A method for preparing and applying CeO2 nanoparticles loaded with squid ink melanin
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-05-11
- Publication Date
- 2026-08-14
AI Technical Summary
这两类方法具备检测灵敏度高、定性定量结果精准可靠的技术优势，但也存在明显的应用局限性：前者具有样品前处理步骤繁琐、仪器设备购置与维护成本高昂、检测过程整体花费较高、对操作人员的专业技能要求严苛等缺点；后者则有生物酶在环境中非常容易失活变性，对存储条件要求极为严格的缺点
[0026]1.本发明制备的墨鱼汁黑色素负载CeO2纳米颗粒克服了传统CeO2纳米颗粒易团聚，导致活性位点少、类酶活性较低的问题。墨鱼汁黑色素稳定的结构为CeO2纳米颗粒提供了一个合适的载体，形成CeO2纳米颗粒包裹黑色素颗粒的结构。
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Figure CN122567528A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of pesticide residue detection technology. This invention provides a squid ink melanin-loaded CeO2 nanoparticle, and also provides a preparation method and application of the squid ink melanin-loaded CeO2 nanoparticle. Background Technology
[0002] Traditional detection methods for organophosphorus pesticides are mainly divided into two categories: instrumental analysis methods and immunoassay methods. Both methods offer advantages such as high detection sensitivity and accurate and reliable qualitative and quantitative results, but they also have significant limitations: the former involves cumbersome sample pretreatment steps, high costs for instrument purchase and maintenance, high overall detection costs, and stringent requirements for operator skills; the latter suffers from the disadvantage of enzymes being easily inactivated and denatured in the environment, requiring extremely strict storage conditions.
[0003] Colorimetric methods, as an emerging rapid detection technology, are currently a hot research topic in organophosphorus pesticide detection. Early colorimetric sensing systems were mostly based on natural enzymes, but the ease of inactivation and high difficulty in extraction and purification of natural enzymes limited their practical application. In recent years, nanozymes have gradually replaced natural enzymes as materials for the colorimetric detection of organophosphorus pesticides due to their low cost and high stability. However, existing systems still suffer from technical bottlenecks such as insufficient sensitivity to the inhibition of nanozyme activity by organophosphorus pesticides, low nanozyme activity, and poor detection specificity. To balance the catalytic stability of nanozymes with the specific recognition ability of phosphate ester substrates, researchers have begun to focus on cerium-based nanomaterials that possess both enzyme-like activity and phosphate hydrolysis potential, attempting to build a more efficient colorimetric sensing platform for organophosphorus pesticides. However, due to limitations such as low specific surface area and weak adsorption capacity, pure CeO2 nanoparticles are generally difficult to hydrolyze organophosphorus pesticides.
[0004] Squid ink melanin nanoparticles contain numerous functional groups, such as amino, carboxyl, and indole groups. These groups exhibit a high affinity for metal ions, and their surface functional groups (-OH, -COOH) can stabilize metal active sites through coordination interactions. They can serve as biological templates and carriers for composite metal-based nanomaterials. Therefore, loading nanomaterials onto melanin nanoparticles holds promise for increasing binding sites with organophosphorus pesticides, thereby enhancing the sensitivity and specificity of organophosphorus pesticide colorimetric sensing and providing a new technical pathway for the detection of organophosphorus pesticide residues. Summary of the Invention
[0005] To overcome the shortcomings of the prior art, the first objective of this invention is to provide a squid ink melanin-loaded CeO2 nanoparticle, referred to as CeO2-CI, which has peroxide hydrolase-like activity, can specifically catalyze the hydrolysis of para-oxygenated phosphorus, has a larger specific surface area and stronger substrate adsorption capacity, and can effectively improve the detection sensitivity of para-oxygenated phosphorus.
[0006] The second objective of this invention is to provide a method for preparing the squid ink melanin-loaded CeO2 nanoparticles. This method has mild preparation conditions, simple process, is easy to scale up for production, and the resulting product has good dispersibility and stable catalytic activity.
