Method for detecting magnetite nano-enzyme activity and regeneration method thereof

Abstract

The present application relates to the field of environmental catalysis and water treatment engineering, and specifically relates to a method for detecting magnetite nanoscale enzyme activity and a regeneration method thereof. The present application removes the active layer on the surface of the magnetite nanoscale enzyme by physical etching, determines the change in the Fe isotope composition of the sample before and after etching, determines the activity state of the magnetite nanoscale enzyme according to the Fe + is easy to enrich light isotopes, and Fe + is easy to enrich heavy isotopes, analyzes the distribution and migration behavior of the iron species in the inner and outer layers of the magnetite, and determines the migration of Fe + from the inside to the surface, which leads to the continuous regeneration of the active sites, and determines the activity state of the magnetite nanoscale enzyme by using the Fe isotope fractionation results, and establishes a catalyst activity judgment standard by using magnetite nanoscale enzymes in different oxidation states to realize the timely addition of Fe 2+ regenerates the active sites of the catalyst, which not only reflects the activity stability of the nanoscale enzyme in the field of water treatment and environmental remediation, but also avoids the blind addition of Fe 2+ and other pollution such as iron sludge.

Description

Technical Field

[0001] This invention relates to the field of environmental catalysis and water treatment engineering, specifically a method for detecting the activity of magnetite nanozymes and a method for their regeneration. Background Technology

[0002] With the increasing prominence of industrial wastewater discharge and recalcitrant organic pollutants, advanced oxidation technologies based on hydrogen peroxide activation have been widely applied in wastewater treatment and in-situ remediation projects due to their advantages such as mild reaction conditions, strong oxidizing power, and wide applicability to various pollutants. However, traditional Fenton systems generally suffer from harsh reaction conditions, large amounts of iron sludge formation, and insufficient operational stability, limiting their widespread adoption in practical engineering. Magnetite (Fe3O4) nanozymes, as a functional material with peroxidase-like activity, possess Fe... 2+ / Fe 3+ The coexisting anti-spinel structure enables electron transfer and redox cycles within the material, while also possessing advantages such as magnetic separability, high reusability, and strong environmental friendliness, gradually becoming an important catalytic material in water treatment and environmental remediation. During engineering operation, magnetite nanozymes are typically used as fixed-bed packing materials, suspended catalysts, or in-situ remediation materials for long-term operation.

[0003] Theoretically, Fenton-like reactions would lead to surface Fe... 2+ Continuously oxidized to Fe 3+ This leads to the depletion of active sites and a reduction in catalytic efficiency; therefore, Fe is typically added periodically in this field. 2+ To maintain the catalytic efficiency of magnetite nanozymes, or to replace them periodically. However, in actual water treatment operations, magnetite nanozymes can actually maintain high activity for a relatively long time. If Fe is added blindly... 2+ Alternatively, replacing the magnetite nanozyme would increase operating costs and generate pollution such as iron sludge. Therefore, the Fe content of the inner and outer layers of the magnetite nanozyme... 2+ / Fe 3+ Quantitative identification of ⁺ evolution has important engineering guiding significance.

[0004] Current technologies lack effective evaluation methods for the activity stability and degradation mechanism of magnetite nanozymes. Current research on this self-regeneration process mainly relies on valence state analysis, electrochemical testing, or conventional surface characterization methods. However, existing characterization methods generally have significant limitations, making it difficult to determine the impact of the magnetite interior on the regeneration of active sites. Therefore, they cannot provide direct evidence for determining the operational lifespan or making decisions on catalyst regeneration control. For example, XPS and XANES mainly reflect the average valence state information of Fe in the nanometer range of the material surface, making it difficult to distinguish the surface Fe... 2+Source of active sites; Mössbauer spectroscopy can distinguish Fe 2+ and Fe 3+ The percentage is given, but it only reflects the overall composition of magnetite and cannot provide information on Fe. 2+ and Fe 3+ Spatial distribution and migration direction information. It is evident that existing technologies primarily reflect static information about the valence state or local structure of iron, making it difficult to distinguish the source and migration direction of iron with different valence states on the magnetite surface and in the bulk phase. Furthermore, they cannot track the internal iron migration behavior in complex reaction systems, thus failing to clarify the self-regeneration pathway of active sites during magnetite nanozyme catalysis. Additionally, no existing technology has been found that reflects the operating state of the reaction catalyst and guides the regeneration of its active sites. Moreover, in complex water treatment systems, coexisting ions, organic pollutants, and reaction intermediates easily interfere with spectral signals, leading to significant uncertainty in mechanism determination.

