Method for detecting content of quinclorac in grain

Abstract

The application discloses a method for detecting the content of quinclorac in grain, and belongs to the technical field of food safety detection. The detection method comprises the following steps: (1) crushing a sample to be detected, adding anhydrous ethanol to perform ultrasonic extraction, and centrifuging to obtain an extraction liquid; (2) concentrating the extraction liquid to less than 1 mL; (3) adding a purifying agent to the concentrated liquid, vortexing, and centrifuging to obtain a purified extraction liquid; (4) detecting the purified extraction liquid by adopting an Rxi-5Sil MS or VF-1701ms capillary column in combination with GC-NCI-MS selected ion mode, and quantitatively calculating the content of quinclorac by adopting an internal standard method. The method is simple in operation, green and environment-friendly, accurate in qualitative and quantitative determination, and high in sensitivity, can be used for determining quinclorac in grain, and meets the detection requirements.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of food safety detection, and particularly relates to a method for detecting the content of quinclorac in grain. BACKGROUND

[0002] Various pesticides have played a positive role in controlling weeds, preventing and treating pests and diseases, and improving crop yield, and have made outstanding contributions to the rapid development of agriculture and animal husbandry. However, the problem of pesticide residues that has followed has attracted widespread attention, especially the problem of pesticide residues caused by environmental pollution and non-standard use of pesticides. On the one hand, it has an adverse effect on physical health, and on the other hand, it causes huge losses in international trade.

[0003] Herbicides have been applied for several decades, and some have developed resistance. Quinclorac, as a new herbicide, related studies have shown that quinclorac has high biological activity on barnyard grass, crabgrass, earthworm, Chinese violet, isotype sedge, and broken rice sedge, and has a certain inhibitory effect on alligator grass and ear leaf water sedge, and can be applied to rice fields to prevent and control various grasses, broadleaf weeds and sedge weeds.

[0004] In March 2021, the National Health Commission, the Ministry of Agriculture and Rural Affairs and the General Administration of Market Supervision jointly announced the Food Safety National Standard Maximum Residue Limits of Pesticides in Food (GB 2763-2021) (hereinafter referred to as the 2021 version of GB 2763). Compared with the 2019 version, the 2021 version of GB 2763 adds 81 new pesticide varieties, including quinclorac. Therefore, it is urgent to develop a method for detecting the content of quinclorac in food.

[0005] At present, the extraction reagents used by those skilled in the art to detect pesticide residues include methanol, acetonitrile, n-hexane, dichloromethane, acetone, ethyl acetate, etc. Ethanol, as an organic solvent with good environmental compatibility, is rarely used in pesticide residue detection methods. SUMMARY

[0006] In order to solve the problems in the background art, the application provides a method for determining the content of quinclorac in grain by applying ethanol as an extraction solvent and combining gas chromatography-mass spectrometry with negative chemical source method. The method is used to determine the content of quinclorac in grain, and the quantitative limit can reach 1.5 μg / kg. After further concentration of the sample and internal insertion of the sample, the quantitative limit can be as low as 0.15 μg / kg. Through retrieval, it is rare to report the application of ethanol as an extraction solvent and the combination of gas chromatography-mass spectrometry with negative chemical source method for detecting quinclorac.

[0007] In order to achieve the above purpose, the application adopts the following technical solutions:

[0008] A method for determining quinclorac in grain, comprising the following steps:

[0009] (1) crushing the sample to be tested, adding an extraction solvent for ultrasonic extraction, and centrifuging to obtain an extract;

[0010] (2) concentrating the extract to less than 1 mL;

[0011] (3) adding a purification agent to the concentrated solution, vortexing, and centrifuging to obtain a purified extract;

[0012] (4) detecting the purified extract by capillary column combined with GC-NCI-MS in selected ion mode, and quantitatively calculating the quinclorac content by internal standard method.

[0013] The extraction solvent in step (1) is selected from one or more of the following: methanol, anhydrous ethanol, ethyl acetate, acetonitrile, acetone, and dichloromethane.

[0014] Preferably, the extraction solvent is anhydrous ethanol, the amount of anhydrous ethanol added is 4-10 mL / g of sample, the extraction temperature is room temperature-40℃, and the ultrasonic extraction time is 20-40 min.

[0015] The concentration method in step (2) is parallel concentration, and the water temperature is 30-45℃.

