A method for simultaneously determining the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals

Abstract

The present invention provides a method for simultaneously detecting the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals, which belongs to the technical field of pesticide residue detection. First, the cereal sample is soaked in a formic acid aqueous solution and extracted with acetonitrile; secondly, a hydrophilic lipophilic polymer solid phase extraction column is used for purification, methanol and 0.5% ammonia methanol (v / v) are eluted, and the eluent is concentrated and fixed to volume; finally, it is detected by high performance liquid chromatography-tandem mass spectrometry. The present invention has the advantages of being sensitive and accurate, and can simultaneously detect fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals, effectively improving detection efficiency and reducing detection cost.

Description

Technical Field

[0001] The invention belongs to the technical field of pesticide residue detection and relates to a method for simultaneously detecting the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals. Background Art

[0002] Fenpyrazone, bispyribac, cypermethrin, and fenpyrazone are herbicides that inhibit p-hydroxyphenylpyruvate dioxidase (PHD) activity. They work by inhibiting PHD activity, affecting carotenoid biosynthesis, causing bleaching in newly grown weed tissue and ultimately killing the weeds. These herbicides have seen rapid market growth due to their high efficacy, crop safety, unique mechanism of action, and resistance to resistance. With the widespread use of PHD herbicides, their residues have become a concern. GB 2763-2021, "Maximum Residue Limits of Pesticides in Foods," and GB 2763.1-2022, "Maximum Residue Limits of 112 Pesticides, Including 2,4-D Butyrate Sodium Salt, in Foods," stipulate the residue limits for fenpyrazone, bispyribac, cypermethrin, and fenpyrazone in cereals.

[0003] Currently, there are no standardized testing methods for fenpyrazone, bispyribac, cyprodinil, and fenpyrazone; research literature only covers the testing of individual herbicides. Existing testing standards and research fail to meet regulatory requirements for fenpyrazone, bispyribac, cyprodinil, and fenpyrazone residues in cereals. Therefore, establishing a method for the simultaneous detection of fenpyrazone, bispyribac, cyprodinil, and fenpyrazone residues in cereals is crucial for achieving efficient regulation of fenpyrazone, bispyribac, cyprodinil, and fenpyrazone residues and ensuring food safety. Summary of the Invention

[0004] The purpose of the present invention is to provide a method for simultaneously detecting the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals, so as to overcome the shortcomings of the prior art.

[0005] The technical solution adopted in the present invention is:

[0006] A method for simultaneously detecting the residues of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone in cereals includes: first, soaking the cereal sample in a formic acid aqueous solution and extracting it with acetonitrile; second, cleanup using a hydrophilic-lipophilic polymer solid-phase extraction column, eluting with methanol and 0.5% ammonia methanol (v / v), and concentrating the eluate to a fixed volume; finally, detection by high-performance liquid chromatography-tandem mass spectrometry. The specific steps are as follows:

[0007] (1) Sample preparation: Take the grain sample to be tested, crush it so that it can pass through a 425 μm standard mesh sieve, put it into a polyethylene bottle, and store it at -18°C.

[0008] (2) Extraction: Weigh 5.00 g of grain sample into a 50 mL plastic centrifuge tube, add 10 mL of 3% formic acid aqueous solution (v / v), mix well, and let stand for 30 min; add 10 mL of acetonitrile, and extract at 300 r / min for 30 min; add 6 g of anhydrous magnesium sulfate and 1.5 g of sodium chloride, shake vigorously for 1 min, and centrifuge at 10,000 r / min for 3 min; aspirate 4 mL of supernatant and blow nitrogen to near dryness; add 4 mL of 50% methanol solution (containing 1.5% formic acid, v / v), vortex to dissolve the residue, and wait for purification.

[0009] (3) Purification: Activate the balanced hydrophilic and lipophilic polymer solid phase extraction column with 5 mL of methanol and 5 mL of water; add the sample solution to be purified in step (2) to the solid phase extraction column and discard the effluent; elute the solid phase extraction column with 5 mL of 40% methanol solution (containing 1.5% formic acid, v / v) and discard the eluent; first add 5 mL of methanol to elute, collect the eluate, blow nitrogen to nearly dryness, add 1 mL of methanol, and vortex to dissolve the residue; then add 5 mL of 0.5% ammonia methanol (v / v) to elute, collect the eluate, blow nitrogen to nearly dryness, add 1 mL of 2% formic acid methanol (v / v), and vortex to dissolve the residue; combine the reconstituted solutions, vortex to mix, and filter to obtain the sample solution to be tested.

