A rare earth material with peroxidase-like activity and its preparation method and application

Abstract

This invention provides a rare earth material with peroxidase-like activity, its preparation method, and its application. The preparation method includes the following steps: dissolving rare earth oxides in a strong acid solution and dissolving under heating conditions to obtain a rare earth salt mother liquor; diluting the rare earth salt mother liquor with the strong acid solution to obtain a rare earth salt solution; adding N,N,N'-trimethylethylenediamine to the rare earth salt solution to react; adjusting the pH of the reaction solution to 1.5-7.5 after the reaction is complete; allowing the reaction solution to stand overnight at room temperature; centrifuging to collect the precipitate; and washing to obtain the rare earth material with peroxidase-like activity. The rare earth material of this invention can efficiently catalyze the decomposition of hydrogen peroxide, remove reactive oxygen species in diabetic wounds, significantly reduce oxidative stress, and has better chemical stability, enabling it to continue to function in the wound environment.

Description

Technical Field

[0001] This invention belongs to the field of new materials, and in particular relates to a rare earth material with peroxidase-like activity, its preparation method and application. Background Technology

[0002] Diabetic wounds, particularly diabetic foot ulcers, pose a significant challenge globally, with oxidative stress being a major mechanism in chronic diabetic foot ulcers. During the healing process, diabetic wounds activate inflammation, leading to the generation of reactive oxygen species (ROS). Studies have shown that H2O2 levels in chronic wound model mice increase fivefold within 4 hours post-injury, while catalase (CAT) and glutathione peroxidase (GPx) activities are similar to those in healthy mice. Therefore, in the early stages of wound healing, elevated ROS levels, coupled with unchanged antioxidant enzyme activity, result in oxidative stress at the wound site. Currently, traditional antioxidant hydrogels contain polyphenols such as curcumin, dopamine, and tannins. However, these substances are unstable and rapidly released into wound dressings, limiting their sustained effects. Novel nanozymes exhibit excellent antioxidant properties due to their enhanced catalytic function, stability, design flexibility, and extended cycle time. Therefore, the development of rare-earth materials with peroxidase-like activity is crucial for the treatment of diabetic wounds. Summary of the Invention

[0003] In view of this, the present invention aims to overcome the defects in the prior art and proposes a rare earth material with peroxidase-like activity, its preparation method and application.

[0004] To achieve the above objectives, the technical solution of the present invention is implemented as follows: In a first aspect, the present invention provides a method for preparing a rare earth material with peroxidase-like activity, the method comprising the following steps: S1: Dissolve rare earth oxides in a strong acid solution and then dissolve them under heating conditions to obtain rare earth salt mother liquor; S2: Dilute the strong acid solution with the rare earth salt mother liquor to obtain the rare earth salt solution; S3: N,N,N'-trimethylethylenediamine was added to a rare earth salt solution for reaction. After the reaction was completed, the pH of the reaction solution was adjusted to 1.5-7.5. The reaction solution was allowed to stand overnight at room temperature, and the precipitate was collected by centrifugation. After washing, a rare earth material with peroxidase-like activity was obtained.

[0005] Preferably, the rare earth oxide is one or a mixture of several of cerium oxide, lanthanum oxide, and yttrium oxide.

[0006] Preferably, the strong acid is nitric acid and / or hydrochloric acid.

[0007] Preferably, the concentration of the strong acid solution in step S1 is 13.0-15.5 mol / L.

[0008] Preferably, the concentration of the rare earth salt mother liquor in step S1 is 2.1-2.6 mol / L.

[0009] Preferably, the concentration of the strong acid in the rare earth salt solution in step S2 is 2-4 mol / L.

[0010] Preferably, the concentration of the rare earth salt solution in step S2 is 0.17-0.5 mol / L.

[0011] Preferably, in step S3, the volume ratio of N,N,N'-trimethylethylenediamine to the rare earth salt solution is (1~3):(0.3~1.5).

[0012] Secondly, the present invention also provides the application of the above-mentioned rare earth materials with peroxidase-like activity in the preparation of drugs for treating skin inflammation, drugs for repairing diabetic wounds, or dressings.

[0013] Thirdly, the present invention also provides rare earth materials with peroxidase-like activity prepared by the above preparation method.

[0014] Application of rare earth materials with peroxidase-like activity in the preparation of drugs for treating skin inflammation.

