CN114081897A - Selenium-doped Prussian blue nanoenzyme for regulating intestinal cells to treat colitis and preparation method and application thereof - Google Patents
Selenium-doped Prussian blue nanoenzyme for regulating intestinal cells to treat colitis and preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a selenium-doped Prussian blue nano enzyme for regulating intestinal cells to treat colitis and a preparation method and application thereof. The selenium-doped Prussian blue nanoenzyme disclosed by the invention has good active oxygen scavenging capacity and the functions of inhibiting lipid peroxidation of intestinal epithelial cells and T cell differentiation, and can be used for relieving intestinal inflammatory diseases.
Description
Technical Field
The invention belongs to the technical field of nano materials, and particularly relates to a preparation method of selenium-doped Prussian blue nano enzyme for regulating intestinal cells to treat colitis.
Background
Ulcerative Colitis (UC) is a chronic inflammatory bowel disease of unknown etiology affecting the colon and rectum. UC occurs due to multiple factors, including genetic background, environment and mucus immunoregulation imbalance, which are all important causes of intestinal inflammation. Chronic inflammation in patients with colitis can lead not only to abdominal pain and diarrhea, but also to disruption of the gastrointestinal epithelial barrier, dysregulation of cell types and intestinal microflora in the colon, and the appearance of these symptoms can significantly reduce the quality of life of patients with inflammatory bowel disease. UC has now developed as a global burden, with a great impact on developed countries and a greatly increased impact on developing countries. Current clinical interventions for inflammatory bowel disease mainly involve oral or intravenous injection of small molecule drugs (such as salicylates), corticosteroids, and immunosuppressants. Although these treatments may cure a subset of patients, they are not well suited for patients who develop colitis-related colorectal hyperplasia or cancer and develop resistance to the disease. In order to solve this troublesome medical problem, development of a novel therapeutic method for safely and reliably treating inflammatory bowel disease has received high attention from a large number of researchers.
Reactive Oxygen Species (ROS) are oxygen-containing, one-electron by-products produced by oxygen molecules during cellular respiration and metabolism of organisms, including superoxide radicals, hydroxyl radicals, hydrogen peroxide, and the like. Although the etiology and pathogenesis of inflammatory bowel disease is not well defined, numerous studies have demonstrated excessive accumulation of ROS at inflammatory sites in the colon. Many animal and clinical studies have demonstrated that high levels of ROS result in oxidative damage to proteins, lipids and DNA, further leading to increased mucosal damage, mucosal ulceration and exacerbation of intestinal inflammation. Therefore, strategies that are effective in scavenging ROS have received a great deal of attention. Some nano-drugs loaded with ROS-scavenging enzymes have been successfully developed and applied for intracolonic administration for treatment of colitis. Despite achieving a certain therapeutic effect, the inherent disadvantages of natural enzymes have prevented their further development and use. Recently, nanoenzymes with ROS scavenging ability have gained new applications in the field of medical materials, which have diverse mimic enzyme activities, helping to protect tissues from oxidative damage of endogenous activities and promoting tissue regeneration. In addition, the nano enzyme also has the advantages of simple preparation, high stability, good catalytic activity and the like, and the nano enzyme can gradually become a substitute of the traditional enzyme and the natural enzyme. Prussian Blue (PB) is a nano enzyme with more application prospects in recent years, and due to good enzyme simulation properties of Catalase (CAT) and superoxide dismutase (SOD), the Prussian Blue (PB) can effectively remove ROS, and has good application in various inflammatory diseases. However, PB alone scavenges reactive oxygen species, which are difficult to deal with in complex immune cell imbalances at the site of colitis.
Disclosure of Invention
In order to solve the problems of inflammatory reaction of various cells at the inflammatory part of the colon, high preparation cost, low catalytic activity and the like of the traditional nano enzyme, the invention constructs the selenium-doped Prussian blue nano enzyme for regulating intestinal cells to treat enteritis.
