GB 23200.64-2016Food safety national standard -- Determination of Residual Potassium Acetate in Food by Liquid Chromatography -- Mass Spectrometry / Mass Spectrometry (English PDF)

4.1 Reagents

4.1.1 anhydrous magnesium sulfate (MgSO4).

4.1.2 anhydrous sodium acetate (C2H3O2Na).

4.1.3 acetic acid (C2H4O2). chromatography grade.

4.1.4 Acetonitrile (C2H3N). Chromatographic grade.

4.1.5 N-propyl ethylenediamine (PSA) filler. 50 m, chromatographic grade.

4.1.6 Formic acid (CH2O2). Chromatographic grade.

4.1.7 Ammonium acetate (C2H7NO2).

4.2 Solution preparation

4.2.1 0.025% formic acid aqueous solution (containing 5 mmol/L ammonium acetate). accurately absorb 0.25 mL formic acid and weigh 0.386 g ammonium acetate in 1 L volumetric flask, dissolved in water and diluted to 1 L.

4.2.2 1% acetic acid in acetonitrile solution.

4.3 Standards

4.3.1 Pyridine ether standard (Pyriproxyfen, C20H19NO3, CAS. 95737-68-1). purity > 99%.

4.4 Standard solution preparation

4.4.1 Pyridyl ether standard stock solution. accurately weighed the appropriate amount of pyridyl ether standard, dissolved in acetonitrile and diluted with 100 µg/mL standard Quasi-stock solution, stored below 4 °C.

4.4.2 Pyridyl ether standard working solution. If necessary, with 10% acetonitrile aqueous solution diluted to the appropriate concentration of standard working fluid, 4 °C below Save

4.5 Materials

4.5.1 Microporous membrane. 0.2 m, organic phase type.

6.1 Rice, soybeans, spinach, lemon, tea, mushrooms, beef, liver

Take a representative sample of about 500 g, with the tissue crusher fully mashed and mix, are divided into two, respectively, into the clean container as a test kind. Sealed and marked with mark. The sample was stored at -18 ° C and protected from light.

6.2 Milk

Take a representative sample of about 500 g, fully mixed, are divided into two, respectively, into a clean container as a sample, sealed, and marked mark. Store the sample below 4 ° C and store it in dark.

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

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

7.1 Extraction

7.1.1 Tea, soybeans, mushrooms, rice samples

Weigh the sample 5 g (accurate to 0.01 g) in a 50 mL polypropylene plastic centrifuge tube, add 10 mL of water, mix and let stand for 30 min.

The centrifuge tube was placed in an ice bath and then 6 g of anhydrous magnesium sulfate and 1.5 g of anhydrous sodium acetate were added. After returning to room temperature, 15.0 mL 1% acetic acid in acetonitrile solution, shake extraction 4 min, 4000 r/min centrifugation 5 min.

7.1.2 Spinach, lemon and milk samples

Weigh 15 g (accurate to 0.01 g) in 50 mL of polypropylene plastic pre-called 6 g of anhydrous magnesium sulfate and 1.5 g of anhydrous sodium acetate Centrifuge tube, the centrifuge tube placed in an ice bath, returned to room temperature, the accurate addition of 15.0 mL 1% acetic acid acetonitrile solution, the oscillation extraction 4 min, 4000 r/min centrifugation 5 min.

7.1.3 Beef, liver samples

Weigh 15 g (accurate to 0.01 g) in 50 mL polypropylene plastic centrifuge tube, accurately add 15 mL 1% acetic acid acetonitrile solution, shaking Extraction 4 min, 4000 r/min centrifugation 5 min. Also called 50 g of polypropylene plastic with 6 g of anhydrous magnesium sulfate and 1.5 g of anhydrous sodium acetate Centrifuge tube, add 10 mL of water, dissolved until the aqueous solution to room temperature, then the acetonitrile extract transferred to them, oscillation 4 min, 4000 R/min centrifugation 5 min.

7.2 Purification

Take 1 mL of the above acetonitrile extract in a 2 mL polypropylene plastic centrifuge tube pre-called 50 mg PSA and 150 mg anhydrous magnesium sulfate, Oscillation was extracted for 1 min and centrifuged at 4000 r/min for 5 min.

Accurately absorb.200 µL supernatant with water to 1 mL, mix well after 0.2 µm filter. The filtrate was subjected to liquid chromatography-mass spectrometry.

7.3 Determination

7.3.1 High Performance Liquid Chromatography Reference Conditions

A) Column. Column. CAPCELL PAK C18 column, 50 mm x 2.0 mm (id), 3 µm or equivalent.

B) Mobile phase. A. 0.1% formic acid acetonitrile solution, B. 0.025% formic acid aqueous solution (containing 5 mmol/L ammonium acetate).