[0015] Fourthly, the present invention also provides the application of the above-mentioned rare earth materials with peroxidase-like activity in the preparation of drugs or dressings for the repair of diabetic wounds.

[0016] This invention utilizes a chemical precipitation method to generate rare earth-based precipitated materials with nanoscale or specific coordination structures by coordinating and hydrolyzing N,N,N'-trimethylethylenediamine with high oxidation state rare earth ions in an acidic aqueous solution.

[0017] Rare earth oxides dissolve upon heating in concentrated strong acid, generating a high-concentration rare earth salt mother liquor, providing highly active metal centers for the reaction. Upon addition of N,N,N'-trimethylethylenediamine, both tertiary nitrogen atoms in its molecule possess strong coordination capabilities. In an acidic environment, they coordinate with Ce(IV) ions, partially replacing water molecules or nitrate ions surrounding Ce(IV). By adjusting the pH of the reaction system to 1.5-7.5 (weakly acidic to near neutral), a controlled hydrolysis reaction is induced in the final complex, forming a rare earth oxyhydroxyl compound or nanocluster rich in oxygen vacancies, possessing unsaturated coordination sites and specific surface chemistry. During static standing at room temperature, this ultimately forms a stable solid precipitate.

[0018] This invention utilizes a strategy of "strong acid dissolution-ligand regulation-controlled hydrolysis" to prepare a rare-earth nanomaterial with a unique structure and rich in active Ce(III) / Ce(IV) redox pairs. Its peroxidase-like activity originates from the nanosize effect, abundant surface oxygen vacancies, and optimized electronic structure. This activity enables it to intelligently respond to and regulate the pathological oxidative stress microenvironment, thus demonstrating great application potential in treating skin inflammation and promoting the repair of diabetic wounds.

[0019] Compared with the prior art, the present invention has the following advantages: (1) The rare earth material of the present invention can efficiently catalyze the decomposition of hydrogen peroxide (H2O2), remove reactive oxygen species (ROS) in diabetic wounds, and significantly reduce oxidative stress; (2) The rare earth material of the present invention has better chemical stability and can continue to function in the wound environment; (3) The rare earth materials prepared by the present invention are synthesized by solution method, which is simple to operate, mild under mild conditions, and easy to scale up production. Attached Figure Description

[0020] Figure 1 SEM images of the rare earth material prepared in Example 1 at different scales; Figure 2 The figure shows the experimental results of reactive oxygen species generation in bacteria using Example 1. Figure 3 The image shows the results of the Escherichia coli agar plate experiment in Example 2. Figure 4 This is a comparison chart of the survival rates of Escherichia coli in Application Example 2. Detailed Implementation

[0021] The embodiments of the present invention are described in detail below. The embodiments described below are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0022] In this document, unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0023] In this document, when values ​​are described as ranges, it should be understood that such disclosure includes disclosure of all possible subranges within that range, as well as the specific numerical values ​​falling within that range, regardless of whether the specific numerical value or specific subrange is explicitly specified.

[0024] In this article, the terms "multiple" or "more than" are used unless otherwise specified, referring to a quantity greater than or equal to 2. For example, "one or more" means one or more types.

[0025] In this document, the terms "preferred" and "more preferred" are used only to describe implementation methods or embodiments with better effects, and should be understood as not constituting a limitation on the scope of protection of this invention.

[0026] In this document, terms such as "further" are used for descriptive purposes to indicate differences in content, but should not be construed as limiting the scope of protection of this invention.

[0027] In this article, the term "and / or" describes an association between objects, indicating that three relationships can exist. For example, A and / or B means: A or B, or A and B.

[0028] In this document, the term "about" means a specified value of + / - 10%, preferably + / - 5%, and more preferably + / - 1%.

[0029] In this article, the terms “include,” “including,” “have,” “contain,” etc., are all open-ended terms, meaning that they include but are not limited to.

[0030] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar to or equivalent to those described herein may be used in the implementation or testing of this invention.

[0031] Cerium oxide (CeO2), concentrated nitric acid (HNO3), and N,N,N'-trimethylethylenediamine (C5H) 14 N2 was purchased from Aladdin Chemicals Ltd. All chemicals were used without further treatment. Distilled water (ρ=18.2 MΩ·cm, 25℃) came from the Millipore Milli-Q water purification system. Gram-negative bacteria. E. coli All reagents are from Beijing Sihuan Biopharmaceutical Co., Ltd. Unless otherwise specified, all reagents are standard biochemical reagents; all experimental methods, unless otherwise specified, are standard methods.