In order to solve the technical problem, the invention adopts the following technical scheme:
the invention firstly discloses a selenium-doped Prussian blue nano enzyme for regulating intestinal cells to treat enteritis, which is characterized in that: the selenium-doped Prussian blue nano enzyme takes hollow mesoporous Prussian blue as a core, and selenium element is doped on the surface of the hollow mesoporous Prussian blue. The particle size of the selenium-doped Prussian blue nano enzyme is 120-140 nm. On the basis of the existing Prussian blue, the invention further improves the synthesis method to enable the Prussian blue to be in a hollow mesoporous structure, and a layer of selenium is covered on the outer layer through a hard template method, so that the Prussian blue contains glutathione peroxidase (GPx) enzyme activity, and the Prussian blue can play a potential of reversing the lipid peroxidation process of intestinal epithelial cells like GPX4 enzyme of the cells. Meanwhile, selenium is an essential trace mineral, and is incorporated into proteins to form selenoproteins, which can inhibit the differentiation of T cells into CD4+T, relieving intestinal inflammation.
Further, the valence state of the selenium element doped on the surface is 0 valence.
The preparation method of the selenium-doped Prussian blue nano enzyme comprises the following steps:
(1) dissolving 2.5-3.5g PVP in 30mL of HCL solution with the concentration of 0.01-0.05M, adding 100-150mg of potassium ferricyanide powder, and stirring until the mixture is uniformly mixed; placing the obtained mixed solution in an oven to react for 24 hours at the temperature of 80 ℃; after the reaction is finished, centrifuging, washing with water, and freeze-drying to obtain Prussian blue powder;
(2) dissolving 20-30mg Prussian blue powder in 30mL of HCL solution with the concentration of 0.8-1M, adding 200-300mg PVP, stirring at room temperature for 3-3.5h, and then placing in an oven for reaction at 140 ℃ for 3 h; after the reaction is finished, centrifuging, washing and freeze-drying to obtain hollow mesoporous Prussian blue powder;
(3) adding 4-6mL of 2mg/mL sodium selenite solution into 2mL of 2mg/mL hollow mesoporous Prussian blue aqueous dispersion, stirring at normal temperature for reaction for 20-30min, slowly adding 8-12mL of 2mg/mL ascorbic acid solution, and reacting at normal temperature for 3 h; after the reaction is finished, centrifuging, dialyzing in a dialysis bag with Mw (3500 Da) for 24h, and freeze-drying to obtain selenium-doped Prussian blue nano-enzyme powder; wherein the mass ratio of the ascorbic acid to the sodium selenite is 2: 1.
The selenium-doped Prussian blue nano enzyme has the following characteristics: not only can remove hydroxyl free radicals, but also has the function of glutathione peroxidase; no obvious hemolytic behavior and cytotoxicity; has the capability of eliminating various active oxygen; the selenium element can inhibit lipid peroxidation of epithelial cells of the intestinal tract and restore the barrier of the intestinal tract; the selenium can inhibit T cell differentiation and reduce T cell mediated inflammatory reaction.
The selenium-doped Prussian blue nanoenzyme has the function of regulating intestinal cells, has no obvious toxicity to mammalian cells, has good biological safety, and is used for preparing the nanoenzyme for regulating the intestinal cells and treating colitis.
The mechanism of the selenium-doped Prussian blue nanoenzyme used for regulating intestinal cells to treat colitis is as follows: prussian blue has good mimic enzyme properties of Catalase (CAT), superoxide dismutase (SOD) and the like, can effectively remove ROS, and has good application in various inflammatory diseases. Selenium (Se) is used as essential nutrient substance of human body, and has good biological activityAnd (4) compatibility. In addition, selenium, which is an important component of glutathione peroxidase (GPx), has high GPx enzyme activity, and can exert the potential of reversing lipid peroxide processes like GPX4 enzyme of cells. Meanwhile, selenium is an essential trace mineral, and is incorporated into proteins to form selenoproteins, which can inhibit the differentiation of T cells into CD4+T, relieving intestinal inflammation.
