, a pharmaceutically acceptable ester of the above three, and a pharmaceutically acceptable salt of the above three.

[0046] As used in this disclosure, the term "effective amount" refers to the amount or dose of an active ingredient administered to a patient, whether once or multiple times, to provide the intended effect to the diagnosed or treated patient. The effective amount can be determined by the participating diagnostic physician based on knowledge of known techniques and observations obtained in similar circumstances. In determining the effective amount or dose of the administered active ingredient, the participating diagnostic physician should consider a variety of factors, including but not limited to: the species of mammal; size, age, and general health; the specific disease involved; the extent or severity of the disease; the individual patient's response; the specific compound administered; the administration method; the bioavailability properties of the administered formulation; the chosen dosing regimen; the use of concomitant drug therapies; and other relevant factors.

[0047] As used in this disclosure, "fullerene" is a hollow molecule composed entirely of carbon, in the form of spheres, ellipsoids, cylinders, or tubules. Fullerenes are structurally similar to graphite, which is composed of stacked layers of graphene composed of six-membered rings, while fullerenes contain not only six-membered rings but also five-membered rings, and occasionally seven-membered rings. Typically, the general formula for fullerenes is C1. 2n n is an integer, n≥10; in a preferred embodiment, 30≤n≤60; exemplary, the general formula of the fullerene is C 60 C 70 C 76 C 78 C 82 Or C 84 C is preferred. 60 C 70 Or C 84 More preferably, C 60 Or C 70 .

[0048] As used in this disclosure, "intercalated metallofullerene" refers to a metal atom completely enclosed within a closed carbon cage. Common metals used are primarily alkali metals, alkaline earth metals, and most rare earth metals. Intercalated metallofullerenes can contain one, two, or even three metal atoms, with the most common being those containing only one atom. Depending on the type and number of metal atoms within the cage, intercalated metallofullerenes can be represented by a general formula, such as M@C. 2p M2@C2p MA@C 2p M3P@C 2p M2C2@C 2p M2S@C 2p M2O@C 2p and M x A 3-x P@C 2p One or more of the following can be used to represent it, where M represents the metal atom in the cage; C 2p The symbol represents a fullerene molecule; 2p represents the number of carbon atoms, where p is an integer; preferably, 30 ≤ p ≤ 60, 0 ≤ x ≤ 3; @ indicates that the atom on the left is enclosed in the fullerene cage on the right, indicating that the metal atom is inside the carbon cage. M and A are each independently selected from metal elements. In a preferred embodiment, M and A are each independently selected from one or more of Fe, Zn, Cu, Mn, Co, Sr, Cr, Se, Sc, Y, and lanthanides; in another preferred embodiment, M and A are each independently selected from one or more of Sc, Y, and lanthanides.

[0049] As used in this disclosure, the term "water-soluble fullerene" refers to a water-soluble modified fullerene obtained by water-soluble modification of the fullerene matrix, wherein the fullerene may be of the general formula C0. 2n Fullerenes and / or endogenous metal-encapsulated fullerenes. Among them, water-soluble fullerene derivatives include one or more fullerenes selected from the group consisting of: (1) fullerenes with hydrophilic groups modified on their surface; (2) fullerenes encapsulated by hydrophilic biomolecules; (3) fullerenes supported by a biocompatible carrier material; and (4) water-soluble supramolecular fullerene systems formed by self-assembly.

[0050] The hydrophilic groups include one or more of piperidine, hydroxyl, carboxyl, thiol and water-soluble amino acid residues.

[0051] Wherein, water-soluble amino acid residues refer to incomplete amino acids remaining after a portion of the amino acid molecule is lost when a water-soluble amino acid is modified with fullerenes and / or intercalated with metallofullerenes. In other words, an amino acid residue is a part of an amino acid molecule, but it is an incomplete amino acid. The absence of any part of an amino acid molecule constitutes an amino acid residue, such as the loss of hydrogen from the amino group, hydrogen from the carboxyl group, or a hydroxyl group. Optionally, the water-soluble amino acid residue is at least one of alanine, glycine, serine, arginine, lysine, and aspartic acid residues.

[0052] Among them, hydrophilic biomolecules include at least one of isoperidinic acid, cyclodextrin, polyethylene glycol, and amino acids.

[0053] The fullerene with hydrophilic groups modified on its surface is selected from one or more of fullerene amino acid derivatives, fullerene hydroxylated derivatives, fullerene carboxylated derivatives, fullerene carboxylic acid esterified derivatives, fullerene thiolated derivatives, and fullerene pyrrolidine derivatives. The fullerene carboxylated derivatives include one or more carboxylated fullerenes; in a preferred embodiment, the carboxyl group is an isoperidinic acid group and a malonic acid group.

