Method for detecting residues of thiamethoxam and six metabolites thereof on hot peppers and hot pepper products

Abstract

The invention relates to a method for detecting residues of thiamethoxam and six metabolites of thiamethoxam on chili and chili products, and belongs to the technical field of chili pesticide residue detection. The detection method comprises the following steps: 1) sample extraction and purification: extracting fresh chili, dry pickled chili, wet pickled chili, fermented chili, chili sauce and chili leaf samples by adopting 2% acetic acid-acetonitrile, and purifying by adopting C18 and GCB; 2) calculating standard curves and linear correlation coefficients of thiamethoxam and metabolites thereof; and 3) calculating the addition recovery rate, the relative standard deviation and the method quantitation limit. The standard recovery rate of the seven compounds in the six matrixes is 71%-107%, RSD is smaller than 9.53%, it is proved that the repeatability and reproducibility of the detection method are acceptable, and an analysis method can be provided for simultaneous detection and research of thiamethoxam and six metabolites thereof in fresh chilies, dry pickled chilies, wet pickled chilies, fermented chilies, chilli sauce and chilli leaves.

Description

Technical Field

[0001] The invention belongs to the technical field of pesticide residue detection, and specifically relates to a method for detecting the residues of thiamethoxam and six metabolites thereof in peppers and pepper products. Background Art

[0002] Pepper (Capsicum annuum.L) is a plant of the genus Capsicum in the Solanaceae family and is one of the important economic crops in my country. In addition to being eaten as a fresh vegetable, peppers are also processed into products such as pickled peppers, pickled peppers and chili sauce. However, during the pepper planting process, it is vulnerable to a variety of pests such as aphids, whiteflies, mites, thrips, etc., and mainly relies on chemical pesticides for prevention and control to reduce pests and stabilize yields. During the large-scale planting of peppers, some farmers will use a large amount of pesticides for appearance and yield, resulting in excessive pesticide residues in peppers. Pesticide residues may not only endanger human health, but also be one of the important factors affecting the import and export trade of peppers. Therefore, strictly controlling pesticide residues in peppers and strengthening the ability to detect pesticide residues are key steps to ensure the food safety of peppers.

[0003] Thiamethoxam (TMX) is a second-generation nicotinoid insecticide with a completely new structure. It has stomach poison and contact activity against pests, and can act through plant systemic absorption. It has good insecticidal activity against thrips, whiteflies, aphids, etc., and is widely used in pepper pest control. Although thiamethoxam is a highly effective and low-toxic insecticide, there is still a risk of residues to plants and the environment after unreasonable application. Eventually, these residues will enter the human body through food ingestion and other means, causing harm to human health. It is reported that after thiamethoxam is applied to crops, six main metabolites will be produced, namely clothianidin (Clothianidin, CLO), demethylthiamethoxam, and thiamethoxam.

[0004] (Desmethyl-thiamethoxam, DM-TMX), Desmethylthiamethoxam

[0005] Desmethyl-clothianidin (DM-CLO), Desnitro-thiamethoxam (DN-TMX), Desnitro-clothianidin (DN-CLO) and Thiamethoxam-urea (TMX-urea). Studies have shown that the toxicity of clothianidin and thiamethoxam-urea to bees and green algae is 1.27 times and 39.51 times that of thiamethoxam, respectively; the hepatotoxicity and hepatocarcinogenicity of thiamethoxam to mice are attributed to its metabolite desmethylthiamethoxam, and desmethylthiamethoxam can aggravate the hepatotoxicity; in addition, thiamethoxam and thiamethoxam-urea may also be more toxic to mammals than to insects. Since thiamethoxam and its six metabolites have potential risks to humans and the environment, it is necessary to develop a residue analysis method for the simultaneous and rapid analysis of thiamethoxam and its six metabolites in peppers to comprehensively evaluate the safety of thiamethoxam used on peppers.