The invention has the beneficial effects that:
1. according to the invention, the selenite is reduced on the surface of the hollow mesoporous Prussian blue to form the monovalent selenium element, the method is simple, and the obtained selenium-doped Prussian blue nanoenzyme has good stability and dispersibility.
2. Compared with the single prussian blue, the selenium-doped prussian blue nano enzyme further improves the capacity and variety of eliminating active oxygen, and can be used for treating various inflammatory diseases.
3. The selenium-doped Prussian blue nanoenzyme has the potential of inhibiting lipid peroxidation of intestinal epithelial cells and T cell differentiation, and can be used for relieving intestinal inflammatory diseases.
4. The selenium-doped Prussian blue nanoenzyme is used as a novel nanoenzyme capable of regulating intestinal cells to relieve enteritis, and remarkably relieves the enteritis in a DSS (dextran sulfate) induced prevention and delay colitis model and a TNBS (trinitrobenzene sulfonic acid) induced rat Crohn's disease model.
5. The nano enzyme has good biological safety and is beneficial to clinical use.
Drawings
FIG. 1 is a schematic synthesis of the present invention.
FIG. 2 is a transmission electron micrograph of the selenium-doped Prussian blue nanoenzyme prepared in example 1.
FIG. 3 is an X-ray photoelectron spectrum of the selenium-doped Prussian blue nanoenzyme prepared in example 1.
Fig. 4 is a graph of in vitro hydroxyl radical scavenging of the selenium-doped prussian blue nanoenzyme prepared in example 1.
Fig. 5 is a diagram of the selenium-doped prussian blue proteasome external mimic glutathione peroxidase prepared in example 1.
FIG. 6 is a graph showing the reactive oxygen species staining of cells by selenium-doped Prussian blue nanoenzymes prepared in example 1.
FIG. 7 is a graph of cell death and staining of selenium-doped Prussian blue nanoenzymes prepared in example 1.
Fig. 8 is a cell inhibitory lipid peroxide map of the selenium-doped prussian blue nanoenzyme prepared in example 1.
FIG. 9 is a hemolysis experiment diagram of the selenium-doped Prussian blue nanoenzyme prepared in example 1.
Fig. 10 is a MTT experimental graph of the selenium-doped prussian blue nanoenzyme prepared in example 1.
Fig. 11 is a statistical plot of colon length after prevention and treatment of colitis in mice with selenium-doped prussian blue nanoenzyme prepared in example 1.
Fig. 12 is a statistical plot of colon length after delayed treatment of colitis in mice with selenium-doped prussian blue nanoenzymes prepared in example 1.
FIG. 13 is a flow chart of the selenium-doped Prussian blue nanoenzyme prepared in example 1 for treating Crohn's disease in rats.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention more comprehensible, specific embodiments thereof will be described in detail with reference to the following examples. The following is merely exemplary and illustrative of the inventive concept and various modifications, additions and substitutions of similar embodiments may be made to the described embodiments by those skilled in the art without departing from the inventive concept or exceeding the scope of the claims defined thereby.
Example 1
In this example, selenium-doped prussian blue nanoenzyme was prepared as follows:
(1) dissolving 3.0g of PVP in 30mL of HCL solution with the concentration of 0.01M, adding 132mg of potassium ferricyanide powder, and stirring until the mixture is uniformly mixed; placing the obtained mixed solution in an oven to react for 24 hours at the temperature of 80 ℃; after the reaction, the mixture was centrifuged at 14000rpm for 10min, washed with water 3 times, and lyophilized to obtain Prussian blue powder (PB NCs).
(2) Dissolving 30mg of Prussian blue powder in 30mL of HCL solution with the concentration of 1M, adding 300mg of PVP, stirring at room temperature for 3h, and placing in an oven for reaction at 140 ℃ for 3 h; after the reaction is finished, 14000rpm is carried out, centrifugation is carried out for 10min, water washing is carried out for 3 times, and freeze-drying is carried out, so as to obtain hollow mesoporous Prussian blue powder (HMPB NCs).