[0007] The third objective of this invention is to provide the application of the squid ink melanin-loaded CeO2 nanoparticles in the colorimetric detection of oxygen and phosphorus. The colorimetric sensor constructed based on these nanoparticles has a wide detection range, low detection limit, strong selectivity, and strong anti-interference ability, enabling rapid detection of oxygen and phosphorus residues in real samples and meeting the routine detection needs for oxygen and phosphorus residues in agricultural products and environmental water samples. The squid ink melanin-loaded CeO2 nanoparticles are abbreviated as CeO2-CI.
[0008] Therefore, the first technical solution provided by this invention is as follows:
[0009] A method for preparing squid ink melanin-loaded CeO2 nanoparticles comprises the following steps:
[0010] 1) Dissolve cerium trichloride in deionized water, then add squid ink melanin, ammonia and hydrogen peroxide in sequence;
[0011] 2) With vigorous stirring, keep the solution from step 1) at 80-100℃ for 30-90 min;
[0012] 3) After the reaction solution is cooled to room temperature, the precipitate is collected by centrifugation, washed and dried to obtain CeO2-CI nanoparticles loaded with squid ink melanin.
[0013] The mass ratio of cerium trichloride to squid ink melanin is 2.5-10:1.
[0014] Furthermore, in the above-mentioned method for preparing CeO2 nanoparticles loaded with squid ink melanin, the mass ratio of cerium trichloride, squid ink melanin, ammonia, and hydrogen peroxide is 2.5-10 : 1 : 20-30 : 60-80.
[0015] Furthermore, in the above-mentioned method for preparing CeO2 nanoparticles loaded with squid ink melanin, the squid ink melanin is prepared by the following method:
[0016] 1) Cut open the ink sac of the cuttlefish, take the cuttlefish ink and mix it with the mixed solvent, place it in the refrigerator to soak, and obtain a suspension; centrifuge the suspension and take the precipitate, wash it, dry the washed solid, grind it and sieve it to obtain the melanin cuttlefish ink, which is recorded as crude CI.
[0017] 2) Take the sieved crude CI and dissolve it in sodium hydroxide solution. Heat it at 70-90℃ for 1-3 hours. After cooling, centrifuge the mixture and discard the supernatant. Then centrifuge the mixture a second time and take the supernatant. Dry the supernatant to obtain purified CI.
[0018] Furthermore, in the above-mentioned method for preparing CeO2 nanoparticles loaded with squid ink melanin, the ratio of squid ink to mixed solvent in step 1) is 1 g: 4 mL; and the soaking time is 6-18 h.
[0019] Furthermore, in the above-mentioned method for preparing squid ink melanin-loaded CeO2 nanoparticles, the ratio of crude CI to sodium hydroxide solution in step 2) is 1 g: 10 mL; the concentration of sodium hydroxide solution is 1 mM; the initial centrifugation involves centrifuging the cooled mixed solution at 5000 rpm for 20 min, and the secondary centrifugation involves centrifuging the mixture at 150 rpm for 8 min.
[0020] Furthermore, in the above-mentioned method for preparing squid ink melanin-loaded CeO2 nanoparticles, the mixed solvent in step 1 is composed of ethanol and water in a volume ratio of 1:1.
[0021] The second technical solution of the present invention is to provide squid ink melanin-loaded CeO2 nanoparticles, which are prepared by the method described in the first technical solution.
[0022] The present invention also provides the application of the above-mentioned squid ink melanin-loaded CeO2 nanoparticles as a paraoxon detection reagent.
[0023] The present invention also provides a colorimetric sensor for detecting para-oxygen phosphorus, comprising the aforementioned squid ink melanin-loaded CeO2 nanoparticles.
