GB 23200.6-2016Food safety national standard -- Methods for the determination of herbicide residues -- Part 6: Determination of herbicidal residues in food by liquid chromatography-mass spectrometry / mass spectrometry (English PDF)

4.1 Reagents

4.1.1 glacial acetic acid (C2H4O2).

4.1.2 Ammonia (NH4OH).

4.1.3 Dichloromethane (CH2Cl2).

4.1.4 Acetone (C3H6O).

4.2 Solution preparation

4.2.1 1% acetic acid solution. absorb 10 mL of glacial acetic acid, with ultra-pure water volume to 1 L.

4.2.2 25% acetone - water solution. the amount of 250 mL of acetone, with ultra-pure water volume to 1 L.

4.2.3 1% acetic acid - acetone - water solution. absorb 10 mL of glacial acetic acid, with acetone aqueous solution to 1 L.

4.2.4 5% ammoniated methanol solution. absorb 5 mL of ammonia, with methanol volume to 100 mL.

4.3 Standards

4.3.1 Straw standard material. Amitrole, C2H4N4, molecular weight. 84.08, CAS. 61-82-5, purity >= 99.9%.

4.4 Standard solution preparation

4.4.1 to kill grass strong standard stock solution. Weigh the appropriate amount of saccharification standard (accurate to 0.1mg) in 50mL volumetric flask, with acetonitrile prepared into 1.0 Mg/mL standard stock solution; 0 ~ 4 0C preservation.

4.4.2 to kill grass strong standard working fluid. according to the need for mobile phase diluted with the preparation of the preparation of the standard concentration of the standard working fluid. 0-4 0C Save

4.5 Materials

4.5.1 PCX solid phase extraction column. 60 mg/3 mL or equivalent. (Mixed cation exchange column).

4.5.2 ENVI-Carb solid phase extraction column. 500 mg/6 mL or equivalent, (graphitized non-porous carbon).

4.5.3 Filtration. 0.2 µm.

6.1 Preparation of the sample

6.1.1 Wheat, corn, honeysuckle, tea

Approximately 500 g of representative sample, crushed with a pulverizer and passed through a 2.0 mm round hole screen. Mix well, into a clean container, Sealed, marked.

6.1.2 Fruits, vegetables, fish, meat, liver

Approximately 500 g of representative sample was taken and the sample was processed into a slurry with a food masher. Mix well, into a clean container, Sealed, marked.

6.1.3 Ginger powder, pepper powder

Take a representative sample of about 100 g, fully mixed evenly, over 2.0 mm round hole sieve, into a clean container, sealed, marked mark.

6.1.4 Peanuts

Take a representative sample 500 g, all grinding with an attritor. Mix well, into a clean container, sealed, marked mark.

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

6.2 Sample storage

Wheat, corn, peanuts, tea, ginger powder, pepper powder samples stored at 0-4 °C; fish, meat, liver and fruit and vegetable samples Frozen at -18 ° C or lower.

During sample and sample preparation, the sample should be protected from contamination or changes in the residue content.

7.1 Extraction

7.1.1 Apple, pineapple, spinach, carrot, basil leaves

Weigh 10 g of sample (accurate to 0.1 g) in a 100 mL centrifuge tube, add 20 Ml of 1% acetic acid solution and 20 mL of dichloromethane,.20000 R/min homogeneous 1 min, centrifuged at 5000 r/min 5 min, supernatant into 100 mL volumetric flask, discard the lower layer of methylene chloride, residue with 1% acetic acid solution 20 mL and dichloromethane 20 mL repeated extraction time; combined supernatant, with 1% acetic acid solution to the scale, remove 10 ML of the extract to be purified.

7.1.2 Honeysuckle, ginger powder, pepper powder, tea leaves

Weigh 1 g sample (accurate to 0.01 g) in 50 mL centrifuge tube, add 20 mL 1% acetic acid aqueous solution,.20000 r/min homogeneous 1 min, 5000 r/min centrifugal 5 min, the supernatant filtered into the 125 mL separatory funnel, the residue and then 10 mL 1% acetic acid aqueous solution repeated extraction time, the supernatant was combined, 10 mL of dichloromethane was added, shaken for 1 min, left to stand, and the lower layer of methylene chloride was discarded. To be purified.

