Application of self-cascaded cerium-based MOF nanoszyme in detection of profenofos

Abstract

The application discloses application of a self-cascaded cerium-based MOF nanometer enzyme in detection of profenofos, and the cerium-based MOF nanometer enzyme is a cerium-based metal organic framework material HMUiO-66(Ce) nanometer enzyme, which can successively exert the organic phosphorus hydrolysis enzyme and the laccase-like activity of the HMUiO-66(Ce) nanometer enzyme by adjusting pH. First, the HMUiO-66(Ce) nanometer enzyme hydrolyzes profenofos to produce 4-bromo-2-chlorophenol under alkaline conditions; then the solution is adjusted to be acidic, and the HMUiO-66(Ce) nanometer enzyme exerts the laccase-like activity under the acidic conditions, 4-bromo-2-chlorophenol and 4-aminoantipyrine are complexed into quinone imine, the ultraviolet absorbance of the quinone imine at 508 nm is detected, and the concentration of profenofos is calculated. It is found that the nanometer enzyme has good selectivity for profenofos, the operation in detection is extremely simple, the cost is low, and it is possible to rapidly detect profenofos on site.

Description

Technical Field

[0001] This invention belongs to the field of organophosphorus pesticide detection technology, specifically involving the application of self-cascaded cerium-based MOF nanozymes in the detection of profenofos. Background Technology

[0002] Organophosphorus pesticides (OPs) are a class of chemical substances that irreversibly inhibit the activity of acetylcholinesterase. In agricultural practice, organophosphorus pesticides are widely used to control insect pests to improve crop yield and quality. They play a positive role in controlling agricultural diseases and improving the quality and yield of agricultural products. However, the long-term overuse of organophosphorus pesticides has led to serious pesticide residue problems, which not only pollute the environment but also pose potential risks to human health. Profenofos (PFF) is a widely used organophosphorus pesticide, primarily used to control pests and diseases in various crops. Although it has significant effects in agriculture, it is moderately toxic and can significantly inhibit acetylcholinesterase in the human body, causing harm to the nervous system. With the banning of other highly toxic pesticides, the use of profenofos has been increasing, and its residue problem has become increasingly prominent.

[0003] Traditional methods for detecting organophosphorus pesticides, such as high-performance liquid chromatography (HPLC) and gas chromatography (GC), suffer from drawbacks including being time-consuming, costly, and reliant on specialized technical expertise. Immunoassays, on the other hand, are limited by the instability of enzymes or antibodies. Colorimetric analysis, as a simple, rapid, economical, and visual detection method, has become a viable alternative for detecting profenofos residues and has garnered increasing attention in recent years. Yang et al. disclosed a method for preparing hierarchical macroporous-microporous metal-organic frameworks (HMMOFs) with thin macroporous walls by achieving open channels with macropore sizes up to 100 nm and macropore walls of 11-12 nm in a cerium-based MOF (HMUiO-66(Ce)). Specifically, a microemulsion-guided assembly strategy was adopted, using Pluronic P123 and Pluronic F127 as co-stabilizers and toluene as the oil phase to form a columnar microemulsion to template HMUiO-66(Ce). The HMMOFs can act as nuclease mimics, hydrolyzing and cleaving stable phosphodiester bonds in various DNAs (Yang, J.; Li, K.; Gu, J. Hierarchically Macro-Microporous Ce-Based MOFs for the Cleavage of DNA. ACS Mater. Lett. 2022, 4, 385-391.). However, no further information on this cerium-based MOF has been found to date. Reports on the application of HMUiO-66(Ce) nanozyme in the detection of profenofos. Summary of the Invention

[0004] The purpose of this invention is to overcome the above-mentioned defects and deficiencies in the prior art and to provide the application of cerium-based metal-organic framework material HMUiO-66(Ce) nanozyme in the detection of profenofos.

[0005] The second objective of this invention is to provide the application of cerium-based metal-organic framework material HMUiO-66(Ce) nanozyme in the preparation of products for detecting propargite.

[0006] The third objective of this invention is to provide a method for detecting propargite using cerium-based metal-organic framework material HMUiO-66(Ce) nanozymes.

