CN109254046B - Nitrofuran antibiotic sensor and preparation method thereof - Google Patents
Nitrofuran antibiotic sensor and preparation method thereof Download PDFInfo
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Abstract
The invention discloses a preparation method of a nitrofuran antibiotic sensor. Belongs to the technical field of novel nanometer functional materials and biosensing analysis. Firstly, preparing a cobalt-nickel bimetal layered hydroxide nanosheet array on a disposable throwable electrode, and utilizing the large specific surface area and the high-activity hydroxyl functional group of the cobalt-nickel bimetal layered hydroxide nanosheet array and the amino functional group of polydopamine to directly and successively prepare a polydopamine film containing an electronic mediator and a molecularly imprinted polymer taking nitrofuran antibiotics as template molecules on the cobalt-nickel bimetal layered hydroxide nanosheet array in an in-situ growth method, wherein after the template molecules are eluted, the original positions of the template molecules are changed into cavities, namely the molecularly imprinted polymer of the template molecules is eluted, so that the preparation of the nitrofuran antibiotic sensor is completed.
Description
Technical Field
The invention relates to a preparation method and application of an electrochemical analysis sensor. Belongs to the technical field of novel nanometer functional materials and biosensing analysis.
Background
Nitrofuran antibiotics are 5-nitro-2 substituted furan derivatives and once are important anti-infective drugs. The medicines mainly comprise furazolidone, nitrofurantoin, furacilin and the like, and are mainly used for treating urinary system infection, intestinal bacterial infection and skin wound infection and used as food additives for preventing poultry intestinal infectious diseases. However, due to the genotoxicity and carcinogenic effects of furazolidone, its use in humans and animals has been banned by the Food and Drug Administration (FDA) and the European Medicines Administration (EMA) in 2005. Furthermore, the clinical importance of nitro heterocyclic compounds is high due to their cell-induced degeneration and animal carcinogenic toxicity. Therefore, the development of a method for rapidly, highly selectively and sensitively detecting the nitrofuran antibiotics is very important to public health and has wide market application prospect.
The molecular imprinting electrochemical sensor has high specificity selectivity, excellent stability, excellent reproducibility, wide detection range and bottom detection limit. The sensor has the advantages of simple preparation, convenient detection, high sensitivity, low cost and the like, and can be widely applied to the fields of chromatographic separation, membrane separation, solid-phase extraction, drug controlled release, chemical sensing and the like. Molecularly Imprinted Polymers (MIPs), also known as "plastic antibodies", are capable of specifically recognizing and selectively adsorbing a specific target molecule (i.e., template molecule). The molecular imprinting technology has many advantages, such as corrosion resistance of organic reagents, good stability, high temperature resistance and simple preparation. Thus, MIP electrochemical sensors based on MIP in combination with electrochemical sensors (MIP-ECS) have attracted a hot interest in the field of electroanalytical chemistry, especially the detection of small molecule contaminants, over the last few years. However, in the preparation process of the traditional MIP-ECS, the defects of difficult elution of template molecules, difficult control of the thickness of the imprinted membrane, poor reproducibility and the like exist, and the application of the molecularly imprinted membrane in an electrochemical sensor is limited. The problems, especially the technical problems that the sensitivity of the electrochemical sensor is reduced due to the fact that the thickness of the molecularly imprinted membrane is not easy to control, and the stability and the reproducibility are reduced due to the fact that the molecularly imprinted membrane is easy to fall off from the surface of an electrode in the elution process, limit the application of MIP _ ECS, so that the research of a new molecularly imprinted polymer synthesis method, a new molecularly imprinted membrane electrode modification method and a method for combining the molecularly imprinted membrane and a substrate material for solving the preparation and application problems of MIP-ECS has important research significance and market value.