[0006] At present, the research on thiamethoxam mainly focuses on the residues of thiamethoxam and its metabolite clothianidin, while the research on the other five metabolites focuses on the insecticidal activity and toxicity. So far, there has been no report on the method for simultaneously detecting the residues of thiamethoxam and its six metabolites in peppers. The present invention is the first to establish a residue analysis method for simultaneously detecting seven target compounds in peppers and pepper products, providing a detection basis for risk assessment and safe use research on peppers. Summary of the invention

[0007] In view of the deficiencies of the above-mentioned prior art, the purpose of the present invention is to design and provide a method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products. The method improves the QuEChERS pretreatment technology, combines ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS / MS), and establishes a set of residue detection methods for simultaneously detecting thiamethoxam, clothianidin, demethylthiamethoxam, demethylthiamethoxam, thiamethoxam guanidine, clothianidin guanidine and thiamethoxam urea in fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, hot peppers, chili sauces and pepper leaves. The method has a fast and simple pretreatment process, good instrumental analysis sensitivity, and high method accuracy, which can provide a detection basis for conducting risk assessment of thiamethoxam and its metabolites on peppers and research on their safe use.

[0008] This is achieved specifically through the following technical solutions:

[0009] A method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products comprises the following steps:

[0010] 1) Extraction of samples: The pre-treated peppers and pepper products were shaken for the first time for 20 minutes using 2% acetic acid-acetonitrile, and then NaCl and anhydrous magnesium sulfate were added and shaken again for 5 minutes, and centrifuged at 5000 r / min for 5 minutes, and the upper organic phase solution was collected;

[0011] 2) Purification of the sample to be tested: Add the organic phase solution in step 1) to the 18 +GCB+anhydrous magnesium sulfate in a 2 mL centrifuge tube, vortex for 1 min, centrifuge at 5000 r / min for 5 min, take the supernatant and filter it through a 0.22 μm filter membrane into a sample injection bottle to obtain the test solution;

[0012] 3) Detection: Using UPLC HSS T3 as the chromatographic column, ultra-high performance liquid chromatography-mass spectrometry was used to quantify the residues of 7 target compounds, including thiamethoxam and its 6 metabolites, in 6 matrices.

[0013] Furthermore, in step 1), the peppers and pepper products are fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, pepper sauce and pepper leaves, and the pretreatment is specifically to homogenize the fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers and spicy peppers, then weigh 5g of each and add 2mL of pure water, and directly weigh 2g of pepper leaves without adding water.

[0014] Furthermore, in step 1), the amount of acetic acid-acetonitrile added is 10 mL, the amount of NaCl added is 1 g, and the amount of anhydrous magnesium sulfate added is 4 g.

[0015] Further, in step 2), C 18 The amount of addition is 50 mg, the amount of addition of GCB is 5 mg, and the amount of addition of anhydrous magnesium sulfate is 150 mg.

[0016] Further, in step 3), the chromatographic conditions of the ultra performance liquid chromatography-mass spectrometry are as follows: chromatographic column: UPLCHSS T3; column temperature 40°C; injection volume 5 μL; mobile phase A is 0.1% formic acid water, B is acetonitrile; flow rate 0.25 mL / min, and the gradient elution program is: 0-1.5 min, 95% A; 1.5-3.0 min, 95%-5% A; 3-3.5 min, 5% A; 3.5-5.0 min, 5%-95% A; 5.0-6.0 min, 95% A.

[0017] Furthermore, in step 3), the mass spectrometry conditions of the ultra-high performance liquid chromatography-mass spectrometry instrument are: ionization source mode: electrospray ionization ESI; ion source polarity: ESI+; nebulizer gas: nitrogen; sheath gas flow rate 12 L / min, auxiliary gas flow rate 3 L / min; collision gas: argon; spray voltage: 3500 V; capillary temperature: 350°C; ion transfer tube temperature: 300°C.

[0018] Furthermore, thiamethoxam and its six metabolites are: thiamethoxam, clothianidin, demethylthiamethoxam, demethylthiamethoxam, thiamethoxam guanidine, clothianidin guanidine and thiamethoxam urea.