[0010] (4) Determination

[0011] ① Qualitative Analysis: Analyze the sample and matrix-matched mixed standard working solution using liquid chromatography-tandem mass spectrometry. Record the chromatographic retention time of the target compound in both the sample and matrix-matched mixed standard working solution. The relative abundance of the qualitative ion pair is expressed as a percentage of the most intense ion abundance. Record the relative ion abundance of the target compound in the sample and matrix-matched mixed standard working solution at comparable concentrations. The target compound is confirmed to be present in the sample when a chromatographic peak with a retention time identical to that of the target compound in the matrix-matched mixed standard working solution is detected (within ±2.5%), and the tolerance for the relative ion abundance is within the range specified in Table 1.

[0012] Table 1 Maximum allowable deviation of relative ion abundance for qualitative confirmation

[0013] Relative abundance (%) >50％ 20％～50％ 10％～20％ ≤10％ Maximum allowable deviation (%) ±20 ±25 ±30 ±50

[0014] The liquid chromatography-tandem mass spectrometry detection conditions are:

[0015] (I) Chromatographic conditions

[0016] a) Chromatographic column: ACQUITY UPLC BEH C18 (2.1 mm × 50 mm, 1.7 μm);

[0017] b) Mobile phase: A: 0.2% formic acid in water (v / v), B: methanol, gradient elution;

[0018] c) Flow rate: 0.3 mL / min;

[0019] d) Column temperature: 40°C;

[0020] e) Injection volume: 2 μL;

[0021] The gradient elution program is shown in Table 2.

[0022] Table 2 Gradient elution program

[0023]

[0024]

[0025] (II) Mass spectrometry conditions

[0026] a) Ion source: electrospray ionization (ESI);

[0027] b) Detection method: multiple reaction monitoring (MRM);

[0028] c) Scanning mode: positive ion mode;

[0029] d) Ionization voltage: 5500 V;

[0030] e) Ion source temperature: 350°C;

[0031] f) Curtain air pressure: 35psi;

[0032] g) Spray gas pressure: 55psi;

[0033] h) Auxiliary gas pressure: 60psi

[0034] i) Other mass spectrometry parameters are shown in Table 3.

[0035] Table 3 Quantitative ion pairs, qualitative ion pairs and mass spectrometry parameters of four herbicides *Quantitative ion

[0036] The steps for preparing the matrix matching mixed standard working solution are as follows:

[0037] (I) Standard stock solution: Accurately weigh a certain amount of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone standards, dissolve them in acetonitrile, and transfer them to a 10 mL volumetric flask to make up the volume to prepare the standard stock solution.

[0038] (II) Mixed standard solution: Pipette appropriate amount of the above standard stock solutions into 50 mL volumetric flasks, dilute to volume with acetonitrile, and prepare a mixed standard solution with a mass concentration of 1 mg / L for fenpyrazone, cypermethrin, cypermethrin and fenpyrazone.

[0039] (III) Matrix-matched mixed standard working solution: Accurately pipette a certain amount of mixed standard solution and dilute it with blank sample solution to make matrix-matched mixed standard working solutions with mass concentrations of 3.0 ng / mL, 10.0 ng / mL, 30.0 ng / mL, 50.0 ng / mL, 100.0 ng / mL, and 150.0 ng / mL.

[0040] The blank sample solution refers to the sample solution obtained after the sample without fenpyrazone, cypermethrin, cypermethrin and fenpyrazone is treated in steps (2) and (3).

[0041] ②Quantitative determination:

[0042] Analyze the matrix-matched mixed standard working solutions using liquid chromatography-tandem mass spectrometry to obtain the corresponding chromatographic peak areas of the standard solutions. Plot a standard curve using the concentration of the matrix-matched mixed standard working solution as the abscissa and the peak area as the ordinate.

[0043] The content of the target compound in the sample solution was determined using the standard curve external standard method. The content of the target compound in the sample was calculated according to formula (1).

[0044]

[0045] Where: X represents the content of the target substance in the sample, mg / kg; C represents the mass concentration of the target substance in the sample solution read from the standard curve, ng / mL; V1 represents the volume of the extract, mL; V2 represents the volume of the extract taken for nitrogen blowdown, mL; V3 represents the constant volume, mL; m represents the sample mass, g.