Disclosure of Invention
The invention aims to provide a preparation method of a nitrofuran antibiotic sensor, which has the advantages of strong specificity, simple preparation, convenient detection, high sensitivity and low cost. Based on the purpose, firstly, a cobalt-nickel bimetal layered hydroxide nanosheet array is prepared on a disposable throwable electrode, a polydopamine film containing an electron mediator and a molecularly imprinted polymer taking a nitrofuran antibiotic as a template molecule are sequentially and directly prepared on the cobalt-nickel bimetal layered hydroxide nanosheet array in sequence by utilizing the large specific surface area, the high-activity hydroxyl functional group and the amino functional group of the polydopamine through an in-situ growth method, and after the template molecule is eluted, the original position of the template molecule is changed into a cavity, namely the molecularly imprinted polymer of the template molecule is eluted, so that the nitrofuran antibiotic sensor is prepared. When the nitrofuran type antibiotic sensor is used for detecting nitrofuran type antibiotics, the nitrofuran type antibiotic sensor is inserted into a solution to be detected, and the nitrofuran type antibiotics in the solution to be detected can be adsorbed into a cavity of the NIP. The larger the concentration of the nitrofuran antibiotics in the solution to be detected is, the more the nitrofuran antibiotics are adsorbed in the hole of the NIP. When electrochemical detection is carried out, the intensity of detection current is reduced along with the increase of the nitrofuran antibiotics adsorbed in the cavity of the NIP, so that the concentration of the nitrofuran antibiotics in the solution to be detected can be qualitatively and quantitatively determined according to the reduction degree of the current intensity.
The technical scheme adopted by the invention is as follows:
1. a preparation method of a nitrofuran antibiotic sensor is provided, the nitrofuran antibiotic sensor is obtained by in-situ growth of a template-free molecularly imprinted polymer NIP on a cobalt-nickel bimetallic layered hydroxide nanosheet array electrode CoNi LDH-nanoarray; the template-free molecularly imprinted polymer NIP is a molecularly imprinted polymer without template molecules; the molecularly imprinted polymer without the template molecule is obtained by eluting the template molecule from a MIP containing the template molecularly imprinted polymer; the MIP containing the template molecule engram polymer is the MIP containing the template molecule; the template molecule is nitrofuran antibiotics;
2. the preparation method of the cobalt-nickel bimetal layered hydroxide nanosheet array electrode CoNi LDH-nanoarray in the technical scheme 1 comprises the following preparation steps:
(1) carrying out ultrasonic cleaning treatment on the disposable throwable electrode by respectively using dilute hydrochloric acid, absolute ethyl alcohol and deionized water so as to remove an oxide layer and surface impurities of the disposable throwable electrode;
(2) weighing 1-3 mmol of Ni (NO)3)2And Co (NO)3)2And 3 to 9mmol of urea CO (NH)2)2Placing the mixture into a 50mL beaker, adding 30mL deionized water, stirring until the mixture is clear, and then transferring the mixture into a 50mL polytetrafluoroethylene reaction kettle;
(3) putting the disposable throwable electrode processed in the step (1) into the solution in the reaction kettle in the step (2), and reacting at the temperature of 100-130 ℃ for 9-12 hours to prepare a precursor electrode of the cobalt-nickel bimetal layered hydroxide nanosheet array;
(4) inserting the cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode obtained in the step (3) into phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate, reacting for 4-6 hours at the temperature of 20-40 ℃, taking out and washing with deionized water for 2-4 times to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array electrode CoNi LDH-nanoarray;
the disposable and disposable electrode is selected from one of the following electrodes: foam nickel, foam copper, pure nickel sheets, pure copper sheets, pure cobalt sheets, pure silicon sheets and conductive carbon cloth; said Ni (NO)3)2And Co (NO)3)2In a mixture of nickel and cobalt in a molar ratio of 1: 1;
in the phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate: the concentration of dopamine is 2-5 mg/mL, the concentration of ammonium persulfate is 3-8 mg/mL, the concentration of cobalt nitrate is 0.1-0.5 mg/mL, the concentration of phosphate buffer solution PBS is 0.1mol/L, and the pH value is 7.2-8.5;
