CN103926347A - Quantitative detection method for organophosphorus pesticide in soil - Google Patents

Abstract

A quantitative detection method for organophosphorus pesticides in soil, characterized in that: the soil sample is ultrasonically extracted with an acetonitrile solution containing 1% acetic acid by volume fraction, then purified with an appropriate amount of PSA and C18 mixed filler, and the supernatant liquid nitrogen is blown and concentrated To nearly dry, dissolve with acetone to constant volume, separate with DB-1701 quartz capillary column by temperature programming, detect with GC/FPD, and finally quantify with matrix external standard method. It has been confirmed by experiments that the method has a wide linear range, and technical indicators such as detection sensitivity, accuracy and precision all meet the requirements of residue analysis, and the pretreatment operation is simple, convenient and fast, and provides a basis for the analysis and research of the determination of organophosphorus pesticide residues in soil. reliable means.

Description

Quantitative detection method of organophosphorus pesticides in a kind of soil

technical field

The present invention relates to the field of detection methods for pesticide residues, in particular to a quantitative detection method for organophosphorus pesticides in soil. Phosphorus pesticides are detected in order to achieve the purpose of fast, simple and reliable.

Background technique

Organophosphorus pesticides (Organophosphorus pesticides) are a class of chemical pesticides widely used in various regions of my country at present. They have the characteristics of broad spectrum, high efficiency, economy, many varieties, and low dosage. . However, studies have confirmed that such pesticides can have toxic effects on the nervous system of the human body. Some varieties have carcinogenic, teratogenic, mutagenic effects and certain accumulation properties. Excessive use may accumulate in the human body, resulting in chronic accumulation poisoning. In the process of crop pest control, the proportion of pesticides directly scattered on the target organisms is very small, and most of them enter the soil through various channels. Organophosphorus pesticides have strong fat solubility and will be adsorbed in the soil for a long time, causing soil pesticide pollution. It will cause harm to soil organisms, affect soil fertility, and pollute water bodies through runoff, leaching, etc. to affect aquatic organisms, and through the bioaccumulation and amplification effects of the food chain, it will eventually endanger human health. Therefore, the detection and monitoring of such pesticide residues in the soil is helpful to evaluate the residue status in the soil, and is of great significance to take effective measures to remove and reduce pesticide pollution.

Due to the complex composition of the soil itself and many interfering substances, it is easy to cause difficulties in the extraction and purification of pesticides. Therefore, choosing an appropriate sample pretreatment method is the first key step in the detection of soil pesticide residues. In the extraction process of soil pesticide residues, existing reports mainly use traditional Soxhlet extraction, liquid-liquid extraction, microwave extraction, accelerated solvent extraction and other methods. Although Soxhlet extraction has a good extraction effect, it takes a long time (usually 10-24h) and consumes a lot of organic solvents (usually 50-400mL), which is not suitable for the pretreatment of batch samples, and it takes a long time to extract The extraction process is likely to cause the decomposition of some pesticides, resulting in a low recovery rate; although the liquid-liquid extraction equipment is simple and the technology is mature, but the solvent consumption is large, cumbersome and time-consuming, and it is easy to produce a large amount of emulsion during the extraction process, which affects the extraction efficiency. , leading to a decline in recovery; microwave extraction and accelerated solvent extraction use less solvent and have high extraction efficiency, but require expensive extraction equipment and high operating costs, and some extraction processes need to be carried out at high temperatures, which is easy to affect heat-sensitive pesticides , and the follow-up usually requires more tedious purification steps. In the purification process of soil pesticide residues, the current purification methods mostly use solid phase extraction small column method, column chromatography and concentrated sulfuric acid sulfonation method. Solid-phase extraction cartridges use less solvent, are simple to operate, have a high recovery rate, and are easy to operate automatically, but the cost is relatively high; column chromatography is more complete in purification and is suitable for the analysis of pesticide residues in most samples, but the solvent consumption is large. The procedures are cumbersome and require high column packing technology; the purification effect of concentrated sulfuric acid sulfonation is good, but the applicable pesticide varieties are very limited, and there are problems such as the decomposition of acid-unstable pesticides, and the heat released by the dissolution of concentrated sulfuric acid can accelerate organic solvents. Volatile, resulting in quantitative errors.

Contents of the invention

The present invention provides a quantitative detection method for organophosphorus pesticides in soil, the purpose of which is to solve the problem of long time-consuming, cumbersome operation, high consumption of organic solvents and insufficient stability and reliability of the existing detection methods for the determination of organophosphorus pesticides in soil. question.

