CN-122016686-A - Method for quickly detecting sodium dehydroacetate in colorimetric/fluorescent dual-mode by using peroxidase-like laccase nano-enzyme

Abstract

The invention discloses a colorimetric/fluorescent dual-mode rapid detection method of sodium dehydroacetate by using a peroxidase/laccase nano-enzyme, which takes 1-Hydroxybenzotriazole (HOBT) as an organic ligand, selects two transition metals Cu and Zn as central ions, and synthesizes a Cu/Zn-MOF material. The linear range of the colorimetric mode is 0.067-12 mug/mL, the detection limit is 0.020 mug/mL, the linear range of the fluorescent mode is 0.01-22 mug/mL, the detection limit is 0.041 mug/mL, the sensor is applied to the labeling recovery experiment of DHA in food samples, the recovery rate is 92.1% -106.4%, the RSD is not more than 5%, and the analysis method is verified to have good stability, reliability and reproducibility.

Inventors

• ZHANG QIAN
• YANG YALING
• GU YAOHONG
• LI JITAO
• YANG DEZHI

Assignees

• 云南伦扬科技有限公司

Dates

Publication Date

20260512

Application Date

20260414

Claims (3)

1. 1. The method for quickly detecting sodium dehydroacetate in colorimetric/fluorescent dual mode by using the peroxidase/laccase-like nano enzyme is characterized by comprising the following steps of: (1) Dissolving 0.080-0.100 g CuCl 2 ·2H 2 O in 10-15 mL methanol, 1.00-1.20 g of 1-hydroxybenzotriazole in 10-15 mL methanol, dissolving 0.100-0.150 g Zn (OAc) 2 ·2H 2 O in 10-15 mL methanol, respectively performing ultrasonic treatment for 5-10min to completely dissolve, slowly adding CuCl 2 ·2H 2 O solution into 1-hydroxybenzotriazole solution, stirring for 5-10min, slowly adding Zn (OAc) 2 ·2H 2 O solution, continuously stirring at room temperature for 12-15 h, standing for 5-10 h, centrifuging, taking out solid, washing with ethanol for 3-4 times, and vacuum drying to obtain Cu/Zn-MOF nano enzyme; (2) Adding 2, 4-dichlorophenol and 4-aminoantipyrine solution into a buffer solution with the pH of 6.5 MES, adding Cu/Zn-MOF nano enzyme and sodium dehydroacetate standard solutions with different concentrations, uniformly mixing, incubating at room temperature for 20-30 min, measuring absorbance at the wavelength of 505 and nm, and determining the linear relation between the concentration of sodium dehydroacetate and the absorbance value to obtain a regression equation; (3) Adding o-phenylenediamine and H 2 O 2 solution into pH 6.5 MES buffer solution, adding Cu/Zn-MOF nano enzyme and sodium dehydroacetate standard solutions with different concentrations, uniformly mixing, incubating at room temperature for 20-30min, and determining the linear relation between the concentration of sodium dehydroacetate and the fluorescence intensity when the excitation wavelength is 421 nm and the fluorescence intensity is 564 nm, thereby obtaining a regression equation; (4) And (3) measuring the absorbance and the fluorescence intensity of the sample liquid to be measured according to the methods of the steps (2) and (3), and substituting the absorbance and the fluorescence intensity into a regression equation to obtain the concentration of sodium dehydroacetate in the sample liquid to be measured.
2. 2. The method for quickly detecting sodium dehydroacetate by using the similar peroxidase/laccase nanoenzyme colorimetric/fluorescent double mode according to claim 1 is characterized in that the concentration of a Cu/Zn-MOF nanoenzyme solution is 1 mg/mL, the addition amount is 50-100 mu L, the concentration of a2, 4-dichlorophenol solution is 20 mmol/L, the addition amount is 100-200 mu L, the concentration of a 4-aminoantipyrine solution is 20 mmol/L, the addition amount is 100-200 mu L, the concentration of an o-phenylenediamine solution is 20 mmol/L, the addition amount is 100-200 mu L, the concentration of an H 2 O 2 solution is 50 mmol/L, and the addition amount is 100-200 mu L.
3. 3. The method for rapidly detecting sodium dehydroacetate based on nano enzyme according to claim 1 wherein the centrifugation in step (1) is performed at 5000-10000 r/min for 5-10: 10 min.

