GB 23200.38-2016Food safety national standard -- Determination of Cyclohexenone Herbicide Residue in Plant-derived Food by Liquid Chromatography-Mass Spectrometry / Mass Spectrometry (English PDF)

4.1 Reagents

4.1.1 Acetonitrile (CH3CN). Chromatographically pure.

4.1.2 n-hexane (C6H14). pure chromatography.

4.1.3 acetic acid (CH3COOH). pure chromatography.

4.1.4 Acetone (C3H6O). excellent grade pure.

4.1.5 Sodium chloride (NaCl).

4.2 Solution preparation

4.2.1 n-Hexane - Acetone (6 4). Take 60 mL of n-hexane, add 40 mL of acetone and mix well.

4.2.2 acetonitrile - water solution (1 1). measure 500 mL of acetonitrile, water volume to 1 L, mix.

4.2.3 0.1% acetic acid aqueous solution. take 1.0 mL of acetic acid, diluted with water and set to 1000 mL.

4.3 Standards

4.3.1 Reference substance. Pyrrione (CAS No. 79-41-9), Herbicide (CAS No. 565635-13-7), thiadenone (CAS No.

(CAS No. 87820-88-0), olefenil (CAS No. 74051-80-2) and oleokinone (CAS No.

99129-21-2), purity >=99% (see Appendix A, Appendix A.1 for chemical information).

4.4 Standard solution preparation

4.4.1 Cyclohexenone pesticide standard stock solution. respectively, accurately weighed the right amount of cyclohexanone pesticide standard substances, with acetonitrile preparation Concentration of 1000 mg/L standard stock solution, the standard solution to avoid preservation at -18 °C, the shelf life of 12 months.

4.4.2 Cyclohexenone Pesticide Mixed Intermediate Standard Solution. Absorb the appropriate amount of each standard stock solution and dilute with acetonitrile to a concentration of 1.0 mg/L of the mixed standard solution, 0 °C ~ 4 °C dark preservation, the shelf life of 3 months.

4.4.3 matrix mixed standard working solution. absorb the appropriate amount of mixed intermediate standard solution, with blank sample extraction solution concentration of 0µg/L, 10.0 µg/L, 20.0 µg/L, 50.0 µg/L, 100 µg/L and.200 µg/L, respectively. The day of preparation.

4.5 Materials

4.5.1 Graphitized Carbon Black Solid Phase Extraction Column. 3 mL 250 mg, or equivalent. Activated with 3 mL n-hexane-acetone solution before use the column is moist.

4.5.2 C18 solid phase extraction column. Activate with 5 mL acetonitrile before use to keep the column moist.

4.5.3 Microporous membrane. 0.22 µm, organic phase type.

6.1 Preparation of the sample

6.1.1 Corn, soybeans and rice

Take about 500 g of sample, crush and mix thoroughly with a pulverizer. Samples are divided into two copies, into a clean container, sealed, and marked the standard Remember.

6.1.2 Cabbage, potatoes, garlic, grapes and oranges

Take the sample about 500 g, will be (not available to wash) chopped, with a mashed machine processed into a slurry, mix. The samples are divided into two Copies, into a clean container, sealed, and marked with a mark. Frozen at -18 C.

Note. The above sample sampling site according to GB 2763 Appendix A implementation.

6.2 Sample storage

During the operation of the sample preparation, the sample should be protected from contamination or changes in the content of the residue. Corn, soy and rice samples Save at room temperature. Chinese cabbage, potato, garlic, grape and orange samples are stored at -18 C.

7.1 Extraction

7.1.1 Corn, soybeans and rice

Weigh 5 g (accurate to 0.01 g) sample in a 50 mL stoppered centrifuge tube, add 10 mL of water, whirlpool and mix for 1 h. Then add Into the 4.0 g sodium chloride, then add 15 mL of acetonitrile high-speed homogeneous extraction 3 min, shake extraction 15 min, centrifuged at 5 000 r/min 5 min, the acetonitrile layer was transferred to a 25 mL volumetric flask. The residue was extracted again with 10 mL of acetonitrile and the extracts were combined and bound to 25 with acetonitrile ML.

7.1.2 Cabbage, potatoes, garlic, grapes and oranges

Weigh 5 g (accurate to 0.01 g) sample in 50 mL stoppered centrifuge tube, then add 2.0 g of sodium chloride, then add 15 mL of acetonitrile high speed Homogenized extraction for 3 min, shaking extraction for 15 min, centrifugation at 5 000 r/min for 5 min, and the acetonitrile layer was transferred to a 25 mL volumetric flask. Residue again with 10 mL of acetonitrile repeated extraction time, combined extract, and acetonitrile constant volume to 25 mL.

