An analytical method for detecting multiple pesticide residues in tea infusion

Abstract

The present invention discloses an analytical method for detecting the residues of various pesticides in tea leaves and tea soups. The method process is as follows: firstly classify the types of pesticides to be detected, respectively draw the standard curves of the pesticide standards to be detected by gas chromatography or liquid chromatography-tandem mass spectrometry (LC-MS / MS), extract the tea leaves or tea soup samples by solvent extraction, obtain the extracted sample liquid, and obtain the pesticide residue results by the above-mentioned standard curve after testing by gas chromatography GC or LC-MS / MS. The method of the present invention has the advantages of less interference from impurities, convenient sample pretreatment, reliable precision and accuracy, etc. The GC-ECD method can be applied to the trace analysis and detection of chlorpyrifos, bifenthrin, cypermethrin and cypermethrin in tea leaves and tea soup samples; the GC-FPD method can be applied to the analysis and detection of chlorpyrifos and malathion in tea leaves and tea soup samples; the LC-MS / MS method has high sensitivity, accurate qualitative analysis, and very low detection limit, and can be widely used in the trace analysis and detection of neonicotinoid pesticides in tea leaves and tea soup samples.

Description

Technical Field

[0001] The invention relates to an analytical method for detecting various pesticide residues in tea leaves and tea soup. Background Art

[0002] In order to prevent and control pests and diseases, chemicals such as organophosphorus, organochlorine and pyrethroid pesticides are widely used in the tea planting process, resulting in excessive pesticide residues in tea. The tea matrix is ​​complex, and the interference of tea polyphenols and pigments is a difficult problem to solve in the analysis of multiple pesticide residues in tea. Tea is a special drink. People generally do not eat tea directly, but drink the tea soup after brewing. Therefore, the residual pesticides and other harmful substances dissolved in the tea soup may affect human health.

[0003] Tea leaves are rich in organic compounds, such as pigments, sugars, waxes, lipids, alkaloids, organic acids and tea polyphenols. Since these substances will be co-extracted during the extraction of pesticide residues, it is difficult to prepare samples. Common pretreatment methods for tea soup samples include solid phase extraction, accelerated solvent extraction, gel permeation chromatography, etc. In 2003, researchers from the United States Department of Agriculture developed a rapid sample pretreatment technology, which was named QuEChERS because of its advantages of being fast, simple, cheap, effective, reliable and safe. In recent years, in reports on the determination of multiple pesticide residues in tea leaves by gas chromatography technology, the pretreatment methods mostly use solid phase extraction and gel permeation chromatography, while there are fewer reports on QuEChERS-GC methods. Gas chromatography is an instrumental detection method for analyzing pesticide residues in tea soup. Specific detectors are targeted at a specific type of pesticide. Electron capture detectors (ECD) and nitrogen-phosphorus detectors (NPD) are cheaper than mass spectrometers (MS) and can achieve the same level of sensitivity.

[0004] With the increasing use of neonicotinoid pesticides, their residues in fruits, vegetables, tea, etc. have also received more and more attention. In order to meet the requirements of safety testing of tea, etc., a good detection method is also needed for the residues of neonicotinoid pesticides such as "thiamethoxam, thiacloprid, clothianidin, chlorpyrifos, nitenpyram, acetamiprid, dinotefuran and imidacloprid" in tea, which can have the advantages of high sensitivity, low limit of quantification and accurate determination of the pesticide residues. Although the existing technology also uses liquid chromatography-tandem mass spectrometry for detection, on the one hand, it is also necessary to provide a good extraction method for neonicotinoid pesticides in tea samples for subsequent detection processes; on the other hand, the extraction pretreatment process of tea samples is relatively complicated, and the conventional use of external standard method for quantification has large errors and certain unreality. Summary of the invention

[0005] In view of the above technical problems existing in the prior art, the purpose of this application is to provide an analytical method for detecting various pesticide residues in tea leaves.

[0006] The technical solution adopted by the present invention is:

[0007] An analytical method for detecting multiple pesticide residues in tea soup, the method comprising the following steps:

[0008] 1) Drawing of standard curve

[0009] S1: Prepare the pesticide standards into standard stock solutions using n-hexane as solvent;

[0010] S2: Take each standard stock solution, use acetonitrile as solvent to prepare a mixed pesticide standard stock solution, dilute the mixed pesticide standard stock solution step by step to prepare gradient standard solutions of different concentrations, and wait for the standard curve to be analyzed by the machine;

[0011] S3: The pesticide is one or more of chlorpyrifos, bifenthrin, cypermethrin, cypermethrin, and malathion, and is prepared into gradient standard solutions of different concentrations according to steps S1-S2, and analyzed by gas chromatography GC to obtain a chromatogram, and a capillary column: DB-1701, 30m*250μm*0.25μm, is used to determine the matrix matching standard curve by external standard method;

[0012] S4: the pesticide is at least one of thiamethoxam, thiacloprid, clothianidin, chlorpyrifos, nitenpyram, acetamiprid, dinotefuran and imidacloprid, and matrix-matched standard solutions of different concentrations are prepared according to steps S1-S2, and internal standards thiamethoxam-d3 (THI-d3), imidacloprid-d4 (IMI-d4) and clothianidin-d3 (CLO-d3) are added respectively, and liquid chromatography-tandem mass spectrometry LC-MS / MS is used for analysis to obtain a chromatogram, and the matrix-matched standard curve is quantitatively determined by the internal standard method;

[0013] 2) Detection and analysis of pesticide residues

[0014] The tea leaves or tea soup samples are extracted by solvent extraction to obtain an extracted sample solution, and then the chromatogram of the pesticides in the extracted sample solution is detected according to the GC method in step S3 or the LC-MS / MS method in step S4, and the concentrations of different types of pesticides in the tea leaves or tea soup samples are calculated according to the corresponding standard curve.

[0015] In the analytical method for detecting multiple pesticide residues in tea leaves and tea soup, the specific process of preparing the matrix matching standard solution in steps S1 and S2 is as follows:

[0016] M1: Accurately weigh the standard samples of different pesticides, prepare them into 0.5-2.0 mg / mL single pesticide standard stock solutions with n-hexane, and store them at -15 to -25°C;

[0017] M2: Pipette individual pesticide standard stock solutions one by one and inject them into the same volumetric flask respectively, dilute to the scale with acetonitrile, prepare 2.0-8.0 mg / L mixed pesticide standard stock solutions, and store at -15--25℃; dilute the mixed pesticide standard stock solutions step by step with acetonitrile to prepare gradient standard solutions of different concentrations.