C) Flow rate. 300 µL/min, gradient elution program in Table 1.

Table 1 gradient elution program table

D) Column temperature. 35 ° C.

E) Injection volume. 20 µL.

7.3.2 Mass spectrometry reference conditions

A) ion source. electrospray ESI, positive ion.

Gradient time/min mobile phase A /% mobile phase B /% /4.5 20 80 8.5 20 80 B) Scanning mode. Multiple reaction monitoring (MRM).

C) See Appendix A for other parameter conditions.

7.3.3 Quantitative determination

According to the content of the sample in the sample, select the standard response of similar standard working fluid at the same time for analysis. Standard working solution and the test solution the reaction value of the propyl ether should be within the linear response range of the instrument. The reference retention time of pyridine ether under the above chromatographic conditions was 1.4 min the multi-reaction monitoring chromatogram of the quasi-solution is shown in Appendix B.

7.3.4 Qualitative determination

Determination of sample and standard working solution according to liquid chromatography-mass spectrometry/mass spectrometry conditions if the mass retention time and standard the relative abundance of the qualitative ion pair is expressed as a percentage of the intensity relative to the strongest abundance, and should be comparable to the concentration as the relative abundance of the solution is consistent, the relative abundance tolerance does not exceed the range specified in Table 2, it can be judged that there is a corresponding measured Things.

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

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 method of spinach, lemon, beef, liver and milk in the quantitative limit of 5.0 µg/kg;

The limit of quantification in this method for soybean, rice, mushroom and tea is 15.0 µg/kg.

10.2 Recovery rate

The recovery of the addition of pyridyl ether in different matrices is given in Appendix C.

Appendix A

(Informative)

API 4000 Liquid Chromatography - Quad Column Tandem Mass Spectrometer Parameters 1) Monitor ion pairs and voltage, barometric parameters A) Ion source temperature. 350 C;

B) electrospray voltage. 5250 V;

C) Collision pressure. 34.475 kPa (5 psi);

D) air curtain gas pressure. 206.85 kPa (30 psi) E) atomization gas pressure. 344.75 kPa (50 psi);

F) Auxiliary gas pressure. 413.7 kPa (60 psi);

G) pyrolysis ether quantitative ion pair, qualitative ion pair, deblocking voltage (DP), collision gas energy (CE) and retention time are shown in Table A.1.

Table A.1 Pyridine Ether Quantitative Ion Pair, Qualitative Ion Pair, Discharge Voltage (DP), Collision Gas Energy (CE) and Retention Time

Component name Qualitative ion pair m/z Quantitative ion pair m/z to cluster DP/v Collision energy CE/v keep time T/min Pyridine ether 322.2 -> 185.0 322.2 -> 185.0 110 1.4 322.2 -> 227.2 14

1) Non-commercial notices. Appendix Table A.1 The parameters listed in the API 4000 mass spectrometer are listed here. The test instrument model is for reference only, Does not involve commercial purposes, encourage standard users to try different manufacturers and models of equipment Appendix B (Informative) Pyridyl Ether Multi - Reaction Monitoring (MRM) Chromatogram

Figure B.1 Pyridyl ether MRM chromatogram

Appendix C

(Informative)

Method to add concentration and recovery range

Table C.1 Concentration of the concentration of pyridyl ether and recovery range

Sample name Add concentration (µg/kg) Average recovery rate (%) Precision (%) Rice 15.0 89.3 ~ 107.3 9.2 100 83.5 ~ 101.3 6.4 200 86.3 ~ 102.1 6.9 spinach 5.0 81.4 ~ 101.8 7.2 50 86.2 ~ 103.8 7.0 100 79.7 ~ 101.3 8.8 lemon 5.0 70.6 ~ 100.4 10.9 50 73.8 ~ 103.2 6.2 500 80.1 ~ 100.2 8.5 tea 15.0 83.3 ~ 101.3 6.6 100 86.8 ~ 108.9 9.7 200 79.1 to 104.8 8.0 Liver 5.0 77.6 ~ 101.8 8.5 20 81.5 to 113.0 10.5 100 77.7 ~ 107.2 10.9 beef 5.0 80.6 ~ 99.8 7.5 10 83.5 to 106.0 8.3 100 80.5 ~ 108.2 10.9 milk 5.0 81.4 ~ 99.4 6.3 20 81.5 ~ 103.0 7.8 100 78.2 ~ 100.8 9.2 Soybeans 15 83.3 ~ 104.7 7.8 50 75.8 ~ 103.2 8.4 100 84.0 ~ 99.0 8.1 mushroom 15 84.7 ~ 104.0 7.1 100 83.1 to 106.8 8.1 600 82.9 ~ 103.1 8.8 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