Preparation method and application of amino acid derived carbon dot nano-enzyme

Abstract

The invention provides a preparation method and application of amino acid derived carbon dot nano-enzyme, and belongs to the technical field of new biomedical materials. The amino acid comprises one of histidine, proline, cysteine, tryptophan, aspartic acid, arginine, lysine, methionine and phenylalanine; the mass ratio of the amino acid to the citric acid monohydrate is (1: 1)-(1: 5). The amino acid derived carbon dots provided by the invention have free radical scavenging activity and superoxide dismutase-like properties, and show low toxicity and good biological safety at the levels of cells, nematodes and animals. Meanwhile, the amino acid derived carbon dot nano-enzyme also shows excellent anti-inflammatory activity, can down-regulate the inflammatory level of cells, reduce the damage of oxidative stress to nematodes, improve the intestinal flora structure, increase the richness and diversity of intestinal flora, relieve and treat DSS-induced mouse ulcerative colitis, and has a good application prospect. The compound can be further applied to preparation of drugs for treating inflammatory bowel diseases.

Description

Technical Field

[0001] The present invention belongs to the technical field of new biomedical materials, and specifically relates to a preparation method of an amino acid-derived carbon dot nanozyme and application thereof. Background Art

[0002] Inflammatory bowel disease (IBD) is a chronic, idiopathic intestinal inflammatory disease. Clinical and experimental evidence in recent years have shown that the production of excessive free radicals, especially reactive oxygen species (ROS) and reactive nitrogen species (RNS), dominates and aggravates the progression of IBD in the site of enteritis. In recent years, the discovery of antioxidant nanozymes has brought great prospects for the treatment of inflammatory bowel disease (IBD). In order to effectively eliminate reactive oxygen species (ROS), various metal-based nanozymes have been developed for the treatment of IBD. The cobalt oxide nanozymes reported so far use the successful strategy of scavenging ROS to inhibit DSS-induced colitis by targeting the NLRP3 inflammasome (Chen, Ziying et al. "2D Cobalt Oxyhydroxide Nanozymes Inhibit Inflammation by Targeting the NLRP3 Inflammasome." Advanced Functional Materials (2023): 2214693.). This demonstrates the potential role of metal-based nanozymes in the prevention and treatment of related inflammatory diseases, but their unstable elimination activity and potential gastrointestinal metal ion leakage seriously limit the practical application of metal nanozymes in the biomedical field.

[0003] In particular, compared with metal-based nanozymes, carbon-based nanomaterials have the advantages of high chemical stability, high catalytic activity, low toxicity and biosafety, and are expected to play a potential role in the prevention and treatment of inflammatory diseases. Carbon dots (CDs) are a new type of carbon-based nanomaterial with a size of less than 10nm. They have been widely used in the biomedical field due to their inherent nanozyme activity, small size, good biosafety, and easy surface modification and functionalization.

[0004] CDs can be synthesized from different precursors, such as citric acid, glucose, β-cyclodextrin, L-ascorbic acid, graphene oxide and various biomass materials. Studies have shown that precursors are key factors affecting the activity of CDs enzymes (Gao, Wenhui et al. "Deciphering the catalytic mechanism of superoxide dismutase activity of carbon dot nanozyme." Nature Communications (2023) 14: 160). Amino acids, as the basic building blocks of proteins, play an important role in maintaining human nitrogen balance and normal physiological activities. Based on this, the present invention selects various amino acids as precursors, and prepares an amino acid-derived carbon dot nanozyme by hydrothermal method for better application in the biomedical field. Summary of the invention

[0005] In order to solve the shortcomings and deficiencies of the prior art, the primary purpose of the present invention is to quickly obtain a metal-free carbon dot nanozyme with stable chemical properties, high free radical scavenging efficiency, good biocompatibility, and can be applied

[0006] In the preparation of anti-inflammatory drugs.

[0007] The present invention also provides a method for preparing the amino acid-derived carbon dot nanozyme and its application in preparing drugs for treating inflammatory bowel disease (IBD).