[0016] The purification agent used in step (3) is C 18 and PSA, and the amount used is 4-10 mg / g of sample.

[0017] The GC-NCI-MS determination conditions in step (4) are as follows:

[0018] Inlet temperature: 280-300℃; no split high pressure injection; constant linear velocity mode, linear velocity 30-50 cm / sec;

[0019] Column oven temperature program: 40-80℃ for 1-5 min, increased to 280℃ at 20-30℃ / min; maintained for 0-10 min; then increased to 300℃ at 20-30℃ / min, maintained for 5-10 min;

[0020] The capillary column is Rxi-5Sil MS or VF-1701ms, 30 m x 0.25 mm x 0.25 μm;

[0021] Solvent delay time: 4 min, ion source temperature 200℃, interface temperature 280℃; ionization mode: NCI mode.

[0022] The internal standard method of step (4) is to add the same concentration of epoxy heptachlor in the standard solution and the to-be-measured solution, and then calculate the content of quinclorac in the to-be-measured sample according to the peak area of epoxy heptachlor in the to-be-measured solution and the standard solution, and the formula is:

[0023]

[0024] In the formula:

[0025] X is the content of quinclorac in the sample, in mg / kg;

[0026] c is the concentration of quinclorac in the standard solution, in μg / mL;

[0027] A 样 is the peak area of quinclorac in the sample solution;

[0028] A 标准 is the peak area of quinclorac in the standard solution;

[0029] a is the peak area of epoxy heptachlor in the standard solution;

[0030] b is the peak area of epoxy heptachlor in the sample solution;

[0031] V is the final constant volume of the sample solution, in mL;

[0032] m is the mass of the sample, in g.

[0033] The standard solution is prepared by the following method:

[0034] (1) Preparation of standard intermediate solution: accurately take 1.00 mL of standard solution (100 μg / mL) into a 100 mL volumetric flask, and then add dehydrated ethanol to the scale to prepare an intermediate solution with a concentration of 1 μg / mL;

[0035] (2) Preparation of standard use solution: accurately take a certain amount of intermediate solution, and then add dehydrated ethanol to 2 mL to prepare a standard use solution, and then add 100 μL of epoxy heptachlor internal standard solution to 1 mL of standard solution before sampling and analysis, and then mix and filter the membrane for measurement.

[0036] Further, the method also comprises preparation of a blank sample, and the extraction and determination steps are the same as those of the sample determination, except that the sample is not added to the centrifugal tube.

[0037] The grain of the present application includes but is not limited to rice, sorghum, wheat, and soybean.

[0038] Compared with the prior art, the present application has the following beneficial effects: the present application first uses anhydrous ethanol as a pesticide residue extraction solvent, which is friendly to experimental personnel and has excellent environmental compatibility; and the quantitative limit of the gas chromatography-mass spectrometry negative chemical ion source technology can reach 0.15 mu g / kg, and the sensitivity is high. BRIEF DESCRIPTION OF DRAWINGS

[0039] Figure 1 Rxi-5Sil MS capillary column separation effect diagram of example 1.

[0040] Figure 2 Recovery rate statistical diagram after extraction and detection of different extraction solvents in example 2.

[0041] Figure 3 Rxi-5Sil MS capillary column separation effect diagram of example 2.

[0042] Figure 4 VF-1701ms capillary column separation effect diagram of example 3. DETAILED DESCRIPTION

[0043] The technical solutions in the embodiments of the present application will be described below in a clear and complete manner. Obviously, the described embodiments are only some of the embodiments of the present application, rather than all. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0044] Example 1

[0045] The rice sample to be tested was crushed, and 5.00 g was accurately weighed and placed in a centrifuge tube. 20 mL of anhydrous ethanol extraction solvent was added for ultrasonic extraction, and the ultrasonic time was 20 min. After centrifugation at 8000 r / min for 5 min, the supernatant was transferred to a concentration cup, the residue was repeatedly extracted once with 10 mL of anhydrous ethanol, and the two extraction solutions were combined. 40 DEG C was parallelly concentrated to less than 1 mL, and was transferred to a centrifuge tube added with 20 mg of C 18 and 20 mg of PSA purifier, vortexed for 2 min, and after centrifugation, the supernatant was diluted to 1 mL with anhydrous ethanol, 100 mu L of an epoxy heptachlor internal standard solution was added, and after mixing and filtering the membrane, it was tested.