[0019] Furthermore, the present invention uses matrix-matched standard solution curves to quantify seven target compounds, and the linear correlation coefficients of the standard curves of thiamethoxam and its metabolites in different matrices are all greater than 0.99.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] 1) The present invention optimizes the QuEChERS method, extracts the sample with 2% acetic acid-acetonitrile, and uses C18 and GCB for purification, thereby developing a stable, practical and sensitive analytical method that can simultaneously detect the residues of thiamethoxam, clothianidin, demethylthiamethoxam, demethylthiamethoxam, thiamethoxam guanidine, clothianidin guanidine and thiamethoxam urea in fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, hot peppers, hot pepper sauces and pepper leaves;

[0022] 2) The detection method of the present invention makes the average recovery rate of 7 target compounds in fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, chili sauce and pepper leaves range from 71% to 107%, and the relative standard deviation (RSD) of the determination is 0.39% to 9.53%, which proves that the repeatability and reproducibility of the detection method are acceptable;

[0023] 3) The pretreatment process of the method of the present invention is fast and simple, the instrumental analysis sensitivity is good, the method is highly accurate, and can be used for the simultaneous detection and analysis of thiamethoxam and its six metabolite residues in fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, chili sauce and pepper leaves. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 This is a schematic diagram of the solvent standard chromatogram of 0.5 mg / L of 7 compounds of the present invention;

[0025] Figure 2 Schematic diagram of the effect of different extractants on the recovery rate of compound addition in the present invention (A extractant is acetonitrile; B extractant is 1% acetic acid-acetonitrile; C extractant is 2% acetic acid-acetonitrile; D extractant is 1% ammonia water-acetonitrile; E extractant is 2% ammonia water-acetonitrile);

[0026] Figure 3Schematic diagram of the effect of different purifiers on the recovery rate of compound addition in the present invention (purifier A is 50mg PSA + 150mg MgSO4; purifier B is 50mg C18 + 150mg MgSO4; purifier C is 50mg PSA + 5mg GCB + 150mgMgSO4; purifier D is 50mg C18 + 5mg GCB + 150mg MgSO4). DETAILED DESCRIPTION

[0027] The present invention is further described below in conjunction with the accompanying drawings in order to better understand the technical solution.

[0028] Materials and reagents used in the experimental process of the present invention:

[0029] Standard products: thiamethoxam (purity>98.0%), clothianidin (purity>99.0%), Shanghai Yuanye Biotechnology Co., Ltd.; demethylthiamethoxam (purity>98.0%), demethylthiamethoxam (purity 98.2%), thiamethoxam urea (purity 99.0%), Dr. Ehrenstorfer, Germany; thiamethoxam guanidine, clothianidin guanidine, synthesized by Wuxi WuXi AppTec New Drug Development Co., Ltd.; formic acid [chromatographically pure, Thermo Fisher Scientific (China) Co., Ltd.)]; acetonitrile (chromatographically pure, Merck, Germany); acetonitrile (analytical pure, Tianjin Komiou Chemical Reagent Co., Ltd.); octadecyl silica (C 18 , Shenzhen Biocomma Biotechnology Co., Ltd.); N-propylethylenediamine (PSA), graphitized carbon (GCB), 0.22 μm organic filter membrane (Tianjin Bona Aiger Technology Co., Ltd.); water was distilled water.

[0030] The instruments and equipment used in the experimental process of the present invention include: TSQAltis Plus triple quadrupole liquid phase mass spectrometer (UHPLC-MS / MS, Thermo Fisher, USA); UPLC HSS T3 chromatographic column (2.1 mm×100 mm, 1.8 μm), Waters, USA); G16-11 desktop high-speed centrifuge (Hunan Pingfan Technology Co., Ltd.), CK2000 high-throughput tissue grinder (Beijing Tomogen Biotechnology Co., Ltd.).

[0031] Preparation of standard solution and drawing of standard curve during the experiment of the present invention:

[0032] Accurately weigh 7 kinds of compound standards into 10mL volumetric flasks, dissolve them in acetonitrile to make a 1000mg / L standard stock solution, then dilute the standard stock into a 100mg / L mixed standard stock solution, store it at -20℃, and gradiently prepare the 100mg / L mixed stock solution into 10mg / L and 1mg / L intermediate solutions.

[0033] Take blank samples of fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaves respectively, and dilute the above standard stock solutions with blank matrix and acetonitrile solvent to 0.001, 0.005, 0.01, 0.05, 0.1, 0.5 and 1.0 mg / L, and draw a standard curve with concentration as the horizontal axis (x) and peak surface as the vertical axis (y).