[0054] In the case of the fullerene encapsulated by hydrophilic biomolecules, the hydrophilic biomolecules may be selected from piperidine and / or amino acids.

[0055] In the case of the fullerene loaded with a biocompatible carrier material, the biocompatible carrier material may be selected from liposomes and / or cell membranes.

[0056] II. Examples

[0057] The specific embodiments of this disclosure will be described in detail below with reference to the accompanying drawings, but it should be understood that the scope of protection of this disclosure is not limited to the specific embodiments. The raw material C used in the following embodiments... 60 Purchased from Xiamen Funa New Technology Co., Ltd., with a molecular weight of 720 and a purity of 99.9%.

[0058] Example 1: Preparation and characterization of water-soluble carboxylated fullerenes (TIFE)

[0059] Synthetic routes for water-soluble carboxylated fullerenes (TIFE), such as Figure 1 As shown, 100 mg of C 60 Dissolved in 20 mL of o-dichlorobenzene, 5 mL of dimethyl sulfoxide was added under argon protection, and the mixture was bubbled with oxygen for 5 min. 120 mg of methyl isoperidine carboxylate was added, and the mixture was stirred with oxygen for 12 h. The solution was concentrated to obtain a reddish-brown solid. The solid was separated by toluene dissolution column chromatography, with 20% ethyl acetate-toluene as the eluent. The product was concentrated to dryness, and 100 mg of the product was dissolved in 50 mL of toluene. This solution was reacted with an excess of HCl:CH3COOH (3:1) at 120 °C for 24 h. The product was centrifuged (10000 rpm, 5 min), and the precipitate was collected. The precipitate was washed three times with deionized water, and the pH was adjusted to neutral with a low-concentration sodium hydroxide solution. After dialyzing and freeze-drying, water-soluble carboxylated fullerene was obtained.

[0060] A small amount of the solid product was dissolved in ultrapure water, and its molecular ion peak was detected using matrix-assisted laser desorption / ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The corresponding molecular ion peak was detected in negative ion mode. Figure 2 (As shown); 5 mg of the solid product was dissolved in deuterated water, and its composition was detected using a 400 MHz nuclear magnetic resonance spectrometer. 1 H-NMR ( Figure 3 (As shown); Finally, a small amount of the solid product was dissolved in ultrapure water, dropped onto a gold mirror, dried to form a film, and its infrared absorption spectrum was detected using a Fourier transform infrared absorption spectrometer (FTIR) in reflectance mode. Figure 4 (As shown). Based on the characterization data, the solid product was determined to be a water-soluble carboxylated fullerene (TIFE).

[0061] Example 2: Determination of the free radical scavenging ability of water-soluble carboxylated fullerenes (TIFE)

[0062] This disclosure uses electron spin resonance spectroscopy (ESR) to detect the ability of water-soluble carboxy-based fullerenes to scavenge free radicals.

[0063] Detection method: Hydroxyl radicals were generated by the decomposition of hydrogen peroxide under ultraviolet light. 20 μL of 0.1 mM hydrogen peroxide, 40 μL of 0.1 mM dimethylpyridine N-oxide (DMPO, a free radical scavenger), and 20 μL of deionized water were mixed and irradiated under ultraviolet light for 4 min. The free radical electron spin resonance signal was detected, serving as a blank control group. The 20 μL of deionized water was replaced with 20 μL of 50 μM water-soluble carboxy-fullerene solution prepared in Example 1, with all other conditions unchanged. The free radical electron spin resonance (ESR) signal was detected, serving as the experimental group. Figure 5 As shown, the blank control group has obvious signal peaks, indicating that a large number of free radicals are generated; the experimental group has smooth curves, and the free radical signal is significantly reduced, indicating that there are fewer free radicals in the experimental group with added water-soluble carboxylated fullerene (TIFE) material, and TIFE has a strong ability to scavenge free radicals.

[0064] Example 3: Protective effect of water-soluble carboxy-fullerene (TIFE) on HepaRG hepatocytes

[0065] (1) Experimental methods

[0066] The hepatocyte line used in the experiment was HepaRG hepatocytes. This hepatocyte line retains the expression of the CYP2E1 enzyme and can better simulate the process of drug-induced liver injury / liver failure in humans or animals. It was purchased from Peking Union Medical College Cell Bank. HepaRG hepatocytes were cultured in high-glucose DMEM complete medium in an incubator at 37°C and 5% CO2.

[0067] The experiment consisted of three parts: cytotoxicity test, antioxidant stress test, and anti-acetylphenol test.

[0068] (2) Experimental results

[0069] HepaRG hepatocytes were seeded in 96-well plates, and different concentrations (0-50 μM) of water-soluble carboxy-fullerene (TIFE) material synthesized according to Example 1 were added. After co-incubation for 24 h, cell viability was detected by WST-8 assay. Figure 6 As shown, different concentrations of TIFE have no effect on cell viability to a certain extent, indicating that a certain concentration of TIFE has no significant biotoxicity to HepaRG cells.