[0018] In the analytical method for detecting multiple pesticide residues in tea, eight corresponding concentration gradient standard solutions are prepared in step S2, and the concentrations are 5 μg / L, 10 μg / L, 20 μg / L, 30 μg / L, 40 μg / L, 50 μg / L, 100 μg / L, 150 μg / L and 200 μg / L.

[0019] The analytical method for detecting multiple pesticide residues in tea leaves and tea soup, the detector of the gas chromatography GC in step S3 is ECD, which is used for trace analysis and detection of chlorpyrifos, bifenthrin, cypermethrin and cypermethrin in tea leaves or tea soup samples, and the conditions of the GC test are: capillary column: DB-1701, 30m*250μm*0.25μm; carrier gas is high-purity nitrogen, column flow rate is 1.5mL / min; split injection, split ratio is 5:1; injection port temperature is 220℃; heating program: initial temperature is 80℃, maintained for 1min, then increased to 260℃ at a rate of 20℃ / min, maintained for 5min; detector temperature is 280℃.

[0020] The analytical method for detecting multiple pesticide residues in tea leaves and tea soup, the detector of the gas chromatography GC in step S3 is FPD, which is used for the analysis and detection of malathion in tea leaves or tea soup samples, and the conditions of the GC test are: capillary column: DB-1701, 30m*250μm*0.25μm; carrier gas is high-purity nitrogen, column flow rate is 1.5mL / min; split injection, split ratio is 5:1; injection port temperature is 220℃; heating program: initial temperature is 80℃, maintained for 1min, then increased to 260℃ at a rate of 20℃ / min, maintained for 5min; detector temperature is 280℃.

[0021] In the analytical method for detecting the residues of multiple pesticides in tea leaves and tea soup, when the chromatogram of the pesticides in the extracted sample liquid is detected according to the method of step S3, the pesticides are one or more of chlorpyrifos, bifenthrin, cypermethrin, cypermethrin, and malathion, and the method for extracting the tea leaves or tea soup samples is one of the following:

[0022] (1) Tea extraction method: acetonitrile containing a small amount of acetic acid is used as the extraction solution, added to tea leaves, and sodium chloride and sodium acetate are added. After thorough vortexing, ultrasonication, centrifugation, and supernatant addition, GCB, PSA, and anhydrous magnesium sulfate are added. After thorough vortexing, centrifugation, filtration, and injection into GC chromatography detection, the pesticide residue results are obtained through the corresponding standard curve;

[0023] (2) Tea soup extraction method: Tea leaves are brewed with boiling water for 5-30 minutes and then cooled. The extraction solution is added to the tea soup. Acetonitrile containing a small amount of acetic acid is used as the extraction solution, and sodium chloride and sodium acetate are added. After thorough vortexing, ultrasonication and centrifugation, the supernatant is taken and PSA and anhydrous magnesium sulfate are added. After thorough vortexing, centrifugation and filtration, the sample is injected into the GC chromatography for detection. The pesticide residue results are obtained through the corresponding standard curve.

[0024] The analytical method for detecting the residues of multiple pesticides in tea leaves and tea soup is described. The volume fraction of acetic acid in the extraction solution in step (1) is 0.5-2.0%, preferably 1%; the specific process of the tea extraction method in step (1) is: weigh 5.00g±0.5g of tea leaves into a centrifuge tube, add 20mL±2mL of acetonitrile extraction solution containing a small amount of acetic acid, then add 4g±0.5g of sodium chloride and 1g±0.2g of sodium acetate, fully oscillate and vortex, then ultrasonicate, centrifuge, take 2mL of supernatant into a centrifuge tube containing 50mg±10mg GCB, 50mg±10mg PSA and 200mg±50mg anhydrous magnesium sulfate, fully oscillate and vortex, then ultrasonicate, centrifuge, pass through a 0.22μm organic filter membrane, and then inject into GC chromatography for detection, and obtain the pesticide residue result through the corresponding standard curve.

[0025] The analytical method for detecting the residues of multiple pesticides in tea leaves and tea soup, the volume fraction of acetic acid in the extraction solution in step (2) is 0.5-2.0%, preferably 1%; the specific process of the tea soup extraction method in step (2) is: weigh 3.00g±0.5g of tea leaves, add 150mL±20mL of boiling water, brew for 5-30min, weigh 10.00g±2.0g of tea soup in a centrifuge tube after cooling, add 20mL±2mL of acetonitrile extraction solution containing a small amount of acetic acid, then add 4g±0.5g of sodium chloride and 1g±0.2g of sodium acetate, fully oscillate and vortex, then ultrasonicate, centrifuge, add 3-5g of anhydrous magnesium sulfate to the supernatant, fully oscillate and vortex, centrifuge, take 2mL of supernatant and put it in a 50mg±10mg PSA and 200 mg ± 50 mg of anhydrous magnesium sulfate were placed in a centrifuge tube, vortexed thoroughly, ultrasonicated, centrifuged, filtered through a 0.22 μm organic filter membrane, and injected into GC chromatography for detection. The pesticide residue results were obtained through the corresponding standard curve.

[0026] In the analytical method for detecting multiple pesticide residues in tea leaves and tea soup, the conditions for the liquid chromatography-tandem mass spectrometry LC-MS / MS test in step S4 are:

[0027] The chromatographic column is YMC ODS-AQ 100mm*2.1mm*3μm, the mobile phases A and B are 0.1wt% formic acid water and pure acetonitrile respectively, the mobile phase flow rate is 0.3mL / min; the gradient elution conditions are: 0-1.3min, 90%A; 1.3-4min, 70%A; 4-6min, 10%A; 6-6.5min, return to the initial state of 90%A, and keep it unchanged for two minutes at 6.5-8.5min; the mass spectrometry conditions are: electrospray ion source, positive ion mode ESI+.

[0028] In the analytical method for detecting the residues of multiple pesticides in tea leaves and tea soup, when the chromatogram of the pesticide in the extracted sample solution is detected according to the method of step S4, the pesticide is at least one of thiamethoxam, thiamethoxam, clothianidin, chlorpyrifos, nitenpyram, acetamiprid, dinotefuran and imidacloprid, and the method for extracting the tea leaves or tea soup sample is one of the following:

[0029] (A) Tea extraction method: The tea sample is crushed and ground into powder, soaked in water, and then internal standards IMI-d4 and THI-d3 are added, and then acetonitrile containing a small amount of acetic acid is added as an extraction solution. After sufficient vortex mixing, it is filtered, and the supernatant is partially dehydrated with anhydrous sodium sulfate or anhydrous magnesium sulfate, and then allowed to stand for stratification. The supernatant is taken and PVPP and GCB are added for impurity removal, and anhydrous sodium sulfate or anhydrous magnesium sulfate is used to remove water again, vortexed and centrifuged, and the supernatant is taken, gently blown to dryness with nitrogen, re-dissolved with acetonitrile, and CLO-d3 is added. After passing through a 0.22 μm organic filter membrane, the sample is injected into LC-MS / MS for detection, and the pesticide residue result is obtained through the corresponding standard curve; the volume fraction of acetic acid in the extraction solution is 0.5-2.0%, preferably 1%;