In order to achieve the above object, the technical scheme adopted in the present invention is: a quantitative detection method of organophosphorus pesticides in soil, and the quantitative detection method has two parts:

The first part, establishes the standard curve of the measured organophosphorus pesticide of known gradient concentration with gas chromatography; The establishment of described standard curve is made up of the following steps:

The first step is to prepare a blank matrix extract, and the preparation method consists of the following steps:

(1) Weigh 5.0g of the blank soil sample that is determined not to contain the tested organophosphorus pesticide in a 50mL centrifuge tube, then add 2mL of ultrapure water to the centrifuge tube, vortex and mix, infiltrate for 10-20min, and then add 10mL of acetonitrile solution containing 1% acetic acid by volume, mix well, extract by ultrasonic for 10-15min, add 2g of anhydrous sodium acetate and 2g of anhydrous magnesium sulfate, vortex for 2min, centrifuge at 9000r/min for 4min, take the supernatant It is the blank matrix extract to be purified;

(2) Take another 10mL centrifuge tube, add 0.15g C18, 0.15g N-propylethylenediamine and 0.3g anhydrous magnesium sulfate, add 4mL of the blank matrix extract to be purified, and vortex for 1.5min～ 2.5min, centrifuge at 9000r/min for 4.5min-5.5min; after centrifugation, pipette 2.0mL supernatant into a graduated test tube, then slowly blow the supernatant to 50-70μL with nitrogen in a water bath at 50-60℃ Add acetone to the supernatant to be constant to 1.0mL, and after passing through the organic filter membrane, obtain a blank matrix extract for preparing organophosphorus pesticides;

In the second step, using the blank matrix extract obtained in the first step as a solvent to prepare the matrix mixed standard working solution of organophosphorus pesticides, wherein the organophosphorus pesticides refer to dichlorvos, dimethoate, chlorpyrifos, methyl parasulfur Phosphorus, malathion, fenitrothion, triazophos; the concentration range of each organophosphorus pesticide in the matrix mixed standard working solution of the organophosphorus pesticide is:

The 3rd step, measure the chromatographic peak retention time and the chromatographic peak area of each pesticide component in the matrix mixed standard working solution of described organophosphorus pesticide with gas chromatography, qualitative with chromatographic peak retention time, then take mass concentration as abscissa, Take the chromatographic peak area as the ordinate to draw the standard curve of the organophosphorus pesticide;

Wherein, the chromatographic conditions for determining organophosphorus pesticides are:

Chromatographic column: DB-1701 capillary column, specification: 30m×0.32mm×0.25μm; injection volume: 1.0μL, splitless injection; inlet temperature: 220°C, septum purge 3mL/min ;Column temperature: initial temperature 90°C, keep for 1min, rise to 200°C at 20°C/min, keep for 9min, rise to 245°C at 30°C/min, keep for 8min; carrier gas: high-purity nitrogen, purity ≥99.999%; constant flow mode, flow rate 3mL/min; detector: flame photometric detector; detector temperature: 245°C; hydrogen: flow rate is 75mL/min; air: flow rate is 100mL/min; min;

The second part is to measure the 7 kinds of organophosphorus pesticide residues in the first part in the soil sample, and the quantitative detection method comprises the following steps:

The first step is to extract the soil sample;

Weigh the pulverized soil sample, put it in a 50mL centrifuge tube, add 2mL ultrapure water to the centrifuge tube, vortex and mix, soak for 10-20min, then add the acetonitrile solution containing 1% acetic acid by volume, where , the input ratio of the soil sample and the acetonitrile solution is to drop 1mL of the acetonitrile solution into every 0.5g soil sample, mix well, and extract by ultrasonic for 10-15min; then add anhydrous sodium acetate and anhydrous magnesium sulfate, wherein , the mass ratio of the soil sample, anhydrous sodium acetate and anhydrous magnesium sulfate is 5:2:2, vortexed for 1.5min to 2.5min, centrifuged at a speed of 9000r/min for 3.5min to 4.5min, and taken The clear liquid is the soil sample extract to be purified;

The second step is to purify the extract of the soil sample to be purified;

Take another 10mL centrifuge tube, add C18, N-propylethylenediamine and anhydrous magnesium sulfate, and add the extract of the soil sample to be purified, wherein the C18, N-propylethylenediamine and no The content of magnesium sulfate water in the soil sample extract to be purified is 37.5mg/mL, 37.5mg/mL and 75mg/mL respectively, vortex for 1.5min to 2.5min, and centrifuge at 9000r/min for 4.5min to 5.5min ; After centrifugation, transfer 2.0 mL of the supernatant to a graduated test tube, and slowly blow the supernatant to 50-70 μL with nitrogen in a water bath at 50-60 ° C; then add acetone to the supernatant to dilute to 1.0 mL, After passing through an organic filter membrane, the soil sample extraction and purification solution for determining organophosphorus pesticide residues is obtained;