[0046] The method for detecting the residual amounts of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone in cereals, disclosed herein, utilizes solid-phase extraction pretreatment technology in combination with liquid chromatography-tandem mass spectrometry detection technology to simultaneously detect fenpyrazone, bispyribac, cypermethrin, and fenpyrazone in cereals, effectively improving detection efficiency and reducing detection costs. This method is sensitive and accurate.

[0047] The method for detecting the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals described in the present invention can meet the detection requirements for the residue limits of these four herbicides in cereals specified in GB 2763-2021 and GB 2763.1-2022, and can provide strong technical support for efficient supervision of pesticide residues and ensuring food safety. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 The MRM chromatogram of a blank wheat sample that does not contain fenpyrazone, bispyribac, cypermethrin, and fenpyrazone; wherein (a) is the quantification ion of fenpyrazone, (b) is the quantification ion of bispyribac, (c) is the quantification ion of cypermethrin, and (d) is the quantification ion of fenpyrazone;

[0049] Figure 2 The MRM chromatograms of wheat matrix standard solutions containing 10 ng / mL of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone are shown; wherein (a) is the quantification ion of fenpyrazone, (b) is the quantification ion of bispyribac, (c) is the quantification ion of cypermethrin, and (d) is the quantification ion of fenpyrazone;

[0050] Figure 3 The MRM chromatogram of a corn blank sample that does not contain fenpyrazone, bispyribac, cypermethrin, and fenpyrazone; wherein (a) is the quantification ion of fenpyrazone, (b) is the quantification ion of bispyribac, (c) is the quantification ion of cypermethrin, and (d) is the quantification ion of fenpyrazone;

[0051] Figure 4 The MRM chromatograms of corn matrix standard solutions containing 10 ng / mL of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone are shown; wherein (a) is the quantification ion of fenpyrazone, (b) is the quantification ion of bispyribac, (c) is the quantification ion of cypermethrin, and (d) is the quantification ion of fenpyrazone;

[0052] Figure 5 The MRM chromatogram of a blank sample of brown rice that does not contain fenpyrazone, bispyribac, cypermethrin, and fenpyrazone is shown; wherein, (a) is the fenpyrazone quantitative ion, (b) is the bispyribac quantitative ion, (c) is the cypermethrin quantitative ion, and (d) is the fenpyrazone quantitative ion;

[0053] Figure 6 The MRM chromatograms of the brown rice matrix standard solution with 10 ng / mL of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone; wherein (a) is the fenpyrazone quantitative ion, (b) is the bispyribac quantitative ion, (c) is the cypermethrin quantitative ion, and (d) is the fenpyrazone quantitative ion; DETAILED DESCRIPTION

[0054] The instruments and reagents used in the following embodiments of the present invention include:

[0055] AB SCIEX QTRAP 6500 liquid chromatography-mass spectrometry instrument; HS 501 digital display shaker; CR22GⅡ high-speed centrifuge; XW-80A vortex mixer; pro ultrapure water system; acetonitrile (chromatographic grade); methanol (chromatographic grade); formic acid (chromatographic grade); formic acid, ammonia water, sodium chloride, anhydrous magnesium sulfate (analytical grade); HLB solid-phase extraction column (200 mg, 6 mL); bispyribac (99.7%, mass fraction); cypermethrin (1000 μg / mL); fenpyrad-butyl (98.4%, mass fraction); fenpyrad-butyl (100 μg / mL).

[0056] Example 1: Determination of residues of fenpyrazone, cypermethrin, cypermethrin and fenpyrazone in wheat

[0057] (1) Sample preparation: Take the wheat sample to be tested, crush it so that it can pass through a 425 μm standard mesh sieve, put it into a polyethylene bottle, and store it at -18°C.

[0058] (2) Extraction: Weigh 5.00 g of wheat sample into a 50 mL plastic centrifuge tube, add 10 mL of 3% formic acid aqueous solution (v / v), mix well, and let stand for 30 min; add 10 mL of acetonitrile, and extract at 300 r / min for 30 min; add 6 g of anhydrous magnesium sulfate and 1.5 g of sodium chloride, shake vigorously for 1 min, and centrifuge at 10,000 r / min for 3 min; draw 4 mL of supernatant and blow nitrogen to dryness; add 4 mL of 50% methanol solution (containing 1.5% formic acid, v / v), vortex to dissolve the residue, and wait for purification.