[0008] In order to achieve the above purpose, the present invention uses different types of amino acids and monohydrated citric acid as precursors, and then generates carbonized products through a one-step hydrothermal / solvothermal method, and then obtains the amino acid-derived carbon dot nanozymes through separation and purification.

[0009] The above-mentioned preparation method based on different types of amino acid carbon dots specifically comprises the following steps:

[0010] (1) Weighing amino acids and citric acid monohydrate and adding them to a solvent, transferring the solution into a polytetrafluoroethylene-lined reactor, heating to 160-200° C., heating for 8-24 h, and then cooling to room temperature to obtain a yellow carbon dot solution;

[0011] (2) The yellow carbon dot solution is post-treated to obtain amino acid-derived carbon dot nanozymes. The post-treatment includes filtering the yellow carbon dot solution with a 0.22 μm filter membrane, collecting the filtrate, dialyzing the solution with a 500-1000 molecular weight dialysis membrane for 36-72 hours, collecting the dialyzate, and vacuum freeze-drying to obtain a solid powder.

[0012] The amino acid described in step (1) includes one of histidine, proline, cysteine, tryptophan, aspartic acid, arginine, lysine, methionine and phenylalanine.

[0013] Preferably, the amino acid is histidine.

[0014] The amino acid and citric acid monohydrate described in step (1) are dissolved in 50-100 mL of a solvent at a mass ratio of 1 g:1 g to 1 g:5 g; wherein the solvent comprises deionized water and / or an organic solvent; and the organic solvent comprises N,N-dimethylformamide and ethanol.

[0015] In step (2), a 0.22 μm filter membrane is used for the filtration, and the filtration is repeated three times.

[0016] The dialysis external solution used in the dialysis in step (2) is double distilled water, which is replaced every 4 hours until the external solution becomes clear. The dialysis time can be appropriately extended or shortened depending on the sample. A solid powder is obtained after vacuum freeze drying.

[0017] An amino acid-derived carbon dot nanozyme is prepared by the above-mentioned preparation method, and the amino acid-derived carbon dot nanozyme can efficiently remove free radicals.

[0018] The amino acid-derived carbon dot nanozyme can be used in the preparation of anti-inflammatory drugs.

[0019] Furthermore, the amino acid-derived carbon dot nanozyme can be used in the preparation of drugs for treating inflammatory bowel disease.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] (1) The amino acid-derived carbon dot nanozymes prepared by the present invention have a small average particle size, rich oxygen- and nitrogen-containing functional groups on the surface, good water solubility, and can be widely used in the biomedical field.

[0022] (2) The amino acid-derived carbon dot nanozymes prepared by the present invention do not contain metal elements, are non-toxic, have high biosafety, and exhibit good free radical scavenging activity and certain superoxide dismutase-like activity both in vitro and in vivo, as well as good biocompatibility. Based on this, the amino acid-derived carbon dot nanozymes can be effectively used to prepare drugs for the treatment of inflammatory bowel disease (IBD).

[0023] (3) Compared with natural enzymes, the amino acid-derived carbon dot nanozymes prepared by the present invention have low preparation and purification costs, stable chemical properties, and can work efficiently in harsh environments (strong acids and bases) and disease conditions, providing new ideas for the application of nanomaterials in anti-oxidative stress.

[0024] (4) The amino acid-derived carbon dot nanozymes prepared by the present invention can improve the intestinal flora structure of ulcerative colitis, increase the richness and diversity of intestinal flora, and alleviate the damage of the intestinal mucus layer in mice, and have good application prospects in the preparation of drugs for the treatment of inflammatory diseases. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 The DPPH free radical scavenging ability of the amino acid-derived carbon dot nanozymes obtained in Example 2 of the present invention;

[0026] Figure 2 The transmission electron microscopy morphology image (A) and the Fourier transform infrared spectrum image (B) of the histidine-derived carbon dot nanozyme obtained in Example 2 of the present invention;