7.2 Purification

7.2.1 Soybeans and maize

Extraction of 5.0 mL of extract in the C18 solid phase extraction column, at about 1.5 mL/min flow rate of all the sample through the solid phase extraction column, and then the solid phase was extracted with 3 mL of acetonitrile and the whole extract was collected in a 15 mL centrifuge tube and purged with nitrogen at 40 ° C.

The residue was dissolved in 3.0 mL of n-hexane-acetone and the solution was added to a treated Envi-Carb solid phase extraction column at about 1.5 mL/min the flow rate was passed through a solid phase extraction column and the sample tubes were rinsed with 5 mL of n-hexane-acetone and the rinsing solution was added to Envi-Carb the solid extract was collected and the whole effluent was collected in a 15 mL centrifuge tube and the nitrogen was blown to near dryness in a 40 ° C water bath. The residue was washed with 1.0 mL of acetonitrile - water dissolved, vortex after mixing, through the microporous membrane, for liquid chromatography - mass spectrometry/mass spectrometry.

7.2.2 Rice, cabbage, potatoes, garlic, grapes and oranges

Take 5.0 mL of the extract in 15 mL centrifuge tube, 40 °C with N2 blowing to near dry. The residue was dissolved in 2.0 mL of n-hexane-acetone the solution was added to the treated Envi-Carb solid phase extraction column and the sample solution was passed through a solid phase extraction column at a flow rate of about 1.5 mL/min.

The sample tubes were rinsed with 5 mL of n-hexane-acetone and the rinsing solution was added to the Envi-Carb solid phase extraction column to collect the whole effluent in 15 ML centrifuge tube, in a 40 ° C water bath, nitrogen blowing to near dry, the residue with 1.0 mL acetonitrile - water dissolved, vortex mixing, the microporous filter Membrane for liquid chromatography - mass spectrometry/mass spectrometry.

7.3 Determination

7.3.1 Liquid Chromatographic Reference Conditions

A) Column. Waters Acquity BEH C18 column, 50 mm × 2.1 mm (id), 1.7 µm, or equivalent;

B) Column temperature. 40 ° C;

C) Injection volume. 10 µL;

D) The mobile phase, flow rate and gradient elution conditions are shown in Table 1.

Table 1 Mobile phase, flow rate and gradient elution conditions

time

Min

Flow rate

ML/min

0.1% acetic acid aqueous solution Acetonitrile 0 0.30 50 50 5.0 0.30 10 90 6.0 0.30 10 90 7.5 0.30 50 50

7.3.2 Mass spectrometry reference conditions

A) ionization mode. electrospray ion source;

B) Scanning mode. positive ion scanning;

C) Detection method. Multiple reaction monitoring (MRM);

D) Resolution. Unit resolution;

See Appendix B for other reference mass spectrometry conditions.

7.3.3 Determination and confirmation of chromatography

7.3.3.1 Qualitative determination

Each selected component selected one parent ion, two or more child ions, in the same experimental conditions, the sample retention of the test substance and the retention time of the matrix standard solution is within ± 2.5%. The relative abundance of the qualitative ions in each sample is close to the concentration of the matrix of mixed standard working solution corresponding to the relative abundance of qualitative ions were compared, the deviation does not exceed the range specified in Table 2, as there is a corresponding sample in the sample.

Table 2 shows the maximum allowable error for relative ion abundance when qualitatively by liquid chromatography-tandem mass spectrometry

Relative abundance (base) 50% 20% to 50% 10% to 20% <=10% Allowable relative deviation ± 20% ± 25% ± 30% ± 50%

7.3.3.2 Quantitative determination

Under the optimum working conditions of the instrument, the standard working solution of the matrix was mixed with the peak area as the ordinate and the matrix mixed standard work Solution concentration for the abscissa to draw the standard working curve, with the standard working curve to calculate the sample solution concentration (C), sample the response value of the analyte in the solution shall be within the linear range of the instrument determination. If the linear response range is exceeded, the blank matrix solution is applied Line proper dilution. A multi-reaction monitoring (MRM) chromatogram of the six cyclohexene ketone reference materials is shown in Figure C.1 in Appendix C.

7.4 Blank experiment

In addition to the sample, according to the above determination steps.

10.1 Quantitation limits

The limit of quantification of cyclohexyl ketone pesticides is 0.005 mg/kg.

10.2 Recovery rate

The data on the concentration and recovery of the sample are given in Appendix D.

Appendix A

(Informative)

Cyclohexenone pesticides

Table A.1 Related information about cyclohexanone pesticides

Compound name English name Cas No. molecular formula molecular weight molecular structure Pyramidone Teproloxydim 149979-41-9 C17H24ClNO4 341.83.