The third step is to measure the organophosphorus pesticide residues in the soil sample extraction and purification solution by gas chromatography, record the chromatographic peak retention time and chromatographic peak area, and after the chromatographic peak retention time is qualitative, the soil sample is extracted and purified for detection The chromatographic peak area that goes out compares with the standard curve of the organophosphorus pesticide that described first part obtains, obtains the measured value of every kind of organophosphorus pesticide contained in the described soil sample extraction purification liquid; Then described measured value is brought into In the quantitative calculation formula, the organophosphorus pesticide residue to be measured in the soil sample is finally obtained;

Quantitative calculation formula: ω=(ρ×v×f)/m, where: ω is the organophosphorus pesticide residue to be tested in the soil sample, the unit is mg/kg; ρ is the measured value, the unit is mg/L; m is the amount of sample to be weighed, in g; v is the constant volume, in mL; f is the dilution factor;

Wherein, the chromatographic conditions for determining the organophosphorus pesticides in the extraction and purification solution of the soil sample are the same as the chromatographic conditions for the organophosphorus pesticides in the third step of the first part.

The relevant content in the above-mentioned technical scheme is explained as follows:

1. Among the above schemes, the preferred scheme is that the conditions for ultrasonic extraction in the first step of the first part and the first step of the second part are that the ultrasonic power is between 80W and 100W, and the ultrasonic temperature is between 20 and 30 ℃.

2. In the above scheme, the mass concentrations of the standard stock solutions of the seven pesticides of dichlorvos, dimethoate, chlorpyrifos, methyl parathion, malathion, fenitrothion, and triazophos are all 1000 mg/L, Bought from the Environmental Protection Scientific Research and Monitoring Institute of the Ministry of Agriculture;

Acetone is HPLC grade, and the manufacturer is Shanghai Sinopharm Chemical Reagent Co., Ltd.; N-propyl ethylenediamine (abbreviated as PSA in English), the particle size is 40-60 μm, and the manufacturer is AgelaTechnologies Company of the United States; C18, pore size 6nm, particle size 40-60μm, the manufacturer is SepaxTechnologies of the United States; acetonitrile, acetic acid, anhydrous sodium acetate, and anhydrous magnesium sulfate all need to be burned at 620°C for 4 hours, all of which are analytically pure, and the manufacturer is Shanghai Sinopharm Chemical Reagent Co., Ltd.; The experimental water was ultrapure water (18.4MΩ).

3. In the above scheme, before preparing the matrix mixed standard working solution of organophosphorus pesticides, first prepare the mixed standard stock solution of organophosphorus pesticides. After the mixed standard stock solution of pesticides is prepared, store it in a refrigerator at 4°C. When using it, use a blank matrix The extract is prepared as a solvent to prepare a pesticide matrix mixed standard working solution of appropriate concentration;

Among them, the preparation method of the mixed standard stock solution of organophosphorus pesticides is as follows: the concentrations of the standard stock solutions of 7 kinds of organophosphorus pesticides are all 1000mg/L, take 1mL respectively and place them in a 5mL volumetric flask, dilute to volume with acetone, and prepare a concentration of 200mg /L of single standard solution, sealed and stored at -20°C away from light. According to the response value of each pesticide on the FPD detector, take 0.5mL of dichlorvos as a single standard solution, 2mL each of dimethoate and triazophos, and 1mL of each of the other organic phosphorus single standard solutions, put them in a 10mL volumetric flask, and fix them with acetone. Allow. In the mixed standard stock solution of 7 kinds of organophosphorus pesticides: the concentration of dichlorvos is 10 mg/L, the concentration of chlorpyrifos, methyl parathion, fenitrothion, and malathion are respectively 20 mg/L, and the concentrations of dimethoate and triazophos are respectively 40mg/L.

4. In the above scheme, seven kinds of organophosphorus pesticides were determined by gas chromatography, qualitative by retention time, and quantitative by external standard method of peak area.

Working principle of the present invention and beneficial effect are: in 2003, Anastassiades M et al. of U.S. Department of Agriculture's Agricultural Research Service Center proposed a novel pesticide called "QuEChERS" (Quick, Easy, Cheap, Effective, Rugged and Safe) Multi-residue sample preparation method. This method is flexible and has a wide range of applications. According to the characteristics of samples and target analytes, suitable extraction solvents and purification fillers can be selected. It has the advantages of fast, simple, efficient, and safe, and has been widely used in the field of food safety. There are few reports on the application in soil.

In view of the lack of cumbersome pretreatment operations of organophosphorus pesticides, and the fact that the soil matrix is complex, there are many interferences and other factors, and there are few detection standards, the present invention adopts the improved "QuEChERS" method to extract and purify such pesticides. Combining and using capillary gas chromatography to measure the content on the detector FPD (i.e. gas chromatography flame photometric detector), a multi-residue quantitative detection method of organophosphorus pesticides in soil is established. The method is quick and easy, time-saving and reliable, and improves It improves the analysis accuracy, reduces the test cost, provides a scientific monitoring method and basis for soil quality control, and has a certain promotion and use value.