[0027] Figure 3 The superoxide dismutase-like activity of the histidine-derived carbon dot nanozyme obtained in Example 2 of the present invention;

[0028] Figure 4 This is a biosafety evaluation of the histidine-derived carbon dot nanozyme in Example 6 of the present invention. Among them, Figure A is the cytotoxicity of His-CDs to mouse mononuclear macrophage RAW264.7; Figure B is a cell development diagram of His-CDs; Figure C is a micrograph of nematodes after feeding with His-CDs for one week; Figure D is a body length statistics of nematodes after feeding with His-CDs for one week; Figure E is a statistics of the number of movements of nematodes within 15 seconds after feeding with His-CDs for one week; Figure F is an H&E staining picture of the main organ sections of mice after gavage with His-CDs for 14 days;

[0029] Figure 5 This is an evaluation of the anti-inflammatory ability of the histidine-derived carbon dot nanozyme at the cellular and nematode levels in Example 7 of the present invention. Among them, Figure A shows the effect of His-CDs intervention on the content of reactive oxygen species in M1 macrophages; Figures B and C are the effects of His-CDs on the mRNA levels of inflammatory factors TNF-α and IL-1β in M1 macrophages, respectively; Figure D is the MDA content in nematodes in different treatment groups; Figure E is the mRNA level of the oxidative stress gene SOD-1 in nematodes in different treatment groups.

[0030] Figure 6 The figure is a diagram showing the relief of DSS-induced acute colitis by histidine-derived carbon dot nanozymes in Example 8 of the present invention. Among them, Figure A shows the weight changes and total weight loss of mice from day 7 to day 21; Figure B shows the disease activity index (DAI) scores of each group of mice from day 7 to day 21; Figure C shows the colon length of each group of mice; Figure D shows representative colon photos of each group of mice; Figures E and F are the relative expression levels of inflammatory factors IL-1β and TNF-α mRNA in mouse colon tissue, respectively.

[0031] Figure 7 These are photos of hematoxylin and eosin staining and histopathological scores of colon tissues of mice in each group in Example 8 of the present invention.

[0032] Figure 8 The His-CDs in Example 9 of the present invention alleviated the changes in the composition of the intestinal microbiota induced by DSS. The Venn diagram in Figure A shows the overlapping operational taxonomic units in the intestinal microbiota of three groups of mice; Figures B and C are studies on α diversity based on the Simpson and Sobs indices, respectively.

[0033] Fig. 9 It is the average relative abundance of each group of bacteria at the phylum level in Example 9 of the present invention.

[0034] Fig.10 It is the average relative abundance of each group of bacteria at the genus level in Example 9 of the present invention. DETAILED DESCRIPTION

[0035] The present invention is further described by the following embodiments. It should be understood that the following embodiments are only used to illustrate the present invention, not to limit the present invention. A person skilled in the art can and should know that any simple change or replacement based on the essential spirit of the present invention should fall within the scope of protection required by the present invention.

[0036] Example 1

[0037] The preparation of amino acid-derived carbon dot nanozymes is as follows:

[0038] (1) Histidine-derived carbon dot nanozymes (His-CDs): 1 g of histidine and 5 g of citric acid monohydrate were weighed and added to 100 mL of double distilled water. The solution was added to a 100 mL polytetrafluoroethylene-lined reactor and heated in an oven at 160 °C for 8 h to obtain a yellow carbon dot solution.

[0039] (2) Pour the yellow carbon dot solution into a beaker and filter it with a 0.22 μm filter membrane. Then dialyze it with a 500 molecular weight dialysis membrane for 36 hours, filter the dialyzed liquid with a 0.22 μm filter membrane and pour it into a glass petri dish, put it in a -20°C refrigerator, and freeze-dry it with a vacuum freeze dryer to obtain a solid powder;

[0040] (3) adding ultrapure water to the solid powder obtained above, dissolving it by ultrasonication, filtering with a 0.22 μm water filter to remove insoluble components, and obtaining a filtrate which is the histidine-derived carbon dot nanozyme;

[0041] (4) The preparation of the remaining amino acid-derived carbon dot nanozymes is based on the synthesis and post-treatment methods of His-CDs in (1) to (3). According to the names of the precursor amino acids in the present invention, they are named proline carbon dots (Pro-CDs), cysteine ​​carbon dots (Cys-CDs), tryptophan carbon dots (Trp-CDs), aspartic acid carbon dots (Asp-CDs), arginine carbon dots (Arg-CDs), lysine carbon dots (Lys-CDs), methionine carbon dots (Met-CDs) and phenylalanine carbon dots (Phe-CDs).

