GB 23200.50-2016Food safety national standard -- Determination of Pyridine Pesticide Residues in Food by Liquid Chromatography -- Mass Spectrometry / Mass Spectrometry (English PDF)

4.1 Reagents

4.1.1 Acetonitrile (CH3CN). liquid chromatography grade.

4.1.2 Methanol (CH3OH). liquid chromatography grade.

4.1.3 Toluene (C7H8).

4.1.4 acetic acid (CH3COOH).

4.1.5 Sodium chloride (NaCl).

4.1.6 anhydrous sodium sulfate (Na2SO4). 650 °C burning 4 h, placed in the dryer to spare.

4.2 Solution preparation

4.2.1 0.1% acetic acid solution. take 1mL acetic acid, diluted with water and set to 1000 mL.

4.2.2 methanol - water (1 1, V/V). measure 100 mL of methanol, add 100 mL of water, mix well.

4.2.3 toluene - acetonitrile acidic solution. measure 100 mL of toluene, add 300 mL of acetonitrile and 4 mL of acetic acid, mix well.

4.3 Standards

4.3.1 imidacloprid, acetamiprid, imazethapyr, furazolidone, pyridinamide, thiazole niacin and fenpropathrin Standard substance. purity >=98%, see Appendix A.

4.4 Standard solution preparation

4.4.1 standard stock solution. respectively, accurately weighed the appropriate amount of the standard, with methanol prepared into 1.0 mg/mL standard stock solution, 4 °C Save under darkness.

4.4.2 Preparation of mixed standard intermediate liquid. Take the above standard stock solution amount, with methanol prepared into 10 g/mL mixed standard solution, 4 °C save under the dark.

4.4.3 Preparation of standard working solution. the appropriate amount of mixed standard intermediate solution, with a blank sample matrix prepared into the appropriate concentration of the standard Working solution, prepared before use.

4.5 Materials

4.5.1 C18 solid phase extraction cartridge. 500 mg, 3 mL or equivalent.

4.5.2 Graphite Carbon Black Column. 500 mg, 6 mL or equivalent.

6.1 Preparation of the sample

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

6.1.1 Citrus, Potato and Spinach

Take a representative sample 500 g, chopped (not available in water), and the sample is processed into a slurry with a mashed machine. Mix well and put it clean Within the sample container, seal and mark the mark.

6.1.2 Rice, wheat, walnut and tea

Take a representative sample 500 g, crushed with a pulverizer. Mix, fit into a clean container, sealed and marked.

6.1.3 Fish, pork and liver

Take a representative sample 500 g, take the sample by the mashed machine mashed evenly, into the clean sample container, sealed and marked Remember.

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

6.2 Sample storage

Rice, wheat, walnuts and tea leaves at 0 °C 4 °C; citrus, potatoes, spinach, pork, fish, liver and milk-18 °C below the frozen storage.

7.1 Extraction

7.1.1 Rice, wheat, walnut and tea

For rice, wheat, walnut samples, weigh 2 g sample (accurate to 0.01 g). For tea samples, weigh 1 g sample (fine Indeed 0.01 g). Place the weighed sample in a 50 mL centrifuge tube, add 5 mL of water to mix for 30 min, add 2 g of sodium chloride and 10 mL of acetonitrile was added to the vortex mixer for 3 min and centrifuged at 5000 r/min for 5 min. The supernatant was transferred to another centrifuge tube.

The extraction process was repeated with 5 mL of acetonitrile and the extracts were combined. Concentrated to about 2 mL at 40 ° C and purged.

7.1.2 Citrus, milk

Weigh 5 g (accurate to 0.01 g) sample in a 50 mL centrifuge tube, add 2 g of sodium chloride and 10 mL of acetonitrile in a vortex mixer Mixed for 3 min. 5000 r/min centrifugation 5 min, the supernatant transferred to another centrifuge tube. Repeat the above with 5 mL of acetonitrile Take the process, the combined extract, at 40 °C nitrogen blowing concentrated to about 2 mL, to be purified.

7.1.3 Potatoes, spinach, pork, fish and liver

Weigh 2 g (accurate to 0.01 g) sample in a 50 mL centrifuge tube, add 3 mL of water, 2 g of sodium chloride and 10 mL of acetonitrile, Mix the mixture for 3 min, centrifuge at 5000 r/min for 5 min, transfer the supernatant to another centrifuge tube. Re-use 5 mL of acetonitrile Repeat the extraction process above, merge the extract. Concentrated to about 2 mL at 40 ° C and purged.