[0070] Similarly, HepaRG hepatocytes were seeded in 96-well plates. The experimental group was first stimulated with 500 μM hydrogen peroxide for 1 h, then different concentrations (0-50 μM) of water-soluble carboxylated fullerene (TIFE) material synthesized in Example 1 were added. After incubation for 3 h, cell viability was detected using the WST-8 assay. Figure 7 As shown, the cell viability of the experimental group without TIFE decreased significantly, while different concentrations of TIFE increased cell viability to some extent. This indicates that TIFE can reduce the oxidative stress induced by hydrogen peroxide in HepaRG cells to a certain extent.

[0071] Acetaminophen (APAP) is the most common drug causing drug-induced liver injury / failure. HepaRG hepatocytes were seeded in 96-well plates. The experimental groups were incubated with 5 mM APAP and different concentrations (0-50 μM) of water-soluble carboxy-fullerene (TIFE) material synthesized according to Example 1 for 24 h. Cell viability was then assessed using the WST-8 assay. Figure 8 As shown, the cell viability of the experimental group without TIFE was significantly reduced, while different concentrations of TIFE increased cell viability to some extent. This indicates that TIFE can reduce the toxicity of APAP-induced HepaRG cells to a certain extent.

[0072] Example 4: Treatment of acetylphenol-induced liver injury / failure with water-soluble carboxylated fullerene (TIFE) materials

[0073] (1) Experimental methods

[0074] The acetylphenol-induced liver injury / liver failure model mice used in the experiment were male C57BL / 6J mice, purchased from Beijing Huafukang Biotechnology Co., Ltd.

[0075] The experimental animals were divided into three groups, with 6-8 mice in each group. They were fasted overnight but allowed free access to water for 12 hours. Group A consisted of 6 mice, which were injected with an equal volume of physiological saline as a control group. Group B consisted of 6 mice, which were injected with 350 mg / kg APAP as a model group. Group C consisted of mice that were injected intraperitoneally with 10 mg / kg of the water-soluble carboxylated fullerene (TIFE) material synthesized in Example 1, followed by an injection of 350 mg / kg APAP as an experimental group.

[0076] (2) Experimental results

[0077] This disclosure describes the process of euthanizing mice after drug administration, photographing and performing H&E pathological sections on the liver organs, and collecting serum samples to test liver function indicators such as alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Additionally, liver proteins were extracted for testing biochemical indicators such as GSH, MDA, and SOD activity. The results are as follows.

[0078] 1) Optical photography and H&E testing

[0079] Mice were sacrificed 24 hours after modeling. Liver samples were harvested, quickly washed with physiological saline to remove blood stains, and photographed. The livers were then fixed in 4% paraformaldehyde, embedded in paraffin, and subjected to a series of dehydration, sectioning, and staining processes. (See attached image.) Figure 9 , 10 As shown, the blank group consisted of normal mice with normal liver structure and no obvious lesions; the model group consisted of mice with obvious lesions and large areas of necrosis concentrated in the center of the liver lobules; while the experimental group mice treated with the water-soluble carboxy-fullerene (TIFE) material synthesized according to Example 1 showed a significant reduction in liver lesions, a significant reduction in necrotic area, and a significant reduction in the degree of necrosis.

[0080] 2) Liver function index test

[0081] Twenty-four hours after mouse modeling, blood was collected from the orbital cavity. Whole blood was collected, incubated at 4°C for 4 hours, and then centrifuged at 3500 rpm for 15 minutes. The supernatant serum was collected, and the levels of liver function indicators, alanine aminotransferase (ALT) and aspartate aminotransferase (AST), were measured using an automated blood biochemistry analyzer. Figure 11 As shown, compared to the control group, the alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels in the model group were significantly increased, while the ALT and AST levels in the experimental group mice treated with the water-soluble carboxylated fullerene (TIFE) material synthesized in Example 1 were significantly reduced and tended to normal. This indicates that the liver function of mice was significantly improved after treatment with the water-soluble carboxylated fullerene (TIFE) material synthesized in Example 1.

[0082] 3) Biochemical index testing

[0083] Mice were sacrificed 24 hours after modeling, and their livers were harvested and rapidly preserved in liquid nitrogen. Liver proteins were then extracted at low temperature according to the instructions of the GSH, MDA, and SOD activity kits. The total protein content was detected using a BCA kit. The procedures were followed according to the kit instructions, and the corresponding biochemical indicators were calculated using the formulas provided in the kit instructions. (See attached image.) Figure 12As shown, compared with the blank group, the biochemical indicators of the liver of mice in the model group showed significant changes, while the biochemical indicators of the liver of mice in the experimental group treated with the water-soluble carboxyfullerene (TIFE) material synthesized in Example 1 showed a significant tendency to return to normal. This indicates that the biochemical indicators of the liver of mice were significantly improved and the level of oxidative stress was significantly reduced after treatment with the water-soluble carboxyfullerene (TIFE) material synthesized in Example 1.