[0024] This invention also provides a method for detecting paraoxonium by adding Tris-HCl buffer and the cuttlefish juice melanin-loaded CeO2 nanoparticles as described in claim 8 to the sample to be tested, and reacting at 70-90℃ for 10-120 min. After the reaction, qualitative judgment is made by observing the color of the reaction solution and / or quantitative analysis is made by measuring the absorbance of the solution using a UV spectrophotometer.
[0025] Compared with the prior art, the technical solution provided by the present invention has the following technical advantages:
[0026] 1. The cuttlefish juice melanin-loaded CeO2 nanoparticles prepared in this invention overcome the problems of easy aggregation of traditional CeO2 nanoparticles, resulting in fewer active sites and lower enzyme-like activity. The stable structure of cuttlefish juice melanin provides a suitable carrier for CeO2 nanoparticles, forming a structure in which CeO2 nanoparticles encapsulate melanin particles.
[0027] 2. The cuttlefish juice melanin-loaded CeO2 nanoparticles prepared in this invention have good phosphatase (OPH) activity, which is higher than that of CeO2 nanoparticles. The colorimetric sensor constructed based on CeO2-CI OPH activity has a wide detection range for PO, a low detection limit, and good selectivity. In addition, it has a good recovery rate when detecting actual samples, indicating its application potential in detecting actual samples. Attached Figure Description
[0028] Figure 1 (a) and (d) are SEM images of CeO2 prepared in Comparative Example 7, (b) and (e) are SEM images of CI, and (c) and (f) are SEM images of CeO2-CI prepared in Example 1.
[0029] Figure 2 (a) and (d) are TEM images of CeO2 prepared in Comparative Example 7, (b) and (e) are TEM images of CI, and (c) and (f) are TEM images of CeO2-CI prepared in Example 1.
[0030] Figure 3 (a) Evaluation of OPH-like activity of the materials prepared in Example 1 and Comparative Examples 1-7; (b), (c), and (d) are single-factor experiments on reaction time, temperature, and pH of CeO2-CI prepared in Example 1, respectively.
[0031] Figure 4 (a) Michaelis-Menten curve and (b) Lineweaver-Burk curve of OPH-like activity of CeO2-CI prepared in Example 1;
[0032] Figure 5 XPS fine spectra of (a) C 1s, (b) N 1s, (c) O 1s and (d) Ce 3d of CeO2-CI prepared in Example 1;
[0033] Figure 6 For example 1, (a) the absorption spectra of different concentrations of paraoxonium in the wavelength range of 200-600 nm were detected, and (b) the standard curve of paraoxonium was detected.
[0034] Figure 7(a) shows the interference of different anions on oxygen and phosphorus in Example 1, (b) shows the interference of different cations on oxygen and phosphorus in Example 1, and (c) shows the selective detection experiment in Example 1. Detailed Implementation
[0035] The technical solutions of the present invention will be clearly and completely described below with reference to specific embodiments. The present invention will be described in detail below with reference to the accompanying drawings and specific embodiments.
[0036] Example 1
[0037] A method for preparing CeO2 nanoparticles loaded with squid ink melanin includes the following steps:
[0038] (1) Purification of squid ink melanin
[0039] Mix 20 g of squid ink with 80 mL of anhydrous ethanol-water (1:1 volume ratio), stir well, and let stand at 4°C for 12 h. Centrifuge the suspension at 4000 rpm for 10 min, remove the supernatant, wash three times with deionized water, and dry in a 60°C oven for 12 h. After thorough drying, crush and grind the mixture, and record the residue passing through a 200-mesh sieve as crude CI.
[0040] Dissolve 5 g of crude CI in 50 mL of sodium hydroxide solution (1 mM), heat to 80 °C and continue for 2 h. After cooling, centrifuge the mixture at 5000 rpm for 20 min and discard the supernatant. Then centrifuge the mixture at 150 rpm for 8 min and collect the supernatant. Dry the supernatant at 60 °C to obtain purified CI. Unless otherwise specified, CI refers to purified CI.