[0005] Given the above background, there is an urgent need for a method that can directly distinguish surface Fe 2+ A novel method for deriving active sites and using them for evaluating catalyst operational stability and regenerating catalyst activity is proposed. This method is used to track the self-regeneration pathway of active sites in magnetite nanozymes during water treatment and environmental remediation, and to evaluate catalyst operational stability and regulate regeneration. This provides reliable technical support for the performance evaluation, material optimization, and engineering applications of magnetite nanozymes in environmental remediation and water treatment. Summary of the Invention

[0006] In view of this, the technical problem to be solved by the present invention is a method for detecting the activity of magnetite nanozymes and a method for regenerating them. The method provided by the present invention can detect the activity of magnetite nanozymes and thus regenerate them in a timely manner.

[0007] This invention provides a method for detecting the activity of magnetite nanozymes, comprising the following steps: S1) The magnetite nanozyme was dissolved in concentrated hydrochloric acid and evaporated to dryness, then concentrated nitric acid was added and evaporated to dryness again, and then dilute nitric acid was added to dissolve it to obtain the sample before etching. and, The outer layer of the magnetite nanozyme was physically etched, dissolved in concentrated hydrochloric acid and evaporated to dryness, then concentrated nitric acid was added and evaporated to dryness again, and then dilute nitric acid was added to obtain the etched sample. S2) Obtain the δ values ​​of the pre-etched and post-etched samples obtained in step S1), respectively. 56 / 54 The Fe value is obtained by measuring the δ before and after the magnetite nanoenzyme etching in step S1). 56 / 54 The absolute difference of Fe, according to the δ 56 / 54 The absolute difference in Fe values ​​is used to determine the activity of the magnetite nanozyme.

[0008] The method for detecting the activity of magnetite nanozymes provided by this invention is based on the method for determining the active state of magnetite nanozyme catalysts through iron isotope fractionation. It mainly determines the active state of the catalyst by measuring the change of Fe isotopes in oxidized magnetite before and after physical etching and by observing the self-regeneration path of the active sites of magnetite nanozymes through the Fe isotope fractionation phenomenon.

[0009] This invention divides magnetite nanozymes into two parts: one part is used for pretreatment to obtain a sample before etching, and the other part is used for post-etching pretreatment to obtain a sample after etching.

[0010] This invention involves dissolving magnetite nanozymes in concentrated hydrochloric acid, evaporating to dryness, adding concentrated nitric acid, evaporating again to dryness, and then adding dilute nitric acid to dissolve, obtaining the sample before etching. Specifically, the magnetite nanozymes are dissolved in concentrated hydrochloric acid, then evaporated to dryness at 90℃~110℃, concentrated nitric acid is added, and evaporation continues at 90℃~110℃, followed by dissolution with dilute nitric acid to obtain the sample before etching. The concentration of the concentrated hydrochloric acid is 10 mol / L~14 mol / L, preferably 12 mol / L; the concentration of the concentrated nitric acid is 14 mol / L~16 mol / L; and the concentration of the dilute nitric acid is 0.4 mol / L~0.6 mol / L, preferably 0.5 mol / L.

[0011] This invention involves physically etching the outer layer of magnetite nanozymes, dissolving it in concentrated hydrochloric acid, evaporating it to dryness, adding concentrated nitric acid, evaporating again to dryness, and then adding dilute nitric acid to obtain the etched sample. Specifically, the outer layer of the magnetite nanozymes is physically etched, the etched magnetite nanozymes are dissolved in concentrated hydrochloric acid, then evaporated to dryness at 90℃~110℃, concentrated nitric acid is added, and evaporation continues at 90℃~110℃, followed by dissolution with dilute nitric acid to obtain the etched sample. The physical etching etches the outer layer of the magnetite nanozymes by 1 nm~2 nm, preferably 1 nm; the physical etching is specifically ion beam physical etching; the bias voltage of the physical etching is 180 V~220 V, preferably 200 V; the etching time is 6 s~8 s, preferably 7 s; the tilt angle of the physical etching is 8°~12°, preferably 10°; and the rotation speed of the physical etching is 8 rpm~12 rpm, preferably 10 rpm. In some embodiments of the present invention, the ion beam etching machine used for the physical etching is a Leuven Heverlee Lorem R. The concentration of the concentrated hydrochloric acid in the present invention is 10 mol / L to 14 mol / L, preferably 12 mol / L; the concentration of the concentrated nitric acid is 14 mol / L to 16 mol / L; and the concentration of the dilute nitric acid is 0.4 mol / L to 0.6 mol / L, preferably 0.5 mol / L.

