GB 23200.59-2016Food safety national standard -- Determination of residual glycosides in food by gas chromatography-mass spectrometry (English PDF)

4.1 Reagents

4.1.1 Acetonitrile (CH3CN).

4.1.2 Toluene (C7H8).

4.1.3 Acetone (C3H6O).

4.1.4 Diethylene glycol (C4H10O3). Analytical purity.

4.1.5 Sodium chloride (NaCl). Analytical purity.

4.1.6 anhydrous sodium sulfate (Na2SO4). analytical grade. Burned at 650 °C for 4 h, placed in the dryer in the spare.

4.2 Solution preparation

4.2.1 Protective agent. 2% diethylene glycol-acetone solution. Take 100 mL of acetone, add 2 mL of diethylene glycol, mix well.

4.2.2 Eluent. acetonitrile - toluene (3 1, V/V). Take 150 mL of acetonitrile, add 50 mL of toluene, mix well.

4.3 Standards

4.3.1 Dichlobenil. CAS No. 1194-65-6, Molecular formula. C7H3Cl12N, molecular weight 172.02, purity >=99%.

4.4 Standard solution preparation

4.4.1 Dichlorvoside standard stock solution. accurately weighed the amount of diclofenac standard (accurate to 0.1 mg), dissolved in toluene, prepared into 1000 µg/mL of standard stock solution, stored at -18 ° C.

4.4.2 Dichlorvoside standard working solution. dilute to the appropriate concentration of standard working solution with toluene as needed. 0.1 µg/mL standard work the solution was stored at -18 ° C.

4.5 Materials

4.5.1 Graphitized Carbon Solid Phase Extraction Column. ENVI-Carb, 6 mL, 500 mg, or equivalent. Add 1 ~ 2 g of anhydrous sulfur before use Sodium acetate, activated with 10 mL of acetonitrile-toluene.

4.5.2 C18 solid phase extraction column. C18,6 mL, 1000 mg, or equivalent. Add 1 ~ 2 g of anhydrous sodium sulfate before use, add 10 mL Acetonitrile - toluene activation.

4.5.3 Microporous membrane. 0.45µm, organic phase.

6.1 Rice, soybeans, chestnuts

500 g of the sample was taken and pulverized with a pulverizer. Mix, are divided into two as a sample, into the clean Sheng Sheng bag, sealed Stored at 0 ~ 4 °C.

6.2 Mango, blackberry, watermelon, tomato, onion, spinach, oranges, mushrooms

After picking the sample 1000 g, (without water washing), the sample was processed into a slurry by a mashing machine. Mix, are divided into Two as a sample, into the clean Sheng Sheng bag, sealed. The sample was stored at -18 ° C or lower.

6.3 Pork

Take 500 g of pork and chicken liver samples, and then chopped the samples into a slurry and mix them into two pieces.

Sample, into the clean Sheng Sheng bag, sealed. The sample was stored at -18 ° C or lower.

6.4 Milk

Take about 500 g of the sample, mix, are divided into two as a sample, into the clean vial, sealed. Will sample on 0 ~ 4 °C under the conditions of preservation.

Note. During the operation of the sample preparation, the sample should be prevented from being contaminated or the change in the residual content of the diclofenac.

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

7.1 Extraction

7.1.1 Milk, pork, rice, soybeans

Weigh 5 g (accurate to 0.01 g) sample in 50 mL polytetrafluoroethylene centrifuge tube, add 5 mL of water to mix and soak for 10 min. Join B Nitrile 10 mL, homogenized at 10000 r/min 0.5min, add 5 g sodium chloride, shake, ultrasonic extraction at 40 °C for 20min, at 8000 R/min centrifugal 8min, remove the upper organic phase with 2 ~ 3g anhydrous sodium sulfate funnel to the concentration of the bottle; then add 10 mL of acetonitrile, shake Uniform, ultrasonic extraction at 40 °C for 20min, 8,000 r/min centrifugation 8min, remove the upper organic phase with 2 ~ 3g anhydrous sodium sulfate Funnel to the concentrated flask, merge the extracted organic phases twice, then rinse the funnel with about 5 mL of acetonitrile and add 2 mL Curing agent, concentrated below 40 °C to about 1 mL, to be purified.

7.1.2 Mango, Blackberry, Watermelon, Oranges, Tomatoes

Weigh 5 g (accurate to 0.01 g) sample in a 50 mL polytetrafluoroethylene centrifuge tube. Accurately add acetonitrile 10 mL, the following steps with 7.1.1. Concentrated below 40 ° C to about 1 mL, to be purified.

7.1.3 Onions, spinach, mushrooms, chestnuts

Weigh 5 g (accurate to 0.01 g) sample in a 50 mL polytetrafluoroethylene centrifuge tube. Accurately add acetonitrile 10 mL, the following steps with the same 7.1.1. Concentrated below 40 ° C to about 1 mL, to be purified.

7.2 Purification

7.2.1 C18 solid phase extraction column purification

The C18 solid phase extraction column was mounted on a vacuum suction apparatus of a solid phase extraction column, and the solution to be purified in the steps "7.1.1 and 7.1.3" Body column, after sample and then 2 mL eluent repeatedly washed and concentrated bottle, and then with 8 mL eluent for elution, the entire solid phase extraction purification the programmed flow rate does not exceed 6 mL/min. Consolidate the sample solution and eluent about 12 mL in the test tube, 40 °C below concentrated to near dry. With toluene Dissolved and fixed to 1.0 mL, and over 0.45 µm organic phase filter, gas chromatography - mass spectrometry.