[0042] Example 2 In this example, different conditions were used to prepare amino acid-derived carbon dot nanozymes. The specific steps are as follows:

[0043] (1) Histidine-derived carbon dot nanozymes (His-CDs): 1 g of histidine and 1 g of citric acid monohydrate were weighed and added to 50 mL of double distilled water. The solution was added to a 100 mL polytetrafluoroethylene-lined reactor and heated in an oven at 200 °C for 12 h to obtain a yellow carbon dot solution.

[0044] (2) Pour the yellow carbon dot solution into a beaker and filter it with a 0.22 μm filter membrane. Then dialyze it with a 500 molecular weight dialysis membrane for 48 hours, filter the dialyzed liquid with a 0.22 μm filter membrane and pour it into a glass petri dish, put it in a -20°C refrigerator, and freeze-dry it with a vacuum freeze dryer to obtain a solid powder;

[0045] (3) adding ultrapure water to the solid powder obtained above, dissolving it by ultrasonication, filtering with a 0.22 μm water filter to remove insoluble components, and obtaining a filtrate which is the histidine-derived carbon dot nanozyme;

[0046] (4) The preparation of the remaining amino acid-derived carbon dot nanozymes is based on the synthesis and post-treatment methods of His-CDs in (1) to (3). According to the names of the precursor amino acids in the present invention, they are named proline carbon dots (Pro-CDs), cysteine ​​carbon dots (Cys-CDs), tryptophan carbon dots (Trp-CDs), aspartic acid carbon dots (Asp-CDs), arginine carbon dots (Arg-CDs), lysine carbon dots (Lys-CDs), methionine carbon dots (Met-CDs) and phenylalanine carbon dots (Phe-CDs).

[0047] Example 3

[0048] Amino acid-derived carbon dot nanozymes were prepared according to the method in Example 2(1)-(4), except that the solvent in this example was N,N-dimethylformamide (DMF).

[0049] Example 4

[0050] Amino acid-derived carbon dot nanozymes were prepared according to the method in Example 2(1)-(4), except that the solvent in this example was a mixture of DMF and deionized water in a volume ratio of 1:1.

[0051] Example 5

[0052] The basic physicochemical properties of the carbon dot nanozyme prepared in Example 2 were measured.

[0053] (1) DPPH free radical scavenging activity

[0054] A 0.1 mM DPPH ethanol solution was prepared, and then 100 μL of amino acid-derived carbon dot nanozyme aqueous solution (500 μg / mL) was mixed with 1 mL of DPPH solution. After incubation in the dark for 30 min, the absorption curve of each group of solutions was measured using a UV spectrophotometer. Figure 1 This reflects the DPPH free radical scavenging ability of the amino acid-derived carbon dot nanozymes in Example 2. It can be seen that after the addition of the amino acid-derived carbon dot nanozymes, the characteristic absorption peak of the DPPH free radical in the system at 517 nm is reduced, and the histidine-derived carbon dot nanozymes (His-CDs) have the best scavenging effect.

[0055] Subsequent experiments revealed that the DPPH free radical scavenging ability of the amino acid-derived carbon dot nanozymes obtained in Examples 1, 3, 4 and 5 also met the requirements. Figure 1 Therefore, histidine-derived carbon dot nanozymes were selected for subsequent experiments.