[0034] Example

[0035] Weigh 5.00 g of fresh pepper, dried pickled pepper, wet pickled pepper, pickled pepper, and chili sauce samples (2.00 g for pepper leaves) after homogenization into a 50 mL centrifuge tube, add 2 mL of purified water (no water is needed for pepper leaves), then add 10 mL of 2% acetic acid-acetonitrile, shake for 20 min, add 1.0 g of NaCl and 4 g of anhydrous magnesium sulfate, shake for 5 min, and centrifuge at 5000 r / min for 5 min. Take 1.5 mL of the upper organic phase solution and add it into a 50 mg C 18 +5 mg GCB + 150 mg anhydrous magnesium sulfate in a 2 mL centrifuge tube, vortex for 1 min, centrifuge at 5000 r / min for 5 min, take the supernatant and filter it through a 0.22 μm filter membrane into a sample injection bottle for UHPLC-MS / MS testing.

[0036] Chromatographic conditions: UPLC HSS T3 column (2.1×100 mm, 1.8 μm); column temperature 40°C; injection volume 5 μL; flow rate 0.25 mL / min; mobile phase A was 0.1% formic acid water, and mobile phase B was acetonitrile; gradient elution program was: 0-1.5 min, 95% A; 1.5-3.0 min, 95%-5% A; 3-3.5 min, 5% A; 3.5-5.0 min, 5%-95% A; 5.0-6.0 min, 95% A.

[0037] Mass spectrometry conditions: ionization source mode: electrospray ionization (ESI); ion source polarity: ESI+; nebulizer gas: nitrogen; sheath gas flow rate 12 L / min, auxiliary gas flow rate 3 L / min; collision gas: argon; spray voltage: 3500 V; capillary temperature: 350 °C; ion transfer tube temperature: 300 °C; mass spectrometry parameters of thiamethoxam and its metabolites are shown in Table 1. The 7 compounds were finally confirmed according to the established instrumental method. The UHPLC-MS / MS spectra of the 7 compounds are shown in Figure 1 shown.

[0038] Table 1: Mass spectrometry parameters of thiamethoxam and its metabolites

[0039]

[0040]

[0041] Note: * is the quantitative ion

[0042] Addition recovery and precision test: According to the requirements of the agricultural industry standard "Test Guidelines for Pesticide Residues in Crops" (NY / T788-2018), this test was designed with four addition levels. Seven compounds were added to the blank samples of fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaves at four concentrations of 0.01 mg / kg, 0.1 mg / kg, 1.0 mg / kg and 10.0 mg / kg for recovery. Each concentration was repeated 5 times, and the above detection steps were used for analysis to calculate the recovery rate and relative standard deviation (RSD).

[0043] Matrix effect: Matrix effects caused by sample matrix components are common and difficult to avoid during UHPLC-MS / MS detection. The presence of matrix components can enhance or inhibit the response of the analyte during ionization, thus interfering with quantification and producing erroneous results. In this application, the matrix effect (Me) is calculated according to the following formula:

[0044] Me=km / ks

[0045] Where km is the slope of the matrix-matched standard curve, ks is the slope of the solvent standard curve, Me>1.2 is the matrix enhancement effect, 0.8≤Me≤1.2 is the weak matrix effect which can be ignored, and Me<0.8 is the matrix suppression effect.

[0046] Optimization of extraction conditions: The acidity and alkalinity of the extraction solvent directly affect the extraction effect of residual compounds in the sample. This application compares the extraction effect of acetonitrile and acetonitrile with acid and alkali on thiamethoxam and its metabolites. Figure 2 As shown, when acetonitrile, 1% ammonia-acetonitrile and 2% ammonia-acetonitrile were used to extract the target compounds in 6 matrices, the average recoveries of the 7 compounds were 36% to 110%, and the RSDs were 1.16% to 13.71%. According to the NY / T 788-2018 standard, the recoveries of DM-TMX, DN-TMX and DN-CLO in some matrices were lower than 70%, which may be because these three compounds are more soluble in acidic solutions; after extraction with 1% acetic acid-acetonitrile, the recoveries of DM-TMX, DN-TMX and DN-CLO increased, and the average recoveries of the three compounds were 50-105%, but the recoveries of DN-TMX and DN-CLO in some matrices still did not meet the standard requirements (70-120%), which may be due to the insufficient acidity of the extract; when 2% acetic acid-acetonitrile was used as the extraction solvent, the average recoveries of the seven compounds were 73-104%, and the RSDs were 0.52-8.98%, which means that the seven compounds can be extracted simultaneously in fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, chili sauce and pepper leaves, and the added recovery rates and RSDs can meet the requirements.