[0030] (B) Tea soup extraction method: After the tea leaves are fully brewed with boiling water, the brewing liquid is added with sodium chloride, internal standards IMI-d4 and THI-d3, and dichloromethane is added for extraction and layering. The supernatant is taken and evaporated to near dryness, and the residue is dissolved in acetonitrile containing a small amount of acetic acid. The supernatant is taken and added with PVPP and GCB for impurity removal and PSA, vortexed and centrifuged, the supernatant is taken, gently blown to dryness with nitrogen, re-dissolved with acetonitrile, and CLO-d3 is added. After passing through a 0.22 μm organic filter membrane, the sample is injected into LC-MS / MS for detection, and the pesticide residue result is obtained through the corresponding standard curve. The volume fraction of the small amount of acetic acid contained in the acetonitrile is 0.5-2.0%, preferably 1%.

[0031] The beneficial effects achieved by this application are:

[0032] 1) This application selects several types of pesticides that have relevant limit regulations for tea matrix in my country's GB2763-2021 "Maximum Residue Limits of Pesticides in Food", as well as the types of pesticides that are prohibited and restricted on tea in the announcement of the Ministry of Agriculture. When testing for pesticides such as chlorpyrifos, bifenthrin, cypermethrin, chlorpyrifos, malathion, etc., the method for extracting tea or tea soup samples is: tea leaves and tea soup are extracted with acetonitrile (containing 1% acetic acid) solution, purified by QuEChERS rapid pretreatment method, and measured by GC-ECD and GC-FPD instruments. The ECD and FPD detectors under gas chromatography are used to analyze the pesticide residues in tea leaves and tea soup. The instrument and equipment cost is low, and the detection limit specified by the standard limit can be achieved, which improves the detection sensitivity.

[0033] 2) For pesticides such as thiamethoxam, thiamethoxam, clothianidin, chlorothiazolin, nitenpyram, acetamiprid, dinotefuran and imidacloprid, the GC-ECD and GC-FPD methods cannot detect the residual pesticides well, and the present application further uses LC-MS / MS for detection and analysis. In the method of extracting tea leaves or tea soup samples, the optimal sample purification method is finally determined by the different effects of graphitized carbon black (GCB) on the pre-treatment process of tea leaves and tea soup.

[0034] 3) A certain amount of acetic acid is added to the extraction solution of the present application. The acidity of acetic acid helps to dissolve organic matter, pigments and other macromolecular substances (including pesticide substances) in tea leaves. If acetic acid is not added, organic matter, pigments and other macromolecular substances will be very tightly and firmly combined with pesticide substances. Even if acetonitrile with strong polarity is used, it is difficult to dissolve all the pesticide substances. In this case, the test results will basically be undetected or the detection amount will be relatively low, the sample chromatographic peak will be relatively clean, and organic matter, pigments and other macromolecular substances cannot be completely dissolved. The purpose of adding acetic acid is to increase the acidity of the extract, according to the principle of material polarity compatibility, to extract pesticide substances as much as possible and improve the recovery rate of sample extraction.

[0035] 4) The role of NaCl, sodium sulfate or magnesium sulfate in the extraction of this application is salting out. Salts make the pesticides in the tea leaves fully contact and absorb with acetonitrile, which is convenient for acetonitrile extraction. This application has verified through experiments that if sodium chloride is not added, the extraction conversion efficiency is low for the actual sample analysis. Taking the extraction efficiency of the experiment with sodium chloride added in the embodiment of the present invention as 100%, the extraction efficiency of the experiment without sodium chloride is only 25.8-40.2%.

[0036] 5) The method of the present invention has the advantages of less interference from impurities, convenient sample pretreatment, reliable precision and accuracy, etc. The disadvantage of the GC-ECD method is that the qualitative analysis is not accurate enough, there is interference from chlorine-containing impurity peaks, and it is only suitable for the detection of chlorine-containing organic matter. It can be applied to the trace analysis and detection of chlorpyrifos, bifenthrin, cypermethrin and cypermethrin in tea soup samples; the disadvantage of the GC-FPD method is that the detection limit is not low enough, and it is only suitable for the detection of phosphorus-containing and sulfur-containing organic matter, and can be applied to the analysis and detection of chlorpyrifos and malathion in tea soup samples; the advantages of the LC-MS / MS method are high sensitivity, accurate qualitative analysis, and very low detection limit, which can be widely used in the trace analysis and detection of neonicotinoid pesticides in tea soup samples; the above three analysis methods all meet the limit requirements of standard GB 2763-2021. BRIEF DESCRIPTION OF THE DRAWINGS

[0037] Figure 1 A schematic diagram of gas chromatography of four pesticides at a concentration of 100 μg / L under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 1 of the present invention is shown;

[0038] Figure 2 A schematic diagram of gas chromatography of two pesticides at a concentration of 100 μg / L under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup according to Example 2 of the present invention is shown;

[0039] Figure 3a A schematic diagram of total ion chromatogram of clothianidin, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 3 of the present invention.

[0040] Figure 3b A schematic diagram of total ion chromatogram of acetamiprid, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea soup provided in Example 3 of the present invention.

[0041] Figure 3c A schematic diagram of the total ion chromatogram of a clothianidin-d3 internal standard with a concentration of 100 μg / L under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 3 of the present invention is shown.

[0042] Figure 3d A schematic diagram of total ion chromatogram of dinotefuran, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 3 of the present invention.

[0043] Figure 3eA schematic diagram of total ion chromatogram of imidacloprid, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 3 of the present invention.

[0044] Figure 3f A schematic diagram of total ion chromatogram of chlorothiazide, a neonicotinoid pesticide type with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea soup provided in Example 3 of the present invention.

[0045] Figure 3g A schematic diagram of total ion chromatogram of nitenpyram, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea soup provided in Example 3 of the present invention.

[0046] Figure 3h A schematic diagram of total ion chromatogram of thiacloprid, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 3 of the present invention.

[0047] Figure 3i A schematic diagram of total ion chromatogram of thiamethoxam, a neonicotinoid pesticide with a concentration of 100 μg / L, is shown under an analytical method for detecting multiple pesticide residues in tea leaves and tea soup provided in Example 3 of the present invention. DETAILED DESCRIPTION

[0048] The present invention will be further described below in conjunction with specific embodiments, but the protection scope of the present invention is not limited thereto.

