CN105572107A - Method for detecting acetamiprid in waste water by chemiluminiscence - Google Patents

Abstract

The invention discloses a method for detecting acetamiprid in wastewater by chemiluminescence, which uses a luminol-silver nitrate-nanometer silver chemiluminescence analysis system, utilizes the interaction between acetamiprid aptamers and acetamiprid, aptamers The configuration change induces the change of nano-silver morphology, which leads to the reduction of the catalytic performance of nano-silver. The method of static injection is used to realize the detection of acetamiprid in wastewater by chemiluminescence analysis. The instruments used in the present invention are cheap and have almost no requirements for technicians; the luminol-silver nitrate chemiluminescent analysis system catalyzed by nano-silver is used, and the system is stable, so that the method has good reproducibility, and the RSD is between 2.9% and 4.2%. , high sensitivity, and the detection limit reaches 2.0×10 ^–9 g/L.

Description

A method for detecting acetamiprid in wastewater by chemiluminescence

technical field

The invention belongs to the field of organic pesticide detection, in particular to a method for detecting acetamiprid in waste water by chemiluminescence.

Background technique

For a long time, the pollution caused by pesticide residues to the environment has attracted much attention. Acetamiprid, also known as ethiamiprid and acetamiprid, is a new type of broad-spectrum neonicotinoid insecticide after imidacloprid. , low residue and other characteristics, it is widely used and occupies a high share in the market. Acetamiprid can cause neurological disorders and poisoning in humans. Therefore, the residue and accumulation of acetamiprid in discharged wastewater must threaten people's health.

At present, the detection methods of acetamiprid residue mainly include high performance liquid chromatography, gas chromatography, mass spectrometry, liquid chromatography-mass spectrometry, electrochemistry, colorimetry, fluorescence spectroscopy and enzyme-linked immunoassay. These methods generally have the disadvantages of cumbersome sample pretreatment, high cost, long detection time on the machine, low work efficiency, large amount of organic solvent used or the complexity of the matrix itself affecting the accuracy of the analysis; mass spectrometry is expensive due to the , so it is not well popularized; and the enzyme-linked immunoassay method is easily disturbed and has poor stability. Colorimetry and fluorescence spectroscopy have the problem of low sensitivity, and electrochemical methods cannot solve the problem of heterogeneous operation and are difficult to automate. Therefore, the development of a sensitive, rapid and practical method for the analysis and detection of acetamiprid is of great practical significance for environmental monitoring and people's health.

Chemiluminescence analysis has the advantages of high sensitivity, simple instrument, rapid analysis, and easy automation. However, there is no report on the detection of acetamiprid using chemiluminescence analysis. Nano-silver can catalyze the chemiluminescence reaction between luminol and silver nitrate, and the luminol-silver nitrate-nano-silver chemiluminescence analysis system is a stable and sensitive analysis system. The present invention utilizes the luminol-silver nitrate-nanometer silver chemiluminescence analysis system, utilizes the interaction between the acetamiprid aptamer and acetamiprid, and the change of the configuration of the aptamer induces the change of the shape of the nano-silver, thereby leading to the change of the nano-silver reduction in catalytic performance. The method of static injection is used to realize the detection of acetamiprid in wastewater by chemiluminescence analysis. The high specificity of aptamers for target recognition makes this method highly selective.

Contents of the invention

In order to solve the problems existing in the existing technology for measuring acetamiprid, the invention provides a method for detecting acetamiprid in wastewater by chemiluminescence.

The present invention is achieved through the following technical solutions:

A method for chemiluminescent detection of acetamiprid in wastewater, comprising the following steps:

Interact the acetamiprid aptamer with the wastewater containing acetamiprid, add the nano-silver solution, and then move the effected nano-silver-aptamer-acetamiprid solution into the chemiluminescence pool, inject luminol-nitric acid The silver chemiluminescence reagent is used for luminescence reaction and detection of chemiluminescence signal at the same time. After the interaction between acetamiprid and its aptamer, the aptamer changes from single-stranded DNA to a specific conformation, and the conformational change of acetamiprid aptamer induces a change in the shape of nano-silver, which leads to the effect of nano-silver on luminol-silver nitrate. The reduction of the catalytic activity of the chemiluminescence reaction; the concentration of acetamiprid in the wastewater can be obtained by quantitatively measuring the intensity of the luminescence reaction.

As a further improvement, the nano-silver particle size in the nano-silver solution is 15nm, and the concentration is 3.2×10 ^-10 mol/L.

As a further improvement, the concentration of the aptamer is 6×10 ^-6 mol/L, and the interaction time between the aptamer and acetamiprid is 10 minutes.