[0012] After obtaining the pre-etched and post-etched samples, the present invention acquires the δ values ​​of the pre-etched and post-etched samples respectively. 56 / 54 The Fe value is obtained by measuring the δ before and after the magnetite nanoenzyme etching in step S1). 56 / 54 The absolute difference of Fe, according to the δ 56 / 54 The absolute difference in Fe content was used to determine the activity of the magnetite nanozyme. Specifically, the samples before and after etching were diluted, and the Fe isotope composition of the samples before and after etching was measured using an MC-ICP-MS instrument. 56 Fe / 54 Fe), and then the δ of the samples before and after etching is obtained according to the Fe isotopic composition of the samples before and after etching. 56 / 54 Fe value, determined by δ before and after etching with the magnetite nanoenzyme. 56 / 54 The absolute difference analysis of Fe isotope fractionation results of the inner and outer layers of magnetite; in the Fe isotope measurement, the Fe element in the sample is diluted to 1 ppm to 3 ppm, preferably 2 ppm; the Fe isotope measurement standard is the IRMM-019 iron isotope standard. The δ¹⁸O⁻¹ of this invention... 56 / 54 The Fe value is obtained according to Formula 1: Formula 1.

[0013] The method for detecting the activity of magnetite nanozymes provided by this invention utilizes the phenomenon of Fe isotope fractionation. Fe isotope fractionation refers to the uneven distribution of various stable isotopes of iron among different substances or phases due to slight differences in mass during physical, chemical, or biological processes. It can indicate the composition of Fe isotopes. The method for detecting the activity of magnetite nanozymes provided by this invention is based on the Fe isotope fractionation phenomenon of magnetite nanozymes, i.e., Fe... 2+ Fe tends to accumulate light isotopes. 3+ It readily enriches heavy isotopes, δ 56 / 54 The increase in Fe indicates a fractionation pattern where heavier Fe isotopes appear in the measured sample. As the magnetite nanozyme is gradually oxidized, the internal Fe... 2+ As it migrates outwards and partially dissolves, its overall Fe isotope becomes increasingly heavier, δ... 56 / 54 The Fe content will increase until it reaches a threshold. Therefore, the Fe content inside... 2+ The migration ability can be measured by the δ of magnetite. 56 / 54 Fe is used to represent this.

[0014] This invention measures the Fe isotopic composition of magnetite before and after etching, thereby realizing the Fe content of the inner and outer layers of magnetite nanozymes. 2+ and Fe 3+Analysis of the proportion change. If the Fe isotope fractionation result is that the Fe isotope of the etched magnetite nanoenzyme is heavier than that before etching, that is, the measured δ of the etched sample... 56 / 54 An increase in the Fe value indicates that Fe isotope fractionation occurs on the surface and inside the magnetite nanozyme after oxidation, with the internal Fe... 3+ The proportion is higher than that on the surface. This indicates that during the oxidation process of magnetite nanozymes, the internal Fe... 2+ Outward migration causes the magnetite surface to exhibit Fe... 2+ The enriched state makes the surface Fe 2+ The active sites regenerate, thus maintaining the peroxidase-like activity of magnetite nanozymes. Based on this, Fe isotope fractionation results can be used to assess the Fe content within magnetite. 2+ The migration ability is used to assess the operating status of the catalyst.

[0015] This invention also provides a method for regenerating the activity of magnetite nanozymes. Specifically, due to Fe... 2+ The migration ability of nanozymes is the reason why they maintain their activity. Therefore, after establishing the judgment criteria, Fe can be added to the catalyst to be tested in a timely manner based on the judged operating state of the catalyst. 2+ This process replenishes the vacancies of magnetite nanoenzymes, regenerating their active sites and thus extending catalyst lifespan. At the same time, it is less expensive than directly replacing the catalyst, reducing operating costs in engineering.

[0016] This invention provides a method for regenerating the activity of magnetite nanozymes, comprising the following steps: S1) The magnetite nanozymes in different oxidation states were dissolved in concentrated hydrochloric acid and evaporated to dryness. Then, concentrated nitric acid was added and evaporated to dryness again. Then, dilute nitric acid was added to dissolve them to obtain samples in different oxidation states before etching. The outer layers of magnetite nanozymes in different oxidation states were physically etched, dissolved in concentrated hydrochloric acid and evaporated to dryness, then concentrated nitric acid was added and evaporated to dryness again, and then dilute nitric acid was added to obtain etched samples in different oxidation states. S2) Obtain the δ values ​​of the samples before etching and the samples after etching in different oxidation states as described in step S1). 56 / 54 Fe value, to obtain the δ before and after the magnetite nanoenzyme etching in different oxidation states described in step S1). 56 / 54 absolute difference in Fe; S3) Obtain the δ-value of the magnetite nanozyme in the system according to any of the above-described methods for detecting the activity of magnetite nanozymes. 56 / 54 If the absolute difference in Fe is greater than the δ before and after etching of magnetite nanoenzymes in different oxidation states as described in step S2), 56 / 54 The absolute difference in Fe values ​​is δ, which ranges from 0.035 to 0.045. 56 / 54If the absolute difference in Fe is found, then Fe is added to the system. 2+ source.