(3) Adding 6mg/mL sodium selenite solution with the same volume into 2mg/mL hollow mesoporous Prussian blue aqueous dispersion, stirring at normal temperature for reaction for 30 minutes, slowly adding 12mg/mL ascorbic acid solution with the same volume with the hollow mesoporous Prussian blue aqueous dispersion, and reacting at normal temperature for 3 hours; after the reaction, centrifugation, dialysis (dialysis in a dialysis bag with Mw of 3500Da for 24 hours) and lyophilization were performed to obtain selenium-doped prussian blue nanoenzyme powder (Se-HMPB NCs).
FIG. 2 is a transmission electron microscope image of the selenium-doped Prussian blue nanoenzyme obtained in the present embodiment, and it can be seen from the image that the nanoenzyme has a nanosheet structure and a diameter of 120-140 nm.
Fig. 3 is an X-ray photoelectron spectrum of the selenium-doped prussian blue nanoenzyme obtained in this example, and it can be seen from the diagram that the valence of selenium element contained in the nanoenzyme is 0.
Fig. 4 is a hydroxyl radical scavenging diagram of the selenium-doped prussian blue nanoenzyme obtained in this example, which is characterized by the following steps: salicylic Acid (SA) is used as a chromogenic substrate, and the ability of the selenium-doped Prussian blue nanoenzyme to remove hydroxyl radicals is detected in an acetic acid/sodium acetate (HAc/NaAc) buffer solution. First 100. mu.L of 20mM H2O2With 100. mu.L of 10mM FeSO4·7H2O is added into 400. mu.L HAc/NaAc buffer solution and mixed for reaction for 10 min. Then adding 300 mu L of different materials (PB, HMPB or Se-HMPB) into the reaction system (the final concentration of the materials in the system is 25, 50 or 100 mu g/mL) for reaction for 1h, and setting FeSO4+H2O2Is a positive control. Finally, 100. mu.L of 18mM SA was added and the reaction was developed in the dark for 15min, and then the UV absorption at 510nm was measured. As can be seen from the figure, the capacity of eliminating hydroxyl radicals of the selenium-doped Prussian blue nanoenzyme with the concentration of 100 mu g/mL is about 65%.
FIG. 5 shows the results obtained in this exampleThe selenium-doped Prussian blue nano-enzyme in-vitro simulation glutathione peroxidase map is characterized in that: and (3) detecting the GPx enzyme-like activity of the selenium-doped Prussian blue nano-enzyme in a PBS (phosphate buffer solution) by taking dinitrobenzoic acid (DTNB) as a substrate. First, a sample (PB, HMPB or Se-HMPB) with a final concentration of 100. mu.g/mL was mixed with GSH at a final concentration of 1mM and H at a final concentration of 2mM2O2The mixture was reacted for 1h in the dark. Then, 100. mu.L of the reaction mixture was added to 800. mu.L of PBS, and after mixing, 100. mu.L of 1mM DTNB was added and the mixture was developed in the dark for 10 min. The UV absorption at 412nm was determined after centrifugation at 3000rpm for 5 min. As can be seen from the figure, the GSH-consuming capacity of the selenium-doped Prussian blue nanoenzyme of 100. mu.g/mL is about 70%.
Fig. 6 is a cell reactive oxygen species staining chart of the selenium-doped prussian blue nanoenzyme obtained in this example, which is characterized by the following steps: to verify the clear effect of selenium-doped Prussian blue nanoenzymes on intracellular reactive oxygen species, 100. mu.L (1X 10) per well was seeded in 96-well plates5) And (3) incubating for 24 h. The supernatant was aspirated and 3mM H was prepared in DMEM2O2100 mu L of the prepared solution is added into each well for incubation for 4 h. The experiments were divided into the following groups: control (fresh Medium), 3mM H2O2、100μg/mL PB、100μg/mL Se-HMPB、3mM H2O2+100μg/mL PB、3mM H2O2+ 100. mu.g/mL Se-HMPB. And (3) after incubation, sucking the supernatant, washing the supernatant for three times by PBS (phosphate buffer solution), adding 10 mu M of DCFH-DA dye solution prepared by DMEM into each hole, incubating the mixture for 30min at 37 ℃ in the dark, washing the mixture for three times by a serum-free culture medium, and taking a picture by a fluorescence microscope. As can be seen from the figure, H2O2The +100 mu g/mL Se-HMPB group has no obvious green fluorescence, which indicates that the Se-HMPB group has better capacity of eliminating active oxygen in cells.