[0047] Optimization of purification conditions: The matrix of peppers and pepper leaves is relatively complex. Acetonitrile can extract or dissolve a variety of substances therein. These substances may interfere with the detection and affect the residual detection. Therefore, the choice of purification agent is particularly important. PSA, GCB, C18, etc. are commonly used as purification agents in the QuEChERS method. PSA (N-propylethylenediamine) can effectively remove sugars, organic acids and some polar pigments in the sample matrix; C18 (octadecylsilane bonded silica gel) is used to adsorb non-polar impurities such as lipids and lipids from polar samples; GCB (graphitized carbon black) can effectively remove hydrophobic compounds in the matrix such as pigments and sterol impurities. The experiment compared the effects of PSA, C18, PSA+GCB, and C18+GCB on the added recovery rate. The results are shown in Figure 2. Figure 3 The average recoveries of the seven compounds after purification with four purifiers were 68%-113%, with RSDs of 1.07%-15.68%. According to the NY / T788-2018 standard, with PSA+GCB as the purifier, the recoveries of DN-TMX and DN-CLO in the fermented pepper and wet pickled pepper matrices exceeded the standard requirement of 70%-120%, respectively. The recoveries of PSA, C18 and C18+GCB as purifiers were 74%-113%, with RSDs of 1.07%-11.74%, which all met the standard requirements. However, the addition of GCB can make the extract clearer and provide better protection for the instrument.

[0048] Taking into account the recovery rate, experimental cost and protection of the instrument, the seven compounds were extracted with 2% acetic acid-acetonitrile and then 50 mg C18 + 5 mg GCB were used as purifiers.

[0049] Method validation: As shown in Table 2, after optimizing the pretreatment conditions, the peak areas and mass concentrations of the seven compounds in acetonitrile, fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaf matrix had a good linear relationship within the linear range of 0.001-1.0 mg / L, and the correlation coefficient (R 2 ) were all greater than 0.99. There were certain differences in the matrix effects of the seven compounds in fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, chili sauce and pepper leaves. Among them, Me of TMX, CLO, DM-CLO, DN-TMX, DN-CLO and TMX-urea in 6 matrices was less than 0.8, showing a matrix inhibition effect; Me of DM-TMX in fresh peppers, wet pickled peppers, pickled peppers and pepper leaves was greater than 1.2, showing a matrix enhancement effect, and Me in dry pickled peppers and chili sauce was between 0.8 and 1.2, which was a negligible weak matrix effect. In order to reduce the influence of matrix effect on the accuracy of the method, matrix matching standard curve quantification was used to eliminate the influence of matrix effect to ensure the accuracy of the analysis results.

[0050] As shown in Table 2, in the six matrices of fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaves, the average recoveries of the seven compounds at the 0.01 mg / kg addition level were 74%-103%, and the RSDs were 1.15%-9.53%; the average recoveries at the 0.1 mg / kg addition level were 72%-101%, and the RSDs were 0.39%-6.74%; the average recoveries at the 1.0 mg / kg addition level were 71%-99%, and the RSDs were 0.62%-6.98%; the average recoveries at the 10.0 mg / kg addition level were 74%-107%, and the RSDs were 1.10%-7.99%, all of which met the requirements for pesticide residue analysis. According to EU regulations, the method limit of quantification (LOQ) is defined as the lowest added concentration level that meets the requirements of recovery and relative standard deviation. The LOQ of the seven compounds in six matrices, including fresh peppers, dried pickled peppers, wet pickled peppers, pickled peppers, chili sauce and pepper leaves, is 0.01 mg / kg.