[0017] In the method for regenerating the activity of magnetite nanozymes provided by this invention, the δ-values ​​of magnetite nanozymes in different oxidation states before and after etching are analyzed. 56 / 54 The absolute difference in Fe was used to establish a standard for judging the activity of magnetite nanozymes, namely, a δ value of 0.035~0.045. 56 / 54 The absolute difference in Fe, and then the δ of the magnetite nanozyme in the system obtained according to the aforementioned method for detecting magnetite nanozyme activity. 56 / 54 After determining the absolute difference in Fe, the appropriate judgment criteria can be used to determine whether Fe needs to be added to the system. 2+ Source, through the addition of Fe 2+ Regeneration of active sites of magnetite nanozymes in the source regulation system.

[0018] In the method for regenerating the activity of magnetite nanozymes provided by this invention, step S1) involves obtaining samples before etching in different oxidation states and samples after etching in different oxidation states, and step S2) involves obtaining the δ values ​​of the samples before etching in different oxidation states and samples after etching in different oxidation states obtained in step S1). 56 / 54 The Fe values ​​are the same as those described in the aforementioned method for detecting the activity of magnetite nanozymes, and will not be repeated here.

[0019] In the magnetite nanozyme regeneration method provided by this invention, the magnetite nanozymes in different oxidation states mentioned in step S1) refer to magnetite nanozymes in different Fe... 2+ Fe 3+ Magnetite nanozymes with a stoichiometric ratio of Fe 2+ Fe 3+ The magnetite nanozymes have stoichiometric ratios of (0.48~0.52), (0.35~0.40), (0.25~0.30), and (0.15~0.20), respectively. In some embodiments of the present invention, the magnetite nanozymes of different oxidation states are specifically Fe... 2+ Fe 3+ Magnetite nanozymes with stoichiometric ratios of 0.5, 0.39, 0.28 and 0.17, respectively.

[0020] The preparation method of the magnetite nanozyme of the present invention includes the following steps: under anaerobic conditions, Fe... 2+ Source and Fe 3+ The source is added to water, ammonia is added to the solution to initiate a reaction, and then different amounts of H2O2 are added, followed by shaking to obtain magnetite nanozymes in different oxidation states. Preferably, the Fe... 2+ The source is FeCl2·4H2O; the Fe 3+The source is FeCl3·6H2O. Preferably, the Fe... 2+ The concentration of the source is 0.15 mol / L to 0.25 mol / L, preferably 0.2 mol / L; the Fe 3+ The concentration of the source is 0.35 mol / L to 0.45 mol / L, preferably 0.4 mol / L. Preferably, the concentration of the ammonia solution is 5 mol / L to 7 mol / L, preferably 6 mol / L; according to the dissolution of the Fe... 2+ Source and Fe 3+ The amount of source water is calculated to be 200 mL, and the volume of the ammonia water is 240 mL. Preferably, the reaction time is 20 h to 30 h, more preferably 24 h. Preferably, the shaking reaction time is 20 h to 30 h, more preferably 24 h. Preferably, the Fe... 2+ Source and Fe 3+ The stoichiometric ratio of the sources is 1:2.

[0021] In some embodiments of the present invention, the preparation method of magnetite nanozymes of different oxidation states of the present invention includes the following steps: under anaerobic conditions, Fe... 2+ Source and Fe 3+ The source was added to 200 mL of deionized water at a stoichiometric ratio of 1:2. Excess ammonia was slowly added dropwise to the solution, and the reaction was shaken for 24 h. Solid-liquid separation and washing were performed four times using a strong magnet. Subsequently, different amounts of H2O2 solution were added, and the reaction was shaken for 24 h to obtain magnetite nanozymes in different oxidation states.

[0022] To investigate the peroxidase-like activity of magnetite nanozymes in different oxidation states, this invention uses TMB as a chromogenic substrate. H₂O₂ is added to the obtained magnetite nanozymes, and the absorbance at 652 nm is measured using a UV spectrophotometer to reflect the peroxidase-like activity of the magnetite nanozymes. The concentration of TMB added is 0.8 mmol / L to 1.2 mmol / L, preferably 1 mmol / L; the concentration of the magnetite suspension is 0.01 g / L to 0.015 g / L, preferably 0.0125 g / L; and the concentration of H₂O₂ is 400 mmol / L to 600 mmol / L, preferably 500 mmol / L.