3. the preparation method of the template-containing molecularly imprinted polymer MIP by CoNi LDH-nanoarray in-situ growth described in the technical scheme 1 comprises the following preparation steps:
(1) respectively weighing 0.25-0.45 mmol of template molecules and 3-5 mmol of 2-methacrylic acid MAA in an ampoule bottle, adding 8-15 mL of acetonitrile, and carrying out ultrasonic treatment for 30min until all the template molecules and the 2-methacrylic acid MAA are dissolved;
(2) adding 15-25 mmol of ethylene glycol dimethacrylate EDMA into the solution obtained in the step (1), and carrying out ultrasonic treatment for 30min until the mixture is uniformly mixed to obtain a precursor mixed solution;
(3) clamping the CoNi LDH-nanoarray prepared in the technical scheme 2 onto a rotary stirrer, inserting the CoNi LDH-nanoarray into the precursor mixed solution in the step (2), and adding N2Under the temperature of an environment and a water bath of 20-40 ℃, rotationally stirring at the speed of 5-200 r/s, simultaneously dripping 1mmol of azobisisobutyronitrile AIBN into the mixed solution at the speed of 1-20 drops/s to initiate polymerization, and obtaining an in-situ grown MIP containing template molecularly imprinted polymer on CoNi LDH-nanoarray;
4. the preparation steps of the template-free molecularly imprinted polymer NIP grown in situ by CoNi LDH-nanoarray in the technical scheme 1 are as follows: immersing the MIP which is obtained in the technical scheme 3 and grows in situ on the CoNi LDH-nanoarray and contains the template molecularly imprinted polymer into an eluant, eluting the template molecules for 5-20 min at room temperature, and then taking out to obtain the NIP without the template molecularly imprinted polymer; the eluent is a mixed solution of formic acid and methanol, wherein the volume ratio of the formic acid to the methanol is 9 (1-5);
5. the preparation steps of the nitrofuran antibiotic sensor in the technical scheme 1 are as follows: washing the template-free molecularly imprinted polymer NIP which grows on the CoNi LDH-nanoarray in situ prepared in the technical scheme 2-4 with deionized water for 2-4 times, and airing at room temperature to prepare the nitrofuran antibiotic sensor;
6. the nitrofuran antibiotic sensor prepared by the technical scheme 1-5 is applied to detection of nitrofuran antibiotics, and comprises the following application steps:
(1) preparing a standard solution: preparing a group of nitrofuran antibiotic standard solutions with different concentrations including blank standard samples;
(2) modification of a working electrode: inserting the nitrofuran antibiotic sensor serving as a working electrode into the nitrofuran antibiotic standard solutions with different concentrations prepared in the step (1), incubating for 10min, taking out, and washing with deionized water for 3 times;
(3) drawing a working curve: taking a saturated calomel electrode as a reference electrode, taking a platinum wire electrode as a counter electrode, forming a three-electrode system with the modified working electrode in the step (2), connecting the three-electrode system with an electrochemical workstation, and sequentially adding 15mL phosphate buffer solution PBS into an electrolytic bath; detecting a current response of the assembled working electrode by Differential Pulse Voltammetry (DPV); the response current intensity of the blank sample is recordedI 0The response current intensity of the standard solution containing different concentrations of nitrofurans antibiotics is recorded asI iThe difference in response to the decrease in current intensity is ΔI = I 0-I i，ΔIMass concentration of the nitrofuran antibiotic standard solutionCWith a linear relationship therebetween, plotting ΔI－CA working curve; the concentration of the phosphate buffer solution PBS is 10mmol/L, pH, and the value is 7.4; the parameters during DPV detection are set as follows: the range and the direction are 0-1V, the step is 0.05V, the pulse time is 0.05s, the sampling time is 0.016s, and the pulse period is 0.5 s;
(4) detecting nitrofuran antibiotics in a sample to be detected: replacing the nitrofuran antibiotics standard solution in the step (1) with a sample to be detected, detecting according to the methods in the steps (2) and (3), and detecting according to the difference delta of the reduction of the response current intensityIAnd working curve to obtain the content of nitrofuran antibiotics in the sample to be detected;
7. the nitrofuran antibiotics in technical schemes 1-6 are one of the following nitrofuran antibiotics: furazolidone, nitrofurantoin, furacilin.