[0046] The instrument determination conditions are as follows:

[0047] The injection port temperature was 280 DEG C, the non-split high pressure injection was used, the constant linear velocity mode was used, the linear velocity was 40 cm / sec, the column oven temperature rising program was that 70 DEG C was kept for 1 min, and then increased to 300 DEG C at the rate of 30 DEG C / min, and kept for 5 min. The solvent delay time was 4 min, the ion source temperature was 200 DEG C, the interface temperature was 280 DEG C, the ionization mode was NCI mode, and the detection ions are shown in Table 1.

[0048] Table 1 target retention time and SIM parameters

[0049] SEQ ID NO: Compound Name Retention Time (min) Quant Ion Qual Ion 1 Qual Ion 2 Qual Ion 3 1 Epoxychloridone 8.074 354 356 352 358 2 Dichloroquinolinone 10.996 291 293 138 112

[0050] Example 2

[0051] The sample to be tested was crushed, 5.00 g was accurately weighed and placed in a centrifuge tube, 6 identical operation groups were set, 30 mL of methanol, anhydrous ethanol, ethyl acetate, acetonitrile, acetone and dichloromethane were added as extraction solvents for ultrasonic extraction, the ultrasonic time was 40 min, and after centrifugation at 10000 r / min for 5 min, the supernatant was moved into a concentration cup, the residue was repeatedly extracted once with 10 mL of the corresponding extraction solvent, and the two extraction liquids were combined. Concentrate at 40°C to less than 1 mL, transfer to a centrifuge tube with 30 mg of C 18 18 and 30 mg of PSA purifier, vortex for 2 min. After centrifugation, the supernatant was diluted to 1 mL with anhydrous ethanol, 100 μL of epoxy heptachlor internal standard solution was added, and the mixed solution was filtered and measured.

[0052] The instrument determination conditions are as follows:

[0053] Inlet temperature: 280℃; no split high pressure injection; constant linear velocity mode, linear velocity 40 cm / sec. Column oven temperature program: 70℃ for 1 min, increased to 300℃ at 30℃ / min, and kept for 5 min. Solvent delay time: 4 min, ion source temperature 200℃, interface temperature 280℃; ionization mode: NCI mode, detection ions as shown in Table 2, and detection results as shown in Table 3. From the figure, it can be seen that the extraction effect is better and the recovery rate is higher when anhydrous ethanol is used as the extraction solvent. Figure 2

[0054] Anhydrous ethanol was selected as the extraction solvent, and the sample was treated with the purifier after extraction according to the above method. After centrifugation, the supernatant was diluted to 1 mL with anhydrous ethanol, 100 μL of epoxy heptachlor internal standard solution was added, and the mixed solution was filtered and measured a ; After centrifugation at 12000 r / min, the supernatant was nearly dried by nitrogen blowing, 200 μL of anhydrous ethanol and 20 μL of epoxy heptachlor internal standard solution were added, and the mixture was mixed and measured b . The method validation data are shown in Table 3.

[0055] Table 2 target retention time and SIM parameters

[0056]

[0057]

[0058] Table 3 method validation data

[0059] ​

[0060] By concentrating the solution to be tested to 200 μL of injection, the sensitivity of the detection can be further improved, and the detection limit of the method can reach 0.046 μg / kg, and the quantification limit of the method reaches 0.15 μg / kg.

[0061] Example 3

[0062] The wheat sample to be tested was crushed, and 5.00 g was accurately weighed into a centrifuge tube, 20 mL of anhydrous ethanol extraction solvent was added for ultrasonic extraction, the ultrasonic time was 40 min, and after centrifugation at 8000 r / min for 5 min, the supernatant was transferred to a concentration cup, the residue was extracted with 10 mL of ethanol once more, and the two extraction solutions were combined. Concentrate at 40°C to less than 1 mL, transfer to a centrifuge tube with 20 mg of C 18 and 20 mg of PSA purifier, vortex for 2 min, and after centrifugation, dilute the supernatant to 1 mL with anhydrous ethanol, add 100 μL of epoxy heptachlor internal standard solution, mix, filter, and then test. The preparation of the blank sample is the same as the sample determination except that the sample is not added to the centrifuge tube.