[0049] The graphitized carbon in the present invention is formed by heating carbon black to about 2800°C in an inert gas flow and calcining for 2 hours. At high temperature, the original uneven surface structure is graphitized to obtain a uniform graphitized surface. The present invention directly uses graphitized carbon material to treat tea samples, which can be used to purify organic extracts of plants in pesticide residue analysis and remove pigments in the pretreatment of samples such as fruits and vegetables.

[0050] Embodiment 1

[0051] 1. Green tea and chrysanthemum tea samples were purchased from supermarkets and tea shops in Hangzhou, Zhejiang Province;

[0052] 2. Sample pretreatment

[0053] 2.1 Tea leaves: Weigh (5.00 g ± 0.01 g) tea leaves into a 50 mL centrifuge tube, add 20 mL acetonitrile (containing 1% volume fraction of acetic acid) extraction solution, then add 4 g sodium chloride and 1 g sodium acetate, fully vortex for 2 min, then ultrasonic for 20 min, centrifuge at 8000 r / min for 5 min. Take 2 mL of supernatant into a 10 mL centrifuge tube containing 50 mg GCB, 50 mg PSA and 200 mg anhydrous magnesium sulfate, fully vortex for 2 min, centrifuge at 8000 r / min for 5 min, and pass through a 0.22 μm organic filter membrane before GC-ECD detection.

[0054] 2.2 Tea soup: Weigh tea leaves (3.00 g ± 0.01 g), prepare tea soup according to the tea sensory evaluation method, add 150 mL of boiling water, brew for 10 min, and after cooling, weigh (10.00 g ± 0.01 g) of tea soup in a 50 mL centrifuge tube, add 20 mL of acetonitrile (containing 1% volume fraction of acetic acid) extraction solution, then add 4 g of sodium chloride and 1 g of sodium acetate, fully vortex for 2 min, then ultrasonic for 20 min, and centrifuge at 8000 r / min for 5 min. Transfer all the centrifugal supernatant, add 4 g of anhydrous magnesium sulfate, fully vortex for 2 min, centrifuge at 8000 r / min for 5 min, take 2 mL of supernatant in a 10 mL centrifuge tube containing 50 mg PSA and 200 mg of anhydrous magnesium sulfate, fully vortex for 2 min, centrifuge at 8000 r / min for 5 min, and pass through a 0.22 μm organic filter membrane before GC-ECD detection.

[0055] 3. Detection of four pesticide residues in tea leaves under optimal gas chromatography conditions

[0056] Capillary column: DB-1701 (30m×250μm×0.25μm); carrier gas: high-purity nitrogen (purity 99.999%), column flow rate: 1.0mL / min; split injection, split ratio of 5:1; inlet temperature: 220℃; injection volume: 2μL; septum purge flow: 3mL / min; heating program: initial temperature 80℃, hold for 1min, then increase to 260℃ at 20℃ / min, hold for 15min; detector temperature (ECD): 280℃; tail gas flow: 60mL / min.

[0057] 4. Under the experimental conditions of step 3, the linear regression equation, correlation coefficient and detection limit of four pesticides (chlorpyrifos, bifenthrin, cypermethrin and cypermethrin) were obtained:

[0058] 4.1 Preparation of standard solutions: Accurately weigh the four pesticide standards and prepare a 1.0 mg / mL single pesticide standard stock solution with n-hexane, and store it at -20°C. Pipette a certain volume of the single pesticide standard stock solution one by one and inject it into the same volumetric flask, dilute it to the scale with acetonitrile, and prepare a 5.0 mg / L mixed pesticide standard stock solution, and store it at -20°C.

[0059] 4.2 Preparation of standard curve: The mixed standard stock solution was diluted step by step (the diluent was acetonitrile) to prepare 8 matrix-matched standard solutions of corresponding concentrations (5 μg / L, 10 μg / L, 20 μg / L, 30 μg / L, 40 μg / L, 50 μg / L, 100 μg / L, 150 μg / L and 200 μg / L) for determination.

[0060] 4.3 Draw a standard curve: Inject samples in order from low to high concentration, use the concentration of the standard solution series (μg / L) as the horizontal axis and the peak area of ​​the chromatographic peaks of the four pesticides as the vertical axis to draw a standard curve and obtain their linear regression equation and correlation coefficient;

[0061] 4.4 Calculation of detection limit: Calculation of detection limit of four pesticides: Take 1mL of four pesticide solutions with a concentration of 10μg / L under gas chromatography GC-ECD, repeat the test 8 times according to the detection limit test method, calculate its standard deviation S, and calculate the detection limit of four pesticide samples according to the "Technical Guidelines for the Formulation and Revision of Environmental Testing and Analysis Methods" (HJ 168-2020): Among them, the calculation formula of HJ168-2020 detection limit is: MDL=t (n-1,0.99) *S, where MDL is the method detection limit; t is the t distribution with (n-1) degrees of freedom and 99% confidence level (n=8, t=2.998); S is the standard deviation of n replicates; n is the number of replicates of the sample;

[0062] Table 1 Experimental results of spiked determination of tea matrix sample-1 (n=8) μg / L

[0063]

[0064]

[0065] Table 1 shows the test results of using pesticide-free chrysanthemum tea leaves, the sample solution obtained by the treatment method in step 2.1, and then adding pesticides. As can be seen from Table 1, the detection limits of the four pesticides (chlorpyrifos, bifenthrin, cypermethrin, and cypermethrin) under gas chromatography are 3.4μg / L, 2.6μg / L, 3.1μg / L, and 6.7μg / L, respectively; the lower limits of determination are 13.6μg / L, 10.4μg / L, 12.4μg / L, and 26.8μg / L, respectively.

[0066] 5. Under the experimental conditions of steps 3 and 4, conduct precision and accuracy tests on the spiked tea matrix samples within the linear range:

[0067] 5.1 Take the recovery rate experiment of three levels of addition in tea matrix samples (low, medium and high) (10, 20, 100 μg / kg) and the recovery rate experiment of three levels (10, 20, 100 μg / kg) in tea soup respectively, and measure three times according to the full procedure of steps 3 and 4, and measure the recovery rate and precision of the matrix samples after spiked. The limit of quantification is 10 times the standard deviation of the test results of 8 blank samples; among them, the precision and accuracy test results of tea and tea soup matrix samples after spiked are shown in Tables 2 and 3:

[0068] Table 2 Linear range, correlation coefficient, accuracy, precision and quantification limit of four pesticides in tea

[0069]

[0070] Table 2 shows the test results obtained by using pesticide-free chrysanthemum tea leaves, obtaining the sample solution according to the treatment method in step 2.1, and then adding pesticides.