[0051] Table 2: Regression equations and correlation coefficients (R) of 7 compounds in different matrices 2 ), matrix effect (Me), average recovery (%) and relative standard deviation (RSD%)

[0052]

[0053] The present invention optimizes the extraction and purification conditions, uses 2% acetic acid-acetonitrile extraction for fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaves, C18 and GCB purification, and uses UHPLC-MS / MS matrix external standard method to quantify thiamethoxam and its 6 metabolites, and establishes a method for simultaneously detecting thiamethoxam and its 6 metabolites in 6 matrices of fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaves. The linear correlation coefficients of the standard curves of all compounds in different matrices are all above 0.99, the average addition recovery rate is 71%-107%, the relative standard deviation is less than 9.53%, and the method quantitative limit is 0.01 mg / kg. The results show that the method can meet the requirements of residue analysis and can provide a method for the simultaneous detection of thiamethoxam and its 6 metabolites in fresh pepper, dry pickled pepper, wet pickled pepper, pickled pepper, chili sauce and pepper leaves.

Claims

1. A method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products, characterized in that: The method comprises the following steps: 1) Extraction of samples: The pre-treated peppers and pepper products were shaken for the first time for 20 minutes using 2% acetic acid-acetonitrile, and then NaCl and anhydrous magnesium sulfate were added and shaken again for 5 minutes, and centrifuged at 5000 r / min for 5 minutes, and the upper organic phase solution was collected; 2) Purification of the sample to be tested: Add the organic phase solution in step 1) to the 18 +GCB+anhydrous magnesium sulfate in a 2 mL centrifuge tube, vortex for 1 min, centrifuge at 5000 r / min for 5 min, take the supernatant and filter it through a 0.22 μm filter membrane into a sample injection bottle to obtain the test solution; 3) Detection: Using UPLC HSS T3 as the chromatographic column, ultra-high performance liquid chromatography-mass spectrometry was used to quantify the residues of 7 target compounds, including thiamethoxam and its 6 metabolites, in 6 matrices.

2. The method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to claim 1, characterized in that In step 1), the peppers and pepper products are fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers, pepper sauce and pepper leaves. The pretreatment is to weigh 5g of fresh peppers, dry pickled peppers, wet pickled peppers, pickled peppers and spicy peppers and add 2mL of pure water after homogenization, and weigh 2g of pepper leaves directly without adding water.

3. The method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to claim 1, characterized in that In step 1), the amount of acetic acid-acetonitrile added is 10 mL, the amount of NaCl added is 1 g, and the amount of anhydrous magnesium sulfate added is 4 g.

4. The method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to claim 1, characterized in that In step 2), C 18 The amount of addition is 50 mg, the amount of addition of GCB is 5 mg, and the amount of addition of anhydrous magnesium sulfate is 150 mg.

5. The method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to claim 1, characterized in that In step 3), the chromatographic conditions of the ultra performance liquid chromatography-mass spectrometry are as follows: chromatographic column: UPLC HSS T3; column temperature 40°C; injection volume 5 μL; mobile phase A is 0.1% formic acid water, and mobile phase B is acetonitrile; flow rate 0.25 mL / min, and the gradient elution program is: 0-1.5 min, 95% A; 1.5-3.0 min, 95%-5% A; 3-3.5 min, 5% A; 3.5-5.0 min, 5%-95% A; 5.0-6.0 min, 95% A.

6. The method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to claim 1, characterized in that In step 3), the mass spectrometry conditions of the ultra-high performance liquid chromatography-mass spectrometry instrument are as follows: ionization source mode: electrospray ionization ESI; ion source polarity: ESI+; nebulizer gas: nitrogen; sheath gas flow rate 12 L / min, auxiliary gas flow rate 3 L / min; collision gas: argon; spray voltage: 3500 V; capillary temperature: 350° C.; ion transfer tube temperature: 300° C.

7. The method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to claim 1, characterized in that Thiamethoxam and its six metabolites are: thiamethoxam, clothianidin, demethylthiamethoxam, demethylthiamethoxam, thiamethoxam guanidine, clothianidin guanidine and thiamethoxam urea.

8. A method for detecting the residues of thiamethoxam and its six metabolites in peppers and pepper products according to any one of claims 1 to 7, characterized in that: The seven target compounds were quantified using the corresponding matrix-matched standard solution curves.

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