[0041] (2) Preparation of CeO2-CI
[0042] 250 mg of cerium trichloride was dissolved in 37 mL of deionized water. Then, 1.25 mL of H₂O₂, 3.75 mL of ammonia, and 50 mg of CI were added sequentially with rapid stirring. The temperature was raised to 100 °C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO₂-CI, denoted as CeO₂-CI₅:1.
[0043] Example 2
[0044] This embodiment provides another type of squid ink melanin-loaded CeO2 nanoparticles, which were prepared by the following steps: 500 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the sequential addition of 1.25 mL of H2O2, 3.75 mL of ammonia, and 50 mg of CI prepared in Example 1, and the mixture was rapidly stirred. The temperature was raised to 100°C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60°C to obtain CeO2-CI-10:1.
[0045] Example 3
[0046] This embodiment provides another type of squid ink melanin-loaded CeO2 nanoparticles, which are prepared by the following steps:
[0047] 125 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the addition of 1.25 mL of H₂O₂, 3.75 mL of ammonia, and 50 mg of the Cl prepared in Example 1. The mixture was stirred rapidly, and the temperature was raised to 100 °C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO₂-Cl₂.5:1.
[0048] Example 4
[0049] This embodiment provides another type of squid ink melanin-loaded CeO2 nanoparticles, which are prepared by the following steps:
[0050] 250 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the addition of 1.25 mL of H₂O₂, 3.75 mL of ammonia, and 50 mg of the Cl prepared in Example 1, with rapid stirring. The temperature was raised to 80 °C and maintained for 90 min. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO₂-Cl₅:1-80.
[0051] Example 5
[0052] This embodiment provides another type of squid ink melanin-loaded CeO2 nanoparticles, which are prepared by the following steps:
[0053] 250 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the addition of 1.25 mL of H₂O₂, 3.75 mL of ammonia, and 50 mg of the Cl prepared in Example 1, with rapid stirring. The temperature was raised to 90 °C and maintained for 30 min. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO₂-Cl₅:1-90.
[0054] Comparative Example 1
[0055] This comparative example provides another squid ink melanin-loaded CeO2 nanoparticle, which was prepared by the following steps: 50 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the sequential addition of 1.25 mL of H2O2, 3.75 mL of ammonia, and 500 mg of Cl, with rapid stirring. The temperature was raised to 100 °C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO2-Cl-1:10.
[0056] Comparative Example 2
[0057] This comparative example provides another squid ink melanin-loaded CeO2 nanoparticle, which was prepared by the following steps: 50 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the sequential addition of 1.25 mL of H2O2, 3.75 mL of ammonia, and 250 mg of Cl, with rapid stirring. The temperature was raised to 100 °C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO2-Cl-1:5.
[0058] Comparative Example 3
[0059] This comparative example provides another squid ink melanin-loaded CeO2 nanoparticle, which was prepared by the following steps: 50 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the sequential addition of 1.25 mL of H2O2, 3.75 mL of ammonia, and 125 mg of Cl, with rapid stirring. The temperature was raised to 100 °C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO2-Cl-1:2.5.
[0060] The CIs described in Comparative Examples 1-3 were prepared using the CI preparation method described in Example 1.
[0061] Comparative Example 4
[0062] This comparative example provides CeO2 nanoparticles loaded without squid ink melanin, which were prepared by the following steps: 50 mg of cerium trichloride was dissolved in 37 mL of deionized water, followed by the sequential addition of 1.25 mL of H2O2 and 3.75 mL of ammonia water, and the mixture was stirred rapidly. The temperature was raised to 100 °C and maintained for 1 h. The reaction mixture was then centrifuged and washed several times with deionized water until the pH was neutral. Finally, the brown precipitate was washed with methanol and thoroughly dried at 60 °C to obtain CeO2.
[0063] To verify the OPH-like activity of the squid ink melanin-loaded CeO2 nanoparticles provided in this application, the following are the OPH-like activity experimental results of the squid ink melanin-loaded CeO2 nanoparticles prepared in Example 1 and the above comparative examples.