7.1.3 Corn, peanuts

Weigh 2 g sample (accurate to 0.01 g) in 50 mL centrifuge tube, add 20 mL 1% acetic acid-acetone aqueous solution,.20000 r/min 1 min, 5000 r/min centrifugation 5 min, the supernatant filter to 125 mL separatory funnel, the residue and then 10 mL 1% acetic acid - acetone the aqueous solution was repeated once, the supernatant was added, 10 mL of methylene chloride was added, shaken for 1 min, and the layers were separated and the lower layer of methylene chloride was discarded.

To be purified.

7.1.4 Wheat, fish, meat, liver

Wheat, fish, meat, weighed 2 g sample, the liver weighed 1 g sample (accurate to 0.01 g) in 50 mL centrifuge tube, add 20 mL of acetone Aqueous solution,.20000 r/min homogeneous 1 min, 5000 r/min centrifugation 5 min, the extract was filtered to 125 mL separatory funnel, the residue with 10 ML aqueous solution of acetone was repeated once, the supernatant was added, 10 mL of methylene chloride was added, shaken for 1 min, and the layers were separated and discarded.

Methane layer. The solution was added with 0.25 mL of glacial acetic acid to be purified.

7.2 Purification

7.2.1 Apple, pineapple, spinach, carrot, basil leaves, corn, peanuts, wheat, fish, meat, liver

The PCX cartridge was pre-eluted with 3 mL of methanol and 5 mL of water, and the effluent was discarded. Inject the extract, then 3 mL of water, 3 mL of methanol Rinse and discard the effluent. The eluent was eluted with 2 mL of 5% ammoniated methanol solution and concentrated at 40 ° C to near dryness. The residue was washed with 1.0 mL the mobile phase was dissolved and passed through a 0.2 µm membrane for liquid chromatography - mass spectrometry/mass spectrometry.

7.2.2 Honeysuckle, ginger powder, pepper powder, tea

Envi-Carb was pre-eluted with 3 mL of methanol and 3 mL of water. The effluent was discarded and the extract was filled and the effluent was collected.

The PCX cartridge was pre-eluted with 3 mL of methanol and 5 mL of water, and the effluent was discarded. The Envi-Carb purified sample was transferred to a PCX purification column, and then with 3 mL of water, 3 mL of methanol leaching, with 2 mL 5% ammoniated methanol solution elution, the eluent collected at 40 ° C concentrated to near dry, the residue was dissolved in 1.0 mL mobile phase and passed through a 0.2 µm filter for liquid chromatography-mass spectrometry/mass spectrometry.

7.3 Determination

7.3.1 Liquid Chromatography - Mass Spectrometry/Mass Spectrometry Reference Conditions

A. Column. CAPCELL PAK (1. 4) 2.00mmI.D. * 100mm 5µm or equivalent B. Mobile phase. 10 mmol acetic acid amine, 0.1% formic acid water PH = 3 (A) acetonitrile (B)

Table 1 Mobile phase conditions

Condition Sample name Flow rate/mL/min% A% B Condition 1 fish, meat, ginger, pepper, liver, tea 0.20 20 80 Condition 2 apple, pineapple, spinach, carrot, Basil leaves, honeysuckle, wheat, corn, peanut 0.20 50 50 C. Column temperature. 40 ° C.

D. Flow rate. 0.2 mL/min.

E. Injection volume. 10 µL.

F. Mass spectrometry conditions. see Appendix A, A.1

7.3.2 Determination and confirmation of chromatography

According to the sample content of the sample, the selected concentration of similar standard working solution, the standard working solution and sample solution volume into the sample the response values of dinduazites in the standard working solution and the sample to be tested shall be within the linear range of the instrument.