7.2 Purification

7.2.1 Rice, wheat, potatoes, citrus, pork, fish and milk

In the top of the C18 solid phase extraction column into the 1 cm high anhydrous sodium sulfate, first with 5 mL of acetonitrile pre-leaching column, discard the eluent. Then the resulting extract was passed through the column and eluted with 4 mL of acetonitrile to control the flow rate at 0.5 mL/min. All effluents were collected. At 40 ° C and the residue was filtered through a 1.0 mL methanol-water solution and passed through a 0.45 µm organic membrane for liquid chromatography-mass spectrometry/mass spectrometry Determination.

7.2.2 Spinach, liver, walnut and tea

In the top of the graphitized carbon black solid phase extraction column into the 1cm high anhydrous sodium sulfate, first with 10mL toluene - acetonitrile acid solution pre-leaching Small column, abandoned eluent. The column was then added to the extract and eluted with 25 mL of toluene-acetonitrile acid solution to control the flow rate of 0.5 ML/min, collecting all effluent. In 40 ° C water bath rotation concentrated to about 1 ~ 2mL, and then blown to the near-dry nitrogen flow. With 1.0 mL Methanol - aqueous solution to dissolve the residue, 0.45 µm of the organic phase microporous membrane, for liquid chromatography - mass spectrometry/mass spectrometry.

7.3 Determination

7.3.1 Liquid Chromatographic Reference Conditions

A) Column. C18 column, 150 x 4.6 mm (inner diameter), particle size 5 µm or equivalent;

B) mobile phase. gradient elution program in Table 1;

C) Flow rate. 0.5 mL/min;

D) Column temperature. 30 ° C;

E) Injection volume. 10 µL.

Table 1 Mobile phase gradient elution program

time

Min

0.1% acetic acid

Methanol

7.3.2 Mass spectrometry reference conditions

A) ion source. electrospray ion source;

B) scanning mode. positive ions;

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

D) atomization gas, air curtain gas, auxiliary heating gas, collision gas are high purity nitrogen; before use should adjust the gas flow to mass spectrometry Sensitivity to meet the test requirements, the reference conditions see Appendix B;

E) Parameters such as spray voltage, de-cluster voltage, and collision energy should be optimized to optimum sensitivity. See Appendix B for reference conditions.

7.3.3 Quantitative determination

According to the sample in the sample content, select the appropriate response value of the standard working solution for analysis. Standard working fluid and sample solution to be tested the response value of the pyridine pesticide should be within the linear response range of the instrument. If the content exceeds the standard curve range, dilute to the appropriate concentration Post analysis. The reference retention time of pyridine pesticides under these chromatographic conditions was imidacloprid (8.5 min), acetamiprid (9.0 min) (11.6 min), paclitaxel (11.9 min), thiazolyl nicotinic acid (12.8 min) and fluorosulfide (14.0 min). The multi-reaction monitoring chromatogram of the standard solution is shown in Appendix C in Figure C.1.

7.3.4 Qualitative determination

Under the same experimental conditions, the chromatographic peak retention time of the sample in the sample solution was the same as that of the standard working solution, and after subtracting the background , The relative abundance of each qualitative ion is compared with the abundance of the standard ion, and the deviation Does not exceed the scope specified in Table 2, you can determine the existence of the corresponding sample in the sample.

Table 2 The maximum allowable deviation of relative ion abundance when qualitative confirmation

Relative ion abundance 50% 20% to 50% 10% to 20% <=10% Allowable relative deviation ± 20% ± 25% ± 30% ± 50%

7.4 Blank test

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

10.1 Quantitation limits

Quantitative limits for pyridine pesticides in rice, wheat, potatoes, spinach, citrus, walnut, pork, fish, liver and milk 0.005 mg/kg; the limit of quantification of pyridine pesticides in tea was 0.01 mg/kg.

10.2 Recovery rate

Rice, potato,, spinach, citrus, walnut, tea, pork, fish, liver and milk.

The data on the level and recovery are shown in Appendix D.

Appendix A

(Informative)

The basic information of the standard substance

Table A.1 Basic information on reference materials

Chinese name English name CAS number Molecular formula Molecular weight Imidacloprid 138261-41-3 C9H10ClN5O2 255.7 Acetamiprid 160430-64-8 C10H11ClN4 222.7 Imazethapyr 81385-77-5 C15H19N3O3 289.3 Thiazole niacin thiazopyr 117718-60-2 C16H17F5N2O2S 396.4 Flutidone fluridone 59756-60-4 C19H14F3NO 329.3 Pyridinamide boscalid 188425-85-6 C18H12Cl2N2O 343.2 Fisulfuride dithiopyr 97886-45-8 C15H16F5NO2S2 401.4 Appendix B (Informative) Reference to mass spectrometry conditions Reference mass spectrometry conditions.