[0084] 4) Protein electrophoresis test

[0085] Mice were sacrificed 24 hours after modeling, and their livers were harvested. The livers were rapidly stored in liquid nitrogen, and proteins were extracted at low temperature using RIPA lysis buffer. The concentration of the extracted proteins was determined using a BSA kit, followed by electrophoresis, transfer, and imaging procedures as per protein electrophoresis assays. Figure 13 As shown, compared to the control group, the expression of p-JNK protein in the liver of mice in the model group was significantly increased, while the expression of p-JNK protein in the liver of mice in the experimental group treated with the water-soluble carboxyfullerene (TIFE) material synthesized in Example 1 was significantly decreased and tended to normal. This indicates that the physiological response regulated by p-JNK protein in mice was inhibited after treatment with the water-soluble carboxyfullerene (TIFE) material synthesized in Example 1, indirectly protecting mitochondria.

Claims

1. A water-soluble fullerene derivative, characterized in that, The water-soluble fullerene derivative is a fullerene isoperidinic acid derivative; wherein, in each molecule of the fullerene isoperidinic acid derivative, the ratio of fullerene molecules to isoperidinic acid molecules is 1:(1-6).

2. The water-soluble fullerene derivative according to claim 1, wherein, In each molecule of the fullerene isoperidinic acid derivative, the ratio of fullerene molecules to isoperidinic acid molecules is 1:(1-4).

3. The method for preparing the water-soluble fullerene derivative according to claim 1 or 2, characterized in that, The method includes the following steps: (1) Dissolve fullerene or endogenous metal-embedded fullerene in a solvent; (2) Add isopiridine ester, and after purification, acid hydrolysis and drying, the water-soluble fullerene derivative is obtained.

4. The preparation method according to claim 3, wherein, The isoperidine ester is methyl isoperidine carboxylate or tert-butyl isoperidine carboxylate.

5. The preparation method according to claim 3 or 4, wherein, The method includes the following steps: (1) C 60 Dissolved in o-dichlorobenzene, dimethyl sulfoxide was added under inert gas protection, and the mixture was bubbled with oxygen to replace it. (2) Add isoperidine ester, stir with oxygen, and concentrate to obtain a reddish-brown solid; (3) The reddish-brown solid was separated by column chromatography to obtain the elution product; (4) The elution product is hydrolyzed with hydrochloric acid-acetic acid or trifluoroacetic acid, centrifuged, the precipitate is collected, washed with dilute acid, neutralized, and finally dialyzed and freeze-dried to obtain the water-soluble fullerene derivative.

6. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises the water-soluble fullerene derivative as described in claim 1 or 2.

7. The use of the water-soluble fullerene derivative according to claim 1 or 2, or the pharmaceutical composition according to claim 6, in the preparation of a medicament for the prevention and / or treatment of liver injury / liver failure.

8. The application according to claim 7, wherein, The liver damage / liver failure mentioned refers to any drug that can cause liver damage / liver failure.

9. The application according to claim 8, wherein, The drug includes one or more of acetylphenol, isoniazid, cyclosporine A, tetracycline, and tripterygium wilfordii.

10. The application according to any one of claims 7-9, wherein, The prevention and / or treatment of liver injury / liver failure includes improving liver function.

11. The application according to any one of claims 7-9, wherein, The prevention and / or treatment of liver injury / liver failure includes reducing the necrotic area of ​​the liver.

12. The application according to any one of claims 7-9, wherein, The prevention and / or treatment of liver injury / liver failure includes reducing the degree of liver necrosis.

13. The application according to any one of claims 7-9, wherein, The prevention and / or treatment of liver injury / liver failure includes improving liver biochemical indicators.

14. The application according to claim 10, wherein, The improvement in liver function brought liver function biochemical indicators closer to normal.

15. The application according to claim 14, wherein, The liver function biochemical indicators are selected from one or more of the following: alanine aminotransferase (ALT) and aspartate aminotransferase (AST).

16. The application according to claim 13, wherein, The improvement in liver biochemical indicators brings one or more of the following biochemical indicators, such as GSH, MDA, and SOD, closer to normal:

17. The application according to any one of claims 7-9, wherein, The prevention and / or treatment of liver injury / liver failure includes one or more methods that significantly reduce drug-induced hepatocyte death and significantly reduce the level of oxidative stress within hepatocytes.

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