[0032] The present invention will be described in detail below with reference to embodiments.

[0033] Example 1 (1) Preparation of Ce(IV) solution First, 20 g of CeO2 was dissolved in 50 mL of concentrated nitric acid (14.5 mol / L) and heated at 60 °C to obtain a Ce(IV) mother liquor of 2.32 mol / L. Next, 200 mL of 0.5 mol / L nitric acid was added to the above solution to dilute it, preparing a Ce(IV) solution of 0.46 mol / L, at which point the nitric acid concentration was 3.3 mol / L.

[0034] (2) Preparation of cerium oxide particles Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 1 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 0.6 mL of 0.46 mol / L Ce(IV) solution to the beaker; the resulting solution is orange. After stirring for another 15 minutes, the solution turns pale yellow. Adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) with 30-33% NH3·H2O. Subsequently, Ce particles form, and the solution becomes pale white and turbid. Let the solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain a rare earth material with peroxidase-like activity. SEM images of the prepared rare earth material at different resolutions are shown below. Figure 1 As shown.

[0035] Example 2 (1) Preparation of Ce(IV) solution First, 18.9 g of CeO2 was dissolved in 50 mL of concentrated nitric acid (15 mol / L) and heated at 60 °C to obtain a 2.2 mol / L Ce(IV) mother liquor. Next, 183 mL of 0.5 mol / L nitric acid was added to the above solution to dilute it, preparing a 0.47 mol / L Ce(IV) solution. At this point, the nitric acid concentration was 3.6 mol / L.

[0036] Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 3 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 1 mL of 0.47 mol / L Ce(IV) solution to the beaker; the resulting solution is orange. After stirring for another 15 minutes, the solution turns pale yellow. Adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) using 30-33% NH3·H2O. Subsequently, Ce particles form, and the solution becomes pale white and turbid. Let the solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain rare earth material with peroxidase-like activity.

[0037] Example 3 (1) Preparation of Ce(IV) solution First, 21.5 g of CeO2 was dissolved in 50 mL of concentrated nitric acid (13 mol / L) and heated at 60 °C to obtain a 2.5 mol / L Ce(IV) mother liquor. Next, 327 mL of 0.5 mol / L nitric acid was added to the above solution to dilute it, preparing a 0.33 mol / L Ce(IV) solution. At this point, the nitric acid concentration was 2.16 mol / L.

[0038] Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 2 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 0.5 mL of 0.33 mol / L Ce(IV) solution to the beaker; the resulting solution is orange. After stirring for another 15 minutes, the solution turns pale yellow. Adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) using 30-33% NH3·H2O. Subsequently, Ce particles form, and the solution becomes pale white and turbid. Let the solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain rare earth material with peroxidase-like activity.

[0039] Example 4 (1) Preparation of Ce(IV) solution First, 18 g of CeO2 was dissolved in 50 mL of concentrated nitric acid (15.5 mol / L) and heated at 60 °C to obtain a 2.1 mol / L Ce(IV) mother liquor. Next, 565 mL of 0.8 mol / L nitric acid was added to the above solution to dilute it, preparing a 0.17 mol / L Ce(IV) solution. At this point, the nitric acid concentration was 2 mol / L.

[0040] Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 2 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 0.3 mL of 0.17 mol / L Ce(IV) solution to the beaker; the resulting solution is orange. After stirring for another 10 minutes, the solution turns pale yellow. Adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) using 30-33% NH3·H2O. Subsequently, Ce particles form, and the solution becomes pale white and turbid. Let the solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain rare earth material with peroxidase-like activity.

[0041] Example 5 (1) Preparation of Ce(IV) solution First, 22.4 g of CeO2 was dissolved in 50 mL of concentrated nitric acid (15.5 mol / L) and heated at 60 °C to obtain a 2.6 mol / L Ce(IV) mother liquor. Next, 210 mL of 1.26 mol / L nitric acid was added to the above solution to dilute it, preparing a 0.5 mol / L Ce(IV) solution. At this point, the nitric acid concentration was 4 mol / L.