Description

Method for quickly detecting sodium dehydroacetate in colorimetric/fluorescent dual-mode by using peroxidase-like laccase nano-enzyme Technical Field The invention relates to the technical field of chemical analysis and detection, in particular to a method for quickly detecting sodium dehydroacetate in a colorimetric/fluorescent dual-mode by using a peroxidase/laccase-like nano enzyme. Background Sodium dehydroacetate (DHA-S) is a food additive, has antibacterial effect, and can be used as antiseptic for food and product, and for inhibiting mold growth and prolonging shelf life of food. The substance is difficult to be discharged through metabolism of human body, excessive intake can accumulate in the body, and adverse reactions such as emesis, confusion, liver and kidney function injury and the like are caused. The use of sodium dehydroacetate in foods such as starch products, breads, pastries and baked fillings has been eliminated by the use of national food additive standards for food safety (GB 2760-2024) implemented in 2.8.2025. Therefore, the determination of the content of sodium dehydroacetate in the food has great significance for human health. At present, the reported methods for measuring DHA-S include high performance liquid chromatography, gas chromatography, electrochemiluminescence, etc. The colorimetric method and the fluorescent method have important positions in the detection technology because of simple and rapid operation and no need of large-scale instruments and equipment, and colorimetric and fluorescent detection of DHA-S are freshly reported. The Metal Organic Frameworks (MOFs) are supermolecule assemblies composed of metal ions and organic ligands, are candidate materials capable of selectively generating Reactive Oxygen Species (ROS) and high in utilization rate of reactive free radicals, and doped metal-nitrogen-carbon (Me-N-C) structures constructed by the MOFs can promote uniform distribution of heteroatoms in the materials, and can enhance the synergistic effect between adjacent non-bonded metal atoms, and simultaneously can regulate and control catalytic activity and nano enzyme types by selecting proper ligands and central ions. Disclosure of Invention The invention provides a colorimetric/fluorescent dual-mode rapid detection method for sodium dehydroacetate by using a peroxidase/laccase nano-enzyme, which takes 1-Hydroxybenzotriazole (HOBT) as an organic ligand, selects two transition metals Cu and Zn as central ions, and synthesizes a Cu/Zn-MOF material. Cu/Zn-MOF shows excellent laccase-like performance, 2, 4-dichlorophenol (2, 4-DP) is oxidized and coupled with 4-aminoantipyrine (4-AP) to generate red quinone imine, characteristic absorption peak is generated at 505 nm, meanwhile, cu/Zn-MOF has peroxidase-like activity, o-phenylenediamine (OPD) is oxidized to generate fluorescent product DAP, characteristic fluorescent emission peak is generated at 564 nm, and introduction of sodium dehydroacetate (DHA-S) can specifically inhibit laccase-like activity and POD-like enzyme activity of Cu/Zn-MOF, so that red color is reduced and fluorescent intensity is reduced, and a rapid and high-sensitivity colorimetric-fluorescent DHA-S dual-mode sensing detection method is constructed. The linear range of the colorimetric mode is 0.067-12 mug/mL, the detection limit is 0.020 mug/mL, the linear range of the fluorescent mode is 0.01-22 mug/mL, the detection limit is 0.041 mug/mL, the sensor is applied to the standard adding and recycling experiment of DHA-S in food samples, the recycling rate is 92.1% -106.4%, the RSD is not more than 5%, and the analysis method is verified to have good stability, reliability and reproducibility. The method has the characteristics of high sensitivity, strong specificity, simple and quick operation and the like. The invention relates to a colorimetric/fluorescent dual-mode rapid detection method for sodium dehydroacetate by using peroxidase-like/laccase nano-enzyme, which comprises the following steps: (1) Dissolving 0.080-0.100 g CuCl 2·2H2 O in 10-15 mL methanol, 1.00-1.20 g of 1-hydroxybenzotriazole in 10-15 mL methanol, dissolving 0.100-0.150 g Zn (OAc) 2·2H2 O in 10-15 mL methanol, respectively performing ultrasonic treatment for 5-10min to completely dissolve, slowly adding CuCl 2·2H2 O solution into 1-hydroxybenzotriazole solution, stirring for 5-10min, slowly adding Zn (OAc) 2·2H2 O solution, continuously stirring at room temperature for 12-15 h, standing for 5-10 h, centrifuging, taking out solid, washing with ethanol for 3-4 times, and vacuum drying to obtain Cu/Zn-MOF nano enzyme; (2) Adding 2, 4-dichlorophenol and 4-aminoantipyrine solution into a buffer solution with the pH of 6.5 MES, adding Cu/Zn-MOF nano enzyme and sodium dehydroacetate standard solutions with different concentrations, uniformly mixing, incubating at room temperature for 20-30 min, measuring absorbance at the wavelength of 505 and nm, and determining the linear relation