Fig. 7 is a cell death and viability staining chart of the selenium-doped prussian blue nanoenzyme obtained in this example, which is characterized by the following steps: to demonstrate that selenium-doped prussian blue nanoenzymes can protect NCM460 cells by scavenging ROS. After 24H incubation in the above experiment, the supernatant was aspirated and 3mM H in DMEM2O2100 mu L of the prepared solution is added into each well for incubation for 4 h. The experiments were divided into the following groups: control (fresh Medium), 3mM H2O2、50μg/mL PB、100μg/mL PB、50μg/mL Se-HMPB、100μg/mL Se-HMPB、3mM H2O2+50μg/mL PB、3mM H2O2+100μg/mL PB、3mM H2O2+50μg/mL Se-HMPB、3mM H2O2+ 100. mu.g/mL Se-HMPB. After the incubation is finished, the Calcein-AM/PI dye is prepared by PBS, 100 mu L of dye is added into each hole after the supernatant is absorbed, the incubation is carried out for 30min under the dark condition at 37 ℃, the PBS is washed for 2 times, and 100 mu L of PBS is added for carrying out fluorescence photography. As can be seen from the figure, 3mM H2O2The +100 mu g/mL Se-HMPB group has no obvious red fluorescence, which indicates that the Se-HMPB group has a better protective effect on intestinal epithelial cells.
Fig. 8 is a cell-inhibitory lipid peroxide map of the selenium-doped prussian blue nanoenzyme prepared in this example, which is characterized by: to demonstrate that selenium-doped prussian blue nanoenzyme can inhibit lipid peroxidation in cells. 1mL (5X 10) of the seed was inoculated into each well of a 24-well plate5) And (3) incubating for 24 h. After the incubation, the supernatant was aspirated, 1. mu.g/mL of LPS was prepared in DMEM, and 1mL of the prepared solution was added to each well for incubation for 12 hours. The experiments were divided into the following groups: control (fresh medium), 1. mu.g/mL LPS, 100. mu.g/mL PB, 100. mu.g/mL Se-HMPB, 1. mu.g/mL LPS + 100. mu.g/mL PB, 1. mu.g/mL LPS + 100. mu.g/mL Se-HMPB. After incubation, 100. mu.L of IP cell lysate was added to each well and lysed for 30min on ice. After complete lysis of the cells, the supernatant was collected by centrifugation (12000g, 10min, 4 ℃). And detecting the MDA level of the cell lysis supernatant by adopting a lipid oxide MDA detection kit. As can be seen from the figure, Se-HMPB NCs are effective in reducing intracellular lipid peroxide levels by only 9.2nmol/105cell。
Fig. 9 is a hemolysis experiment chart of the selenium-doped prussian blue nanoenzyme obtained in this example, which is characterized by the following steps: diluting Se-HMPB NCs aqueous dispersion to 25 mug/mL, 50 mug/mL, 100 mug/mL, 150 mug/mL and 200 mug/mL, adding 0.2mL of the solutions with different concentrations into 0.2mL of treated blood (500 mug L of fresh blood and 4.5mL of physiological saline for centrifugal washing for 5-8 times, wherein the centrifugal rotation speed is 3000rpm, the centrifugal time is 10min, discarding supernatant after supernatant is clear and transparent, fixing the volume to 5mL by using physiological saline), mixing the solutions with 0.6mL of physiological saline, incubating for 4h at 37 ℃, centrifuging for 10min at 3000rpm, sucking the supernatant, measuring the absorbance at OD 541nm, and calculating the hemolysis rate. From the figure, the hemolysis rate of the selenium-doped prussian blue nano-enzyme is lower than 5%, which indicates that the biocompatibility of the material is good.