Advantageous results of the invention
(1) The nitrofuran antibiotic sensor is simple to prepare, convenient to operate, low in cost, applicable to portable detection and has market development prospect, and the sample can be quickly, sensitively and selectively detected;
(2) according to the invention, the molecularly imprinted polymer is grown in situ on the cobalt-nickel bimetal layered hydroxide nanosheet array electrode CoNi LDH-nanoarray for the first time, on one hand, more and more uniform molecularly imprinted polymers can be grown by utilizing the large specific surface area of the CoNi LDH-nanoarray, and the CoNi LDH-nanoarray has excellent electron transfer capacity, so that the detection sensitivity is greatly improved; on the other hand, when dopamine is polymerized on a cobalt-nickel bimetallic layered hydroxide nanosheet array in situ, cobalt ions are creatively doped as an electronic mediator, and electrochemical response current is directly generated during detection, so that the sensor can directly detect in a buffer solution without adding other mediator substances, thereby further reducing the signal background, improving the detection sensitivity, greatly reducing the detection cost and reducing the environmental pollution;
(3) according to the invention, the dopamine is combined with the high-activity hydroxyl functional group rich in the cobalt-nickel double-metal layered hydroxide nanosheet array and the large specific surface area, so that when dopamine is polymerized in situ on the surface of the cobalt-nickel double-metal layered hydroxide nanosheet array, a thin enough polydopamine film is formed and simultaneously the polydopamine film is uniformly covered on the cobalt-nickel double-metal layered hydroxide nanosheet array, thereby well laying more and better polymerized molecularly imprinted polymers for the next step; then utilizing strong connection effect of polydopamine on hydroxyl functional groups and amino groups rich in the molecularly imprinted polymer, skillfully using CoNi LDH-nanoarray as a stirrer, immersing and stirring in the molecularly imprinted precursor mixed solution, and directly growing the molecularly imprinted polymer with the film thickness in situ on the surface of the CoNi LDH-nanoarray by controlling the stirring speed, the dropping speed of a polymerization reaction initiator and the polymerization reaction temperature, so that the CoNi LDH-nanoarray can firmly load the molecularly imprinted polymer on one hand, and the stability and the reproducibility of the prepared electrochemical sensor are obviously improved; on the other hand, the film forming thickness of the molecularly imprinted polymer on the surface of the electrode can be effectively controlled, and the technical problem of poor reproducibility caused by the fact that the film forming thickness of the molecularly imprinted film on the surface of the electrode cannot be controlled is solved; in addition, the preparation method of the invention has important scientific significance and application value for effectively controlling the thickness of the formed film and coating the electronic mediator in situ, and fully improving the sensitivity and detection limit of the electrochemical sensor based on the molecular imprinting.
Detailed Description
Example 1 preparation of CoNi LDH-nanoarray
(1) Carrying out ultrasonic cleaning treatment on the disposable throwable electrode by respectively using dilute hydrochloric acid, absolute ethyl alcohol and deionized water so as to remove an oxide layer and surface impurities of the disposable throwable electrode;
(2) weighing 1mmol Ni (NO)3)2And Co (NO)3)2And 3mmol of urea CO (NH)2)2Put it into a 50mL beaker, add 30mL of deionized water, stir until clear, then transfer to 50mLIn a polytetrafluoroethylene reaction kettle;
(3) putting the disposable throwable electrode processed in the step (1) into the solution in the reaction kettle in the step (2), and reacting at the temperature of 100 ℃ for 12 hours to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode;
(4) inserting the cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode obtained in the step (3) into phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate, reacting for 4 hours at the temperature of 20 ℃, taking out and washing with deionized water for 2 times to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array electrode CoNi LDH-nanoarray;
wherein the disposable throwable electrode is foamed nickel; said Ni (NO)3)2And Co (NO)3)2In a mixture of nickel and cobalt in a molar ratio of 1: 1; the concentration of dopamine is 2 mg/mL, the concentration of ammonium persulfate is 3 mg/mL, the concentration of cobalt nitrate is 0.1 mg/mL, the concentration of phosphate buffer solution PBS is 0.1mol/L, and the pH value is 7.2.
Example 2 preparation of CoNi LDH-nanoarray
(1) Carrying out ultrasonic cleaning treatment on the disposable throwable electrode by respectively using dilute hydrochloric acid, absolute ethyl alcohol and deionized water so as to remove an oxide layer and surface impurities of the disposable throwable electrode;
(2) weighing 2 mmol of Ni (NO)3)2And Co (NO)3)2And 6 mmol of urea CO (NH)2)2Placing the mixture into a 50mL beaker, adding 30mL deionized water, stirring until the mixture is clear, and then transferring the mixture into a 50mL polytetrafluoroethylene reaction kettle;
(3) putting the disposable throwable electrode processed in the step (1) into the solution in the reaction kettle in the step (2), and reacting for 11 hours at the temperature of 110 ℃ to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode;
(4) inserting the cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode obtained in the step (3) into phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate, reacting at the temperature of 30 ℃ for 5 hours, taking out and washing with deionized water for 3 times to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array electrode CoNi LDH-nanoarray;
wherein the disposable throwable electrode is a pure copper sheet; said Ni (NO)3)2And Co (NO)3)2In a mixture of nickel and cobalt in a molar ratio of 1: 1; the concentration of dopamine is 3.5 mg/mL, the concentration of ammonium persulfate is 6.2 mg/mL, the concentration of cobalt nitrate is 0.3 mg/mL, the concentration of phosphate buffer solution PBS is 0.1mol/L, and the pH value is 8.0.