[0023] This invention obtains the δ values ​​of magnetite nanoenzymes in different oxidation states before and after etching in step S1). 56 / 54 After determining the absolute difference in Fe, the internal Fe content of magnetite nanozymes in different oxidation states was analyzed. 2+ Based on the migration ability, a criterion for judging the activity of magnetite nanozymes was established, namely, a δ value of 0.035~0.045. 56 / 54The absolute difference in Fe. Specifically, the δ value of the magnetite nanozyme in the system is obtained according to any of the methods described above for detecting the activity of magnetite nanozymes. 56 / 54 The absolute difference in Fe, if the δ of the magnetite nanozyme in the system 56 / 54 The absolute difference in Fe is greater than the δ value in the range of 0.035 to 0.045. 56 / 54 The absolute difference in Fe indicates that the activity of the magnetite nanozyme has decreased to a certain extent, and the addition of Fe to the system... 2+ The source can restore the activity of the catalyst, thereby increasing its lifespan and reducing engineering operating costs. Among these, the added Fe... 2+ The source is FeSO4·7H2O, with added Fe 2 + The concentration of the source was measured using the o-phenanthroline spectrophotometric method, and the total Fe content in the system was measured using inductively coupled plasma.

[0024] This invention provides a method for detecting the activity of magnetite nanozymes and a method for their regeneration. This invention utilizes physical etching to remove the active layer on the surface of the magnetite nanozyme, measures the change in Fe isotopic composition of the samples before and after etching, and then uses Fe... 2+ It readily accumulates light isotopes and Fe. 3+ The fractionation pattern of easily enriched heavy isotopes was analyzed to determine the distribution and migration behavior of iron species in the inner and outer layers of magnetite, thereby determining the internal Fe content of magnetite. 2+ Surface migration leads to continuous regeneration of active sites. Simultaneously, Fe isotope fractionation results are used to determine the active state of magnetite nanozymes, and a criterion for judging catalyst activity is established based on magnetite nanozymes in different oxidation states to achieve timely Fe addition. 2+ The regenerated catalyst active sites not only reflect the activity stability of nanozymes in water treatment and environmental remediation, but also avoid the blind addition of Fe. 2+ This results in pollution such as iron sludge. Compared with traditional valence state or surface characterization methods, this invention utilizes iron isotope fractionation. Through the Fe isotope fractionation generated by the redox transformation of iron in the inner and outer layers of magnetite nanozymes, the migration path and transformation mechanism of iron are precisely tracked. This accurately reveals the self-regeneration behavior of active sites in the hydrogen peroxide activation and water treatment reaction processes, thereby achieving the analysis of Fe in the inner and outer layers of magnetite nanozymes. 2+ / Fe 3+ The quantitative identification of evolution lays the foundation for the stability evaluation, structural optimization, and application of magnetite nanozymes in environmental remediation and water treatment. Attached Figure Description

[0025] Figure 1 This is a flowchart illustrating the preparation process of the magnetite nanozyme described in this invention. Figure 2 XRD patterns of magnetite nanozymes in different oxidation states prepared in Example 1 of this invention; Figure 3 X-ray photoelectron spectra of magnetite nanozymes in different oxidation states of the present invention; Figure 4 The graph shows the reaction curves of magnetite nanozymes in different oxidation states and maghemite-like peroxidases prepared in Example 1 of the present invention. Figure 5 The images show the activity maps of magnetite nanozymes and hematite-like peroxidases prepared in different oxidation states in Example 1. Figure 6 This is a schematic diagram showing the depth of the magnetite nanoenzyme etching of the present invention; Figure 7 These are actual images of the magnetite nanoenzyme before and after etching according to the present invention; Figure 8 These are atomic force microscopy images of the magnetite nanozyme of this invention before and after etching. Figure 9 The image shows the Fe isotope fractionation results measured by magnetite nanozymes before and after etching in this invention. Figure 10 Fe in the solution during the active site regeneration experiment in Example 5 of this invention 2+ Concentration graph; Figure 11 This is a graph showing the stoichiometry of the active sites of the oxidized magnetite nanozyme before and after regeneration in Example 5 of this invention. Detailed Implementation

[0026] This invention discloses a method for detecting the activity of magnetite nanozymes and a method for regenerating them. Those skilled in the art can refer to the content of this document and appropriately modify the process parameters to achieve the desired results. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included in this invention. The methods and applications of this invention have been described through preferred embodiments, and those skilled in the art can obviously make modifications or appropriate alterations and combinations to the methods and applications described herein without departing from the content, spirit, and scope of this invention to realize and apply the technology of this invention.