[0071] Table 3 Linear range, correlation coefficient, accuracy, precision and quantification limit of four pesticides in tea soup

[0072]

[0073] Table 3 shows the test results obtained by using pesticide-free chrysanthemum tea leaves, obtaining the sample solution according to the treatment method in step 2.2, and then adding pesticides.

[0074] As shown in Tables 2 and 3, the relative standard deviations of the four pesticide determination results of tea leaves and tea soup samples after spiked were 1.1% to 18.6% and 2.1% to 19.5%, respectively, and the spiked recovery ranges were 85.9% to 110% and 80.1% to 119%, respectively, indicating that the tea leaves and tea soup methods have good precision and accuracy and meet the quality control requirements;

[0075] 6. Test results of actual samples of tea leaves and tea soup:

[0076] Using the detection method developed by this study, two kinds of tea samples sold on the market were screened for the above four pesticides, and the results are shown in Table 4. The experimental results show that pesticide residues were detected in both types of tea, but the residues were far lower than the maximum residue limits of pesticides in tea stipulated in GB2763-2021. The above tea samples were brewed according to the method in 2.2, and the tea soup samples were processed and analyzed according to the established pretreatment method. The results of the on-machine determination showed that the pesticide residues detected in both green tea and chrysanthemum tea soups were far lower than the maximum residue limits of pesticides in tea stipulated in GB 2763-2021.

[0077] Table 4 Detection results of pesticides in tea samples

[0078]

[0079] Note: ND means not detected;

[0080] The method of the present invention has the advantages of low detection limit, convenient sample pretreatment, reliable precision and accuracy, etc. It can be applied to the trace analysis and detection of four pesticides in tea and tea soup samples, and the method meets the limit requirements of standard GB 2763-2021.

[0081] A gas chromatography diagram of four pesticides at a concentration of 100 μg / L under an analytical method for detecting multiple pesticide residues in tea soup provided in Example 1 of the present invention is shown in FIG. Figure 1 As shown, Figure 1 Chinese: 1 - chlorpyrifos; 2 - bifenthrin; 3 - chlorpyrifos; 4 - cypermethrin.

[0082] Embodiment 2

[0083] 1. Green tea and chrysanthemum tea samples were purchased from supermarkets and tea shops in Hangzhou, Zhejiang Province;

[0084] 2. Sample pretreatment

[0085] 2.1 Tea leaves: Weigh (5.00 g ± 0.01 g) tea leaves into a 50 mL centrifuge tube, add 20 mL acetonitrile (containing 1% volume fraction of acetic acid) extraction solution, then add 4 g sodium chloride and 1 g sodium acetate, fully vortex for 2 min, then ultrasonic for 20 min, centrifuge at 8000 r / min for 5 min. Take 2 mL of supernatant into a 10 mL centrifuge tube containing 50 mg GCB, 50 mg PSA and 200 mg anhydrous magnesium sulfate, fully vortex for 2 min, centrifuge at 8000 r / min for 5 min, and pass through a 0.22 μm organic filter membrane before GC-ECD detection.

[0086] 2.2 Tea soup: Weigh tea leaves (3.00 g ± 0.01 g), prepare tea soup according to the tea sensory evaluation method, add 150 mL of boiling water, brew for 10 min, and after cooling, weigh (10.00 g ± 0.01 g) of tea soup in a 50 mL centrifuge tube, add 20 mL of acetonitrile (containing 1% volume fraction of acetic acid) extraction solution, then add 4 g of sodium chloride and 1 g of sodium acetate, fully vortex for 2 min, then ultrasonic for 20 min, and centrifuge at 8000 r / min for 5 min. Transfer all the centrifugal supernatant, add 4 g of anhydrous magnesium sulfate, fully vortex for 2 min, centrifuge at 8000 r / min for 5 min, take 2 mL of supernatant in a 10 mL centrifuge tube containing 50 mg PSA and 200 mg of anhydrous magnesium sulfate, fully vortex for 2 min, centrifuge at 8000 r / min for 5 min, and pass through a 0.22 μm organic filter membrane before GC-FPD detection.

[0087] 3. Detection of two pesticide residues (chlorpyrifos and malathion) in tea leaves under the best gas chromatography conditions

[0088] Capillary column: DB-1701 (30m×250μm×0.25μm); carrier gas: high-purity nitrogen (purity 99.999%), column flow rate: 1.5mL / min; split injection, split ratio of 5:1; inlet temperature: 220℃; injection volume: 2μL; septum purge flow: 3mL / min; heating program: initial temperature 80℃, hold for 1min, then increase to 260℃ at 20℃ / min, hold for 15min; detector temperature (FPD): 280℃; hydrogen flow: 75mL / min; air flow: 100mL / min; tail gas flow: 60mL / min.

[0089] 4. Under the experimental conditions of step 3, obtain the linear regression equation, correlation coefficient and detection limit of the two pesticides:

[0090] 4.1 Preparation of standard solution: Accurately weigh the standard products of two pesticides, prepare a 1.0 mg / mL single pesticide standard stock solution with n-hexane, and store it at -20°C. Pipette a certain volume of the single pesticide standard stock solution one by one and inject it into the same volumetric flask, dilute it to the scale with acetonitrile, prepare a 5.0 mg / L mixed pesticide standard stock solution, and store it at -20°C.

[0091] 4.2 Preparation of standard curve: Dilute the mixed standard stock solution step by step to prepare 6 matrix-matched standard solutions of corresponding concentrations (30 μg / L, 40 μg / L, 50 μg / L, 100 μg / L, 150 μg / L and 200 μg / L) for determination.

[0092] 4.3 Draw a standard curve: Inject samples in order from low to high concentration, use the concentration of the standard solution series (μg / L) as the horizontal axis and the chromatographic peak area of ​​the two pesticides as the vertical axis to draw a standard curve and obtain their linear regression equation and correlation coefficient;

[0093] 4.4 Calculation of detection limit: Calculation of detection limit of two pesticides: Take 1mL of two pesticide solutions with a concentration of 20μg / L under GC-FPD, repeat the test 7 times according to the detection limit test method, calculate its standard deviation S, and calculate the detection limit of four pesticide samples according to the "Technical Guidelines for the Formulation and Revision of Environmental Testing and Analysis Methods" (HJ 168-2020): Among them, the calculation formula of HJ 168-2020 detection limit is: MDL=t (n-1,0.99) *S, where MDL is the method detection limit; t is the t distribution with (n-1) degrees of freedom and 99% confidence level (n=7, t=3.143); S is the standard deviation of n replicates; n is the number of replicates of the sample;

[0094] Table 5 Experimental results of spiked determination of tea matrix sample-1 (n=7) μg / L

[0095]

[0096] As can be seen from Table 5, the detection limits of the two pesticides (chlorpyrifos and malathion) under gas chromatography are 6.0 μg / L and 5.1 μg / L, respectively; the determination lower limits are 24.0 μg / L and 20.4 μg / L, respectively.