A) Scanning mode. positive ion scanning;

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

C) air curtain gas (CUR). 10 Psi;

D) Atomizing gas (GS1). 40 Psi;

E) Auxiliary heated gas (GS2). 50 Psi;

F) collision gas (CAD). 7 Psi;

G) electrospray voltage (IS). 5000 V;

H) ion source temperature (TEM). 450 C;

I) Qualitative ion pair, quantitative ion pair, collision energy, de-cluster voltage, contact chamber inlet and outlet voltage, and collision chamber outlet voltage see

Table B.1.

Table B.1 Multi-reaction monitoring conditions

Compounds

Mother ion

Ion

Dwell time

To cluster voltage

(DP)/V

Collision energy

(CE)/V

Collision room

Inlet voltage

(EP)/V

Collision room

Outlet voltage

(CXP)/V

Imidacloprid 256.0

209.0

175.0.200 46 33 10 6 Acetamidine 223.0 126.0 90.0.200 36 45 10 12 Imazethapyr 290.1 245.6 200 62 26 10 16 177.4.200 62 37 10 15 Fluorothiazone 330.5 309.5 200 124 46 10 15 259.2.200 124 60 10 15 Pyridinamide 343.0 307.2 200 98 27 10 15 140.2.200 98 30 10 6 Thiazole niacin 397.6 377.2 200 94 33 10 11 335.2.200 94 44 10 18 Fluoride Sulfide 402.3 354.0 500 86 26 10 15 272.4 500 98 410 10 15 Note. The ions with "*" are quantitative ions.

1) Non-commercial notices. The parameters listed in Appendix B are performed on the API4000 mass spectrometer, where the test instrument model is listed for reference only and Do not involve commercial purposes, encourage standard users to try to use different manufacturers or models of equipment.

Appendix C

(Informative)

Multi - reaction monitoring (MRM) chromatograms for standard solutions Pyridinamides 343.4/307.2 Pyridinamides 343.4/140.2 Acetamidine 223.0/126.0 Acetamidine 223.0/90 Imazethapyr 290.1/245.6 Imitation grass smoke 290.1/177.4 Imidacloprid 256.0/209.0 Imidacloprid 256.0/175.0

Figure C.1 Multi-reaction monitoring (MRM) chromatograms for standard solutions

Fluorothiazone

330.5/259.2 Fluorothiazone 330.5/309.5 Thiazole niacin 397.4/335.2 Thiazole niacin 397.4/377.2 Fenbutetine 402.2/354.0 Fenbutetine 402.2/272.4 Appendix D (Informative) the recovery of pyridine pesticides in different sample substrates

Table D.1 Recovery of Pyridine Pesticides in Different Sample Substrates

sample

Matrix

Add to

Level

/ (Mg/kg)