Example 3 preparation of CoNi LDH-nanoarray
(1) Carrying out ultrasonic cleaning treatment on the disposable throwable electrode by respectively using dilute hydrochloric acid, absolute ethyl alcohol and deionized water so as to remove an oxide layer and surface impurities of the disposable throwable electrode;
(2) weighing 3mmol Ni (NO)3)2And Co (NO)3)2And 9mmol of urea CO (NH)2)2Placing the mixture into a 50mL beaker, adding 30mL deionized water, stirring until the mixture is clear, and then transferring the mixture into a 50mL polytetrafluoroethylene reaction kettle;
(3) putting the disposable throwable electrode processed in the step (1) into the solution in the reaction kettle in the step (2), and reacting at the temperature of 130 ℃ for 9 hours to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode;
(4) inserting the cobalt-nickel bimetal layered hydroxide nanosheet array precursor electrode obtained in the step (3) into phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate, reacting at the temperature of 40 ℃ for 6 hours, taking out and washing with deionized water for 4 times to prepare a cobalt-nickel bimetal layered hydroxide nanosheet array electrode CoNi LDH-nanoarray;
wherein the disposable throwable electrode is a conductive carbon cloth; said Ni (NO)3)2And Co (NO)3)2In a mixture of nickel and cobalt in a molar ratio of 1: 1; the concentration of dopamine is 5mg/mL, the concentration of ammonium persulfate is 8mg/mL, and the concentration of cobalt nitrateThe concentration is 0.5mg/mL, the concentration of phosphate buffer solution PBS is 0.1mol/L, and the pH value is 8.5.
EXAMPLE 4 preparation of Nitrofuran antibiotic sensor
(1) Respectively weighing 0.25 mmol of template molecules and 3mmol of 2-methacrylic acid MAA in an ampoule bottle, adding 8 mL of acetonitrile, and performing ultrasonic treatment for 30min until all the template molecules and the 3mmol of 2-methacrylic acid MAA are dissolved;
(2) adding 15 mmol of ethylene glycol dimethacrylate EDMA into the solution obtained in the step (1), and carrying out ultrasonic treatment for 30min until the mixture is uniformly mixed to obtain a precursor mixed solution;
(3) the CoNi LDH-nanoarray prepared in example 1 was clamped to a rotary stirrer, inserted into the precursor mixed solution in step (2), and subjected to N2Under the temperature of environment and water bath 20 ℃, stirring in a rotating way at the speed of 200 r/s, and simultaneously dripping 1mmol of azobisisobutyronitrile AIBN into the mixed solution at the speed of 1 d/s to initiate polymerization to obtain an in-situ grown MIP containing template molecularly imprinted polymer on CoNi LDH-nanoarray;
(4) immersing the MIP which is obtained in the step (3) and grows in situ on the CoNi LDH-nanoarray and contains the template molecularly imprinted polymer into an eluant, eluting the template molecules for 5 min at room temperature, and then taking out to obtain the NIP which is the template-free molecularly imprinted polymer; continuously washing with deionized water for 2 times, and air drying at room temperature to obtain the nitrofuran antibiotic sensor;
the eluent is a mixed solution of formic acid and methanol, wherein the volume ratio of the formic acid to the methanol is 9: 1.
EXAMPLE 5 preparation of Nitrofuran antibiotic sensor
(1) Respectively weighing 0.35mmol of template molecules and 4 mmol of 2-methacrylic acid MAA in an ampoule bottle, adding 12 mL of acetonitrile, and carrying out ultrasonic treatment for 30min until all the template molecules and the 2-methacrylic acid MAA are dissolved;
(2) adding 18 mmol of ethylene glycol dimethacrylate EDMA into the solution obtained in the step (1), and carrying out ultrasonic treatment for 30min until the mixture is uniformly mixed to obtain a precursor mixed solution;
(3) the CoNi LDH-nanoarray prepared in the technical scheme 2 is clamped to a rotary stirrerOn the device, inserting the precursor mixed solution in the step (2) into a reactor, and adding the precursor mixed solution into the reactor to obtain a solution N2Under the temperature of environment and water bath 30 ℃, stirring in a rotating way at the speed of 60 r/s, and simultaneously dripping 1mmol of azobisisobutyronitrile AIBN into the mixed solution at the speed of 10 d/s to initiate polymerization to obtain an in-situ grown MIP containing template molecularly imprinted polymer on CoNi LDH-nanoarray;
(4) immersing the MIP which is obtained in the step (3) and grows in situ on the CoNi LDH-nanoarray and contains the template molecularly imprinted polymer into an eluant, eluting the template molecules for 10min at room temperature, and then taking out to obtain the NIP which is the template-free molecularly imprinted polymer; continuously washing with deionized water for 3 times, and air drying at room temperature to obtain the nitrofuran antibiotic sensor;
the eluent is a mixed solution of formic acid and methanol, wherein the volume ratio of the formic acid to the methanol is 9: 3.