CH2

CH2 CH3

Herbs off. Alloxydim 55635-13-7 C17H25NO5 323.37 CC CH2 CH2 CH3 O CH2 CH CH2 OH OCH3 CH3 CH3 Thioketone Cycloxydim 101205-02-1 C17H27NO3S 325.47 CH2 CH2 CH3 CH2 OH CH3 Oxalcone Clethodim 99129-21-2 C17H26ClNO3S 359.91 OH CH2 CH3 O CH2 C Cl CH2CHCH3 CH2CH3 Olefinic acid Sethoxydim 74051-80-2 C17H29NO3S 327.48 OH CH3 O CH3 CH3 CH3 Phenacetone Methoxypheno Ne 87820-88-0 C20H27NO3 329.43 CH2 CH2 CH3 CH3 OH CH3 H3C CH3 Appendix B (Informative) Mass spectrometry conditions A) electrospray voltage. 3000 V;

B) Auxiliary gas flow rate. 750 L/h;

C) collision gas. argon;

D) curtain air flow rate. 50 L/h;

E) ion source temperature. 105 ° C;

F) Auxiliary gas temperature. 350 ° C;

G) Qualitative ion pair, quantitative ion pair, acquisition time, cone hole voltage and collision energy are shown in Table B.1.

Table B.1 Mass Spectrometric Parameters of Cyclohexenone Pesticide Reference Material

The name of the compound Reference Reserved time Min Qualitative ion pair / (M/z) Quantitative ion pair / (M/z) Acquisition time / S Collision gas Volume/eV Cone hole voltage/V Pyrrione 1.85 0.05 30 32 342/250 0.05 19 32 Herbs off 2.58 0.05 30 30 324/234 0.05 21 30 Thiadenone 3.18 0.05 29 30 326/280 0.05 18 30 Oxalate 3.38 0.05 29 32 360/268 0.05 18 32 Olefinic acid 3.59 0.05 29 30 328/220 0.05 22 30 Bromethrin 3.78 0.05 28 30 330/284 0.05 18 30 Note. * for the quantitative ions, for different mass spectrometry equipment, instrument parameters may be different, should be measured before the mass spectrometry parameters to optimize the best.

Non-commercial declaration. The reference mass spectrometry conditions listed in B.1 are performed on a Waters Quattro Premier LC/MS, where the test instrument Model only for reference, does not involve commercial purposes, to encourage standard users to try different manufacturers or models of equipment.

Appendix C

(Informative)

Cyclohexanone Pesticide Standard Quality Spectrum

Figure C.1 Standard spectrum of cyclohexanone pesticides

Enherofen

Appendix D

(Informative)