[0027] The present invention will be further described below with reference to the embodiments: Example 1 Preparation of magnetite nanozymes in different oxidation states: Under anaerobic conditions, Fe... 2+ Fe 3+ A magnetite nanozyme was added to 200 mL of deionized water at a molar ratio of 1:2. Excess ammonia was slowly added dropwise, and the reaction was allowed to proceed with shaking for 24 h. The mixture was then washed four times using a strong magnet for solid-liquid separation, and a certain amount of deionized water was added to obtain a magnetite suspension. Subsequently, different amounts of H₂O₂ solution were added, and the reaction was allowed to proceed with shaking for 24 h to obtain magnetite nanozymes in different oxidation states. The stoichiometric ratio x (Fe) was [not specified in the original text].2+ :Fe 3+ =0.50, 0.39, 0.28, 0.17.

[0028] A certain amount of magnetite nanozyme was placed in a centrifuge tube, centrifuged at 10,000 rpm, the supernatant was discarded, and the mixture was freeze-dried for 24 h, followed by calcination at 200 °C for 2 h to obtain hematite (γ-Fe₂O₃). Figure 1 As shown, Figure 1 This is a flowchart illustrating the preparation process of the magnetite nanozyme described in this invention.

[0029] Magnetite nanozymes and hematite in different oxidation states were characterized by XRD, such as... Figure 2 As shown, Figure 2 The images show the XRD patterns of magnetite nanozymes and hematite in different oxidation states prepared in Example 1 of this invention. The results indicate that magnetite nanozymes in different oxidation states were successfully synthesized using the above method.

[0030] Example 2

[0031] Measurement of peroxidase-like activity of magnetite nanozymes: 1.8 mL of pH 5.5 200 mM sodium acetate buffer was added to a centrifuge tube, followed by 50 μL of 10 g / L TMB, then 10 μL of 2.5 g / L magnetite nanozyme suspensions in different oxidation states, and finally 100 μL of 10 mol / L H₂O₂ solution. After thorough mixing, the absorbance at 652 nm was measured using a UV spectrophotometer to reflect the peroxidase-like activity of magnetite. Subsequently, the magnetite nanozymes in different oxidation states were characterized by X-ray photoelectron spectroscopy to analyze their surface valence states. Results are as follows: Figure 3 As shown, Figure 3 The X-ray photoelectron spectra of magnetite nanozymes in different oxidation states of the present invention are shown.

[0032] According to GB / T 37966-2019, the absorbance of magnetite nanozymes and hematite in different oxidation states in Example 1 was obtained using a UV spectrophotometer, thereby obtaining peroxidase-like reaction curves, such as... Figure 4 , Figure 4 The graphs show the reaction curves of magnetite nanozymes in different oxidation states and maghemite-like peroxidases prepared in Example 1 of this invention. Figure 4 Further obtain Figure 5 , Figure 5The images show the peroxidase activity of magnetite nanozymes and maghemite nanozymes prepared in different oxidation states in Example 1. The results show that the peroxidase activity of magnetite nanozymes decreases with increasing oxidation state, while the activity of maghemite nanozymes approaches zero. Furthermore, compared to maghemite, oxidized magnetite nanozymes maintain high activity in the initial stage after H₂O₂ oxidation, indicating the sustainability of the peroxidase activity of magnetite nanozymes and suggesting that its activity is not solely derived from the Fe₂O₃ surface of magnetite. 2+ It also comes from the Fe inside magnetite. 2+ Meanwhile, X-ray photoelectron spectroscopy showed that during the oxidation process of magnetite nanozymes with H2O2, Fe was continuously enriched in the outer layer. 2+ This may be the direct reason why magnetite nanozymes retain peroxidase-like activity.

[0033] Example 3

[0034] Physical etching of oxidized magnetite nanozymes, with an expected etching depth of approximately 1 nm on the outer layer of the magnetite nanozymes, such as... Figure 6 As shown, Figure 6 The schematic diagram illustrating the etching depth of the magnetite nanozyme of this invention is as follows: 0.1 mL of a magnetite nanozyme suspension with a stoichiometric ratio of 0.28 and 0.50 was placed in a centrifuge tube, and 3.9 mL of anhydrous ethanol was added and mixed thoroughly. 200 μL was dropped onto a silicon wafer and allowed to air dry. Magnetite was etched using ion beam etching technology under the following conditions: a bias voltage of 200V, a tilt angle of 10°, a rotation speed of 10 rpm, and an etching time of 7 s. Figure 7 As shown, Figure 7 These are actual images of the magnetite nanoenzyme before and after etching according to the present invention. The left image shows the magnetite nanoenzyme before etching, and the right image shows the magnetite nanoenzyme after etching.