[0064] 1. Evaluation of CeO2-CI type OPH activity
[0065] This experiment used disodium p-nitrophenyl phosphate (p-NPP) as a hydrolysis substrate to determine the activity of OPH-like enzymes. The strength of the OPH-like activity was evaluated by comparing the ability of different materials to hydrolyze p-NPP into p-nitrophenol.
[0066] The prepared CeO2-CI was dispersed in deionized water to prepare a nanozyme solution with a concentration of 1 mg / mL, and Tris-HCl (0.1 M) at pH 9 was prepared. In a 2 mL centrifuge tube, 600 μL of Tris-HCl buffer (0.1 M pH 9), 100 μL of p-NPP (1 mM), and 300 μL of nanozyme solution (1 mg / mL) were added, and the reaction was carried out at 70℃ for 60 min. Because the hydrolysis product p-nitrophenol has an absorption peak at 400 nm in the ultraviolet light, after the reaction, the absorbance spectrum of the solution in the centrifuge tube in the wavelength range of 200-600 nm was measured using a UV spectrophotometer. The intensity of the absorption peak near 400 nm can be used to determine the OPH-like activity of the material.
[0067] from Figure 3As shown in Figure a, the OPH-like activity of the material is mainly contributed by CeO2. In materials with CI as the main component, such as CeO2-CI-1:10, CeO2-CI-1:5, and CeO2-CI-1:2.5, the OPH-like activity is very low, but it increases with the increase of CeO2. In materials with CeO2 as the main component, the OPH-like activity of the material shows a trend of first increasing and then decreasing with the increase of CI, and all of them are higher than the OPH-like activity of pure CeO2. Among them, CeO2-CI-5:1 has the highest OPH-like activity. Unless otherwise specified, CeO2-CI in subsequent experiments refers to CeO2-CI-5:1.
[0068] Single-factor experiments were conducted on the single factors affecting the activity of CeO2-CI type OPH, such as reaction time, temperature and pH.
[0069] Reaction time: The experimental protocol was similar to that for the enzyme activity evaluation described above, with reaction times set at 10 min, 20 min, 30 min, 40 min, 50 min, and 60 min.
[0070] Temperature: The experimental protocol was similar to that for the enzyme activity evaluation described above, with reaction temperatures set at 25℃, 40℃, 50℃, 60℃, 70℃ and 80℃.
[0071] pH: The experimental protocol was similar to that for the enzyme activity evaluation described above, with the pH of the Tris-HCl buffer set at 7, 8, 9, 10, 11, 12, and 13.
[0072] like Figure 3 As shown in b, the intensity of the absorption peak increases with time between 10 and 50 minutes, but there is no significant increase between 50 and 60 minutes, indicating that the reaction has ended. Therefore, a reaction time of 50 minutes was chosen for subsequent experiments. Figure 3 As shown in Figure c, experiments were conducted between 25-80℃. With increasing temperature, the absorption peak at 400 nm became increasingly higher, indicating that the OPH-like activity of CeO2-CI increases with temperature and is significantly affected by temperature. Figure 3 As shown in d, experiments were conducted under pH conditions of 7-13. As pH increased, the absorption peak at 400 nm became higher and higher, indicating that the OPH-like activity of CeO2-CI increased with increasing pH and was greatly affected by pH.
[0073] Furthermore, the catalytic kinetic parameters (KL) of CeO2-CI similar to OPH were calculated using the Michaelis-Menten equation. m and V max ).like Figure 4As shown in figures a and b, the reaction rate initially increases and then plateaus with increasing substrate concentration, consistent with the Michaelis-Menten model (R² = 0.999). The double reciprocal curves of substrate concentration and reaction rate also show a good linear relationship (R² = 0.995). The steady-state kinetic parameter K was calculated. m =0.875 mM, V max =0.088 μM / s.
