GB 29703-2013Determination of Sodium nifurstyrenate residues in animal derived food by Liquid Chromatography-tandem Mass Spectrometric method (English PDF)

6.1 Preparation of the sample

Fresh or frozen blank or test tissue, minced, and homogenized.

--- the test sample taken after homogenization, as the test material.

--- blank sample taken after homogenization, as a blank material.

--- blank sample taken after homogenization, adding a suitable concentration of the standard working solution, is added as a blank sample.

Save 6.2 sample

Or less at -20 °C.

Determination Step 7

7.1 Extract

Take Sample 2g ± 0.05g, in 50mL centrifuge tubes, add 0.5% formic acid - ethyl acetate solution 15mL, vortexed, shaken 5min, 8000r/min centrifugal 10min, the ethyl acetate layer eggplant-shaped flask, the residue was added 0.5% formic acid - ethyl acetate solution 10mL, heavy Re-extracted once, twice with ethyl acetate layers were combined, rotary evaporated at 45 °C to dryness, treated with 50% aqueous acetonitrile and the residue was dissolved 3mL, go Centrifuge tube with 3mL washed with 50% aqueous acetonitrile eggplant-shaped flask, two washings were combined, hexane was added 3mL, shaking after mixing, 5000r/min centrifugal 5min, the lower layer was taken and set aside.

7.2 Purification

HLB column eluted sequentially with methanol and 3mL 3mL water activation, taking stock solution through the column. 6mL water rinsed, drained, washed 3mL of acetonitrile was added Off, the eluate was collected, dried under nitrogen 50 °C, the mobile phase the residue was dissolved 1.0mL, 1.5mL transferred to centrifuge tube, 10000r/min Centrifugal 10min, supernatant, membrane filtration, liquid chromatography - tandem mass spectrometry.

7.3 Preparation of standard curve

They were the precise amount of 10µg/mL sodium acid benzyl furan-standard working fluid amount, diluted with mobile phase to prepare a concentration of 0.2,0.5,2, 5 and 8µg/L series of standard solutions, the liquid chromatography - tandem mass spectrometry. A characteristic ion mass chromatogram peak area for the vertical axis, standard solutions Concentration as the abscissa, the standard curve.

7.4 Determination

7.4.1 LC Conditions

7.4.1.1 Column. C18 (150mm × 2.1mm, particle size 5µm), or equivalent person.

7.4.1.2 Mobile phase. aqueous acetonitrile (40 60, volume ratio).

7.4.1.3 Column temperature. 40 °C.

7.4.1.4 Injection volume. 10µL.

7.4.2 MS conditions

7.4.2.1 ion source. electrospray ionization source.

7.4.2.2 Scan mode. positive ion scan.

7.4.2.3 Detection mode. selective reaction monitoring.

7.4.2.4 Spray voltage. 4000V.

7.4.2.5 Ion transfer tube temperature. 330 °C.

7.4.2.6 Sheath Gas Pressure. 40arb.

7.4.2.7 Auxiliary gas pressure. 6arb.

7.4.2.8 optimization parameters selected reaction monitoring. Table 1.

Table 1 Sodium Benzene furanyl acid selected reaction monitoring of optimization parameters

Drug Name

Relative retention time min Qualitative ion pair m/z Quantitative ion pair m/z Collision energy eV Furan benzene acid 6.5 Sodium 290.18 > 202.43 290.18 > 228.26 290.18 > 228.26

7.4.3 Assay

Take a sample and standard solutions, multi-point calibration is calculated, the external standard method. The sample solution and the standard solution of sodium acid benzyl furan-average peak area it should be in the linear range of the detection instrument. Ion in the sample solution as compared to the relative abundance of the standard relative abundance of ions in solution, in line with

Table 2 is required. Added standard solution and blank sample solution ion mass chromatogram characteristic Appendix A.

The maximum allowable deviation relative ion abundance qualitative determination of Table 2 the relative ion abundances > 50 20 ~ 50 10 ~ 20 <=10 Permissible relative deviation ± 20 ± 25 ± 30 ± 50

7.5 Blank test

But without addition of the sample, the same steps employed in parallel operation.

Calculation and Expression of Results 8 from a standard curve calibrated by. AS = acS b to obtain a and b, then c = (Ab)/a (1) Residues in the specimens furan benzene acid sodium (µg/kg) according to formula (2).

A × cS × V

AS × m

Where.

--- the AS acid in a standard solution of sodium benzene furan-peak area;

Furan benzene concentration of sodium cS --- acid standard solution, units of nanograms per milliliter (ng/mL);

A --- furan sample solution of sodium benzene acid peak area;

--- the specimens residues X-furanyl benzene sodium acid micrograms per kilogram (µg/kg);

V --- The residue was dissolved in mobile phase volume in milliliters (mL);

m --- try supply feed mass in grams (g).

Note. The blank value should be subtracted from the results, expressed as the arithmetic mean of the measurement result measured parallel to three significant figures.

9.1 Sensitivity

The detection limit of the method was 0.2µg/kg, the limit of quantitation was 0.5µg/kg.

9.2 Accuracy

This method of adding 0.5µg/kg ~ 2µg/kg on recovery levels of 60% to 120%.

9.3 Precision

<=20% relative standard deviation in this method and inter - batch relative standard deviation of <=25%.

Appendix A

Chromatogram

Figure A.1 furan benzene acid sodium ion chromatogram of the standard solution (0.5µg/L)

Figure A.2 pig liver tissue sample blank ion chromatogram

Figure A.3 blank add pig liver tissue sample furan benzene acid sodium ion chromatogram (0.5µg/kg)