CN-121978069-A - Preparation of iron-copper bimetallic nano-enzyme and application of iron-copper bimetallic nano-enzyme in detection of tetracycline antibiotics in food

Abstract

The invention provides a preparation method of iron-copper bimetallic nano-enzyme and application thereof in detection of tetracycline antibiotics in food. The nano-enzyme is synthesized by a simple coordination-assisted polymerization assembly method, and has excellent colorimetric performance (activity of similar peroxidase and similar laccase) and fluorescence performance. The three-channel colorimetric fluorescence sensing array constructed based on the performance can realize the accurate identification of five tetracycline antibiotics (tetracycline, terramycin, doxycycline, minocycline and metacycline) and can effectively distinguish binary and ternary mixed tetracycline antibiotics with different proportions. In order to break through the equipment and cost limitations of the traditional detection method, the invention also develops a portable detection device with a built-in test paper sheet, combines the smart phone photographing and Image J software analysis, realizes the rapid and on-site detection of tetracycline antibiotics in actual samples such as milk, chicken farm sewage and the like, and has good practical application prospect.

Inventors

• NIU XIAODI
• WANG XINYAN
• WANG HONGSU
• WANG YIRU

Assignees

• 吉林大学

Dates

Publication Date

20260505

Application Date

20260205

Claims (7)

1. 1. The Fe-Cu bimetallic nano-enzyme is characterized by comprising the following preparation method: Firstly adding ethanol and distilled water with equal volume into a round bottom flask, uniformly mixing, then adding 1 g Pluronic F127, 1 mL of 1,3, 5-Trimethylbenzene (TMBE) and 0.75 g dopamine hydrochloride (DA) into the system, uniformly stirring, then dissolving ferrous ammonium sulfate hexahydrate ((NH 4 ) 2 Fe(SO 4 ) 2 ·6H 2 O) and cupric ammonium hexahydrate ((NH 4 ) 2 Cu(SO 4 ) 2 ·6H 2 O)) with the molar concentration of 1:1 into the solution, then dropwise adding a certain amount of ammonia water into the solution, reacting for 3 hours to obtain a FeCu/PDA/F127 composite material, centrifuging, washing and freeze-drying the collected product, calcining for 3 hours at 350 ℃ under nitrogen, and calcining for 5 hours at 800 ℃ to obtain the FeCu nano enzyme.
2. 2. The application of the iron-copper bimetallic nano-enzyme in detecting tetracycline antibiotics in foods is characterized in that a three-channel colorimetric fluorescence sensing detection array is constructed by utilizing the colorimetric performance and the fluorescence performance of the FeCu bimetallic nano-enzyme, so that the detection and the distinction of five common tetracycline antibiotics in foods are realized, and meanwhile, a portable detection device is constructed, so that the rapid and accurate detection of the tetracycline antibiotics in actual samples is realized.
3. 3. The use according to claim 2, wherein the three-channel colorimetric fluorescence sensing detection array consists of a peroxidase-like colorimetric channel, a laccase-like colorimetric channel and a fluorescent channel, wherein the array is configured as a three-channel x five tetracycline antibiotics to be tested x five sets of parallel tests, and the five tetracycline antibiotics are tetracycline, oxytetracycline, doxycycline, minocycline and metacycline.
4. 4. The application of the three-channel kit according to claim 3, wherein the three-channel reaction system comprises a first channel and a laccase-like colorimetric channel, wherein the first channel and the laccase-like colorimetric channel comprise 2-morpholinoethanesulfonic acid buffer (MES buffer) +FeCu nano-enzyme+tetracycline antibiotics+2, 4-dichlorophenol (2, 4-DP) +4-amino-thiolin (4-AP), the second channel and the peroxidase-like colorimetric channel comprise acetic acid-sodium acetate buffer (HAc-NaAc buffer) +FeCu bimetallic nano-enzyme+tetracycline antibiotics+hydrogen peroxide (H 2 O 2 ) +3,3', 5' -tetramethyl benzidine (TMB), and the third channel and the fluorescent channel comprise phosphoric acid buffer (PBS buffer) +FeCu bimetallic nano-enzyme+tetracycline antibiotics.
5. 5. The use according to claim 2, wherein said detection and differentiation comprises detection and differentiation of single tetracyclines at different concentrations, tetracyclines of different species, binary/ternary mixed tetracyclines of different proportions.
6. 6. The application of claim 2, wherein the portable detection device is prepared by 3D printing after drawing design and three-dimensional modeling, the device body is a cuboid box body with a reserved view finding port, a drawable test paper sheet bearing assembly is arranged in the cuboid box body, LED light sources are assembled at two ends of the box body, and a sensing test paper sheet can be accurately fixed in a groove formed in the drawing assembly.
7. 7. The application of the portable detection device according to claim 2, wherein the detection process of the portable detection device is that after a test paper sheet loaded with FeCu nano enzyme reacts with an actual sample, the test paper sheet is placed in the portable detection device for photographing, an Image J software is used for extracting an Image gray value, and the Image gray value is substituted into a standard curve for calculation, so that the concentration of tetracycline antibiotics in the actual sample is obtained, and the actual sample is milk and chicken farm sewage.