Recovery rate/%

Imidaclopridamphamidine imidazolium niacin fenfluramine ketophenadiazole n -Rice 0.005 87.8 to 103.0 89.4 to 97.2 73.2 to 90.2 86.2 to 97.6 82.4 to 95.6 64.6 to 83.6 61.2 to 73.4 0.01 97.6 to 105.0 95.5 to 100.5 86.5 to 92.3 89.5 to 103.5 80.8 to 89.9 74.9 to 82.0 60.9 to 78.7 0.025 89.8 to 100.6 92.2 to 99.4 79.0 to 97.6 88.7 to 98.8 86.6 to 99.6 77.8 to 87.5 67.9 to 87.0 potato 0.005 85.0 to 100.2 86.6 to 100.4 65.0 to 86.0 77.0 to 91.2 65.0 to 82.4 69.6 to 79.6 60.6 to 79.6 0.01 88.0 to 96.1 92.9 to 98.5 83.5 to 99.5 79.7 to 87.8 80.5 to 89.0 75.6 to 88.0 65.9 to 76.0 0.025 81.0 to 99.0 82.6 to 100.0 79.6 to 94.7 86.6 to 94.6 79.4 to 88.7 75.2 to 88.1 66.2 to 75.2 Walnut 0.005 82.0 to 95.0 80.2 to 93.2 63.4 to 77.0 79.0 to 95.0 63.6 to 81.0 60.8 to 69.6 66.4 to 74.6 0.01 83.0 to 92.4 89.7 to 96.1 72.5 to 80.3 82.5 to 93.4 78.5 to 89.7 68.0 to 83.4 69.9 to 80.1 0.025 76.4 to 96.6 76.4 to 100.8 69.6 to 77.8 75.8 to 86.2 84.2 to 93.6 64.9 to 76.9 66.2 to 75.2 pork 0.005 76.4 ~ 87.6 75.4 ~ 101.0 60.4 ~ 77.6 75.2 ~ 90.0 61.6 ~ 86.0 60.2 ~ 77.8 60.8 ~ 76.6 0.01 96.7 to 100.3 91.5 to 101.0 70.6 to 101.0 88.5 to 93.0 76.1 to 88.6 71.0 to 81.5 63.4 to 70.1 0.025 87.2 to 99.2 87.1 to 100.0 72.4 to 91.8 87.4 to 95.0 73.4 to 84.3 77.8 to 87.6 63.6 to 68.6 Fish 0.005 91.0 to 102.0 84.8 to 99.4 67.8 to 90.6 79.6 to 91.2 74.2 to 87.4 69.6 to 81.2 64.6 to 79.8 0.01 96.6 to 102.5 94.2 to 102.0 78.9 to 83.4 76.5 to 85.2 80.7 to 89.5 65.5 to 78.9 62.4 to 80.6 0.025 86.5 to 96.6 91.2 to 101.2 80.3 to 91.6 75.8 to 86.2 84.2 to 102.0 66.2 to 83.1 64.9 to 76.9 wheat 0.005 87.0 to 100.2 79.8 to 102.4 81.0 to 93.0 79.4 to 87.8 73.4 to 86.4 73.2 to 83.0 60.2 to 73.0 0.01 89.4 to 104.5 86.8 to 99.9 79.0 to 90.4 85.5 to 95.0 79.5 to 95.5 81.5 to 90.8 62.3 to 78.7 0.025 91.4 to 100.5 86.8 to 99.9 76.6 to 87.9 79.4 to 94.1 71.6 to 87.5 84.0 to 93.0 69.1 to 73.5 Tangerine 0.005 78.0 to 91.2 84.4 to 101.2 75.6 to 95.6 81.2 to 100.0 77.4 to 97.2 75.6 to 94.4 61.4 to 77.8 0.01 87.4 to 97.0 89.0 to 95.6 72.6 to 99.2 81.2 to 100.0 77.2 to 97.8 71.6 to 86.2 79.1 to 85.9 0.025 83.4 to 91.6 84.0 to 99.2 79.6 to 91.2 82.9 to 89.1 77.5 to 87.2 71.8 to 85.8 71.0 to 78.0 milk 0.005 76.4 to 95.2 77.0 to 108.0 69.0 to 93.0 64.6 to 85.0 73.8 to 92.2 65.0 to 82.6 60.0 to 68.8 0.01 81.6 to 97.2 82.4 to 98.0 78.9 to 88.8 83.6 to 91.5 68.4 to 88.4 67.2 to 84.2 63.2 to 81.2 0.025 82.4 to 94.0 81.9 to 92.8 80.8 to 97.2 85.8 to 106.8 78.2 to 87.6 71.6 to 87.6 67.2 to 77.7 spinach 0.005 72.4 to 88.4 76.6 to 97.8 68.6 to 80.4 77.2 to 91.4 64.6 to 82.4 63.6 to 77.6 60.4 to 69.0 0.01 81.4 to 92.0 87.6 to 98.5 74.2 to 87.6 79.8 to 88.8 77.8 to 84.5 67.9 to 82.3 61.3 to 80.1 0.025 76.6 to 98.0 78.6 to 89.2 78.6 to 87.0 82.8 to 91.6 77.4 to 92.8 67.4 to 79.5 64.0 to 76.6 Liver 0.005 75.0 to 89.0 70.6 to 89.2 62.8 to 74.0 72.6 to 80.8 74.6 to 82.6 61.6 to 71.8 64.4 to 75.6 0.01 72.7 to 84.6 84.4 to 91.5 60.0 to 80.4 82.5 to 92.9 71.2 to 81.4 72.3 to 80.1 60.3 to 73.8 0.025 72.8 ~ 90.4 78.8 ~ 99.9 72.9 ~ 76.9 77.7 ~ 95.0 74.4 ~ 85.6 68.5 ~ 82.3 63.0 ~ 76.4 tea 0.01 70.0 to 82.0 83.2 to 94.1 66.6 to 77.8 79.6 to 94.6 68.4 to 80.2 62.6 to 78.2 62.0 to 78.8 0.02 78.9 to 94.5 76.8 to 94.5 71.7 to 84.8 78.6 to 101.5 67.8 to 91.0 73.5 to 86.5 62.5 to 84.5 0.05 70.2 to 85.2 78.8 to 93.2 68.0 to 75.8 76.5 to 91.9 65.4 to 73.4 71.1 to 79.8 70.2 to 85.2 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