[0056] (2) Characterization of morphology and surface functional groups

[0057] The morphology of histidine-derivatized carbon dot nanozymes (His-CDs) was characterized by JEM-F200 transmission electron microscopy. Fourier transform infrared (FT-IR) spectra were collected using a Bruker Hyperion 2000 spectrometer. Figure 2 Figure A shows the morphology of the histidine-derived carbon dot nanozyme. The His-CDs are evenly distributed with an average diameter of 2.5 ± 0.4 nm. As shown in the HRTEM image (inset), the lattice spacing is 0.21 nm, corresponding to the (100) crystal plane of graphite. Figure 2 B is the Fourier transform infrared spectrum of His-CDs, where 3460 cm-1 is the stretching vibration peak of the hydroxyl group and 1576 cm-1 is the asymmetric stretching vibration peak of the carboxyl group C=O.

[0058] (3) Determination of superoxide dismutase (SOD) activity

[0059] The SOD enzyme activity of His-CDs was determined using the Beyotime total superoxide dismutase activity detection kit (WST-8 method). Figure 3 The results reflect the superoxide dismutase (SOD) activity of His-CDs. Figure 3 As can be seen in Figure A, the SOD inhibition rate of His-CDs is 50%. Figure 3 In Figure B, we can calculate that the SOD enzyme activity of His-CDs is approximately 125U / mg.

[0060] Example 6

[0061] Biosafety evaluation of amino acid-derived carbon dot nanozymes:

[0062] Cell level: The His-CDs with a final concentration of 0-1.25 mg / mL in Example 2 were co-cultured with mouse mononuclear macrophages RAW264.7 for 24 h, and the survival rates of cells in different treatment groups were determined by CCK-8 method. Figure 4 As shown in Figure A, when the concentration of His-CDs was as high as 0.5 mg / mL, the survival rate of RAW264.7 was still greater than 80%, showing good biocompatibility. After co-culturing RAW264.7 with His-CDs at a final concentration of 0.2 mg / mL for 1 hour, the cells were observed using a laser confocal microscope. Figure 4 As shown in B, after 1 h of co-culture, His-CDs can enter the cells and emit bright blue light under the 405 nm channel. The cells are round and in good condition.

[0063] Nematode level: Caenorhabditis elegans was used as an experimental animal to explore the effects of feeding His-CDs on the body length and movement frequency of nematodes. The experimental groups were as follows: the control group used Escherichia coli OP50 to feed the nematodes, and the experimental group used OP50 mixed with 100μg / mL His-CDs to feed the nematodes. Use a picker to transfer the synchronized L4 nematodes to the corresponding NGM plates, with at least 3 plates per group and about 200 nematodes per plate, which were inverted and cultured at 20°C. This was recorded as day 0. Transfer the nematodes on each culture plate to a new NGM every one or two days. On the 7th day, randomly pick at least 10 live nematodes in each group using a picker, gently place them on a water agar plate, observe them under a microscope, and measure and record the body length of the nematodes using LAZ EZ software. The experiment was repeated three times independently. The body length of the nematodes reflects the growth status and growth rate of the nematodes. Figure 4 Center C shows microscopic photographs of different groups of Caenorhabditis elegans. Figure 4 D and E in the figure respectively counted the body length and movement times of nematodes in each group. The results showed that there was no significant difference in body length and movement times of nematodes fed with mixed diet compared with those of the control group.

[0064] Animal level: After the His-CDs in Example 2 were continuously gavaged into mice for 14 days, H&E staining of the main organ sections was as follows: Figure 4 As shown in F, no obvious pathological damage was observed compared with the control group, demonstrating the safety of His-CDs.

[0065] Example 7

[0066] The cell experiment was divided into three groups: the control group was RAW264.7 cells cultured normally; the model group (LPS group) was RAW264.7 cells induced by lipopolysaccharide to transform into M1 macrophages that can produce pro-inflammatory factors; the CDs group was based on the model group, and His-CDs prepared in Example 2 were added to co-culture for 24 hours. The reactive oxygen detection kit was used to determine the level of intracellular reactive oxygen in the presence or absence of His-CDs. Figure 5 As shown in Figure A, compared with the model group, the level of intracellular reactive oxygen species was significantly reduced after His-CDs intervention. Real-time fluorescence quantitative PCR was used to detect the relative expression of M1 macrophage markers (TNF-α and IL-1β) in cells. Figure 5 As shown in Figures B and C, the His-CDs prepared in Example 2 can reduce the expression of M1 macrophage markers and exhibit a good anti-inflammatory effect at the cellular level.