As a further improvement, the nano-silver solution is prepared by the following method: 25 mL of silver nitrate solution with a concentration of 1×10 ^-3 mol/L is added dropwise to 75 mL of silver nitrate solution with a concentration of 2×10 ^-3 mol/L In the sodium borohydride solution, react for 10 minutes, and add 5 mL of 1% sodium citrate solution as a stabilizer.

As a further improvement, the luminol-silver nitrate chemiluminescent reagent is mixed with 5×10 ^-5 mol/L luminol solution and 1.0×10 ^-4 mol/L silver nitrate solution at a volume ratio of 1:1 get.

As a further improvement, the pH value of the luminol-silver nitrate chemiluminescence reagent is 12.0.

Compared with the prior art, the present invention has the following advantages:

(1) The chemiluminescence analysis technology is adopted, the instruments used are cheap, and there is almost no requirement for technicians; (2) The luminol-silver nitrate chemiluminescence analysis system catalyzed by nano-silver is used, and the system is stable, which makes the method reproducible. , the RSD is between 2.9% and 4.2%, the sensitivity is high, and the detection limit reaches 2.0×10 ^–9 g/L; (3) The high specificity of the nucleic acid aptamer for target recognition makes the method selective; (4) The whole experiment operation is mild, and it is all carried out in a liquid phase system, which is easy to operate and is conducive to automatic analysis operation; (5) The experiment operation is simple and fast, and the analysis and test time is shortened to less than 30 minutes. The method established by the present invention undoubtedly fills the gap in the research field of chemiluminescence analysis and detection of acetamiprid.

Description of drawings

Fig. 1 is the response graph of luminol-silver nitrate-nanometer silver chemiluminescent signal, wherein a is the luminescence signal of aptamer, b is the luminescence signal of acetamiprid+aptamer;

Fig. 2 is the sensitivity analysis of detection of acetamiprid by chemiluminescence analysis.

Specific implementation cases

The main equipment and reagents used:

The acetamiprid aptamer is a nucleic acid capable of selectively binding to acetamiprid, which was purchased from Shanghai Sangon Biotechnology Co., Ltd., and acetamiprid was purchased from Aladdin Chemical Reagent Co., Ltd. The experimental water was secondary deionized water. Silver nitrate was purchased from Shanghai Chemical Reagent Company. The sewage samples used were taken from the sewage treatment plant in Fuzhou University City. After filtering the waste water samples with medium-speed filter paper, they were diluted with secondary deionized water and fixed in a 100mL volumetric flask. IFFL-D flow injection chemiluminescence analyzer and its supporting equipment (Xi'an Ruimai Analytical Instrument Co., Ltd.), TU-1901 ultraviolet-visible spectrophotometer (Beijing General Instrument Co., Ltd.), transmission electron microscope (Hitachi, Specification model: H-600).

The acetamiprid aptamer was prepared by dissolving in Tris-HCl buffer solution (pH 7.5) and stored in the refrigerator. Before use, it should be heated at a high temperature of 80°C for 5 minutes and then cooled to room temperature for use. Use secondary water to dissolve acetamiprid to prepare a standard solution, and then use the standard solution to dilute acetamiprid standard series solutions.

Preparation of nano-silver: sodium borohydride was used to reduce silver ions, and trisodium citrate was used as a stabilizer. The glassware used in the preparation process was soaked and cleaned with aqua regia (1:3HNO ₃ -HCl), then thoroughly rinsed with secondary deionized water, and dried for later use. Add 25mL of 1×10 ^-3 mol/L silver nitrate solution dropwise into 75mL of 2×10 ^-3 mol/L sodium borohydride solution stirred at high speed, react for 10 minutes, add 5mL of 1% Sodium citrate solution was used as a stabilizer, and the stirring was continued for 10 minutes, and the stirring was stopped. Put the obtained nano-silver sol in a brown reagent bottle, store it in a refrigerator at 4°C, and ripen it for two days before use.

Dissolve 4.43g of luminol solid powder in 20mL of 0.10Mmol/L solution and dilute to 1L to prepare a 2.5×10 ^-2 mol/L luminol stock solution. Store in the dark for one week before use to ensure the stability of the reagent. Working solutions of luminol were obtained by dilution of stock solutions. Silver nitrate was dissolved in secondary water to prepare a stock solution, and the working solution was obtained by diluting the stock solution. The luminol-silver nitrate reagent is obtained by mixing 5×10 ^-5 mol/L luminol solution and 1.0×10 ^-4 mol/L silver nitrate solution at a volume ratio of 1:1, and its pH value is 12.0.