Description

Preparation of iron-copper bimetallic nano-enzyme and application of iron-copper bimetallic nano-enzyme in detection of tetracycline antibiotics in food Technical Field The invention belongs to the technical field of nanotechnology and analysis and detection, and particularly relates to preparation of iron-copper bimetallic nano-enzyme and application thereof in detection of tetracycline antibiotics in food. Background The tetracycline antibiotics have the characteristics of broad-spectrum antibiosis and low cost, are widely applied to the field of food production, and are commonly used for treating and preventing animal diseases especially in the links of livestock and poultry cultivation and aquaculture. However, part of production subjects lack of standard medication consciousness, have problems of overdose use, even illegal addition and the like, cause tetracycline antibiotics to remain in foods such as meat, eggs, milk products, aquatic products and the like, and flow into the consumer market through a food supply chain, thus constituting a potential threat to food safety and human health. Therefore, the establishment of an effective tetracycline antibiotic detection technology has important practical significance. Currently, methods for detecting tetracycline antibiotics mainly include high performance liquid chromatography, liquid chromatography-tandem mass spectrometry, capillary electrophoresis, enzyme-linked immunosorbent assay, and the like. The method has the advantages of high detection accuracy, high instrument cost, complex sample pretreatment and long detection time consumption, is difficult to meet the field rapid screening requirement, is difficult to realize the accurate distinction of various tetracycline antibiotics in a single signal response mode, and cannot adapt to complex detection scenes of coexistence of various tetracycline antibiotics in food. The nano enzyme is used as an artificial enzyme with unique physicochemical properties of nano materials and catalytic activity of natural enzyme, and has become a research hot spot in the field of analysis and detection by virtue of the advantages of simple preparation, high stability, adjustable catalytic activity and the like. The bimetallic nano-enzyme is used as an important branch of nano-enzyme, and through the synergistic effect of two metal elements, the electronic structure of the material can be effectively regulated, the number of active sites can be increased, and the catalytic efficiency can be obviously improved. The Fe-Cu bimetallic nano enzyme has high catalytic activity, high selectivity and high stability due to excellent synergistic effect among bimetallic, is an ideal material for constructing a detection system, can be flexibly regulated and controlled by preparation conditions, and can better adapt to the detection requirement of a target object. The multichannel sensing array technology provides an effective scheme for solving the problem of accurately distinguishing various targets. According to the technology, a multichannel sensing array is constructed, and characteristic fingerprint signals are generated by utilizing interaction differences between different sensing units and target objects, so that efficient and accurate detection and differentiation of various target objects can be realized. Compared with a single signal detection mode, the technology has the characteristics of high flux, high distinction and strong anti-interference capability, and can adapt to the detection scene of coexistence of various target objects in a complex matrix. The invention synthesizes the Fe-Cu bimetallic nano-enzyme (FeCu nano-enzyme), and the material has excellent colorimetric performance (laccase-like activity and peroxidase-like activity) and fluorescence performance. Based on the above properties, a three-channel colorimetric fluorescence sensing array is constructed, and when five tetracycline antibiotics (tetracycline (TC), oxytetracycline (OTC), doxycycline (DOX), minocycline (MINO), metacycline (MET)) and FeCu nano enzymes act, colorimetric and fluorescence signals with different intensities can be triggered due to the difference of molecular structures, so that respective unique signal fingerprints are formed. Through the collection and pattern recognition analysis of the multidimensional signals, the accurate distinguishing and quantitative detection of the five tetracycline antibiotics can be realized rapidly, and the technical problem that the similar antibiotics are difficult to distinguish in the traditional method is effectively solved. In order to break through the limit of expensive equipment and complicated operation of the traditional detection method, the invention also develops a portable detection device with a built-in test paper sheet, and can realize the rapid and on-site detection of tetracycline antibiotics in actual samples such as milk, chicken farm sewage and the like by combining the photograp