The soil sample was ultrasonically extracted with an acetonitrile solution containing 1% acetic acid by volume, and then purified with an appropriate amount of PSA and C18 mixed filler. The supernatant was concentrated and nearly dry by blowing liquid nitrogen, dissolved in acetone to constant volume, and the temperature of DB-1701 quartz capillary column was programmed. Separation, GC/FPD detection, and final quantification by matrix external standard method. The matrix-matching standard solution is used for calibration and quantification, which reduces the matrix effect of the pesticide to be tested, and the determination result is more accurate. It is confirmed by experiments that the method has a wide linear range, and the technical indicators such as detection sensitivity, accuracy and precision all meet the requirements of residue analysis, which provides a reliable means for the analysis and research of organophosphorus pesticide residues in soil, especially in tea garden soil.

Compared with the existing soil pesticide residue detection method and the national standard method, the pretreatment operation of the present invention is simple, the extraction and purification can be completed in only a few steps, and the ultrasonic method is used for extraction, the result is stable, the repeatability is good, and it is convenient for subsequent batch samples The treatment is more practicable; the purification process avoids the use of purification columns and a large amount of solvent elution, the loss of pesticides is small, the recovery rate is high, and the time and organic solvents are greatly reduced at the same time, which saves testing costs and has little environmental pollution. Also safer. The gas chromatograph (equipped with FPD) commonly used in laboratories is used for detection and analysis. The price is moderate, the operation and maintenance are simple, and the technical requirements are not as strict as those of chromatography-mass spectrometry. The quantitative method adopts the matrix matching standard solution to calibrate the quantification, which reduces the matrix effect of the pesticide to be tested, and the determination result is more accurate. The quantitative detection method of the present invention is convenient and fast, and meets the requirements of pesticide multi-residue analysis in terms of accuracy, precision, sensitivity and other indicators, provides a scientific monitoring method and basis for soil quality control, and has certain popularization and use value.

Description of drawings

Accompanying drawing 1 is the matrix mixed standard working solution chromatogram of 7 kinds of organophosphorus pesticides in the embodiment of the present invention;

Accompanying drawing 2 is the matrix effect figure of 7 kinds of pesticides in soil in the embodiment of the present invention;

Accompanying drawing 3 is the standard chromatogram of adding 7 kinds of organophosphorus pesticides in the soil used for recovery and precision determination in the embodiment of the present invention.

Detailed ways

The present invention will be further described below in conjunction with accompanying drawing and embodiment:

Example: A Quantitative Detection Method of Organophosphorus Pesticides in Soil

The soil for testing is yellow loam, collected from tea gardens in Dongshan District, Suzhou, and the sampling depth is 0-15cm. Remove grass roots, stones and other sundries from the soil sample, mash it manually, mix thoroughly in a pot or other suitable container, and take a certain amount by quartering method for later use. The prepared soil sample should be analyzed as soon as possible, otherwise put it in -20 Stored in a refrigerator at ℃, the basic physical and chemical properties of the soil are: pH 4.1-4.7, clay content 14%-17%, organic matter content 23.41-59.83g/kg. The quantitative detection method consists of two parts:

The first part, establishes the standard curve of the measured organophosphorus pesticide of known gradient concentration with gas chromatography; The establishment of described standard curve is made up of the following steps:

The first step is to prepare a blank matrix extract, and the preparation method consists of the following steps:

(1) Weigh 5.0g of blank soil sample that is determined not to contain the tested organophosphorus pesticide in a 50mL centrifuge tube, then add 2mL of ultrapure water to the centrifuge tube, vortex and mix, infiltrate for 15min, then add 10mL containing Acetonitrile solution with a volume fraction of 1% acetic acid, mix well, extract by ultrasonic for 10-15min, ultrasonic power is between 80W-100W, ultrasonic temperature is 20-30℃, add 2g of anhydrous sodium acetate and 2g of anhydrous magnesium sulfate, vortex Spin for 2 minutes, centrifuge at 9000r/min for 4 minutes, and take the supernatant as the blank matrix extract to be purified;

(2) Take another 10mL centrifuge tube, add 0.15g C18, 0.15g N-propylethylenediamine and 0.3g anhydrous magnesium sulfate, add 4mL of the blank matrix extract to be purified, and vortex for 1.5min～ Centrifuge at 9000r/min for 2.5min for 5min; transfer 1.5mL of the supernatant to a graduated test tube after centrifugation, then slowly blow the supernatant to 50-70μL with nitrogen in a water bath at 50-60°C; Acetone was added to the clear liquid to make it constant to 0.75 mL, and after passing through a 0.22 μm organic filter membrane, a blank matrix extract for preparing organophosphorus pesticides was obtained;