[0067] Caenorhabditis elegans was used as the experimental animal, and 0.2 mM Juglone was used to induce the oxidative stress model. After 7 days of culture of the synchronized L4 nematodes on NGM plates containing or without His-CDs, the nematodes were collected and operated according to the instructions of the MDA content detection kit. The amount of MDA reflects the degree of peroxidation in the body and indirectly reflects the degree of cell damage. Figure 5 As shown in D, the MDA content in the nematodes in the model group was significantly increased, and the MDA content after His-CDs intervention was significantly reduced compared with the model group, indicating that His-CDs intervention can effectively alleviate the damage of cells in nematodes in the oxidative stress model. Figure 5 The mRNA levels of the oxidative stress gene SOD-1 in nematodes in different treatment groups in E also proved that His-CDs intervention can promote the expression of the SOD-1 gene and regulate oxidative stress.

[0068] Example 8

[0069] The His-CDs in Example 2 were used to treat mild acute inflammatory bowel disease in mice induced by dextran sulfate sodium salt (DSS). C57BL / 6 mice were randomly divided into a Control group, a DSS model group, and a His-CDs group (DSS+CDs, 25 mg / kg). The mouse colitis model was established by drinking 2.5% (w / v) DSS for 7 consecutive days, and the Control group was given an equal amount of drinking water. After modeling, the mice were gavaged for 14 consecutive days. The weight of the mice was recorded daily, the feces and blood in the stool of the mice were observed, and the colon tissue of each group of mice was collected on the 14th day.

[0070] like Figure 6 As shown in Figure A, His-CDs intervention alleviated the weight loss of colitis mice. After modeling, the DAI scores of the DSS and His-CDs groups were both around 8. After 14 days of intragastric administration, the DAI score of the DSS group fluctuated around 8, and intragastric administration of sterile water could not reduce the DAI score. However, after 14 days of intragastric administration of His-CDs, the DAI score showed a downward trend ( Figure 6 The colon length of each group of mice was measured with a ruler, and the statistical results showed that the intervention of His-CDs alleviated the shortening of the colon in colitis mice ( Figure 6 Middle C). Figure 6 The representative colon photo in Figure D also proves this conclusion. Real-time fluorescence quantitative PCR was used to detect the relative expression of inflammatory factors in the colon tissue of each group of mice. The results showed that His-CDs could downregulate the pro-inflammatory factor IL-1β ( Figure 6 Middle E), TNF-α( Figure 6 F) mRNA expression level. Figure 7 The HE staining results also showed that the colon tissue of the model group mice showed goblet cell loss, inflammatory cell infiltration, mucosal damage and other phenomena. In contrast, the colon tissue of the His-CDs intervention group almost maintained its integrity, and the histopathological score was significantly lower than that of the model group. The above results all indicate that His-CDs intervention can alleviate DSS-induced colitis in mice and relieve the damage of the intestinal mucus layer in mice.

[0071] Example 9

[0072] The His-CDs in Example 2 were administered intragastrically to mice with DSS-induced ulcerative colitis. After 14 days, the feces of mice in each group were collected to explore the changes in the intestinal microbiota of mice. As shown in the Venn diagram, the control group, DSS group, and His-CDs group contained 114, 75, and 99 different microorganisms, respectively. The number of OTUs in the DSS group was lower than that in the control group and CDs group. This indicates that CDs intervention alleviated the reduction in the number of OTUs induced by DSS and enhanced the diversity of intestinal microorganisms in mice ( Figure 8In addition, the community diversity (Simpson index) and community richness (SOBs) of the His-CDs group were significantly higher than those of the DSS group ( Figure 8 Therefore, intervention with His-CDs alleviated the reduction in gut microbiota diversity and richness induced by DSS.