Fig. 10 is an MTT experimental graph of the selenium-doped prussian blue nanoenzyme obtained in this example, which is characterized by: each well was inoculated with 100. mu.L (1X 10)4) And (3) incubating for 24 h. The supernatant was aspirated, Se-HMPB (0, 25, 50, 100, 150, 200, 300. mu.g/mL) was added at various concentrations in DMEM, 100. mu.L/well, and incubated for 24 h. After the incubation was complete, 20. mu.L of MTT (5mg/mL) was added to each well and the incubator was incubated for 4 h. Finally, the supernatant was aspirated and the culture was stopped by adding 150 μ L DMSO per well. Incubating at 37 ℃ in dark for 10min, and measuring the absorbance at 490 nm. The graph shows that the cell survival rates of the selenium-doped Prussian blue nano-enzyme are higher than 85%, which indicates that the biological safety of the material is good.
To verify the ability of the selenium-doped prussian blue nanoenzyme obtained in this example to alleviate colitis, the following tests were performed: in the prophylactic treatment colitis model, normal mice were randomized into 6 groups (n-7 per group): (1) PBS + Normal Drinking Water (2) PBS + 3% DSS (3) PB (10mg/kg) + 3% DSS (4) Se-HMPB (10mg/kg) + 3% DSS (5) PB (20mg/kg) + 3% DSS (6) Se-HMPB (20mg/kg) + 3% DSS. Mice in the experimental group were fed with 3% DSS for 7 consecutive days, the administration group was given through the tail vein on days 1, 3, and 5, the mice were sacrificed on the ninth day, and the colon length of each group of mice was recorded. As can be seen from FIG. 11, the colon length of the Se-HMPB (20mg/kg) + 3% DSS group mice is close to 7cm, which is comparable to that of the normal group mice.
To further verify the ability of the selenium-doped prussian blue nanoenzyme obtained in this example to alleviate colitis, the following tests were performed: in the delayed treatment colitis model, normal mice were randomized into 6 groups (n-7 per group): (1) PBS + Normal Drinking Water (2) PBS + 3% DSS (3) PB (10mg/kg) + 3% DSS (4) Se-HMPB (10mg/kg) + 3% DSS (5) PB (20mg/kg) + 3% DSS (6) Se-HMPB (20mg/kg) + 3% DSS. Mice in the experimental group were fed with 3% DSS for 7 consecutive days, the administration group was administered via the tail vein on days 9, 11, and 13, the mice were sacrificed on day 16, and the colon length of each group of mice was recorded. As can be seen from FIG. 12, the colon length of the Se-HMPB (20mg/kg) + 3% DSS group mice is close to 7.3cm, which is not much different from that of the normal group mice.
To further verify the effect of the selenium-doped prussian blue nanoenzyme obtained in this example in inhibiting T cell differentiation to treat crohn's disease, the following tests were performed: in the rat crohn's disease model, normal mice were randomly divided into 3 groups (n-7 per group): (1) PBS + TNBS (2) PB (10mg/kg) + TNBS (3) Se-HMPB (10mg/kg) + TNBS. Rats in experimental groups were subjected to TNBS enema every other day for 14 consecutive days, and the administration groups were administered through the tail vein on days 3, 6, 9, 12, and 14, and sacrificed on day 16. Cells were extracted from the ends of the colon by dissection and were detected on the machine after labelling the cells with Anti-CD3, Anti-CD45 and Anti-CD 4. As can be seen from FIG. 13, Se-HMPB (10mg/kg) + CD4 of TNBS group rats+The number of T positive cells accounts for about 11.3% of the total immune cells, which is significantly lower than that of the control group.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.