[0074] 2. Morphological and structural analysis of CeO2-CI
[0075] SEM image of CeO2-CI prepared in Example 1 is shown below. Figure 1 As shown, the three-dimensional structure of CeO2 under SEM is irregular granular, while that of CI is spherical, and the three-dimensional structure of CeO2-CI is a combination of granular and spherical shapes. TEM images are shown below. Figure 2 As shown, CeO2 appears as irregular granules under TEM, while CI appears as black spheres. The CeO2-CI morphology is a ring of granular material surrounding the black spheres, demonstrating the combination of CeO2 and CI.
[0076] like Figure 5 As shown in figure a, the C 1s spectral peaks of CeO2-CI appear near 287.98 eV and 284.80 eV, corresponding to OC=O and CC, respectively. Figure 5 As shown in b, a peak in the N 1s spectrum of CeO2-CI appears near 399.70 eV, corresponding to C-NH2. Peaks in the O 1s spectrum of CeO2-CI appear near 532.70 eV, 531.39 eV, and 529.44 eV. Figure 5 c), corresponding to C=O, CO, and Ce-O. For example... Figure 5 As shown in d, the Ce 3d fine spectrum of CeO2-CI shows a total of ten energy peaks, consistent with the simultaneous presence of Ce in Ce. 3+ and Ce 4+ The energy peaks near 910 eV are Ce 3d3 / 2, and the energy peaks near 890 eV are Ce 3d5 / 2. These belong to Ce. 3+ The spectral peaks appear at 903.99 eV, 898.12 eV, 885.21 eV, and 880.52 eV, belonging to Ce. 4+ The energy spectrum peaks appear at 916.61 eV, 907.57 eV, 901.07 eV, 898.3 eV, 888.83 eV, and 882.4 eV. Among them, Ce... 4+ Ce accounts for 9.82% of all elements.3+ Ce accounts for 3.49% of all Ce elements. 4+ It constitutes the majority of Ce elements. This aligns with previous literature mentioning Ce... 3+ and Ce 4+ The synergistic effect of Ce can help improve the natural phosphatase activity of materials, and Ce 4+ The conclusion is that it plays a leading role in the catalytic process.
[0077] Application Example 1
[0078] This application example provides a method for detecting paraoxonium using squid ink melanin-loaded CeO2 nanoparticles. 490 μL of Tris-HCl buffer (0.1 M pH 10), 500 μL of CeO2-CI (1 mg / mL) prepared in Example 1, and 10 μL of a 20 μM paraoxonium solution are added to a 2 mL centrifuge tube. The reaction is carried out at 80°C for 50 min. After the reaction, the absorbance of the solution at 400 nm is measured using a UV spectrophotometer, with three parallel tests performed. A linear curve is obtained by plotting the paraoxonium concentration on the x-axis and the absorbance at 400 nm on the y-axis; this curve serves as the standard curve for paraoxonium detection. Application Examples 2-8
[0079] Application Example 2-8
[0080] Application Examples 2-8 provide a method for detecting paraoxonium using CeO2 nanoparticles loaded with squid ink melanin. This method is basically the same as the method provided in Application Example 1, except that the concentration of the paraoxonium solution is different, as detailed in Table 1.
[0081] Table 1
[0082]
[0083] See the test results in Application Examples 1-8 Figure 6 ,like Figure 6 As shown in Figure a, when the concentration of paraoxonium increased from 0.09 μM to 144 μM, the absorbance at 400 nm gradually increased, indicating that paraoxonium was hydrolyzed by CeO2-CI to p-nitrophenol. A standard curve was plotted using CeO2-CI to detect paraoxonium from 0.9 to 144 μM, with the absorbance at 400 nm as the ordinate and the paraoxonium concentration as the abscissa, as shown in Figure a. Figure 6 As shown in b, the standard curve equation is Y=0.0073X+0.04135 (R²=0.9993), the linear detection range is 0.9-144 μM, and the limit of detection (LOD) is 0.314 μM.