[0007] The above-mentioned objective of this invention is achieved through the following technical solution:

[0008] This invention synthesizes cerium-based metal-organic framework material HMUiO-66(Ce) nanozymes according to the literature (Yang, J.; Li, K.; Gu, J. Hierarchically Macro-Microporous Ce-Based MOFs for the Cleavage of DNA. ACS Mater. Lett. 2022, 4, 385-391.). This invention found that under pH=9 conditions, HMUiO-66(Ce) nanozymes can exhibit laccase-like activity, hydrolyzing propargyl phosphonate (PFF) to produce colorless 4-bromo-2-chlorophenol (BCP). After complete hydrolysis, the pH of the solution is adjusted to pH=6 by adding phosphate buffer, allowing HMUiO-66(Ce) nanozymes to exhibit laccase-like activity, catalyzing the complexation of 4-bromo-2-chlorophenol with the chromogenic agent 4-aminoantipyrine (4-AP) to form a red quinone imine (QI). This red product exhibits strong ultraviolet absorption at a wavelength of 508 nm. As the concentration of profenofos increases, the color of the quinone imine solution gradually changes from colorless to pink. Therefore, when the concentration of profenofos is high, it can be quantitatively analyzed using a UV-Vis spectrophotometer or by using the RGB mode of a smartphone based on the solution color.

[0009] Therefore, this invention provides the application of cerium-based metal-organic framework material HMUiO-66(Ce) nanozyme in the detection of propargite.

[0010] This invention also provides the application of cerium-based metal-organic framework material HMUiO-66(Ce) nanozyme in the preparation of products for detecting propargite.

[0011] Furthermore, the products include, but are not limited to, reagent kits or colorimetric sensors.

[0012] Furthermore, the preparation of the HMUiO-66(Ce) nanozyme includes the following steps:

[0013] S1. Pluronic P123 and Pluronic F127 are added to water and stirred to form linear micelles;

[0014] S2. Add toluene, acetic acid, and sodium perchlorate monohydrate, mix well to form a milky white microemulsion;

[0015] S2. Add terephthalic acid and cerium ammonium nitrate, stir, centrifuge, wash and dry to obtain the product.

[0016] In the preparation method of HMUiO-66(Ce) nanozyme of the present invention, Pluronic P123 and Pluronic F127 are used as co-stabilizers, and toluene is used as an oil phase template to form columnar microemulsions to synthesize HMUiO-66(Ce). First, Pluronic P123 binds with a small portion of Pluronic F127 to form linear micelles in the aqueous phase; upon the addition of toluene, the transparent micelle solution rapidly transforms into a milky white microemulsion. This microemulsion can form crown ether complexes with multivalent metals, thus each microemulsion is surrounded by Ce metal clusters, forming a metal-organic framework (MOF); secondly, the triblock polymer surfactant (such as Pluronic P123 and Pluronic F127) can be activated in C1O 4- With the help of [unclear], the metal-organic framework was guided to grow into an ordered mesoporous structure in aqueous solution, forming open, continuous, thin-walled large channels. Among them, the ratio of Pluronic P123 and Pluronic F127 can greatly affect the morphology and stability of the microemulsion, and thus affect the structure of MOF, forming a self-cascaded cerium-based MOF nanozyme HMUiO-66(Ce).

[0017] Furthermore, in step S1, the mass ratio of Pluronic P123 to Pluronic F127 is 1.5 to 2.5:1.

[0018] Furthermore, in step S1, the mass ratio of Pluronic P123 to Pluronic F127 is 2:1.

[0019] Furthermore, in step S2, the mass-to-volume ratio of toluene, acetic acid, and sodium perchlorate monohydrate is 2-3 mL: 1 mL: 2-4 g.

[0020] Furthermore, in step S2, the mass-to-volume ratio of toluene, acetic acid, and sodium perchlorate monohydrate is 2.4 mL: 1 mL: 3 g.

[0021] Furthermore, in step S3, the mass ratio of phthalic acid to cerium ammonium nitrate is 10-15:53-57.

[0022] Furthermore, in step S3, the mass ratio of phthalic acid to cerium ammonium nitrate is 30:137.

[0023] Further, the washing method in step S3 is to wash with water 1 to 3 times, wash with dimethylformamide 1 to 2 times, soak in acidic dimethylformamide solution for 10 to 14 hours, and soak in ethanol for 2 to 3 days.

[0024] Furthermore, the immersion temperature is 55–65°C.

[0025] Furthermore, the immersion temperature is 60°C.