In the second step, using the blank matrix extract obtained in the first step as a solvent to prepare the matrix mixed standard working solution of organophosphorus pesticides, wherein the organophosphorus pesticides refer to dichlorvos, dimethoate, chlorpyrifos, methyl parasulfur Phosphorus, malathion, fenitrothion, triazophos; the concentration range of each organophosphorus pesticide in the matrix mixed standard working solution of the organophosphorus pesticide is:

The 3rd step, measure the chromatographic peak retention time and the chromatographic peak area of each pesticide component in the matrix mixed standard working solution of described organophosphorus pesticide with gas chromatography, qualitative with chromatographic peak retention time, then take mass concentration as abscissa, Take the chromatographic peak area as the ordinate to draw the standard curve of the organophosphorus pesticide;

Wherein, the chromatographic conditions for determining organophosphorus pesticides are:

Chromatographic column: DB-1701 capillary column, specification: 30m×0.32mm×0.25μm; injection volume: 1.0μL, splitless injection; inlet temperature: 220°C, septum purge 3mL/min ;Column temperature: initial temperature 90°C, keep for 1min, rise to 200°C at 20°C/min, keep for 9min, rise to 245°C at 30°C/min, keep for 8min; carrier gas: high-purity nitrogen, purity ≥99.999%; constant flow mode, flow rate 3mL/min; detector: flame photometric detector; detector temperature: 245°C; hydrogen: flow rate is 75mL/min; air: flow rate is 100mL/min; min;

The second part is to measure the 7 kinds of organophosphorus pesticide residues in the first part in the soil sample, and the quantitative detection method comprises the following steps:

The first step is to extract the soil sample;

Weigh 5.0g of crushed soil sample, put it in a 50mL centrifuge tube, add 2mL of ultrapure water to the centrifuge tube, vortex and mix, soak for 15min, then add 10mL of acetonitrile solution containing 1% acetic acid by volume fraction, Mix well, ultrasonically extract for 10-15min, the ultrasonic power is between 80W-100W, and the ultrasonic temperature is 20-30℃; then add 2g of anhydrous sodium acetate and 2g of anhydrous magnesium sulfate, vortex for 2min, at a speed of 9000r/min Centrifuge for 4 minutes, and take the supernatant as the soil sample extract to be purified;

The second step is to purify the extract of the soil sample to be purified;

Take another 10mL centrifuge tube, add 0.15g C18, 0.15g N-propylethylenediamine (PSA for short) and 0.3g anhydrous magnesium sulfate, add the soil sample extract to be purified, vortex for 2min, Centrifuge at a speed of 9000r/min for 5min; after centrifugation, transfer 2.0mL supernatant to a graduated test tube, blow the supernatant slowly with nitrogen to 50-70μL in a water bath at 50-60°C; then add Acetone was fixed to 1.0mL, and after passing through a 0.22μm organic filter membrane, the soil sample extraction and purification solution for the determination of organophosphorus pesticide residues was obtained;

The third step is to measure the organophosphorus pesticide residues in the soil sample extraction and purification solution by gas chromatography, record the chromatographic peak retention time and chromatographic peak area, and after the chromatographic peak retention time is qualitative, the soil sample is extracted and purified for detection The chromatographic peak area that goes out compares with the standard curve of the organophosphorus pesticide that described first part obtains, obtains the measured value of every kind of organophosphorus pesticide contained in the described soil sample extraction purification liquid; Then described measured value is brought into In the quantitative calculation formula, the organophosphorus pesticide residue to be measured in the soil sample is finally obtained;

Quantitative calculation formula: ω=(ρ×v×f)/m, where: ω is the organophosphorus pesticide residue to be tested in the soil sample, the unit is mg/kg; ρ is the measured value, the unit is mg/L; m is the amount of sample to be weighed, in g; v is the constant volume, in mL; f is the dilution factor;

Wherein, the chromatographic conditions for determining the organophosphorus pesticides in the extraction and purification solution of the soil sample are the same as the chromatographic conditions for the organophosphorus pesticides in the third step of the first part.

In the above examples, the instruments and equipment used are: 7890A gas chromatograph equipped with flame photometric detector (FPD), 7693 autosampler and Chemstation chromatographic workstation (Agilent company of the United States); KQ-500DE numerical control ultrasonic instrument (Kunshan City Ultrasonic Instrument Co., Ltd.); TG16-WS desktop high-speed centrifuge (Hunan Xiangyi Laboratory Instrument Development Co., Ltd.); HSC-24B nitrogen blowing instrument (Tianjin Hengao Technology Development Co., Ltd.); K600 pulverizer (Germany Braun company); VM-10 vortex oscillator (Korean Daihan Scientific); SX2-4-10 muffle furnace (Shanghai Yuejin Medical Instrument Co., Ltd.); EXCEED-AD-24 ultrapure water machine (Chengdu Tangshi Corning Technology Development Co., Ltd).