[0097] 5. Under the experimental conditions of steps 3 and 4, conduct precision and accuracy tests on the spiked tea matrix samples within the linear range:

[0098] 5.1 Take the recovery rate experiment of three levels of addition in tea matrix samples (low, medium and high) (10, 20, 100 μg / kg) and the recovery rate experiment of three levels (10, 20, 100 μg / kg) in tea soup respectively, and measure three times according to the full procedure of steps 3 and 4, and measure the recovery rate and precision of the matrix samples after spiked. The quantitative limit is 10 times the standard deviation of the test results of 8 blank samples; Among them, the precision and accuracy test results of tea and tea soup matrix samples after spiked are shown in Tables 6 and 7:

[0099] Table 6 Linear range, correlation coefficient, accuracy, precision and quantification limit of two pesticides in tea

[0100]

[0101] Table 7 Linear range, correlation coefficient, accuracy, precision and quantification limit of two pesticides in tea soup

[0102]

[0103] As shown in Tables 6 and 7, the relative standard deviations of the four pesticide determination results of tea leaves and tea soup samples after spiked were 4.4% to 17.6% and 2.5% to 25.2%, respectively, and the spiked recoveries were 84.3% to 115% and 79.9% to 94.5%, respectively, indicating that the tea leaves and tea soup methods have good precision and accuracy and meet the quality control requirements. However, the detection limit of chlorpyrifos pesticide by FPD method is higher than that by ECD method, and the sensitivity is not high.

[0104] 7. Test results of actual samples of tea leaves and tea soup:

[0105] Using the detection method developed by this research institute, two kinds of tea samples sold on the market were screened for the above four pesticides, and the results are shown in Table 8. The experimental results show that pesticide residues were detected in both types of tea, but the residues were far lower than the maximum residue limit of tea pesticides stipulated in GB2763-2021 (the maximum residue limit of chlorpyrifos in tea is 2 mg / kg). The above tea samples were brewed according to the method in 2.2, and the tea samples were processed and analyzed according to the established pretreatment method. The results of the machine determination showed that except for a little malathion detected in Chrysanthemum Tea Soup 2, chlorpyrifos and malathion were not detected in the other samples of green tea and chrysanthemum tea soup, which was also due to the different detection limits of the ECD method and the FPD method.

[0106] Table 8 Detection results of pesticides in tea samples

[0107]

[0108] Note: ND means not detected.

[0109] A gas chromatography diagram of two pesticides at a concentration of 100 μg / L under an analytical method for detecting multiple pesticide residues in tea soup provided in Example 2 of the present invention is shown as follows: Figure 2 As shown, Figure 2 Chinese: 1-chlorpyrifos; 2-malathion.

[0110] Embodiment 3

[0111] 1. Green tea, white tea, oolong tea, chrysanthemum tea, black tea, yellow tea, jasmine tea, and honeysuckle tea samples were purchased from supermarkets and tea shops in Hangzhou, Zhejiang Province;

[0112] 2. Sample pretreatment

[0113] 2.1 Tea: The tea samples were crushed, ground into powder, and passed through a 60-mesh sieve; 1 g of the crushed tea sample was placed in a 50 mL centrifuge tube, and 2 mL of ultrapure water was added first, and the mixture was soaked for 30 minutes; 20 μL of 5 ppm IMI-d4 and THI-d3, as well as 20 mL of acetonitrile (containing 1% volume fraction of acetic acid) were added to the centrifuge tube, vortexed for 2 minutes to mix them evenly, and filtered through Whatman No. 1 filter paper; the supernatant was transferred to a 50 mL centrifuge tube, 2 g of anhydrous sodium sulfate was added, vortexed for 1 minute, and then allowed to stand to separate the acetonitrile phase and the aqueous phase; 2 mL of the supernatant was taken, and 160 mg PVPP, 20 mg GCB, 150 mg anhydrous magnesium sulfate, vortex for 2 minutes, centrifuge at 4000 rpm for 8 minutes; take 1 mL of supernatant, blow gently with nitrogen to dryness, reconstitute with 960 μL acetonitrile, add 40 μL CLO-d3 with a concentration of 5 ppm, transfer to the injection vial after filtering through a 0.22 μm filter membrane, and wait for injection;

[0114] 2.2 Tea soup: Take 3g of tea leaves, rinse with 50mL of ultrapure water heated to 100℃, quickly rotate for 20 seconds to get the rinse liquid, then add 150mL of 100℃ ultrapure water into a 200mL glass to soak the tea leaves, cover and seal, put into a conical flask after 5 minutes for later use. The above is the first brewing. Repeat the above tea brewing steps to obtain the second, third and fourth brewing liquids, and the fifth soaking time is one hour; the rinse liquid and brewing liquid are vacuum filtered into a conical flask through a microporous filter paper, and 5g sodium chloride, 5ppm IMI-d4, THI-d3, 20μL each, are added, and the mixture is transferred to a separatory funnel after shaking; 30mL of dichloromethane is added, shaken for 2 minutes, and allowed to stand for 5 minutes to wait for stratification, and liquid-liquid extraction is performed, and the lower supernatant (dichloromethane phase) is placed in a round-bottom flask, and the above extraction steps are repeated twice; the emulsified layer is collected in a 50mL centrifuge tube, centrifuged at 8000rpm for 5 minutes, and the lower supernatant is transferred to the round-bottom flask in step 3, and rotary evaporated to near dryness; the residue is dissolved with 10mL acetonitrile (containing 1% volume fraction of acetic acid), 2mL of the supernatant is taken to a 10mL centrifuge tube, and 150mg PVPP (polyvinyl polypyrrolidone) and 50mg PSA (PSA is a solid phase adsorbent of N-propylethylenediamine, mainly used to adsorb target substances in tea), 10 mg GCB (graphitized carbon), vortex for two minutes, centrifuge at 8000 rpm for 10 minutes; take 1 mL of the supernatant after centrifugation, blow it to dryness with gentle nitrogen, re-dissolve it with 960 μL of acetonitrile, add 40 μL of CLO-d3 with a concentration of 5 ppm, and transfer it to the injection vial through a 0.22 μm filter membrane for injection;

[0115] 3. Detection of 8 neonicotinoid pesticide residues in tea soup under optimal LC-MS / MS conditions

[0116] The chromatographic column is YMC ODS-AQ 100mm*2.1mm*3μm, the mobile phases A and B are 0.1wt% formic acid water and pure acetonitrile respectively, and the mobile phase flow rate is 0.3mL / min. Injection volume: 5μL, gradient elution setting: 0-1.3min, set to 90% mobile phase A and 10% mobile phase B, 1.3-4min set to 70% mobile phase A and 30% mobile phase B, 4-5min set to 10% mobile phase A and 90% mobile phase B, remain unchanged within 5-6min, 6-6.5min return to the initial state of 90% mobile phase A and 10% mobile phase B, and remain unchanged for two minutes at 6.5-8.5min.