[0063] The instrument determination conditions are as follows:

[0064] Inlet temperature: 280°C; no split high pressure injection; constant linear velocity mode, linear velocity 45 cm / sec. Column oven temperature program: 40°C for 1 min, increased to 280°C at 30°C / min, maintained for 10 min, increased to 300°C at 20°C / min, maintained for 5 min. Solvent delay time: 4 min, ion source temperature 200°C, interface temperature 280°C; ionization mode: NCI mode, detection ions are shown in Table 4.

[0065] Table 4 Target retention time and SIM parameters

[0066]

[0067]

[0068] As can be seen from the above examples, the applicant has established a method with the advantages of simple operation, green environmental protection, accurate qualitative and quantitative, high sensitivity, etc. for determining quinclorac in grain through optimization of multiple experimental conditions and comparison of results, which meets the detection requirements.

[0069] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present application, and are not intended to limit the present application; although the present application has been described in detail with reference to the above embodiments, those skilled in the art should understand that the technical solutions recorded in the above embodiments can be modified, or some or all of the technical features can be replaced by equivalents; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present application.

Claims

1. A method for determining quinclorac in grains, the method comprising the following steps: (1) The sample to be tested is crushed, anhydrous ethanol is added for ultrasonic extraction, and the extract is obtained by centrifugation; (2) Concentrate the extract to less than 1 mL; (3) Add a purifying agent to the concentrate, vortex, and centrifuge to obtain the purified extract; (4) The purified extract was detected by using a capillary column combined with GC-NCI-MS selected ion mode, and the content of dichloroquinoline was quantitatively calculated by internal standard method; The GC-NCI-MS measurement conditions in step (4) are as follows: Inlet temperature: 280-300℃; splitless high-pressure injection; constant linear velocity mode, linear velocity 30-50cm / sec; Column oven heating program: 40-80℃, hold for 1-5 minutes, then increase to 280℃ at 20-30℃ / min; hold for 0-10 minutes; then increase to 300℃ at 20-30℃ / min and hold for 5-10 minutes; The capillary column is an Rxi-5Sil MS or a VF-1701ms, 30m × 0.25mm × 0.25μm; Solvent delay time: 4 min, ion source temperature 200℃, interface temperature 280℃; ionization mode: NCI mode.

2. The method according to claim 1, characterized in that, The amount of anhydrous ethanol added was 4-10 mL / g of sample, the extraction temperature was room temperature to 40℃, and the ultrasonic extraction time was 20-40 min.

3. The method according to claim 1, characterized in that, The concentration method in step (2) is parallel concentration, and the water temperature is 30-45℃.

4. The method according to claim 1, characterized in that, The purifying agent used in step (3) is C. 18 The dosage of PSA was 4-10 mg / g of the sample.

5. The method according to claim 1, characterized in that, The internal standard method described in step (4) involves adding the same concentration of heptachlor epoxide to both the standard solution and the test solution. Then, based on the peak areas of heptachlor epoxide in the test solution and the standard solution, the content of quinclorac in the sample is calculated using a formula. The formula is: In the formula: The content of quinclorac in sample X, in mg / kg; The concentration of quinclorac in the c-standard solution is expressed in μg / mL. A 样 - The peak area of ​​quinclorac in the test solution; A 标准 - Peak area of ​​quinclorac in standard solution; The peak area of ​​heptachlor epoxide in the standard solution; b - Peak area of ​​heptachlor epoxide in the test solution; V - The final volume of the solution to be tested, in mL; m - the mass of the sample, in grams.

6. The method according to claim 5, characterized in that, The standard solution was prepared by the following method: (1) Preparation of standard intermediate solution: Accurately pipette 1.00 mL of 100 μg / mL standard solution into a 100 mL volumetric flask, dilute to the mark with anhydrous ethanol, mix well, and obtain an intermediate solution with a concentration of 1 μg / mL; (2) Preparation of standard working solution: Accurately pipette an appropriate amount of intermediate solution, dilute to 2 mL with blank matrix solution, mix well to obtain standard working solution. Before injection analysis, add 100 μL of heptachlor epoxide internal standard solution to each 1 mL standard solution, mix well and filter membrane before testing.

7. The method according to claim 1, characterized in that, The method also includes the preparation of blank samples, except that no sample is added to the centrifuge tube, and the other extraction and determination steps are the same as the sample determination.

8. The method according to claim 1, characterized in that, The grains mentioned include, but are not limited to, rice, sorghum, wheat, and soybeans.

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