Table D.1 Recovery of Cyclohexenone Drugs in Different Substrates

Compounds

name

Add level

Mg/kg

Rice

Recovery rate range

corn

Recovery rate range

Soybeans

Recovery rate range

Cabbage

Recovery rate range

garlic

Recovery rate range

potato

Recovery rate range

orange

Recovery rate range

grape

Recovery rate range

Pyranone

5 83.8 to 104 84.8 to 94.8 82.6 to 96.4 84.1 to 103 72.9 to 82.7 77.3 to 93.8 88.3 to 103 81.7 to 98.7 10 80.0 to 89.9 86.0 to 97.7 75.8 to 89.7 84.6 to 101 71.5 to 77.7 79.2 to 94.9 85.2 to 94.8 80.1 to 101 50 78.5 to 87.0 77.5 to 89.0 82.0 to 91.0 95.3 to 113 70.0 to 77.4 79.5 to 86.2 80.9 to 87.4 84.4 to 99.9 1000 92.6 to 106 81.1 to 90.2 80.4 to 84.4 89.3 to 92.2 92.9 to 97.4 83.2 to 89.4 80.5 to 86.2 80.4 to 84.4 10000 99.3 ~ 112 80.6 ~ 90.7 85.8 ~ 86.2 90.0 ~ 96.1 86.3 ~ 88.4 98.1 ~ 112 80.4 ~ 88.1 85.8 ~ 87.3 Grass off 5 86.6 to 107 87.8 to 97.6 87.8 to 99.8 70.0 to 84.0 75.5 to 82.9 92.4 to 110 84.0 to 90.1 80.9 to 92.3 10 84.9 to 93.8 83.2 to 93.4 84.7 to 93.8 81.2 to 95.6 68.4 to 73.8 105 to 116 80.4 to 87.5 82.0 to 97.5 50 79.5 to 89.0 75.0 to 84.0 83.0 to 94.2 89.0 to 95.0 70.3 to 78.6 87.2 to 97.2 74.3 to 80.6 88.3 to 106 1000 101 to 120 90.7 to 97.4 81.7 to 86.0 89.8 to 90.0 87.1 to 91.5 87.0 to 109 80.4 to 88.0 81.7 to 86.0 10000 87.8 to 114 88.6 to 90.2 78.5 to 86.2 80.2 to 82.3 83.1 to 96.5 99.6 to 106 86.6 to 88.8 81.5 to 86.2 Thiadenone 5 70.0 to 81.2 70.0 to 77.8 70.0 to 74.8 70.0 to 82.9 70.2 to 82.6 70.3 to 82.7 88.1 to 92.6 85.0 to 100 10 72.4 to 76.6 70.0 to 75.5 70.0 to 76.0 72.8 to 81.3 72.8 to 79.4 77.6 to 80.0 83.6 to 94.3 84.9 to 95.6 50 70.0 to 75.5 70.0 to 78.0 70.0 to 76.5 70.0 to 78.7 86.1 to 92.9 70.4 to 74.7 78.8 to 89.0 86.8 to 101 1000 80.7 to 88.8 82.4 to 89.9 80.0 to 82.3 80.3 to 86.6 82.2 to 87.4 84.7 to 89.3 87.1 to 87.4 80.0 to 82.3 10000 80.1 to 82.1 84.3 to 87.9 80.0 to 82.8 84.8 to 89.2 80.0 to 86.0 80.8 to 90.2 86.4 to 95.8 83.4 to 87.8 Oxadiazone 5 70.4 to 79.0 71.0 to 79.0 70.0 to 87.0 75.0 to 91.3 79.1 to 84.9 71.7 to 84.7 75.0 to 91.3 70.6 to 83.2 10 70.0 to 78.4 70.0 to 77.4 71.0 to 78.6 89.7 to 105 86.2 to 91.3 84.7 to 91.8 89.7 to 105 77.0 to 85.5 50 70.0 to 78.0 70.0 to 74.0 70.3 to 78.5 88.9 to 104 92.5 to 101 71.8 to 78.7 88.9 to 104 83.9 to 94.1 1000 80.5 to 88.1 81.8 to 85.9 80.9 to 84.3 88.0 to 91.1 86.5 to 99.4 88.6 to 90.3 92.2 to 108 78.5 to 82.8 10000 80.1 to 89.7 84.8 to 88.4 85.4 to 89.6 83.3 to 95.5 96.7 to 102 81.1 to 88.1 97.4 to 107 85.4 to 89.6 Phenacetone 5 85.4 to 110 85.2 to 94.6 91.8 to 107 75.9 to 90.8 83.8 to 96.4 79.0 to 83.3 89.2 to 92.3 68.3 to 84.8 10 78.4 to 94.8 74.8 to 79.7 80.9 to 93.6 86.0 to 103 88.4 to 95.4 90.2 to 98.5 88.9 to 97.1 82.5 to 96.5 50 87.0 to 104 84.5 to 87.5 83.0 to 94.0 92.0 to 106 88.7 to 93.5 81.5 to 87.9 87.6 to 91.7 83.7 to 101 1000 81.8 ~ 120 83.9 ~ 90.0 85.8 ~ 90.9 80.5 ~ 87.9 86.6 ~ 90.0 84.3 ~ 88.3 86.9 ~ 88.3 85.8 ~ 90.9 10000 99.3 to 112 80.9 to 82.0 81.0 to 89.0 80.8 to 81.0 80.1 to 83.0 81.8 to 90.4 87.1 to 89.4 83.1 to 89.0 Enanthrene 5 73.4 to 83.2 70.0 to 75.0 70.0 to 79.0 73.3 to 82.3 76.5 to 83.6 74.1 to 83.4 73.3 to 82.3 81.4 to 94.4 10 70.3 to 77.0 71.1 to 76.8 70.0 to 80.4 86.5 to 103 83.2 to 91.3 78.4 to 90.1 86.5 to 103 80.5 to 97.8 50 70.0 to 75.0 70.0 to 77.0 70.0 to 78.0 87.3 to 104 87.7 to 95.5 72.7 to 76.6 87.3 to 104 83.5 to 101 1000 82.6 to 106 80.6 to 85.2 92.6 to 97.8 83.9 to 84.6 80.1 to 81.8 80.9 to 88.2 94.4 to 111 82.8 to 97.8 10000 91.7 to 97.4 80.6 to 85.3 87.5 to 100 81.4 to 88.7 90.4 to 98.9 80.0 to 89.2 99.1 to 110 97.5 to 101 Appendix E (Normative appendix) Laboratory repeatability requirements

Table E.1 Laboratory repeatability requirements

Measured component content Mg/kg Precision <=0.001 36 > 0.01 > 1 14 Appendix F (Normative appendix) Inter-laboratory reproducibility requirements

Table F.1 Inter-laboratory reproducibility requirements

Measured component content Mg/kg Precision <=0.001 54 > 0.01 > 1 19