[0059] (3) Purification: Activate the balanced hydrophilic and lipophilic polymer solid phase extraction column with 5 mL of methanol and 5 mL of water; add the sample solution to be purified in step (2) to the solid phase extraction column and discard the effluent; elute the solid phase extraction column with 5 mL of 40% methanol solution (containing 1.5% formic acid, v / v) and discard the eluent; first add 5 mL of methanol to elute, collect the eluate, blow nitrogen to nearly dryness, add 1 mL of methanol, and vortex to dissolve the residue; then add 5 mL of 0.5% ammonia methanol (v / v) to elute, collect the eluate, blow nitrogen to nearly dryness, add 1 mL of 2% formic acid methanol (v / v), and vortex to dissolve the residue; combine the reconstituted solutions, vortex to mix, and filter to obtain the sample solution to be tested.

[0060] (4) Preparation of standard solution

[0061] ① Standard stock solution: Accurately weigh a certain amount of bispyribac-methyl and fenpyrazone standard respectively, dissolve them in acetonitrile, transfer to a 10 mL volumetric flask and make up to volume to prepare the standard stock solution.

[0062] ②Mixed standard solution (1 mg / L): Pipette appropriate amounts of the above standard stock solution and cypermethrin and fenpyrazone standards into a 50 mL volumetric flask, dilute to volume with acetonitrile, and prepare a mixed standard solution with a mass concentration of fenpyrazone, bispyribac, cypermethrin and fenpyrazone of 1 mg / L.

[0063] ③ Blank sample solution: Take a wheat sample that does not contain fenpyrazone, cypermethrin, cypermethrin and fenpyrazone, and treat it according to steps (2) and (3) to obtain a blank sample solution. The MRM chromatogram of the wheat blank sample is shown in Figure 1 .

[0064] ④ Matrix-matched mixed standard working solution: Accurately pipette a certain amount of mixed standard solution and dilute it with blank sample solution to make matrix-matched mixed standard working solutions with mass concentrations of 3.0 ng / mL, 10.0 ng / mL, 30.0 ng / mL, 50.0 ng / mL, 100.0 ng / mL and 150.0 ng / mL. The MRM chromatograms of wheat matrix standard solutions of 10 ng / mL of fenpyrazone, cypermethrin, cyproconazole and fenpyrazone are shown in Figure 2. Figure 2 .

[0065] (5) Liquid chromatography-mass spectrometry / mass spectrometry (HPLC-MS / MS) determination

[0066] The matrix-matched mixed standard working solution, wheat blank sample solution and sample solution were injected into HPLC-MS / MS for determination, and the matrix-matched calibration curve was established. The external standard method was used for quantification.

[0067] ①Chromatographic conditions are:

[0068] Chromatographic column: ACQUITY UPLC BEH C18 (2.1 mm × 50 mm, 1.7 μm);

[0069] Mobile phase: A: 0.2% formic acid in water (v / v), B: methanol, gradient elution;

[0070] Flow rate: 0.3 mL / min;

[0071] Column temperature: 40°C;

[0072] Injection volume: 2 μL;

[0073] The gradient elution program is shown in Table 4.

[0074] Table 4 Gradient elution program

[0075] Time / min Mobile phase A / % Mobile phase B / % 0 80 20 0.5 80 20 3.5 35 65 5 5 95 5.1 80 20 7 80 20

[0076] ②Mass spectrometry conditions are:

[0077] Ion source: electrospray ionization (ESI);

[0078] Detection method: multiple reaction monitoring (MRM);

[0079] Scanning mode: positive ion mode;

[0080] Ionization voltage: 5500V;

[0081] Ion source temperature: 350°C;

[0082] Air curtain pressure: 35psi;

[0083] Spray gas pressure: 55psi;

[0084] Auxiliary gas pressure: 60psi

[0085] Other mass spectrometry parameters are shown in Table 5.

[0086] Table 5 Quantitative ion pairs, qualitative ion pairs and mass spectrometry parameters of four herbicides *Quantitative ion

[0087] The obtained calibration curve is shown in Table 6.