Drugs and reagents used: dichlorvos, dimethoate, chlorpyrifos, methyl parathion, malathion, fenitrothion, triazophos, the mass concentration is 1000mg/L, purchased from the Ministry of Agriculture Environmental Protection Research and Monitoring Institute Acetone, HPLC grade (Shanghai Sinopharm Chemical Reagent Co., Ltd.); N-propyl ethylenediamine (PSA), 40-60 μm (U.S. Agela Technologies); C18, pore size 6nm, particle size 40-60 μm (U.S. Sepax Technologies ); acetonitrile, acetic acid, anhydrous sodium acetate, and anhydrous magnesium sulfate (burned at 620°C for 4h) were all analytically pure (Shanghai Sinopharm Chemical Reagent Co., Ltd.); experimental water was ultrapure water (18.4MΩ).

Preparation of mixed standard stock solution:

The preparation method of the mixed standard stock solution of organophosphorus pesticides is as follows: the concentrations of the standard stock solutions of 7 kinds of organophosphorus pesticides are all 1000mg/L, take 1mL respectively and place them in a 5mL volumetric flask, dilute to volume with acetone, and prepare a concentration of 200mg/L The single standard solution was stored at -20°C in the dark and sealed. According to the response value of each pesticide on the FPD detector, take 0.5mL of dichlorvos as a single standard solution, 2mL each of dimethoate and triazophos, and 1mL of each of the other organic phosphorus single standard solutions, put them in a 10mL volumetric flask, and fix them with acetone. Allow. In the mixed standard stock solution of 7 kinds of organophosphorus pesticides: the concentration of dichlorvos is 10 mg/L, the concentration of chlorpyrifos, methyl parathion, fenitrothion, and malathion are respectively 20 mg/L, and the concentrations of dimethoate and triazophos are respectively 40mg/L.

The above-mentioned pesticide mixed standard stock solutions were stored in a refrigerator at 4°C. When used, the blank matrix extract was used as a solvent to prepare matrix mixed standard working solutions of appropriate concentrations.

The test result of this embodiment:

1. Gas chromatographic separation of test pesticides

The matrix mixed standard working solutions of seven organophosphorus pesticides were measured by the above gas chromatography conditions. The separation effect of the standard samples was good, the peak shape was symmetrical, and the baseline was stable, indicating that the instrument conditions were suitable. Under this chromatographic condition, the retention times of various organic phosphorus pesticides are about 5.941min for dichlorvos, 13.099min for dimethoate, 15.787min for chlorpyrifos, 15.915min for methyl parathion, 16.479min for malathion, and 16.607min for fenitrothion. Triazophos 22.684min (referring to shown in accompanying drawing 1, label 1 in accompanying drawing 1 is dichlorvos, and concentration is 0.25mg/L, and label 2 is dimethoate, and concentration is 1mg/L, and label 3 is chlorpyrifos, and concentration is 0.5mg /L, label 4 is methyl parathion, the concentration is 0.5mg/L, label 5 is malathion, the concentration is 0.5mg/L, label 6 is fenitrothion, the concentration is 0.5mg/L, label 7 is triazophos, the concentration is 1mg/L).

2. Matrix effect

Matrix effect refers to the influence of other matrix components in the sample other than the analyte on the measured value of the analyte. Matrix effects vary for different samples and analytes. In gas chromatographic analysis, most pesticides show different degrees of matrix enhancement effect, that is, the response value of the same concentration of pesticides in the matrix is higher than that in pure solvents. It is generally believed that the presence of matrix components in the sample reduces the chromatographic system. The opportunity for the active site to interact with the analyte molecule increases the detection signal of the analyte. The methods to eliminate the influence of matrix effect include matrix matching standard solution method, standard addition method, multiple purification method, application of analytical protective agent and statistical method correction, etc. In the present invention, the ratio of the response value of the analyte in the matrix mixed standard working solution and the pure solvent mixed standard solution is used to represent the matrix effect, and the closer the ratio is to 1, the smaller the matrix effect is.

The test results showed that the 7 kinds of pesticide compounds analyzed had different degrees of matrix enhancement effects, and the 7 kinds of organophosphorus pesticides had strong matrix effects, as shown in Figure 2. Therefore, when the present invention is quantified by the external standard method, a blank matrix extract is used as a solvent to prepare a standard sample to eliminate matrix interference and reduce errors. (in accompanying drawing 2, label 1 represents matrix mixed standard working solution containing dichlorvos and pure solvent mixed standard solution, and concentration is 0.05mg/L; Label 2 is matrix mixed standard working solution containing dimethoate and pure solvent mixed standard solution , the concentration is 0.2mg/L; similarly, the label 3 indicates that it contains chlorpyrifos, and the concentration is 0.1mg/L; the label 4 indicates that it contains methyl parathion, and the concentration is 0.1mg/L; the label 5 indicates that it contains malathion, The concentration is 0.1mg/L; the number 6 is fenitrothion, the concentration is 0.1mg/L; the number 7 is triazophos, the concentration is 0.2mg/L; A _m represents the response value of each pesticide in the matrix matching mixed standard solution ; A _s represents the response value of each pesticide in the pure solvent mixed standard solution).