EXAMPLE 6 preparation of Nitrofuran antibiotic sensor
(1) Respectively weighing 0.45mmol of template molecules and 5mmol of 2-methacrylic acid MAA in an ampoule bottle, adding 15mL of acetonitrile, and carrying out ultrasonic treatment for 30min until all the template molecules and the 5mmol of 2-methacrylic acid MAA are dissolved;
(2) adding 25mmol of ethylene glycol dimethacrylate EDMA into the solution obtained in the step (1), and carrying out ultrasonic treatment for 30min until the mixture is uniformly mixed to obtain a precursor mixed solution;
(3) clamping the CoNi LDH-nanoarray prepared in the technical scheme 2 onto a rotary stirrer, inserting the CoNi LDH-nanoarray into the precursor mixed solution in the step (2), and adding N2Under the temperature of environment and water bath 40 ℃, rotationally stirring at the speed of 5 r/s, simultaneously dripping 1mmol of azobisisobutyronitrile AIBN into the mixed solution at the speed of 20 drops/s to initiate polymerization, and obtaining the in-situ grown MIP containing the template molecularly imprinted polymer on CoNi LDH-nanoarray;
(4) immersing the MIP which is obtained in the step (3) and grows in situ on the CoNi LDH-nanoarray and contains the template molecularly imprinted polymer into an eluant, eluting the template molecules for 20min at room temperature, and then taking out to obtain the NIP which is the template-free molecularly imprinted polymer; continuously washing with deionized water for 4 times, and air drying at room temperature to obtain the nitrofuran antibiotic sensor;
the eluent is a mixed solution of formic acid and methanol, wherein the volume ratio of the formic acid to the methanol is 9: 5.
Example 7 the nitrofuran antibiotic sensor prepared in examples 1 to 6 is applied to detection of nitrofuran antibiotics, and comprises the following steps:
(1) preparing a standard solution: preparing a group of nitrofuran antibiotic standard solutions with different concentrations including blank standard samples;
(2) modification of a working electrode: inserting the nitrofuran antibiotic sensor serving as a working electrode into the nitrofuran antibiotic standard solutions with different concentrations prepared in the step (1), incubating for 10min, taking out, and washing with deionized water for 3 times;
(3) drawing a working curve: taking a saturated calomel electrode as a reference electrode, taking a platinum wire electrode as a counter electrode, forming a three-electrode system with the modified working electrode in the step (2), connecting the three-electrode system with an electrochemical workstation, and sequentially adding 15mL PBS into an electrolytic bath; detecting a current response of the assembled working electrode by Differential Pulse Voltammetry (DPV); the response current intensity of the blank sample is recordedI 0The response current intensity of the standard solution containing different concentrations of nitrofurans antibiotics is recorded asI iThe difference in response to the decrease in current intensity is ΔI = I 0-I i，ΔIMass concentration of the nitrofuran antibiotic standard solutionCWith a linear relationship therebetween, plotting ΔI－CA working curve; the PBS is 10mmol/L phosphate buffer solution, and the pH value of the phosphate buffer solution is 7.4; the parameters during DPV detection are set as follows: the range and the direction are 0-1V, the step is 0.05V, the pulse time is 0.05s, the sampling time is 0.016s, and the pulse period is 0.5 s;
(4) detecting nitrofuran antibiotics in a sample to be detected: replacing the nitrofuran antibiotics standard solution in the step (1) with the sample to be detected, detecting according to the method in the steps (2) and (3), and responding to the reduction of the current intensityDifference deltaIAnd working curve to obtain the content of nitrofuran antibiotics in the sample to be detected.