[0042] Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 2 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 1.5 mL of 0.5 mol / L Ce(IV) solution to the beaker; the resulting solution is orange. After stirring for another 15 minutes, the solution turns pale yellow. Adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) using 30-33% NH3·H2O. Subsequently, Ce particles form, and the solution becomes pale white and turbid. Let the solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain rare earth material with peroxidase-like activity.

[0043] Comparative Example 1 In this comparative example, the same preparation method as in Example 1 was used, but the step of adding N,N,N'-trimethylethylenediamine was omitted. The prepared rare earth material mainly consisted of cerium oxide and concentrated nitric acid.

[0044] Comparative Example 2 Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 1 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 0.6 mL of 0.48 mol / L cerium nitrate solution to the beaker. After the reaction is complete, adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) with 30-33% NH3·H2O. Let the above solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain the rare earth material.

[0045] Comparative Example 3 (1) Preparation of Ce(IV) solution First, 20 g of CeO2 was dissolved in 50 mL of concentrated nitric acid (14.5 mol / L) and heated at 60 °C to obtain a 2.32 mol / L Ce(IV) stock solution. Next, deionized water was added to dilute the above solution to prepare a 0.46 mol / L Ce(IV) solution. Simultaneously, the pH of the Ce(IV) solution was adjusted to neutral using 30-33% NH3·H2O.

[0046] (2) Preparation of cerium oxide particles Add 5 mL of water and ethanol (1:1) to a 20 mL beaker and stir at 800 rpm for 5 minutes. Add 1 mL of N,N,N'-trimethylethylenediamine to the beaker and stir for at least 5 minutes. Then, add 0.6 mL of 0.48 mol / L Ce(IV) solution to the beaker; the resulting solution is orange. After stirring for another 15 minutes, the solution turns pale yellow. Adjust the pH of the reaction solution to the desired synthesis pH (1.5 < pH < 7.5) using 30-33% NH3·H2O. Subsequently, Ce particles form, and the solution becomes pale white and turbid. Let the solution stand overnight at room temperature while stirring at 800 rpm. Collect the particles by centrifugation and wash three times with ethanol to obtain rare earth material with peroxidase-like activity.

[0047] Comparative Example 4: Traditional Antioxidant Hydrogel This comparative example uses a commercially available antioxidant hydrogel (purchased from Jilin Boshijia Pharmaceutical Co., Ltd.).

[0048] Test Example 1: Experiment on the generation of reactive oxygen species in bacteria Escherichia coli were treated with the rare earth materials prepared in Examples 1-4, Comparative Examples 1, 3, and 4, and the antioxidant hydrogel in Comparative Example 2, respectively. After 9 hours of culture, E. coli were collected, and intracellular reactive oxygen species (ROS) were measured using standard detection methods. Bacterial samples were treated with 30 μL of 1.05 mmol of 2',7'-dichlorofluorescein diacetate (DCFH-DA), a specific indicator that oxidizes non-fluorescent DCFH-DA to fluorescent DFC. The fluorescence intensity of the bacterial suspension at 10 minutes of reaction was recorded using a Shimadzu RF-5301PC fluorescence spectrophotometer (excitation wavelength = 488 nm; emission wavelength = 525 nm). Since fluorescence intensity represents the free radical scavenging rate, the lower the fluorescence intensity, the faster the reactive oxygen species scavenging rate.

[0049] The results are as follows Figure 2As shown, the results indicate that the reactive oxygen species scavenging efficiency of Comparative Example 1 and traditional antioxidant hydrogels is significantly lower than that of the material of this invention, indicating that traditional antioxidant hydrogels cannot effectively inhibit the accumulation of free radicals during the inflammatory phase.

[0050] Test Example 2: Antibacterial Test (1) Bacterial culture Gram-negative *Escherichia coli* were cultured on Luria-Bertani (LB) agar at 37°C. Additionally, single colonies of *E. coli* were inoculated into LB medium and cultured at 37°C and 180 rpm for 14–16 hours to obtain bacterial suspensions. Freshly harvested cells were used for each experiment. *E. coli* growth was assessed by measuring the optical density (OD600) at 600 nm. Subsequent experiments used bacterial suspensions with OD600 = 0.4–0.6.