[0026] Preferably, in step S3, after adding terephthalic acid and cerium ammonium nitrate, the mixture is stirred at 40°C and 400 rpm for 40 minutes. The resulting solid is separated by centrifugation, washed twice with water, and once with dimethylformamide. The centrifuged solid is then soaked in 4 mL of a dimethylformamide solution containing 0.1 wt% HCl for 12 hours at 60°C. Next, the sample is soaked in ethanol at 60°C for two days, with the ethanol changed daily. Finally, the product is dried overnight in a vacuum drying oven at 60°C.

[0027] This invention also provides a method for detecting propargite using cerium-based metal-organic framework material HMUiO-66(Ce) nanozymes, comprising the following steps:

[0028] S1. Known solutions of profenofos at different concentrations were incubated with HMUiO-66(Ce) nanozyme under alkaline conditions, then the pH was adjusted to acidic conditions, 4-aminoantipyrine was added, incubated, centrifuged, and the supernatant was separated.

[0029] S2. Detect the absorbance value at 508 nm, and construct a standard curve of profenofos concentration-UV absorbance with the concentration of profenofos as the x-axis and the UV absorbance as the y-axis; or analyze the RGB values ​​based on the color of the supernatant, and construct a standard curve of profenofos concentration-RGB values ​​with the concentration of profenofos as the x-axis and the RGB values ​​as the y-axis.

[0030] S3. After treating the sample with HMUiO-66(Ce) nanozyme according to the conditions in step S1, detect the absorbance value of the sample at 508nm, and calculate the concentration of profenofos based on the standard curve of profenofos concentration-UV absorbance; or detect the RGB value of the sample and calculate the concentration of profenofos based on the standard curve of profenofos concentration-RGB value.

[0031] Furthermore, the alkaline condition in step S1 is pH = 8.5–9.5;

[0032] Furthermore, the alkaline condition in step S1 is adjusted using a Tris-HCl buffer solution.

[0033] Furthermore, the acidic conditions in step S1 are pH = 5.5 to 6.5.

[0034] Furthermore, the acidic condition in step S1 is adjusted using phosphate buffer.

[0035] Furthermore, in step S1, the alkaline condition is pH = 9; in step S2, the acidic condition is pH = 6.

[0036] Furthermore, the method for detecting the absorbance value at 508 nm in step S2 is to perform quantitative analysis using a UV-Vis spectrophotometer.

[0037] Furthermore, the method for detecting RGB values ​​in step S2 is to perform quantitative analysis using the RGB mode of a smartphone.

[0038] Compared with the prior art, the present invention has the following beneficial effects:

[0039] This invention discloses the application of a self-cascaded cerium-based MOF nanozyme in the detection of profenofos. The cerium-based MOF nanozyme is the cerium-based metal-organic framework material HMUiO-66(Ce) nanozyme, which can sequentially exhibit organophosphorus hydrolase-like and laccase-like activities by adjusting the pH. First, under alkaline conditions, the HMUiO-66(Ce) nanozyme hydrolyzes profenofos to produce 4-bromo-2-chlorophenol. Then, the solution is adjusted to acidic conditions, and under acidic conditions, the HMUiO-66(Ce) nanozyme exhibits laccase-like activity, complexing 4-bromo-2-chlorophenol and 4-aminoantipyrine to synthesize a quinone imine. The absorbance at 508 nm is measured, and the concentration of profenofos is calculated. This invention reveals that the HMUiO-66(Ce) nanozyme has good selectivity for profenofos. This invention is extremely simple to operate, the detection process is rapid and inexpensive, and the requirements for detection instruments are low. Quantitative analysis can be performed using the RGB mode of a smartphone, making it possible to detect profenofos rapidly on-site, which is convenient for widespread promotion and application. Attached Figure Description

[0040] Figure 1 This is a scanning electron microscope image of the HMUiO-66(Ce) nanozyme in this invention.

[0041] Figure 2 This is a scanning electron microscope image of the HMUiO-66(Ce) nanozyme in this invention.

[0042] Figure 3 This is a schematic diagram of the EDS Mapping characterization of the HMUiO-66(Ce) nanozyme in this invention.

[0043] Figure 4 This is a schematic diagram of the HMUiO-66(Ce) nanozyme detection principle in this invention.

[0044] Figure 5 This is a standard curve of propargite-UV absorbance in this invention. The horizontal axis represents propargite concentration (ppm), and the vertical axis represents UV absorbance (au).

[0045] Figure 6 This is a graph showing the relationship between RGB values ​​and propargite concentration in this invention.

[0046] Figure 7 The image shows the detection results of different organophosphorus pesticides in this invention. Detailed Implementation

[0047] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but the embodiments do not limit the present invention in any way. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in this technical field.