Example 8 the nitrofuran antibiotic sensors prepared in examples 1 to 6 were applied to the detection of different nitrofuran antibiotics according to the detection procedure of example 7, and the linear range and detection limit are shown in table 1:
TABLE 1 technical index for detection of nitrofurans antibiotics
Example 9 detection of nitrofurans antibiotics in milk
Accurately transferring a milk sample, adding a nitrofuran antibiotic standard solution with a certain mass concentration, taking milk without nitrofuran antibiotics as a blank, performing a labeling recovery experiment, detecting by using the nitrofuran antibiotic sensors prepared in examples 1-6 according to the steps of example 7, determining the recovery rate of the nitrofuran antibiotics in the milk sample, wherein the detection result is shown in table 2:
TABLE 2 detection results of nitrofurans antibiotics in milk
The detection results in table 2 show that the Relative Standard Deviation (RSD) of the results is less than 2.5%, the average recovery rate is 99.0-100.2%, and the method can be used for detecting various nitrofuran antibiotics in milk, and is high in sensitivity, strong in specificity, and accurate and reliable in result.
Example 10 detection of nitrofurans antibiotics in Water samples
Accurately transferring a certain water sample, adding a standard solution of nitrofurans antibiotics with a certain mass concentration, taking the water sample without the nitrofurans antibiotics as a blank, performing a labeling recovery experiment, detecting the nitrofurans antibiotics sensor prepared in the embodiments 1-6 according to the steps of the embodiment 7, determining the recovery rate of the nitrofurans antibiotics in the water sample, and obtaining a detection result shown in table 3:
TABLE 3 detection results of nitrofurans antibiotics in water samples
The detection results in table 3 show that the Relative Standard Deviation (RSD) of the results is less than 2.5%, the average recovery rate is 99.0-100.2%, and the method can be used for detecting multiple nitrofuran antibiotics in a water sample, and is high in sensitivity, strong in specificity, and accurate and reliable in result.
Claims (4)
1. A preparation method of a nitrofuran antibiotic sensor is characterized in that the nitrofuran antibiotic sensor is obtained by in-situ growth of a template-free molecularly imprinted polymer NIP on a cobalt-nickel bimetallic layered hydroxide nanosheet array CoNi LDH-nanoarray; the template-free molecularly imprinted polymer NIP is a molecularly imprinted polymer without template molecules; the molecularly imprinted polymer without the template molecule is obtained by eluting the template molecule from a MIP containing the template molecularly imprinted polymer; the MIP containing the template molecule engram polymer is the MIP containing the template molecule; the template molecule is nitrofuran antibiotics, the MIP containing the molecular engram polymer of the template molecule is directly grown on the CoNi LDH-nanoarray in situ, and the preparation method comprises the following preparation steps:
(1) respectively weighing 0.25-0.45 mmol of template molecules and 3-5 mmol of 2-methacrylic acid MAA in an ampoule bottle, adding 8-15 mL of acetonitrile, and carrying out ultrasonic treatment for 30min until all the template molecules and the 2-methacrylic acid MAA are dissolved;
(2) adding 15-25 mmol of ethylene glycol dimethacrylate EDMA into the solution obtained in the step (1), and carrying out ultrasonic treatment for 30min until the mixture is uniformly mixed to obtain a precursor mixed solution;
(3) clamping CoNi LDH-nanoarray on a rotary stirrer, inserting the CoNi LDH-nanoarray into the precursor mixed solution in the step (2), and adding N2Rotating and stirring at the speed of 5-200 r/s under the temperature of 20-40 ℃ in the environment and water bath, and simultaneouslyDropwise adding 1mmol of azobisisobutyronitrile AIBN into the mixed solution at the speed of 1-20 drops/second to initiate polymerization, and obtaining an in-situ grown MIP containing template molecular imprinting polymer on CoNi LDH-nanoarray;
the preparation method of the template-free molecularly imprinted polymer NIP comprises the following steps: immersing the obtained MIP which grows in situ on the CoNi LDH-nanoarray and contains the template molecularly imprinted polymer in an eluant, eluting the template molecules for 5-20 min at room temperature, and then taking out to obtain the NIP which is the template-free molecularly imprinted polymer; the eluent is a mixed solution of formic acid and methanol, wherein the volume ratio of the formic acid to the methanol is 9 (1-5);
the preparation steps of the nitrofuran antibiotic sensor are as follows: and (3) washing the prepared template-free molecularly imprinted polymer NIP growing in situ on the CoNi LDH-nanoarray for 2-4 times by using deionized water, and airing at room temperature to obtain the nitrofuran antibiotic sensor.