Claims (10)
1. A selenium-doped Prussian blue nano-enzyme for regulating intestinal cells to treat colitis is characterized in that: the selenium-doped Prussian blue nano-enzyme takes hollow mesoporous Prussian blue as a core, and selenium element is doped on the surface of the hollow mesoporous Prussian blue.
2. The selenium-doped prussian blue nanoenzyme of claim 1, wherein: the particle size of the selenium-doped Prussian blue nano-enzyme is 120-140 nm, and the valence state of the selenium element doped on the surface is 0 valence.
3. A method for preparing the selenium-doped prussian blue nanoenzyme of claim 1 or 2, comprising the steps of:
(1) dissolving 2.5-3.5g PVP in 30mL of HCL solution with the concentration of 0.01-0.05M, adding 100-150mg of potassium ferricyanide powder, and stirring until the mixture is uniformly mixed; placing the obtained mixed solution in an oven to react for 24 hours at the temperature of 80 ℃; after the reaction is finished, centrifuging, washing with water, and freeze-drying to obtain Prussian blue powder;
(2) dissolving 20-30mg Prussian blue powder in 30mL of HCL solution with the concentration of 0.8-1M, adding 200-300mg PVP, stirring at room temperature for 3-3.5h, and then placing in an oven for reaction at 140 ℃ for 3 h; after the reaction is finished, centrifuging, washing and freeze-drying to obtain hollow mesoporous Prussian blue powder;
(3) adding 4-6mL of 2mg/mL sodium selenite solution into 2mL of 2mg/mL hollow mesoporous Prussian blue aqueous dispersion, stirring at normal temperature for reaction for 20-30min, slowly adding 8-12mL of 2mg/mL ascorbic acid solution, and reacting at normal temperature for 3 h; after the reaction is finished, centrifuging, dialyzing and freeze-drying to obtain selenium-doped Prussian blue nano-enzyme powder;
wherein the mass ratio of the ascorbic acid to the sodium selenite is 2: 1.
4. The production method according to claim 3, characterized in that: the dialysis was performed in a dialysis bag with Mw of 3500Da for 24 h.
5. The use of the selenium-doped prussian blue nanoenzyme of claim 1 or 2, wherein: used for preparing the nano enzyme for regulating intestinal cells and treating the colitis.
6. Use according to claim 5, characterized in that: the selenium-doped Prussian blue nanoenzyme can not only eliminate hydroxyl free radicals, but also has the function of glutathione peroxidase.
7. Use according to claim 5, characterized in that: the selenium-doped Prussian blue nanoenzyme has no hemolytic behavior and cytotoxicity.
8. Use according to claim 5, characterized in that: the selenium-doped Prussian blue nanoenzyme has the capability of eliminating active oxygen.
9. Use according to claim 5, characterized in that: the selenium element contained in the selenium-doped Prussian blue nanoenzyme can inhibit lipid peroxidation of epithelial cells of the intestinal tract and restore the barrier of the intestinal tract.
10. Use according to claim 5, characterized in that: the selenium element contained in the selenium-doped Prussian blue nanoenzyme can inhibit T cell differentiation and reduce T cell-mediated inflammatory reaction.
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| CN116983328A (en) * | 2023-09-26 | 2023-11-03 | 中国人民解放军军事科学院军事医学研究院 | Nanometer enzyme complex and application thereof |
| CN115753649B (en) * | 2022-10-09 | 2024-04-19 | 河北师范大学 | Silicon-based Prussian blue nano-enzyme, preparation method and application thereof in colorimetric detection of glutathione |
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| CN115753649B (en) * | 2022-10-09 | 2024-04-19 | 河北师范大学 | Silicon-based Prussian blue nano-enzyme, preparation method and application thereof in colorimetric detection of glutathione |
| CN116983328A (en) * | 2023-09-26 | 2023-11-03 | 中国人民解放军军事科学院军事医学研究院 | Nanometer enzyme complex and application thereof |
| CN116983328B (en) * | 2023-09-26 | 2023-12-12 | 中国人民解放军军事科学院军事医学研究院 | Nanometer enzyme complex and application thereof |
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