[0117] The mass spectrometer ion source was selected as ESI, and multiple reaction monitoring (MRM) was performed in positive ion mode, with the acceleration voltage set to 5 V, the sheath gas temperature to 250 ° C, the sheath gas flow rate to 11 L / min, the nozzle voltage to 450 V, the nebulizer pressure to 35 psi, the capillary voltage to 4000 V, the high-voltage Funnel RF and the low-voltage Funnel RF to 150 V and 60 V, respectively. The parent ions and daughter ions of eight neonics, dimethomorph, thiamethoxam, thiamethoxam, clothianidin, chlorothiazolin, nitenpyram, acetamiprid and imidacloprid, were set to 203.1 / 129.1, 292.0 / 211.0, 253.0 / 126.1, 250.0 / 169.0, 262.0 / 181.0, 271.0 / 225.1, 223.1 / 126.1, 256.0 / 209.1, respectively. The MRM parameters of internal standards CLO-d3, IMI-d4 and THI-d3 were 253.0 / 172.0, 260.0 / 213.2 and 295.0 / 214.0, respectively.

[0118] 4. Under the experimental conditions of step 3, the linear regression equations, correlation coefficients and detection limits of 8 neonicotinoid pesticides were obtained:

[0119] 4.1 Preparation of standard solution: Weigh the standard products of 8 neonicotinoid pesticides (thiamethoxam, thiacloprid, clothianidin, chlorothiazolin, nitenpyram, acetamiprid, dinotefuran and imidacloprid) respectively, prepare 0.5 mg / mL pesticide standard stock solution with acetonitrile, and use clothianidin-D3, imidacloprid D4 and thiacloprid-D3 as internal standards, and store at -20℃. Pipette a certain volume of individual pesticide standard stock solution one by one and inject it into the same volumetric flask, dilute to the mark with acetonitrile, prepare 5.0 mg / L mixed pesticide standard stock solution, and store at -20℃.

[0120] 4.2 Preparation of standard curve: The mixed standard stock solution was diluted stepwise with acetonitrile-water solution (acetonitrile-water volume ratio 9:1) to prepare 6 matrix-matched standard solutions of corresponding concentrations (0.025 μg / L, 0.5 μg / L, 1.25 μg / L, 2.5 μg / L, 12.5 μg / L and 25 μg / L) for determination.

[0121] 4.3 Draw a standard curve: Inject samples in order from low to high concentration, use the concentration of the standard solution series (μg / L) as the horizontal axis and the mass spectrometry peak area of ​​the 8 pesticides as the vertical axis to draw a standard curve and obtain their linear regression equation and correlation coefficient;

[0122] 4.4 Calculation of detection limit: Calculation of detection limit of 8 pesticides: Take 1mL of 8 pesticide solutions with a concentration of 0.005μg / L under LC-MS / MS, repeat the measurement 7 times according to the detection limit test method, calculate its standard deviation S, and calculate the detection limit of 8 pesticide samples according to the "Technical Guidelines for the Formulation and Revision of Environmental Testing and Analysis Methods" (HJ 168-2020): Among them, the calculation formula of HJ 168-2020 detection limit is: LOD=t (n-1,0.99) *S, where LOD is the limit of detection; t is the t distribution with (n-1) degrees of freedom and 99% confidence level (n=7, t=3.143); S is the standard deviation of n replicates; n is the number of replicates of the sample;

[0123] Table 9 Linear regression equation, correlation coefficient, LOD, LOQ and recovery rate of neonicotinoid pesticides

[0124]

[0125] 5. Under the experimental conditions of steps 3 and 4, the precision and accuracy tests were conducted on the spiked tea matrix samples within the linear range: the recovery rate experiments at three levels (0.5, 1.0, 5.0 μg / kg) in the tea matrix samples and the recovery rate experiments at three levels (0.5, 1.0, 5.0 μg / kg) in the tea soup were conducted respectively. The whole procedure of steps 3 and 4 was followed for three times to measure the recovery rate and precision of the matrix samples after spiked. The precision and accuracy test results of the spiked tea and tea soup matrix samples are shown in Table 9:

[0126] As shown in Table 9, the relative standard deviations of the determination results of 8 neonicotinoid pesticides in tea leaves and tea soup samples after spiked were 3.6% to 13.8% and 7.5% to 14.8%, respectively, and the spiked recoveries were 78.0% to 98.0% and 77.7% to 106.7%, respectively, indicating that the tea leaves and tea soup methods have good precision and accuracy and meet the quality control requirements;

[0127] According to an analytical method for detecting multiple pesticide residues in tea leaves provided in Example 3 of the present invention, the total ion chromatograms of neonicotinoid pesticides with a concentration of 100 μg / L are clothianidin, acetamiprid, clothianidin-d3 (internal standard), dinotefuran, imidacloprid, chlorthiazolin, nitenpyram, thiacloprid and thiamethoxam, respectively. Figure 3a-3i shown.

[0128] 8. Test results of actual samples of tea leaves and tea soup:

[0129] Using the detection method developed by this study, tea leaves and brewed tea samples were processed and analyzed according to the established pretreatment method, and the above 8 pesticides were screened and tested for 6 kinds of tea samples sold on the market. The results are shown in Tables 10 and 11. The experimental results show that pesticide residues were detected in all 8 kinds of tea leaves and tea soups. The average concentration range of tea leaves was ND-1093.33μg / kg, and the average concentration range of tea soup was ND-312.33μg / kg. The residues were lower than the maximum residue limits of tea pesticides in GB 2763-2021. (Among them, the maximum residue limits of thiamethoxam, nitenpyram, chlorpyrifos, imidacloprid, thiamethoxam, clothianidin, acetamiprid and dinotefuran in GB 2763-2021 are 10, 1, 3, 0.5, 10, 10, 10 and 20mg / kg respectively).

[0130] Table 10 Detection results of pesticides in tea samples

[0131]

[0132] Note: ND means not detected.

[0133] Table 11 Detection results of pesticides in tea samples

[0134]

[0135] Note: ND means not detected.