[0084] To explore the practical application value of CeO2-CI detection for oxygen and phosphorus, it is necessary to evaluate the performance of CeO2-CI detection in resisting interference from anions and cations and its selectivity for oxygen and phosphorus.
[0085] 1. Anti-interference and selectivity experiments of a colorimetric sensor constructed based on CeO2 nanoparticles loaded with squid ink melanin.
[0086] Anti-interference test: 20 μL of phosphorus paraoxygenate (500 μg / mL), 500 μL of CeO2-Cl (1 mg / mL), 470 μL of Tris-HCl (pH 10), and 10 μL of different types of anions and cations (1 mg / mL) were added to 2 mL centrifuge tubes. A blank control group without added anions and cations was set up. The reaction system remained unchanged, and all reactions were carried out at 80℃ for 50 min. The relative activity of the anion and cation experimental groups was calculated with the absorbance at 400 nm of the blank group as 100% relative activity.
[0087] Selectivity: 500 μL CeO2-Cl (1 mg / mL), 470 μL Tris-HCl (pH 10), and 20 μL of different organophosphorus pesticides (1 mg / mL) were added to 2 mL centrifuge tubes. A blank control group with the same concentration of paraoxon was also added. The reaction system remained unchanged, and all reactions were carried out at 80℃ for 50 min. The relative activity of the other organophosphorus pesticide experimental groups was calculated using the absorbance at 400 nm of the blank group as 100% relative activity.
[0088] See results Figure 7 ,like Figure 7 As shown in Figure a, at the same concentration, Cl- and CO3... 2- SO4 2- and NO3 - These four anions did not significantly interfere with the detection of oxygen and phosphorus in CeO2-Cl, and PO42- 3- The CeO2-CI assay showed a decrease of approximately 16% in relative activity against oxygen and phosphorus, possibly because some of the OPH-like active sites of CeO2-CI were converted by PO4. 3- Occupancy leads to a decrease in relative activity. For example... Figure 7 As shown in b, at the same concentration, Na + Ca 2+ and K + These three cations did not significantly interfere with the detection of oxygen and phosphorus in CeO2-Cl, while Mg 2+ The approximately 10% reduction in the relative activity of CeO2-CI for oxygen and phosphorus may be due to the interaction between some OPH-like active sites of CeO2-CI and Mg. 2+ This combination leads to a decrease in relative activity.
[0089] like Figure 7 As shown in c, at the same concentration, the CeO2-CI-based detection method exhibits high selectivity for phosphorus oxygenation. Other common organophosphorus pesticides do not produce p-nitrophenol after hydrolysis and do not show a significant absorption peak at 400 nm. The relative activity of profenofos based on CeO2-CI detection is 12.7%, while that of glyphosate, glufosinate, isoprocarb, and phoxim is all below 10%, demonstrating that the CeO2-CI-based detection method for phosphorus oxygenation has good selectivity.
[0090] To evaluate the application of this method for detecting paraoxonium in actual samples, paraoxonium standard solutions of different concentrations were added to tap water filtered through a 0.22 μm aqueous membrane using a spiked method. Finally, the glyphosate content was determined according to the method described in Application Example 1.
[0091] 2. Detection of actual samples by colorimetric sensors
[0092] The results of the spiked recovery test are shown in Table 2. Under the conditions of spiked values of 18, 36 and 72 μM, the recoveries were 104.63%, 97.27% and 99.94%, respectively, and the RSDs were all below 3%, indicating that the detection method can sensitively detect oxygen and phosphorus in tap water.
[0093] Table 2. Determination of paraoxon in tap water by spiking recovery method (n=9)
[0094]
[0095] In summary, this invention is the first to introduce CI into the CeO2 synthesis process, preparing a novel catalyst, CeO2-CI. This catalyst exhibits good OPH-like activity. A colorimetric sensor constructed based on the OPH-like activity of CeO2-CI can specifically detect paraoxygenated phosphorus within a detection range of 0.09-144 μM, with a detection limit of 0.363 μM. Spiking analysis showed that this detection method achieved a recovery rate of 97-105% (RSD < 3%) in tap water samples, indicating that this method can sensitively detect residual paraoxygenated phosphorus in tap water and has good prospects for practical application.