3. Linear range, regression equation and detection limit of the method

Preparation mass concentration is 7 kinds of organophosphorus pesticide matrix mixed standard solutions of 0.01～4.0mg/L, measure according to the chromatographic condition of the organophosphorus pesticide in the 3rd step of said first part respectively, with mass concentration (expressed in ρ, The unit is mg/L) as the abscissa, and the peak area (y) as the ordinate to draw the standard curve. The 7 pesticides have good linearity within their concentration range, and the correlation coefficients (r ² ) are all greater than 0.996, as shown in Table 1. The limit of detection (LOD) and limit of quantification (LOQ) of each pesticide were calculated as 3 times and 10 times of the noise signal in the chromatogram of the lowest added level, respectively. After calculation, the LOD of the seven organophosphorus pesticides is between 0.0038-0.022 mg/kg, and the LOQ is between 0.013-0.072 mg/kg, as shown in Table 1 below.

Table 1 The linear range, regression equation, correlation coefficient, detection limit and quantification limit of the tested pesticides

4. The recovery rate and precision of the method

The recovery and precision of the method were determined by the blank sample spiked method. Weigh several parts of 5.0g blank soil samples that are determined not to contain the tested pesticide, add three kinds of pesticide mixed standard working solution with different concentration levels, the addition level of organophosphorus pesticide is 0.025～1.0mg/kg, each concentration level Parallel 3 samples, after vortex mixing, let stand for 1 h to allow the standard solution to be fully absorbed by the sample, and perform sample pretreatment and chromatographic condition determination according to the second part. The matrix external standard method was used for quantification, and the average recovery rate and relative standard deviation (ie RSD) of each pesticide were calculated, as shown in Table 2, and the sample spiked chromatogram is shown in Figure 3.

It can be seen from Table 2 that the average recovery rate of organophosphorus pesticides in soil is 73.9%-96.6%, and the RSD (n=3) is 1.3%-9.4%, which shows that the method has good accuracy and repeatability.

The addition recovery result (n=3) of table 214 kinds for testing pesticide in soil

Wherein, label 1 in accompanying drawing 3 is dichlorvos, and concentration is 0.25mg/L, and label 2 is dimethoate, and concentration is 1mg/L, and label 3 is chlorpyrifos, and concentration is 0.5mg/L, and label 4 is methyl parathion , the concentration is 0.5mg/L, label 5 is malathion, the concentration is 0.5mg/L, label 6 is fenitrothion, the concentration is 0.5mg/L, label 7 is triazophos, the concentration is 1mg/L .

5. Actual sample analysis

Using this method, 16 soil samples from a local tea plantation in Suzhou were tested for 7 organophosphorus pesticide residues, and no organophosphorus pesticides were detected. The results showed that this method can be used for the detection of seven organophosphorus pesticide residues in soil.

The above-mentioned embodiments are only to illustrate the technical concept and characteristics of the present invention, and the purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly, and not to limit the protection scope of the present invention. All equivalent changes or modifications made according to the spirit of the present invention shall fall within the protection scope of the present invention.

Claims (2)