[0035] Magnetite nanozymes were characterized using atomic force microscopy before and after etching. The nanozymes were then dissolved in hydrochloric acid, and the Fe element concentration was measured using ICP. The etching surface layer was then calculated to be approximately 1 nm thick. Figure 8 As shown, Figure 8 These are atomic force microscopy images of the magnetite nanozyme of this invention before and after etching. The left image shows the magnetite nanozyme before etching, and the right image shows the magnetite nanozyme after etching. Figure 8 It is evident that the increased roughness after etching indicates that the outer layer of the magnetite nanoenzyme was etched.

[0036] Example 4

[0037] Fe isotope fractionation: Magnetite nanozymes in different oxidation states before and after etching were dissolved in 1 mL of concentrated hydrochloric acid to obtain samples; the samples obtained by dissolving in 1 mL of concentrated hydrochloric acid were evaporated to dryness at 100℃, 1 mL of concentrated nitric acid was added, and the samples were evaporated to dryness at 100℃ again, followed by the addition of 1 mL of dilute nitric acid to dissolve, obtaining the processed samples; the samples were diluted by a certain factor, and the Fe isotope composition in the solution was measured using MC-ICP-MS. δ 56 / 54 Fe is an important indicator of Fe isotope fractionation. An increase in the value after etching indicates that the Fe isotope in the solution after the reaction is heavier, and Fe... 3+ It is more likely to accumulate heavy isotopes, which indicates that compared to Fe... 2+ Fe in the solution 3+ The proportion has increased.

[0038] like Figure 9 As shown, Figure 9 This image shows the Fe isotope fractionation results measured by the magnetite nanozyme before and after etching, as per the present invention. Experimental results indicate that the δ¹⁸O₂ of the oxidized magnetite (x=0.28) after etching... 56 / 54 The Fe concentration was 0.47, significantly higher than the pre-etching concentration of 0.43, with an absolute difference of 0.04. This indicates that Fe isotopes underwent fractionation, with the inner Fe isotopes being heavier. 3+ A higher proportion indicates that the outer Fe of the magnetite nanoenzyme 2+ The magnetite nanozyme is enriched and exhibits an oxidized state internally. Meanwhile, compared to the unoxidized magnetite nanozyme (x=0.5), the difference between the inner and outer layers of the unoxidized magnetite nanozyme is not significant, with an absolute difference of only 0.01, indicating that the Fe content during the oxidation process of the magnetite nanozyme is low. 2+ The outward migration causes Fe to... 2+ The magnetite nanozyme is enriched on the surface, which enables the active sites of the magnetite nanozyme to regenerate and continuously react with H2O2, thus maintaining the peroxidase-like activity of the magnetite nanozyme.

[0039] Example 5

[0040] Regeneration experiment of active sites of magnetite nanozymes: 10 mM Fe was added to the magnetite nanozyme suspensions of different oxidation states obtained in Example 1. 2+ The reaction was carried out on a shaker. 1 mL of the suspension was collected at 0 h, 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h, 48 h, and 72 h, respectively, and filtered. The Fe content in the filtrate was measured using the o-phenanthroline spectrophotometric method. 2+ Content. After 7 days, the prepared solid was centrifuged and washed to remove adsorbed Fe. 2+ The magnetite was dissolved using concentrated hydrochloric acid, and the Fe and Fe content in the regenerated magnetite were determined using inductively coupled plasma (ICP) and o-phenanthroline spectrophotometry. 2+ The content of Fe was calculated. 2+ Fe3 + The stoichiometric ratio.

[0041] like Figure 10 , 11 As shown, Figure 10 Fe in the solution during the active site regeneration experiment in Example 5 of this invention 2+ Concentration graph; Figure 11 This is a graph showing the stoichiometry of the active sites of magnetite nanozymes in different oxidation states before and after regeneration in this invention. Experimental results show that with the addition of Fe... 2+ In this case, oxidized magnetite can restore its stoichiometry, thereby restoring its surface active sites. This not only improves catalyst life but also avoids catalyst replacement and reduces engineering operating costs.