[0073] At the phylum level, Firmicutes and Bacteroidetes were the dominant phyla in the fecal microbiota ( Fig. 9 ). Studies have shown that an increase in the ratio of Firmicutes to Bacteroidetes indicates a high level of inflammation in the body. Compared with the DSS group, the ratio of Firmicutes to Bacteroidetes in the intestines of mice in the His-CDs group was significantly reduced. At the genus level, compared with the DSS group, the relative abundance of Bacteroidetes norank_f_Muribaculaceae in the intestines of mice in the His-CDs group was significantly increased, and the relative abundance of Allobaculum and Dubosiella were significantly increased ( Fig.10 ). norank_f_Muribaculaceae can reduce inflammation, inhibit harmful bacteria and oxidative stress, and improve intestinal mucosal inflammation. Allobaculum and Dubosiella can produce short-chain fatty acids to provide energy sources for intestinal mucosal cells, promote intestinal epithelial cell proliferation, maintain intestinal barrier, and resist inflammation. Based on this, the amino acid-derived carbon dot nanozyme prepared by the present invention can improve the intestinal flora structure of ulcerative colitis, increase the richness and diversity of intestinal flora, and alleviate the damage of the intestinal mucus layer in mice, and has good application prospects in the preparation of drugs for the treatment of inflammatory diseases.

Claims

1. A method for preparing an amino acid-derived carbon dot nanozyme, characterized in that: The following steps are involved: (1) Weighing amino acids and citric acid monohydrate and adding them to a solvent, transferring the resulting solution into a polytetrafluoroethylene-lined reactor, and heating the reaction to obtain a yellow carbon dot solution; (2) Post-treating the yellow carbon dot solution to obtain amino acid-derived carbon dot nanozymes.

2. The method for preparing the amino acid-derived carbon dot nanozyme according to claim 1, characterized in that: The amino acid includes one of histidine, proline, cysteine, tryptophan, aspartic acid, arginine, lysine, methionine and phenylalanine.

3. The method for preparing the amino acid-derived carbon dot nanozyme according to claim 1, characterized in that: The amino acid is histidine.

4. The method for preparing the amino acid-derived carbon dot nanozyme according to claim 1, characterized in that: The amino acid and citric acid monohydrate are dissolved in 50-100 mL of a solvent at a mass ratio of 1 g:1 g to 1 g:5 g; wherein the solvent comprises double distilled water and / or an organic solvent; and the organic solvent comprises N,N-dimethylformamide and ethanol.

5. The method for preparing the amino acid-derived carbon dot nanozyme according to claim 1, characterized in that: The temperature of the heating reaction in step (1) is 160-200° C., and the reaction time is 8-24 hours.

6. The method for preparing the amino acid-derived carbon dot nanozyme according to claim 1, characterized in that: The post-treatment in step (2) includes filtering the yellow carbon dot solution with a 0.22 μm filter membrane and collecting the filtrate, then dialyzing it with a 500-1000 molecular weight dialysis membrane for 36-72 hours, and vacuum freeze-drying to obtain a solid powder.

7. An amino acid-derived carbon dot nanozyme, characterized in that: The preparation method according to claims 1 to 6 is used to obtain the product.

8. The amino acid-derived carbon dot nanozyme according to claim 7, characterized in that: The amino acid-derived carbon dot nanozyme does not contain any metal elements and has a particle size of less than 10 nm. The carbon dot nanozyme has DPPH free radical scavenging activity and superoxide dismutase-like activity and exhibits biocompatibility at both the cellular and animal levels.

9. Use of an amino acid-derived carbon dot nanozyme prepared by the preparation method according to any one of claims 1 to 6 in the preparation of anti-inflammatory drugs.

10. The use according to claim 9, characterized in that The amino acid-derived carbon dot nanozyme is used in the preparation of drugs for treating inflammatory bowel disease.

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