7.2.2 Graphitized carbon solid phase extraction column purification

A graphitized carbon solid phase extraction cartridge was mounted on a vacuum suction apparatus of a solid phase extraction column, and the solution to be purified in the step "7.1.2" Body column, after sample and then 2 mL eluent repeatedly washed and concentrated bottle, and then with 8 mL eluent for elution, the entire solid phase extraction purification the programmed flow rate does not exceed 6 mL/min. Collect the sample solution and eluent about 12 mL in the test tube, 40 °C below concentrated to near dry. With toluene Dissolved and fixed to 1.0mL, and over 0.45 µm organic phase filter, gas chromatography - mass spectrometry.

7.3 Determination

7.3.1 Gas Chromatography - Mass Spectrometry Reference Conditions

A column. DB-5 ms quartz elastic capillary column, 30 m × 0.25 mm (id), film thickness 0.25 µm, or equivalent by.

B column temperature. 110 °C for 1min, 10 °C/min heating to 160 °C for 2min, 20 °C/min temperature to.200 °C, the most After running at 300 ° C for 5 min.

C Inlet temperature. 280 ° C.

D Chromatography - Mass Spectrometer Interface Temperature. 280 ° C.

E Carrier gas. helium, purity greater than or equal to 99.999%, flow 1.0 mL/min.

F Injection volume. 2 µL.

G Injection method. splitless injection, opening time. 1.5min.

H ion source. EI.

I ionization energy. 70 eV.

J ion source temperature. 230 ° C.

K quadrupole temperature. 150 ° C.

L Data acquisition mode. Select the ion monitoring mode (SIM).

M Select the ion (m/z). Quantitative ion. 171; Qualitative ion (abundance ratio). 173 (62.4), 136 (30.8), 100 (31.4) (Select only 173 and 136 as qualitative ions for onions and tomatoes).

N solvent delay time. 5.8 min.

7.3.2 Quantitative determination

According to the sample solution of the enemy in the case of nitrile, select the concentration of similar standard working solution. Standard working solution and solution of dextran the value should be within the linear range of the instrument detection. Standard working solution and sample volume and other volume interspersed injection determination. Under the above chromatographic conditions, the retention time of the oxalonitrile is about 6.0 min. The total ion chromatogram of the standard is shown in Figure A.1 in Appendix A.

7.3.3 Qualitative determination

The standard solution and sample solution are checked according to the above conditions, if the sample solution and the standard solution in the same retention time peak Now, it is confirmed by mass spectrometry, after deducting the background of the sample spectrum, the selected ions all appear, while the selected ion the abundance of ions is consistent with the relative abundance of the ions associated with the standard, and the relative ion abundance deviation does not exceed the specified range of Table 1, in the presence of dioxin. The total ion flow chart and mass spectrum of the Energoxide standard are shown in Figures A.1 and A.2 in Appendix A.

Table 1 qualitative determination of the relative ion abundance of the maximum allowable deviation

7.4 Blank test

In addition to no sample, according to the above steps.

10.1 Quantitation limits

The limit of quantification of this method is 5 µg/kg.

10.2 Recovery rate

The range of concentration and recovery of this method is given in Appendix B.

Relative abundance (base) 50% 20% to 50% 10% to 20% <=10% Allowable relative deviation ± 20% ± 25% ± 30% ± 50% Appendix A (Informative) Comparison of standard solution of

Figure A.1 Selective ionization of standard solution GC-MS/EI for dinitriles Total ion chromatogram

Figure A.2 Total Scanning Mass Spectrometry of GC-MS/EI

Appendix B

(Informative)

Sample concentration and recovery of the experimental data

Table B.1 Addition Concentration and Recovery Range of Dietoxan in Different Substrate Samples (%) (n = 6)

Sample concentration (µg/kg) Recovery rate range (%) Rice 5 83.2 ~ 103.3 10 81.5 ~ 101.4 50 72.9 to 102.2 Soybeans 5 80.6 ~ 101.2 10 83.3 ~ 96.5 50 70.6 ~ 102.5 spinach 5 83.9 ~ 100.5 10 72.9 ~ 101.6 50 72.9 ~ 97.4 onion 5 64.1 to 94.4 10 71.6 ~ 95.8 50 71.1 to 102.1 Mushrooms 5 81.7 ~ 100.1 10 85.9 ~ 98.5 50 71.6 ~ 95.0 pork 5 82.2 ~ 96.2 10 75.7 to 103.9 50 72.3 ~ 92.3 milk 5 77.1 to 99.2 10 75.2 to 96.1 50 75.2 ~ 97.4 tomato 5 73.9 ~ 103.3 10 81.8 ~ 103.0 100 80.4 to 92.1 Mango 5 81.6 ~ 107.8 10 75.0 ~ 100.7 200 80.1 to 97.1 watermelon 5 72.4 ~ 96.3 10 70.5 to 108.3 200 80.8 ~ 92.2 orange 5 70.5 to 92.8 10 71.4 ~ 105.0 200 80.1 to 95.8 chestnut 5 84.6 ~ 103.6 10 72.6 ~ 104.7 200 81.2 ~ 96.2 blackberries 5 71.8 ~ 94.8 10 70.5 to 95.6 200 80.2 to 93.5 Appendix C (Normative appendix) Laboratory repeatability requirements

Table C.1 Laboratory repeatability requirements

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

Table D.1 Inter-laboratory reproducibility requirements

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