1. a quantitative detection method of organophosphorus pesticides in soil, characterized in that: said quantitative detection method has two parts: The first part, establishes the standard curve of the measured organophosphorus pesticide of known gradient concentration with gas chromatography; The establishment of described standard curve is made up of the following steps: The first step is to prepare a blank matrix extract, and the preparation method consists of the following steps: (1) Weigh 5.0 g of a blank soil sample that has been determined to be free of the tested organophosphorus pesticide in a 50 mL centrifuge tube, then add 2 mL of ultrapure water to the centrifuge tube, vortex and mix, soak for 10-20 minutes, Then add 10 mL of acetonitrile solution containing 1% acetic acid by volume, mix well, extract by ultrasonic for 10-15 min, add 2 g of anhydrous sodium acetate and 2 g of anhydrous magnesium sulfate, vortex for 2 min, and set the temperature at 9000 r/min Centrifuge at a rotational speed for 4 minutes, and take the supernatant as the blank matrix extract to be purified; (2) Take another 10 mL centrifuge tube, add 0.15g C18, 0.15g N-propylethylenediamine and 0.3g anhydrous magnesium sulfate, add 4mL of the blank matrix extract to be purified, and vortex for 1.5min ~2.5min, centrifuge at 9000 r/min for 4.5min~5.5min; after centrifugation, pipette 2.0 mL of supernatant into a graduated test tube, then slowly blow the supernatant to 50 ~70μL; then add acetone to the supernatant to make the volume to 1.0mL, and pass through an organic filter to obtain a blank matrix extract for preparing organophosphorus pesticides; In the second step, using the blank matrix extract obtained in the first step as a solvent to prepare the matrix mixed standard working solution of organophosphorus pesticides, wherein the organophosphorus pesticides refer to dichlorvos, dimethoate, chlorpyrifos, methyl parasulfur Phosphorus, malathion, fenitrothion, triazophos; the concentration range of each organophosphorus pesticide in the matrix mixed standard working solution of the organophosphorus pesticide is: The 3rd step, measure the chromatographic peak retention time and the chromatographic peak area of each pesticide component in the matrix mixed standard working solution of described organophosphorus pesticide with gas chromatography, qualitative with chromatographic peak retention time, then take mass concentration as abscissa, Take the chromatographic peak area as the ordinate to draw the standard curve of the organophosphorus pesticide; Wherein, the chromatographic conditions for determining organophosphorus pesticides are: Chromatographic column: DB-1701 capillary column, specification: 30m×0.32mm×0.25μm; injection volume: 1.0μL, splitless injection; inlet temperature: 220°C, septum purge 3mL/min ;Column temperature: initial temperature 90°C, keep for 1min, rise to 200°C at 20°C/min, keep for 9min, rise to 245°C at 30°C/min, keep for 8min; carrier gas: high-purity nitrogen, purity ≥99.999%; constant flow mode, flow rate 3mL/min; detector: flame photometric detector; detector temperature: 245°C; hydrogen: flow rate is 75 mL/min; air: flow rate is 100 mL/min; makeup gas: high-purity nitrogen, flow rate is 60mL/min; The second part is to measure the 7 kinds of organophosphorus pesticide residues in the first part in the soil sample, and the quantitative detection method comprises the following steps: The first step is to extract the soil sample; Take the pulverized soil sample, place it in a 50 mL centrifuge tube, and then add an acetonitrile solution containing 1% acetic acid by volume fraction, wherein the input ratio of the soil sample to the acetonitrile solution is 0.5 g of the soil sample Put 1mL of the acetonitrile solution into the solution, mix well, and extract by ultrasonic for 10-15 min; then add anhydrous sodium acetate and anhydrous magnesium sulfate, wherein, the mass of the soil sample, anhydrous sodium acetate and anhydrous magnesium sulfate The ratio is 5:2:2, vortex for 1.5min~2.5min, centrifuge at 9000 r/min for 3.5min~4.5min, take the supernatant as the soil sample extract to be purified; The second step is to purify the extract of the soil sample to be purified; Take another 10 mL centrifuge tube, add C18, N-propylethylenediamine and anhydrous magnesium sulfate, and add the extract of the soil sample to be purified, wherein the C18, N-propylethylenediamine and The contents of anhydrous magnesium sulfate in the soil sample extract to be purified were 37.5mg/mL, 37.5mg/mL and 75mg/mL respectively, vortexed for 1.5min~2.5min, and centrifuged at 9000 r/min for 4.5min~ 5.5min; after centrifugation, pipette 1.5 mL of the supernatant into a graduated test tube, and slowly blow the supernatant to 50~70μL with nitrogen in a water bath at 50-60°C; then add acetone to the supernatant to dilute to 0 . 75 mL, after passing through an organic filter membrane, the soil sample extraction and purification solution for determining organophosphorus pesticide residues was obtained; The third step is to measure the organophosphorus pesticide residues in the soil sample extraction and purification solution by gas chromatography, record the retention time of the chromatographic peak and the area of the chromatographic peak, and after qualitative determination by the retention time of the chromatographic peak, the extraction and purification solution of the soil sample is detected The chromatographic peak area that goes out compares with the standard curve of the organophosphorus pesticide that described first part obtains, obtains the measured value of every kind of organophosphorus pesticide contained in the described soil sample extraction purification liquid; Then described measured value is brought into In the quantitative calculation formula, the organophosphorus pesticide residue to be measured in the soil sample is finally obtained; Quantitative calculation formula: ω=(ρ×v×f)/m, where: ω is the organophosphorus pesticide residue to be tested in the soil sample, the unit is mg/kg; ρ is the measured value, the unit is mg/L; m is the sample volume to be weighed, in g; v is the constant volume, in mL; f is the dilution factor; Wherein, the chromatographic conditions for determining the organophosphorus pesticides in the extraction and purification solution of the soil sample are the same as the chromatographic conditions for the organophosphorus pesticides in the third step of the first part. 2. the quantitative detection method of organophosphorus pesticides in a kind of soil according to claim 1, is characterized in that: the condition of ultrasonic extraction in the first step of described first part and the first step of described second part is The ultrasonic power is between 80W and 100W, and the ultrasonic temperature is between 20 and 30°C.