GB 5009.22-2016National food safety standard - B-group and G-group aflatoxins in foods (English PDF)

3.1 Reagents

3.2 Preparation of reagents

3.3 Standard substance

3.3.1 AFT B1 standard substance (C17H12O6, CAS. 1162-65-8). Purity >= 98%, or

3.4 Preparation of standard solution

4.9 Glass-fiber filter paper. Fast, high-load, which retains 1.6 µm particles in

liquid.

4.13 Liquid chromatography column.

4.14 Immunoaffinity column. Capacity of AFT B1 column >= 200 ng, recovery rate

of AFT B1 column >= 80%, cross-reaction rate of AFT G2 >= 80% (see Appendix B for verification method).

4.15 Aflatoxin-specific solid-phase extraction purification column or functionally

equivalent solid-phase extraction column (hereinafter referred to as purification column). which is used for the determination of samples of complex matrix.

4.18 pH meter.

5.1 Preparation of sample

5.1.1 Liquid samples (vegetable oil, soy sauce, vinegar, etc.)

The sampling amount shall be greater than 1 L. For packaging samples such as bags and bottles, it shall collect at least 3 packages (same batch or number).

All liquid samples shall be mixed by a homogenizer in a container, any 100 g (mL) of sample is taken for testing.

5.1.2 Solid samples (cereals and their products, nuts and seeds, cereal

supplements for infants, etc.)

The sampling amount shall be greater than 1 kg. It shall be crushed by high- speed pulverizer and sieved to make the particle size less than the 2 mm aperture test sieve. It is evenly mixed and then reduced to 100 g, stored in the sample bottle, sealed for preservation, to prepare for testing.

5.1.3 Semi-fluid (fermented bean curd, fermented soybean, etc.)

The sampling amount shall be greater than 1 kg (L). For packaging samples such as bags and bottles, it shall collect at least 3 packages (same batch or number). Then it is crushed and mixed uniformly by the tissue crusher, stored in sample bottle, sealed for preservation, to prepare for testing.

5.2 Extraction of sample

5.2.1 Liquid sample

5.2.2 Solid sample

5.2.2.1 General solid samples

WEIGH 5 g of specimen (accurate to 0.01 g) into a 50 mL centrifuge tube; ADD

100 µL of isotope internal standard working solution; OSCILLATE to mix it; LET it be standing for 30 min. ADD 20.0 mL of acetonitrile-water solution (84 + 16) or methanol-water solution (70 + 30); VORTEX to mix it uniformly; PLACE it in the ultrasonic/vortex oscillator or shaker to oscillate it for 20 min (or homogenized by homogenizer for 3 min)

5.2.2.2 Infant formula and infant complementary foods

WEIGH 5 g of specimen (accurate to 0.01 g) into a 50 mL centrifuge tube; ADD

100 µL of isotope internal standard working solution; OSCILLATE to mix it; LET it be standing for 30 min. ADD 20.0 mL of acetonitrile-water solution (50 + 50) or methanol-water solution (70 + 30); VORTEX to mix it uniformly; PLACE it in the ultrasonic/vortex oscillator or shaker to oscillate it for 20 min (or homogenized by homogenizer for 3 min).

5.2.3 Semi-fluid sample

WEIGH 5 g of specimen (accurate to 0.01 g) into a 50 mL centrifuge tube; ADD

100 µL of isotope internal standard working solution; OSCILLATE to mix it; LET it be standing for 30 min. ADD 20.0 mL of acetonitrile-water solution (84 + 16) or methanol-water solution (70 + 30); PLACE it in the ultrasonic/vortex oscillator or shaker to oscillate it for 20 min (or homogenized by homogenizer for 3 min).

CENTRIFUGE it at 6000 r/min for 10 min (or otherwise homogenized and then filtered by glass-fiber filter paper), TAKE the supernatant to prepare for use.

5.3 Purification of sample

5.3.1 Purification of immunoaffinity column

5.3.2 Simultaneous use of aflatoxin solid-phase purification column and

immunoaffinity column (for complex substrates such as Chinese red pepper, black pepper and chili pepper)

5.5 Reference conditions for mass spectrometry

The reference conditions for mass spectrometry are listed below.

5.6 Qualitative determination

The retention time of the chromatographic peak of the target compound in the specimen is compared with the retention time of the corresponding standard chromatographic peak, the variation range shall be within ±2.5%.

The mass spectrometric ions of each compound must appear, including at least one parent ion and two daughter ions. Meanwhile for the same batch under testing and for the same compound, the allowable deviation between the relative abundance ratio of the two daughter ions of the target compound in the sample and that of the standard solution of equivalent concentration shall not exceed the range as specified in Table 3.

5.7 Production of standard curve

Under the analytical conditions of liquid chromatography-tandem mass spectrometry of 5.4 and 5.5, MAKE the standard series solution subject to injection testing in the order from low to high concentration; USE the area ratio of the AFT B1, AFT B2, AFT G1 and AFT G2 chromatogram peaks to each corresponding internal standard chromatogram peak as well as the concentration to make drawing, to obtain the standard curve regression equation, the linear correlation coefficient of which is greater than 0.99.

5.8 Determination of sample solution

TAKE the sample solution to be tested which is processed in 5.3; USE the internal standard method to calculate the mass concentration of the target substance in the tested solution. Follow the provisions of clause 6 to calculate the content of the tested substance in the sample. The response value in the

5.9 Blank test

DO not weigh the specimen; PERFORM the blank test in accordance with the steps of 5.2 and 5.3.It shall be confirmed that it does not contain substances that interfere with the component to be tested.