[0136] This method has the advantages of less impurity interference, convenient sample pretreatment, reliable precision and accuracy. The disadvantage of GC-ECD is that the qualitative analysis is not accurate enough, there is interference from chlorine-containing impurity peaks, and it is only suitable for the detection of chlorine-containing organic matter, and can be applied to the trace analysis and detection of chlorpyrifos, bifenthrin, cypermethrin and chlorpyrifos in tea and tea soup samples; the disadvantage of GC-FPD is that the detection limit is not low enough, and it is only suitable for the detection of phosphorus-containing organic matter, and can be applied to the analysis and detection of chlorpyrifos and malathion in tea and tea soup samples; the advantages of LC-MS / MS are high sensitivity, very low detection limit, accurate qualitative analysis, and can be widely used in the trace analysis and detection of neonicotinoid pesticides in tea and tea soup samples; the above three analysis methods all meet the limit requirements of standard GB 2763-2021.

Claims

1. An analytical method for detecting multiple pesticide residues in tea soup, characterized in that: The method comprises the following steps: 1) Drawing of standard curve S1: Prepare the pesticide standards into standard stock solutions using n-hexane as solvent; S2: Take each standard stock solution, use acetonitrile as solvent to prepare a mixed pesticide standard stock solution, dilute the mixed pesticide standard stock solution step by step to prepare gradient standard solutions of different concentrations, and wait for the standard curve to be analyzed by the machine; S3: The pesticide is at least one of thiamethoxam, thiacloprid, clothianidin, chlorpyrifos, nitenpyram, acetamiprid, dinotefuran and imidacloprid. According to steps S1-S2, matrix-matched standard solutions of different concentrations are prepared, and internal standards thiamethoxam-d3 (THI-d3), imidacloprid-d4 (IMI-d4) and clothianidin-d3 (CLO-d3) are added respectively. Liquid chromatography-tandem mass spectrometry LC-MS / MS is used for analysis to obtain a chromatogram, and the matrix-matched standard curve is quantitatively determined by the internal standard method; 2) Detection and analysis of pesticide residues Extracting the tea leaves or tea soup sample by solvent extraction to obtain an extracted sample solution, then detecting the chromatogram of the pesticides in the extracted sample solution by the LC-MS / MS method in step S3, and calculating the concentrations of different types of pesticides in the tea leaves or tea soup sample according to the corresponding standard curve; The conditions for the liquid chromatography-tandem mass spectrometry LC-MS / MS test in step S3 are: The chromatographic column was YMC ODS-AQ 100mm*2.1mm*3μm, the mobile phases A and B were 0.1wt% formic acid water and pure acetonitrile, respectively, the mobile phase flow rate was 0.3mL / min; the gradient elution conditions were: 0-1.3min, 90%A; 1.3-4min, 70%A; 4-6min, 10%A; 6-6.5min, return to the initial state of 90%A, and rinse for two minutes at 6.5-8.5min; the mass spectrometry conditions were: electrospray ion source, positive ion mode ESI+; When the chromatogram of the pesticide in the extracted sample solution is detected according to the method of step S3, the pesticide is at least one of thiamethoxam, thiamethoxam, clothianidin, chlorpyrifos, nitenpyram, acetamiprid, dinotefuran and imidacloprid, and the method for extracting the tea or tea soup sample is one of the following: (A) Tea extraction method: The tea sample is crushed and ground into powder, soaked in water, and then the internal standards IMI-d4 and THI-d3 are added. Then, acetonitrile containing a small amount of acetic acid is added as the extraction solution. After thorough vortex mixing, the sample is filtered. The supernatant is partially dehydrated with anhydrous sodium sulfate or anhydrous magnesium sulfate, and then allowed to stand for stratification. The supernatant is taken and PVPP and GCB are added for impurity removal, and anhydrous sodium sulfate or anhydrous magnesium sulfate is used to remove water again. The sample is vortexed and centrifuged. The supernatant is taken and blown to dryness with gentle nitrogen. The sample is re-dissolved in acetonitrile and CLO-d3 is added. The sample is filtered through a 0.22 μm organic filter membrane and then injected into LC-MS / MS for detection. The pesticide residue results are obtained through the corresponding standard curve. (B) Tea soup extraction method: After the tea leaves are fully brewed with boiling water, sodium chloride, internal standards IMI-d4 and THI-d3 are added to the brewing liquid, and dichloromethane is added for extraction and the layers are separated. The supernatant is taken and evaporated to near dryness. The residue is dissolved in acetonitrile containing a small amount of acetic acid, and PVPP and GCB are added to the supernatant for impurity removal and PSA. The supernatant is vortexed and centrifuged, and the supernatant is taken and gently blown to dryness with nitrogen. The supernatant is re-dissolved in acetonitrile and CLO-d3 is added. The sample is filtered through a 0.22 μm organic filter and then injected into LC-MS / MS for detection. The pesticide residue results are obtained through the corresponding standard curve.

2. The analytical method for detecting multiple pesticide residues in tea soup according to claim 1, characterized in that: The specific process of preparing the matrix matching standard solution in steps S1 and S2 is as follows: M1: Accurately weigh the standard samples of different pesticides, prepare them into 0.5-2.0 mg / mL single pesticide standard stock solutions with n-hexane, and store them at -15 to -25°C; M2: Pipette individual pesticide standard stock solutions one by one and inject them into the same volumetric flask respectively, dilute to the scale with acetonitrile, prepare 2.0-8.0 mg / L mixed pesticide standard stock solutions, and store at -15--25℃; dilute the mixed pesticide standard stock solutions step by step with acetonitrile to prepare gradient standard solutions of different concentrations.

3. The analytical method for detecting multiple pesticide residues in tea soup according to claim 1, characterized in that: In step S2, eight corresponding concentration gradient standard solutions are prepared, and the concentrations are 5 μg / L, 10 μg / L, 20 μg / L, 30 μg / L, 40 μg / L, 50 μg / L, 100 μg / L, 150 μg / L and 200 μg / L, respectively.

4. The analytical method for detecting multiple pesticide residues in tea soup according to claim 1, characterized in that: In the tea extraction method of step (A), the volume fraction of acetic acid in the extraction solution is 0.5-2.0%.

5. The analytical method for detecting multiple pesticide residues in tea soup as claimed in claim 4, characterized in that: In the tea extraction method of step (A), the volume fraction of acetic acid in the extraction solution is 1%.

6. The analytical method for detecting multiple pesticide residues in tea soup according to claim 1, characterized in that: In the tea soup extraction method of step (B), the volume fraction of the small amount of acetic acid contained in the acetonitrile is 0.5-2.0%.

7. The analytical method for detecting multiple pesticide residues in tea soup according to claim 6, characterized in that: In the tea soup extraction method of step (B), the volume fraction of the small amount of acetic acid contained in the acetonitrile is 1%.

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