[0048] Unless otherwise specified, all reagents and materials used in the following examples are commercially available.

[0049] The technical solution of the present invention will now be described in detail and completely with reference to the accompanying drawings, aiming to enable those skilled in the art to more deeply understand the advantages and characteristics of the present invention, thereby providing a clearer definition of the scope of protection of the present invention. It should be noted that the embodiments described in this invention are merely exemplary examples of the present invention, and not exhaustive of all possible implementation methods. All other embodiments that can be deduced by those skilled in the art based on the embodiments disclosed in this invention without creative effort are considered to fall within the scope of protection of this invention.

[0050] Example 1: Preparation of HMUiO-66(Ce) nanozymes

[0051] HMUiO-66(Ce) nanozymes were prepared according to the method disclosed in (Yang, J.; Li, K.; Gu, J. Hierarchically Macro-Microporous Ce-Based MOFs for the Cleavage of DNA. ACS Mater. Lett. 2022, 4, 385-391.). The specific preparation method is as follows:

[0052] First, 100mg of Pluronic P123 (PEO) 20 PPO 70 PEO 20 ), 50mg Pluronic F127 (PEO) 106PPO 70 PEO 106 Dissolving in 6 mL of deionized water, Pluronic P123 and Pluronic F127 bind to form linear micelles in the aqueous phase. Next, 240 μL of toluene is added to the solution. Then, 0.1 mL of acetic acid and 300 mg of sodium perchlorate monohydrate (NaClO4·H2O) are added, and the mixture is vigorously stirred for 15 minutes to form a white microemulsion. Subsequently, 120 mg of phthalic acid (H2BDC) and 548 mg of cerium ammonium nitrate ((NH4)2Ce(NO3)6) are added to the microemulsion, and the mixture is stirred at 400 rpm for 40 minutes at 40 °C. The PEO fragments on the Pluronic P123 and Pluronic F127 molecules can form crown ether complexes with polyvalent metal species, thus each microemulsion is surrounded by Ce metal clusters. Subsequently, metal-organic frameworks (MOFs) crystallize along the anchored Ce metal clusters and in ClO2. 4- In the presence of these ligands, the mixture assembled into an ordered mesoporous structure. After the reaction was complete, the resulting solid was separated by centrifugation, washed twice with water, and once with dimethylformamide (DMF). The centrifuged solid was then immersed in 4 mL of a dimethylformamide solution containing 0.1 wt% HCl at 60°C for 12 hours to remove residual ligands. The sample was then immersed in ethanol at 60°C for two days, with the ethanol changed daily. Finally, the product was dried overnight in a vacuum drying oven at 60°C.

[0053] Scanning electron microscope image of HMUiO-66(Ce) nanomaterials as shown below Figure 1 As shown, HMUiO-66(Ce) particles have highly open macropores and uniform thin walls.

[0054] Transmission electron microscopy image of HMUiO-66(Ce) nanomaterials as shown below Figure 2 As shown, the images further confirm that each HMUiO-66(Ce) particle has a uniform distribution of macroporous channels.

[0055] A schematic diagram of the EDS mapping characterization of HMUiO-66(Ce) nanomaterials is shown below. Figure 3 As shown, C, O and Ce elements are uniformly distributed on HMUiO-66(Ce) particles.

[0056] Example 2: Construction of the standard curve of profenofos concentration-UV absorbance

[0057] Profenofos stock solutions with concentrations of 0.002 mg / mL, 0.01 mg / mL, 0.04 mg / mL, 0.1 mg / mL, 0.2 mg / mL, 0.4 mg / mL, 0.6 mg / mL, 0.8 mg / mL, and 1 mg / mL were prepared using methanol and stored at 4°C. 200 μL of HMUiO-66(Ce) (1 mg / mL) was added to a 2 mL centrifuge tube, followed by 700 μL of deionized water and 50 μL of Tris-HCl buffer solution (1 M, pH = 9). Finally, 50 μL of different concentrations of profenofos were added, and the mixture was stirred at 40°C for 20 minutes. The profenofos concentrations were 0.1 μg / mL, 0.5 μg / mL, 1 μg / mL, 2 μg / mL, 5 μg / mL, 10 μg / mL, 20 μg / mL, 30 μg / mL, 40 μg / mL, and 50 μg / mL, respectively. Next, 50 μL of phosphate buffer (1M, pH=2) was added to adjust the pH of the solution to pH=6, followed by 20 μL of 4-aminoantipyrine solution (2 mg / mL). The mixture was stirred at 40 °C for 10 minutes. The sample was then centrifuged at 10,000 rpm for 3 minutes. 800 μL of the supernatant was collected, and the absorbance at 508 nm was measured using a UV spectrophotometer. A standard curve was constructed by plotting the profenofos concentration on the x-axis and the UV absorbance on the y-axis.