[0051] (2) Agar plate experiment The rare earth materials prepared in Examples 1-4, Comparative Examples 1-3, and the antioxidant hydrogel in Comparative Example 4 were mixed with *E. coli* suspension (OD600=0.2). After incubation at 37°C for 3 hours, 0.2 mL of the diluted bacterial suspension was spread onto LB agar, and the bacterial cell viability was determined by the standard plate count method. After culturing for 16-18 hours, the cells were photographed and colonies were counted at 37°C.

[0052] Similarly, LB liquid medium containing E. coli at a density of OD600 = 0.1~0.2 was selected as a blank control. 100 μL of the solution was transferred to a blank culture dish and incubated in a constant temperature incubator according to the above procedure. Three groups of experiments were performed in parallel.

[0053] The agar coating effects obtained from different immersion times were analyzed. For coating results with the same incubation time, images with uniform distribution were selected for display. For example... Figure 3 As shown, the E. coli in the blank control group grew better and was evenly distributed on the agar plate. By counting the number of colonies in the three parallel experiments, the average number of colonies in the three plates was 684. When E. coli was incubated with rare earth materials, the number of colonies decreased significantly. After statistical analysis, the average number of colonies in Examples 1-4 was 0 CFU. The average number of colonies in Comparative Examples 1-4 were 360 ​​CFU, 134 CFU, 148 CFU, and 68 CFU, respectively.

[0054] like Figure 4As shown, the bacterial survival rate of three parallel experiments was statistically analyzed. The blank group was used as a control. The survival rate of E. coli in the blank group was 100%. After incubation with rare earth materials, the survival rate of E. coli decreased to 0%, which shows that rare earth materials can effectively inhibit bacteria, causing all E. coli to die in the end.

[0055] The rare earth material in Comparative Example 1, without the addition of N,N,N'-trimethylethylenediamine, showed an inhibition rate of only about 47.4% against Escherichia coli. The inhibition rates of other comparative examples were also significantly lower than the 99.9% of the rare earth material in Example 1. This indicates that N,N,N'-trimethylethylenediamine plays a key synergistic role in the rare earth material system of the present invention, significantly improving the antibacterial performance of the material.

[0056] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for producing a rare earth material having a peroxidase-like activity, characterized by: The preparation method includes the following steps: S1: Dissolve rare earth oxides in a strong acid solution and then dissolve them under heating conditions to obtain rare earth salt mother liquor; S2: Dilute the strong acid solution with the rare earth salt mother liquor to obtain the rare earth salt solution; S3: N,N,N'-trimethylethylenediamine was added to a rare earth salt solution for reaction. After the reaction was completed, the pH of the reaction solution was adjusted to 1.5-7.

5. The reaction solution was allowed to stand overnight at room temperature, and the precipitate was collected by centrifugation. After washing, a rare earth material with peroxidase-like activity was obtained.

2. The method of producing a rare earth material having a peroxidase-like activity according to claim 1, characterized by: The rare earth oxide is one or a mixture of several of cerium oxide, lanthanum oxide, and yttrium oxide.

3. The method of claim 1, wherein the method is characterized by: The strong acid is nitric acid and / or hydrochloric acid.

4. The method for preparing rare earth materials with peroxidase-like activity according to claim 1, characterized in that: The concentration of the strong acid solution in step S1 is 13.0-15.5 mol / L.

5. The method for preparing rare earth materials with peroxidase-like activity according to claim 1, characterized in that: The concentration of the rare earth salt mother liquor in step S1 is 2.1-2.6 mol / L.

6. The method for preparing rare earth materials with peroxidase-like activity according to claim 1, characterized in that: In step S2, the concentration of the strong acid in the rare earth salt solution is 2-4 mol / L.

7. The method for preparing rare earth materials with peroxidase-like activity according to claim 1, characterized in that: In step S2, the concentration of the rare earth salt solution is 0.17-0.5 mol / L.

8. The method for preparing rare earth materials with peroxidase-like activity according to claim 1, characterized in that: In step S3, the volume ratio of N,N,N'-trimethylethylenediamine to the rare earth salt solution is (1~3): (0.3~1.5).

9. The rare earth material with peroxidase-like activity prepared by the preparation method according to any one of claims 1-8.

10. The use of the rare earth material with peroxidase-like activity as described in claim 9 in the preparation of drugs for treating skin inflammation, drugs for repairing diabetic wounds, or dressings.

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