[0088] Table 6 Wheat matrix matching calibration curves for four herbicides

[0089] herbicide Regression equation Linear range (ng / mL) Correlation coefficient (r) Fenpyrazone y=28124.33335x+7449.43655 3～150 0.9999 Bifenazolin y=27274.28809x+5700.82915 3～150 0.9994 Cyclopyrazone y=27964.39481x-6224.16680 3～150 0.9996 Benpyrazone y=21432.08163x-8515.38954 3～150 0.9998

[0090] (6) Spike recovery and repeatability

[0091] Wheat samples free of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone were spiked with mixed standard solutions at three levels: 0.01 mg / kg, 0.03 mg / kg, and 0.1 mg / kg. After 30 minutes of spiking, the contents were determined according to the above procedure. Six replicates were run for each spike level. The results were compared with the theoretical spike concentrations to obtain the average recoveries and relative standard deviations (RSDs). The results are shown in Table 7.

[0092] Table 7 Recovery and precision of four herbicides in wheat (n=6)

[0093]

[0094] As shown in Table 7, the average recoveries of fenpyrazone, cypermethrin, cypermethrin, and fenpyrazone at the three spiked levels ranged from 73.9% to 87.8%, and the relative standard deviations ranged from 2.2% to 10.0%, meeting the requirements for recovery and precision in Announcement No. 2386 of the Ministry of Agriculture and Rural Affairs, "Guidelines for the Preparation of National Standards for Pesticide Residue Detection Methods."

[0095] (7) Limit of quantification

[0096] The limit of quantification (LOQ) was determined by adding recovery to the blank sample matrix, with the minimum recovery level meeting the methodological requirements of Ministry of Agriculture and Rural Affairs Announcement No. 2386, "Guidelines for the Preparation of National Standards for Pesticide Residue Detection Methods." In this example, the limit of quantification for fenpyrazone, bispyribac, cypermethrin, and fenpyrazone in wheat was 0.01 mg / kg.

[0097] Example 2: Determination of residues of fenpyrazone, cypermethrin, cypermethrin and fenpyrazone in corn

[0098] (1) Take the corn sample to be tested, and the sample preparation, extraction, purification, standard solution preparation and determination methods are the same as in Example 1. The MRM chromatogram of the corn blank sample is shown in Figure 3 The MRM chromatograms of corn matrix standard solutions of 10 ng / mL of fenpyrazone, cypermethrin, cypermethrin and fenpyrazone are shown in Figure 4 The calibration curve obtained is shown in Table 8.

[0099] Table 8 Corn matrix matching calibration curves for four herbicides

[0100] herbicide Regression equation Linear range (ng / mL) Correlation coefficient (r) Fenpyrazone y=28389.42770x-5056.00046 3～150 0.9999 Bifenazolin y=28510.80578x+6544.92285 3～150 0.9991 Cyclopyrazone y=29086.50439x+3933.64789 3～150 0.9997 Benpyrazone y=23896.86810x+1486.73766 3～150 0.9989

[0101] (2) Spike recovery and repeatability

[0102] Corn samples free of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone were spiked with mixed standard solutions at three levels: 0.01 mg / kg, 0.03 mg / kg, and 0.1 mg / kg. After 30 minutes of spiking, the concentrations were determined according to the above procedure. Six replicates were run for each spike level. The results were compared with the theoretical spike concentrations to obtain the average recoveries and relative standard deviations (RSDs). The results are shown in Table 9.

[0103] Table 9 Recovery and precision of four herbicides in corn (n=6)

[0104]

[0105]

[0106] As shown in Table 9, the average recoveries of fenpyrazone, cypermethrin, cypermethrin, and fenpyrazone at the three spiked levels ranged from 75.9% to 87.1%, and the relative standard deviations ranged from 1.6% to 6.5%, meeting the requirements for recovery and precision in Announcement No. 2386 of the Ministry of Agriculture and Rural Affairs, "Guidelines for the Preparation of National Standards for Pesticide Residue Detection Methods."

[0107] (3) Limit of quantification

[0108] The limit of quantification (LOQ) was determined by adding recovery to the blank sample matrix, with the minimum recovery level meeting the methodological requirements of Ministry of Agriculture and Rural Affairs Announcement No. 2386, "Guidelines for the Preparation of National Standards for Pesticide Residue Detection Methods." In this example, the limit of quantification for fenpyrad, bispyribac, cypermethrin, and fenpyrad in corn was 0.01 mg / kg.

[0109] Example 3: Determination of the Residues of Fenpyrazone, Bifenazolin, Cyclopyrazone and Fenpyrazone in Brown Rice

[0110] (1) Take the brown rice sample to be tested, and the sample preparation, extraction, purification, standard solution preparation and determination method are consistent with Example 1. Brown rice blank sample MRM chromatogram is shown in Figure 5 The MRM chromatograms of the brown rice matrix standard solutions of 10 ng / mL of fenpyrazone, cypermethrin, cypermethrin and fenpyrazone are shown in Figure 6 The calibration curve obtained is shown in Table 10.