[0058] A schematic diagram illustrating the principle of HMUiO-66(Ce) nanozyme detection of profenofos is shown below. Figure 4 As shown, under alkaline conditions (pH=9), HMUiO-66(Ce) nanozyme hydrolyzes profenofos to produce 4-bromo-2-chlorophenol; then, under acidic conditions (pH=6), HMUiO-66(Ce) nanozyme exerts laccase-like activity to synthesize quinone imine from 4-bromo-2-chlorophenol and 4-aminoantipyrine. The UV absorbance at 508 nm was then measured, and the concentration of profenofos was calculated.

[0059] The standard curve of profenofos concentration-UV absorbance is as follows: Figure 5 As shown, the fitted linear equation is: y = 0.0154x + 0.0082. Where x is the concentration of profenofos in μg / mL, and y is the UV absorbance.

[0060] The relationship between RGB values ​​and profenofos concentration is as follows: Figure 6 As shown, the RGB mode of a smartphone can be used for quantitative analysis of profenofos concentration. RGB values ​​represent the brightness of red, green, and blue colors, ranging from 0 to 255. Different combinations of RGB values ​​can produce a variety of colors. By comparing the RGB values ​​of each color, the concentration of profenofos can be determined.

[0061] Example 3: Selectivity verification of HMUiO-66(Ce) hydrolysis / detection of profenofos.

[0062] The following sample solutions were prepared using methanol: 5 mg / L of profenofos, phoxim, trichlorfon, fenthion, fenitrothion, and chlorpyrifos. The HMUiO-66(Ce) nanozyme was incubated using the method described in Example 2 to verify the selectivity of the HMUiO-66(Ce) nanozyme constructed in Example 1 of this invention.

[0063] The test results are as follows Figure 7 As shown, the absorbance value of profenofos is much higher than that of other organophosphorus pesticides, indicating that the HMUiO-66(Ce) nanozyme of the present invention has a high selectivity for profenofos.

[0064] In summary, the HMUiO-66(Ce) nanozyme prepared by this invention can recognize and hydrolyze profenofos, and can be used for the concentration detection of profenofos. Moreover, the detection method is simple to operate, has low detection cost, fast detection speed, and low requirements for detection instruments.

Claims

1. Application of cerium-based metal-organic framework material HMUiO-66(Ce) nanoszyme in detection of profenofos, characterized in that, The application includes the following steps: S1. Profenofos solutions of known concentrations were incubated with HMUiO-66(Ce) nanozyme under alkaline conditions, then the pH was adjusted to acidic conditions, 4-aminoantipyrine was added, incubated, centrifuged, and the supernatant was separated; the alkaline conditions were pH=9; the acidic conditions were pH=6. S2. Detect the absorbance value of the supernatant at 508 nm, and construct a standard curve of profenofos concentration-UV absorbance with the concentration of profenofos as the x-axis and the UV absorbance as the y-axis; or analyze the RGB values ​​of the supernatant based on the color, and construct a standard curve of profenofos concentration-RGB values ​​with the concentration of profenofos as the x-axis and the RGB values ​​as the y-axis. S3. After treating the sample with HMUiO-66(Ce) nanozyme according to the conditions in step S1, detect the absorbance value of the sample at 508 nm, and calculate the concentration of profenofos based on the standard curve of profenofos concentration-UV absorbance; or detect the RGB value of the sample and calculate the concentration of profenofos based on the standard curve of profenofos concentration-RGB value.

2. Use according to claim 1, characterized in that, In step S1, the alkaline condition is adjusted using a Tris-HCl buffer solution.

3. Use according to claim 1, characterized in that, In step S1, the acidic condition is adjusted using phosphate buffer.

4. The use according to claim 1, characterized in that, The method for detecting the absorbance value at 508 nm in step S2 is to perform quantitative analysis using a UV-Vis spectrophotometer.

5. The use according to claim 1, characterized in that, The method for detecting RGB values ​​in step S2 is to perform quantitative analysis using the RGB mode of a smartphone.

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