[0042] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A method for detecting the activity of magnetite nanozymes, characterized in that, Includes the following steps: S1) The magnetite nanozyme was dissolved in concentrated hydrochloric acid and evaporated to dryness, then concentrated nitric acid was added and evaporated to dryness again, and then dilute nitric acid was added to dissolve it to obtain the sample before etching. and, The outer layer of the magnetite nanozyme was physically etched, dissolved in concentrated hydrochloric acid and evaporated to dryness, then concentrated nitric acid was added and evaporated to dryness again, and then dilute nitric acid was added to obtain the etched sample. S2) Obtain the δ values ​​of the samples before and after etching as described in step S1). 56 / 54 The Fe value is obtained by measuring the δ before and after the magnetite nanoenzyme etching in step S1). 56 / 54 The absolute difference of Fe, according to the δ 56 / 54 The absolute difference in Fe values ​​is used to determine the activity of the magnetite nanozyme.

2. The detection method according to claim 1, characterized in that, In step S1), the physical etching etches 1 nm to 2 nm of the outer layer of the magnetite nanoenzyme.

3. The detection method according to claim 1, characterized in that, In step S1), the bias voltage for physical etching is 180 V to 220 V, and the etching time is 6 s to 8 s.

4. The detection method according to claim 1, characterized in that, In step S1), during the process of obtaining the sample before etching, the concentration of the concentrated hydrochloric acid is 10 mol / L to 14 mol / L, the concentration of the concentrated nitric acid is 14 mol / L to 16 mol / L, and the concentration of the dilute nitric acid is 0.4 mol / L to 0.6 mol / L. During the process of obtaining the etched sample, the concentration of the concentrated hydrochloric acid is 10 mol / L to 14 mol / L, the concentration of the concentrated nitric acid is 14 mol / L to 16 mol / L, and the concentration of the dilute nitric acid is 0.4 mol / L to 0.6 mol / L.

5. A method for regenerating the activity of magnetite nanozymes, characterized in that, Includes the following steps: S1) The magnetite nanozymes in different oxidation states were dissolved in concentrated hydrochloric acid and evaporated to dryness. Then, concentrated nitric acid was added and evaporated to dryness again. Then, dilute nitric acid was added to dissolve them to obtain samples in different oxidation states before etching. and, The outer layers of magnetite nanozymes in different oxidation states were physically etched, dissolved in concentrated hydrochloric acid and evaporated to dryness, then concentrated nitric acid was added and evaporated to dryness again, and then dilute nitric acid was added to obtain etched samples in different oxidation states. S2) Obtain the δ values ​​of the samples before etching and the samples after etching in different oxidation states as described in step S1). 56 / 54 Fe value, to obtain the δ before and after the magnetite nanoenzyme etching in different oxidation states described in step S1). 56 / 54 absolute difference in Fe; S3) Obtain the δ-value of the magnetite nanozyme in the system according to the detection method described in any one of claims 1 to 4. 56 / 54 If the absolute difference in Fe is greater than the δ before and after etching of magnetite nanoenzymes in different oxidation states as described in step S2), 56 / 54 The absolute difference in Fe values ​​is δ, which ranges from 0.035 to 0.

045. 56 / 54 If the absolute difference in Fe is found, then Fe is added to the system. 2+ source.

6. The regeneration method according to claim 5, characterized in that, In step S1), the magnetite nanozymes in different oxidation states are specifically Fe... 2+ Fe 3+ Magnetite nanozymes with stoichiometric ratios of (0.48~0.52), (0.35~0.40), (0.25~0.30) and (0.15~0.20), respectively.

7. The regeneration method according to claim 5, characterized in that, In step S1), the magnetite nanozymes in different oxidation states are specifically Fe... 2+ Fe 3+ Magnetite nanozymes with stoichiometric ratios of 0.5, 0.39, 0.28 and 0.17, respectively.

8. The regeneration method according to claim 5, characterized in that, In step S1), the physical etching etches 1 nm to 2 nm of the outer layer of the magnetite nanoenzyme.

9. The regeneration method according to claim 5, characterized in that, In step S1), the bias voltage for physical etching is 180 V to 220 V, and the etching time is 6 s to 8 s.

10. The regeneration method according to claim 5, characterized in that, In step S1), during the process of obtaining the sample before etching, the concentration of the concentrated hydrochloric acid is 10 mol / L to 14 mol / L, the concentration of the concentrated nitric acid is 14 mol / L to 16 mol / L, and the concentration of the dilute nitric acid is 0.4 mol / L to 0.6 mol / L. During the process of obtaining the etched sample, the concentration of the concentrated hydrochloric acid is 10 mol / L to 14 mol / L, the concentration of the concentrated nitric acid is 14 mol / L to 16 mol / L, and the concentration of the dilute nitric acid is 0.4 mol / L to 0.6 mol / L.

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