[0111] Table 10 Brown rice matrix matching calibration curves for four herbicides

[0112] herbicide Regression equation Linear range (ng / mL) Correlation coefficient (r) Fenpyrazone y=26793.94145x-24932.76980 3～150 0.9996 Bifenazolin y=22011.87574x-3210.79438 3～150 0.9996 Cyclopyrazone y=26793.94145x-24932.76980 3～150 0.9997 Benpyrazone y=14963.54985x-7693.55169 3～150 0.9996

[0113] (2) Spike recovery and repeatability

[0114] Brown rice samples free of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone were spiked with mixed standard solutions at three levels: 0.01 mg / kg, 0.03 mg / kg, and 0.1 mg / kg. After 30 minutes of spiking, the contents were determined according to the above procedure. Six replicates were run for each spike level. The results were compared with the theoretical spike concentrations to obtain the average recovery and relative standard deviation (RSD). The results are shown in Table 11.

[0115] Table 11 Recovery and precision of four herbicides in brown rice (n=6)

[0116]

[0117] As shown in Table 11, the average recoveries of fenpyrazone, cypermethrin, cypermethrin, and fenpyrazone at the three spiked levels ranged from 77.1% to 90.8%, and the relative standard deviations ranged from 1.9% to 6.3%, meeting the requirements for recovery and precision in Announcement No. 2386 of the Ministry of Agriculture and Rural Affairs, "Guidelines for the Preparation of National Standards for Pesticide Residue Detection Methods."

[0118] (3) Limit of quantification

[0119] By adding recovery to the blank sample matrix, the limit of quantification was determined to be the minimum level of recovery required to meet the methodological requirements of Ministry of Agriculture and Rural Affairs Announcement No. 2386, "Guidelines for the Preparation of National Standards for Pesticide Residue Detection Methods." In this example, the limit of quantification for fenpyrazone, bispyribac, cypermethrin, and fenpyrazone in brown rice was 0.01 mg / kg.

[0120] The above-described embodiments merely express the implementation methods of the present invention, but should not be understood as limiting the scope of the patent of the present invention. It should be pointed out that for those skilled in the art, several variations and improvements can be made without departing from the concept of the present invention, and these all fall within the scope of protection of the present invention.

Claims

1. A method for simultaneously detecting the residues of fenpyrazone, cypermethrin, pyrasulfoac and fenpyrazone in cereals, characterized in that: First, the grain sample was soaked in a formic acid aqueous solution and extracted with acetonitrile. Second, it was purified using a hydrophilic-lipophilic polymer solid-phase extraction column, eluted with methanol and ammonia methanol, and the eluate was concentrated to a fixed volume. Finally, it was detected by high-performance liquid chromatography-tandem mass spectrometry. The specific steps include the following: (1) Sample preparation: Take the grain sample to be tested, crush it so that it can pass through a 425 μm standard mesh sieve, place it in a polyethylene bottle, and store it at -18°C; (2) Extraction: Weigh 5.00 g of grain sample into a 50 mL plastic centrifuge tube, add 10 mL of 3% formic acid aqueous solution, mix well, and let stand for 30 min; add 10 mL of acetonitrile, and extract at 300 r / min for 30 min; add 6 g of anhydrous magnesium sulfate and 1.5 g of sodium chloride, shake vigorously for 1 min, and centrifuge at 10,000 r / min for 3 min; draw 4 mL of supernatant and blow nitrogen to dryness; add 4 mL of 50% methanol solution, vortex to dissolve the residue, and wait for purification, wherein the methanol solution contains 1.5% formic acid by volume; (3) Purification: Activate the equilibrated solid phase extraction column with 5 mL of methanol and 5 mL of water; add the sample solution to be purified in step (2) to the solid phase extraction column and discard the effluent; elute the solid phase extraction column with 5 mL of 40% methanol solution and discard the eluent, wherein the methanol solution contains 1.5% by volume of formic acid; first add 5 mL of methanol for elution, collect the eluate, blow nitrogen to near dryness, add 1 mL of methanol, and vortex to dissolve the residue; then add 5 mL of eluent ② for elution, collect the eluate, blow nitrogen to near dryness, add 1 mL of 2% formic acid in methanol, and vortex to dissolve the residue; The reconstituted solutions are combined, vortex-mixed, and filtered to obtain the sample solution to be tested; the solid phase extraction column is a hydrophilic and lipophilic polymer solid phase extraction column; (4) Determination Liquid chromatography-tandem mass spectrometry detection conditions are: (I) Chromatographic conditions a) Chromatographic column: ACQUITY UPLC BEH C18, 2.1 mm × 50 mm, 1.7 μm; b) Mobile phase: A: 0.2% formic acid in water, v / v, B: methanol, gradient elution; c) Flow rate: 0.3 mL / min; d) Column temperature: 40°C; e) Injection volume: 2 μL; The gradient elution program is shown in Table 1; Table 1 Gradient elution program (II) Mass spectrometry conditions a) Ion source: electrospray ionization source ESI; b) Detection method: Multiple reaction monitoring (MRM); c) Scanning mode: positive ion mode; d) Ionization voltage: 5500 V; e) Ion source temperature: 350°C; f) Curtain air pressure: 35psi; g) Spray gas pressure: 55psi; h) Auxiliary gas pressure: 60psi i) Other mass spectrometry parameters are shown in Table 2; Table 2 Quantitative ion pairs, qualitative ion pairs and mass spectrometry parameters of four herbicides *Quantitative ion.