2. The method for preparing a nitrofuran antibiotic sensor as claimed in claim 1, wherein the method for preparing CoNi LDH-nanoarray comprises the following steps:
(1) carrying out ultrasonic cleaning treatment on the disposable throwable electrode by respectively using dilute hydrochloric acid, absolute ethyl alcohol and deionized water so as to remove an oxide layer and surface impurities of the disposable throwable electrode;
(2) weighing 1-3 mmol of Ni (NO)3)2And Co (NO)3)2And 3 to 9mmol of urea CO (NH)2)2Placing the mixture into a 50mL beaker, adding 30mL deionized water, stirring until the mixture is clear, and then transferring the mixture into a 50mL polytetrafluoroethylene reaction kettle;
(3) putting the disposable throwable electrode processed in the step (1) into the solution in the reaction kettle in the step (2), and reacting at the temperature of 100-130 ℃ for 9-12 hours to prepare a precursor electrode of the cobalt-nickel bimetal layered hydroxide nanosheet array;
(4) inserting the precursor electrode of the cobalt-nickel double-metal layered hydroxide nanosheet array obtained in the step (3) into phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate, reacting for 4-6 hours at the temperature of 20-40 ℃, taking out, and performing immersion washing for 2-4 times by using deionized water to prepare cobalt-nickel double-metal layered hydroxide nanosheet array CoNi LDH-nanoarray;
the disposable and disposable electrode is selected from one of the following electrodes: foam nickel, foam copper, pure nickel sheets, pure copper sheets, pure cobalt sheets, pure silicon sheets and conductive carbon cloth; said Ni (NO)3)2And Co (NO)3)2In a mixture of nickel and cobalt in a molar ratio of 1: 1;
in the phosphate buffer solution PBS containing dopamine, ammonium persulfate and cobalt nitrate: the concentration of dopamine is 2-5 mg/mL, the concentration of ammonium persulfate is 3-8 mg/mL, the concentration of cobalt nitrate is 0.1-0.5 mg/mL, the concentration of phosphate buffer solution PBS is 0.1mol/L, and the pH value is 7.2-8.5.
3. The method for producing a nitrofuran-based antibiotic sensor according to any one of claims 1 to 2, wherein the nitrofuran-based antibiotic is one of the following nitrofuran-based antibiotics: furazolidone, nitrofurantoin, furacilin.
4. The nitrofuran antibiotic sensor prepared by the preparation method of any one of claims 1 to 3, which is used for detecting nitrofuran antibiotics, and comprises the following detection steps:
(1) preparing a standard solution: preparing a group of nitrofuran antibiotic standard solutions with different concentrations including blank standard samples;
(2) modification of a working electrode: taking a nitrofuran antibiotic sensor as a working electrode, inserting the nitrofuran antibiotic standard solutions with different concentrations prepared in the step (1), incubating for 10min, taking out, and washing for 3 times by using deionized water;
(3) drawing a working curve: a saturated calomel electrode is used as a reference electrode, a platinum wire electrode is used as a counter electrode, and the three-electrode system is connected with the working electrode modified in the step (2) to perform electrochemical workFirstly, adding 15mL of phosphate buffer solution PBS into an electrolytic bath; detecting a current response of the assembled working electrode by Differential Pulse Voltammetry (DPV); the response current intensity of the blank sample is recorded as I0The response current intensity of the standard solution containing nitrofuran antibiotics with different concentrations is recorded as IiThe difference of response current intensity is Δ I ═ I0-IiThe mass concentration C of the standard solution of nitrofuran antibiotics is in a linear relation with the Delta I, and a Delta I-C working curve is drawn; the concentration of the phosphate buffer solution PBS is 10mmol/L, and the pH value is 7.4; the parameters during DPV detection are set as follows: the range and the direction are 0-1V, the step is 0.05V, the pulse time is 0.05s, the sampling time is 0.016s, and the pulse period is 0.5 s;
(4) detecting nitrofuran antibiotics in a sample to be detected: and (3) replacing the standard solution of the nitrofuran antibiotics in the step (1) with the sample to be detected, detecting according to the methods in the steps (2) and (3), and obtaining the content of the nitrofuran antibiotics in the sample to be detected according to the difference value delta I of the reduction of the response current intensity and the working curve.
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