Determination: Mix 200 μL acetamiprid standard series solution and the sample to be tested with 200 μL acetamiprid aptamer solution respectively, the concentration of acetamiprid aptamer solution is 6×10 ^-6 mol/L, act for 10 minutes; add nano After the silver solution, react for 5 minutes; then add 50 μL of NaCl solution with a concentration of 0.5 mol/L, and mix to obtain a mixed solution; pipette 100 μL of the mixed solution into the chemiluminescent cell, and inject 200 μL of luminol-silver nitrate reagent, The luminescence signal was detected and recorded by the IFFL-D chemiluminescence analyzer, and a standard curve was prepared according to the luminescence intensity and the concentration of the acetamiprid solution, and then the concentration of acetamiprid in the sample to be tested was calculated according to the standard curve. Representative chemiluminescent signal responses are shown in Figure 1.

The concentrations of acetamiprid standard series solutions used in the experiment were: 1.5×10 ^-7 , 6.5×10 ^-7 , 1.5×10 ^-6 , 6.5×10 ^-6 , 1.5×10 ^-5 , 5.8×10 ^-5 g/L. The experimental results showed that under the optimized experimental conditions, the chemiluminescence signal gradually decreased with the increase of the concentration of acetamiprid. A linear correlation was found between the chemiluminescent intensity and the logarithm of the acetamiprid concentration. According to the measured chemiluminescence intensity, draw a standard curve. As shown in Figure 2, the detection limit is 2.0×10 ^–9 g/L (S/N=3). And 6.5×10 ^-6 g/L acetamiprid was measured in parallel for 6 times, and the relative standard deviation was 2.6%. The experimental results show that the method has low detection limit, good precision and easy operation.

At the same time, the sewage samples were measured, and the results are listed in Table 1. Experimental results show that the method established by the present invention has better accuracy and precision, the recoveries are respectively 94.5%-106.13%, the RSD is between 2.9%-4.2%, the analysis time is short, no need to use large instruments, It can be used for rapid detection of acetamiprid in wastewater.

The result of determination of acetamiprid in the waste water of table 1

Selectivity analysis of this method:

Taking chlorpyrifos, parathion and pentachlorophenol as representatives, the selectivity of the method for the determination of acetamiprid was investigated. Aptamer/acetamiprid, aptamer (no acetamiprid), aptamer/chlorpyrifos, aptamer/parathion, aptamer/pentachlorophenol and arbitrary DNA sequence/ Acetamiprid response to chemiluminescence signal. In the experiment, the aptamer and any DNA sequence were used as the recognition DNA sequence, and the concentration was 6×10 ^-6 mol/L. The concentration of other control pesticides was the same as that of acetamiprid, both were 1.5×10 ^-5 g/L, and the rest of the experimental conditions were the same. The experimental results found that the aptamer-acetamiprid showed a significantly quenched chemiluminescent signal compared to several other chemiluminescent responses. The results show that this method has good selectivity.

Claims (6)

1. the method for Acetamiprid in chemiluminescence detection waste water, is characterized in that, comprise the following steps:

Acetamiprid aptamers and the waste water containing Acetamiprid are interacted, adds Nano silver solution; Moved in chemiluminescence pond by Nano Silver-aptamers after effect-Acetamiprid solution, the luminol that reinjects-silver nitrate chemical illuminating reagent, carries out luminescence-producing reaction, carries out quantitative measurement, namely obtain the concentration of Acetamiprid in waste water to the intensity of luminescence-producing reaction.

2. in the chemiluminescence detection waste water according to claims 1, the method for Acetamiprid, is characterized in that, the Nano Silver particle diameter in described Nano silver solution is 15nm, and concentration is 3.2 × 10 ^-10mol/L.

3. in the chemiluminescence detection waste water according to claims 1, the method for Acetamiprid, is characterized in that, the concentration of Acetamiprid aptamers is 6 × 10 ^-6mol/L, the action time of Acetamiprid aptamers and Acetamiprid is 10 minutes.

4. in the chemiluminescence detection waste water according to claims 1, the method for Acetamiprid, is characterized in that, described Nano silver solution is adopted and obtained with the following method: be 1 × 10 by 25mL concentration ^-3the 75mL concentration that the liquor argenti nitratis ophthalmicus of mol/L dropwise joins high-speed stirred is 2 × 10 ^-3in the sodium borohydride solution of mol/L, react 10 minutes, then to add 5mL massfraction be that 1% sodium citrate solution is as stabilizing agent.

5. in the chemiluminescence detection waste water according to claims 1, the method for Acetamiprid, is characterized in that, described Lu meter Nuo – silver nitrate chemical illuminating reagent is by 5 × 10 ^-5the luminol solution of mol/L and 1.0 × 10 ^-4the liquor argenti nitratis ophthalmicus of mol/L is mixed to get with 1:1 volume ratio.

6. in the chemiluminescence detection waste water according to claims 1, the method for Acetamiprid, is characterized in that, the pH value of described luminol-silver nitrate chemical illuminating reagent is 12.0.