If the sample solution is in the mass chromatogram of the standard working solution, the chromatographic peaks appear at the same retention time, the allowable deviation is less than ± 2.5% the abundance ratio of the selected ions is greater than the abundance ratio of the corresponding ions of the standard, and the value is within the allowable range (see Table 2). You can judge the sample There is a corresponding test object in the product. In the case of 6.3.1, the liquid chromatography-mass spectrum of the herbicidal standard is shown in Appendix B.

Table 2 Use Qualitative Liquid Chromatography - Mass Spectrometry Relative Ion Abundance Maximum Allowable Deviation

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

7.4 Blank experiment

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

10.1 Quantitation limits

The limit of quantification for apple, pineapple, spinach, carrot, basil leaves, corn, peanuts, wheat, ginger, fish, meat and liver is 0.01mg/kg, in tea, honeysuckle, pepper quantitative limit of 0.02 mg/kg.

10.2 Recovery rate

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

Appendix A

Table A.1 Mass spectrometry conditions

Table A.2 Multi-reaction monitoring conditions

Note. Add "*" ions for quantitation.

Non-Commercial Notices. The parameters listed in Appendix A are performed on an Agilent 6410A mass spectrometer, where the test instrument is listed for Provide reference, does not involve commercial purposes, to encourage standard users to try to use different manufacturers or models of equipment.

Ionization mode ESI

Electrospray voltage 4000V the source temperature is 350 ° C Atomizer pressure nitrogen, 40 psi Dry gas nitrogen, flow rate 10L/min Solvent gas stream nitrogen, 600 L/h Collision air pressure, 3.10 × 10-6 Pa Monitoring mode multiple reaction monitoring Compound ion ion ion residence time cone hole voltage collision energy Kill grass strong 85 43 * 0.20 s 100 V 25 eV 57 0.20 s 100 V 15eV Appendix B Standard substance chromatogram

Figure B.1 Table 1 conditions 1 under the grass strong liquid chromatography - mass spectrometry/mass spectrometry multi-reaction monitoring chromatogram

Figure B.2 Table 1 under the conditions of 2 under the grass strong liquid chromatography - mass spectrometry/mass spectrometry multi-reaction monitoring chromatogram

0.08 mg/L standard

Appendix C

(Informative)

Sample concentration and recovery of the experimental data

Table C.1 Experimental data on the concentration and recovery of the sample

sample name

Add concentration

(Mg/kg)

Recovery rate(%)

peanut

10 62.50 to 72.00

20 67.00 ~ 80.00

40 75.75 ~ 85.00

corn

10 61.00 ~ 78.00

20 74.50 to 84.30

40 79.20 ~ 88.00

wheat

10 69.50 to 85.70

20 78.45 ~ 90.00

40 76.00 ~ 93.50

Basil

10 79.22 ~ 82.60

20 71.50 to 91.00

40 85.32 ~ 91.10

pork

10 66.70 ~ 73.20

20 68.40 ~ 81.10

40 78.50 to 85.40

spinach

10 87.20 ~ 98.20

20 90.04 ~ 103.23

40 89.54 ~ 98.55

Salmon

10 69.00 ~ 78.00

20 76.50 to 85.20

40 75.00 ~ 87.50

apple

10 78.40 ~ 83.30

20 77.90 ~ 90.90

40 82.05 to 92.41

Pepper

20 75.00 ~ 94.65

40 81.97 ~ 90.92

80 83.82 ~ 88.99

10 62.80 to 85.60

20 80.30 ~ 82.80

40 80.61 ~ 86.69

honeysuckle

20 69.80 to 88.30

40 81.90 ~ 89.00

80 80.49 ~ 85.40

carrot

10 68.40 ~ 78.50

20 74.50 to 85.45

40 79.20 ~ 94.20

pineapple

10 63.80 to 77.10

20 81.20 to 86.50

40 85.21 ~ 98.52

tea

20 63.25 ~ 79.36

40 71.31 ~ 78.63

80 80.65 ~ 88.63

liver

10 71.31 ~ 78.63

20 70.87 to 81.07

40 81.31 ~ 88.63

Appendix D

(Normative appendix) Laboratory repeatability requirements

Table D.1 Laboratory repeatability requirements

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

Table E.1 Inter-laboratory reproducibility requirements

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