[0096] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. For those skilled in the art, any changes or substitutions that can be easily conceived, such improvements and modifications, are also considered to be within the scope of protection of the present invention.
Claims
1. A method for preparing CeO2 nanoparticles loaded with squid ink melanin, characterized in that, The steps are as follows: 1) Dissolve cerium trichloride in deionized water, then add squid ink melanin, ammonia and hydrogen peroxide in sequence; 2) With vigorous stirring, keep the solution from step 1) at 80-100℃ for 30-90 min; 3) After the reaction solution is cooled to room temperature, the precipitate is collected by centrifugation, washed and dried to obtain CeO2-CI nanoparticles loaded with squid ink melanin. The mass ratio of cerium trichloride to squid ink melanin is 2.5-10:
1.
2. The method for preparing CeO2 nanoparticles loaded with squid ink melanin according to claim 1, characterized in that, The mass ratio of cerium trichloride, squid ink melanin, ammonia, and hydrogen peroxide is 2.5-10 : 1 : 20-30 : 60-80.
3. The method for preparing CeO2 nanoparticles loaded with squid ink melanin according to claim 1, characterized in that, The squid ink melanin mentioned above is prepared by the following method: 1) Cut open the ink sac of the cuttlefish, take the cuttlefish ink and mix it with the mixed solvent, place it in the refrigerator to soak, and obtain a suspension; centrifuge the suspension and take the precipitate, wash it, dry the washed solid, grind it and sieve it to obtain the melanin cuttlefish ink, which is recorded as crude CI. 2) Take the sieved crude CI and dissolve it in sodium hydroxide solution. Heat it at 70-90℃ for 1-3 hours. After cooling, centrifuge the mixture and discard the supernatant. Then centrifuge the mixture a second time and take the supernatant. Dry the supernatant to obtain purified CI.
4. The method for preparing CeO2 nanoparticles loaded with squid ink melanin according to claim 3, characterized in that, Step 1) The ratio of squid ink to mixed solvent is 1 g: 4 mL; the soaking time is 6-18 h.
5. The method for preparing CeO2 nanoparticles loaded with squid ink melanin according to claim 1, characterized in that, Step 2) The ratio of crude CI to sodium hydroxide solution is 1 g: 10 mL; the concentration of sodium hydroxide solution is 1 mM; the initial centrifugation is to centrifuge the cooled mixed solution at 5000 rpm for 20 min, and the secondary centrifugation is to centrifuge at 150 rpm for 8 min.
6. The method for preparing CeO2 nanoparticles loaded with squid ink melanin according to claim 3, characterized in that, The mixed solvent described in step 1 consists of ethanol and water in a volume ratio of 1:
1.
7. A type of squid ink melanin-loaded CeO2 nanoparticle, characterized in that, Prepared by the method described in any of claims 1-8.
8. The application of the squid ink melanin-loaded CeO2 nanoparticles as described in claim 7 as a reagent for detecting paraoxonium.
9. A colorimetric sensor for detecting paraoxon and phosphorus, characterized in that, Including the squid ink melanin-loaded CeO2 nanoparticles as described in claim 7.
10. A method for detecting paraoxonium using squid ink melanin-loaded CeO2 nanoparticles, characterized in that, Tris-HCl buffer and CeO2 nanoparticles loaded with squid ink melanin as described in claim 8 were added to the sample to be tested. The mixture was reacted at 70-90℃ for 10-120 min. After the reaction was completed, the color of the reaction solution was observed for qualitative judgment and / or the absorbance of the solution was measured by ultraviolet spectrophotometer for quantitative analysis.