2. The method for simultaneously detecting the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals according to claim 1, characterized in that: Step (4) includes: ① Qualitative analysis: Use liquid chromatography-tandem mass spectrometry to detect the sample and matrix-matched mixed standard working solution. Record the chromatographic retention time of the target in the sample and matrix-matched mixed standard working solution. The relative abundance of the qualitative ion pair is expressed as a percentage of the strongest ion abundance. Record the relative ion abundance of the target in the sample and matrix-matched mixed standard working solution of equivalent concentrations. When a chromatographic peak with the same retention time as the target peak in the matrix-matched mixed standard working solution is detected in the sample, the variation range is within ±2.5%, and the allowable deviation of the relative ion abundance does not exceed the range specified in Table 3, the target can be confirmed to be detected in the sample. Table 3 Maximum allowable deviation of relative ion abundance for qualitative confirmation The steps for preparing the matrix matching mixed standard working solution are as follows: (I) Standard stock solution: Accurately weigh a certain amount of fenpyrazone, bispyribac, cypermethrin, and fenpyrazone standards, dissolve them in acetonitrile, and transfer them to a 10 mL volumetric flask to make up to volume to prepare the standard stock solution; (II) Mixed standard solution: pipette appropriate amounts of the above standard stock solutions into 50 mL volumetric flasks, dilute to volume with acetonitrile, and prepare a mixed standard solution with a concentration of 1 mg / L for each of fenpyrazone, cypermethrin, cypermethrin, and fenpyrazone; (III) Matrix-matched mixed standard working solution: Accurately pipette a certain amount of mixed standard solution and dilute it with blank sample solution to make matrix-matched mixed standard working solutions of 3.0 ng / mL, 10.0 ng / mL, 30.0 ng / mL, 50.0 ng / mL, 100.0 ng / mL, and 150.0 ng / mL; The blank sample solution refers to the sample solution obtained after the sample without fenpyrazone, bispyribac, cypermethrin and fenpyrazone is treated in steps (2) and (3); ②Quantitative determination: The matrix-matched mixed standard working solutions were measured using a liquid chromatography-tandem mass spectrometer to obtain the chromatographic peak areas of the corresponding standard solutions; a standard curve was drawn with the concentration of the matrix-matched mixed standard working solution as the abscissa and the peak area of ​​the chromatographic peak as the ordinate; The content of the target substance in the sample solution is determined by the standard curve external standard method; the content of the target substance in the sample is calculated according to formula (1); Where: X represents the content of the target substance in the sample, mg / kg; C represents the mass concentration of the target substance in the sample solution read from the standard curve, ng / mL; V1 represents the volume of the extract, mL; V2 represents the volume of the extract taken for nitrogen blowdown, mL; V3 represents the constant volume, mL; m represents the sample mass, g.

3. The method for simultaneously detecting the residues of fenpyrazone, bispyribac, cypermethrin and fenpyrazone in cereals according to claim 1, characterized in that: In the step (3), the eluent ② is ammonia methanol with a volume fraction of 0.5%.

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