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GB 5009.243-2016 PDF English

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GB 5009.243-2016: National food safety standard - Determination of Heterocyclic Amine Substances in High-temperature Cooked Foods
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GB 5009.243-2016: National food safety standard - Determination of Heterocyclic Amine Substances in High-temperature Cooked Foods


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National food safety standard - Determination of Heterocyclic Amines in High Temperature Cooked Foods GB NATIONAL STANDARD OF THE PEOPLE’S REPUBLIC OF CHINA National Food Safety Standard - Determination of Heterocyclic Amine Substances in High-temperature Cooked Foods Issued on. August 31, 2016 Implemented on. March 01, 2017 Issued by. National Health and Family Planning Commission of the PRC

Table of Contents

1 Scope ...3 2 Principles ...3 3 Reagents and materials ...3 4 Instruments and apparatuses ...5 5 Analytical procedures ...6 6 Result calculation ...8 7 Recovery rate and precision ...8 8 Other ...8 Appendix A Gradient elution procedures ...9 Appendix B Mass spectrometry reference conditions ...10 Appendix C Standard solution chromatogram ...11 National Food Safety Standard - Determination of Heterocyclic Amine Substances in High-temperature Cooked Foods

1 Scope

This Standard specifies the liquid chromatography-mass spectrometry/mass spectrometry for the determination of heterocyclic amines in high-temperature cooked foods such as 2-amino-3,4-dimethylimidazo [4,5-f] quinoline (MeIQ), 2- amino-3,8-dimethylimidazo [4,5-f] quinoline (MeIQx), 2-amino-3,4,8- trimethylimidazo [4,5-f] quinoline (4,8-DiMeIQx), 2-amino-3,7,8- trimethylimidazo [4,5-f] quinoline (7,8-DiMeIQx), 2-amino-1-methyl-6-phenyl- imidazo [4,5-b] pyridine (PhIP). This Standard applies to the determination of MeIQ, MeIQx, 4,8-DiMeIQx, 7,8- DiMeIQx, PhIP in grilled fish, barbecue, and their products.

2 Principles

The sample is extracted using sodium hydroxide/methanol solution, purified by solid-phase extraction column, detected by liquid chromatography-tandem mass spectrometry, and quantified by internal standard method.

3 Reagents and materials

Unless otherwise specified, the reagents used are analytically pure; and the water is Grade 1 water specified in GB/T 6682. 3.1 Reagents 3.1.1 Sodium hydroxide. 3.1.2 Ammonium acetate. Purity≥98%. 3.1.3 Methanol. chromatographically pure. 3.1.4 Ethanol. chromatographically pure. 3.1.5 N-hexane. chromatographically pure. 3.1.6 Dichloromethane. chromatographically pure. 3.1.7 Acetonitrile. chromatographically pure. 3.1.8 Glacial acetic acid. chromatographically pure. 3.2 Reagents preparation 3.2.1 Sodium hydroxide solution (40 g/L). WEIGH 40.0 g of sodium hydroxide; USE water to dissolve and dilute to 1 L. 3.2.2 Sodium hydroxide solution (4 g/L). MEASURE and TAKE 50 mL of 40 g/L sodium hydroxide solution (3.2.1); ADD 450 mL of water; MIX well. 3.2.3 40 g/L sodium hydroxide-methanol mixed solution (70+30, volume fraction). MEASURE and TAKE 70 mL of 40 g/L sodium hydroxide solution (3.2.1); ADD 30 mL of methanol; MIX well. 3.2.4 4 g/L sodium hydroxide-methanol mixed solution (45+55, volume fraction). MEASURE and TAKE 45 mL of 4 g/L sodium hydroxide solution (3.2.2); ADD 55 mL of methanol; MIX well. 3.2.5 Ethanol-dichloromethane mixed solution (10+90, volume fraction). MEASURE and TAKE 10 mL of ethanol; ADD 90 mL of dichloromethane; MIX well. 3.2.6 Acetonitrile-water solution (5+95, volume fraction). MEASURE and TAKE 5 mL of acetonitrile; ADD 95 mL of water; MIX well. 3.2.7 Acetic acid-ammonium acetate buffer solution. WEIGH 1.155 g of ammonium acetate; USE 450 mL of water to dissolve; USE acetic acid to adjust the pH to 5.0±0.5; ADD water to dilute to 500 mL. 3.2.8 Acetic acid buffer solution-acetonitrile mixed solution (50+50, volume fraction). MEASURE and TAKE 50 mL of acetic acid-ammonium acetate buffer solution (3.2.7); ADD 50 mL of acetonitrile; MIX well. 3.3 Standards 3.3.1 Heterocyclic amine standard substances. MeIQ (C12H12N4, 77094-11-2), MeIQx (C11H11N5, 77500-04-0), 4,8-DiMeIQx (C12H13N5, 95896-78-9), 7,8- DiMeIQx (C12H13N5, 92180-79-5), PhIP (C13H12N4, 105650-23-5).The purity is greater than 99%. 3.3.2 Internal standard standard substance 4,7,8-TriMeIQx (C13H15N5, 132898- 07-8).The purity is greater than 99%.

5 Analytical procedures

5.1 Sample preparation The edible portion from grilled fish, barbecue, and their products is taken, mashed, and mixed; and after being labelled, stored frozen at -18 °C. 5.2 Sample processing 5.2.1 Extraction. WEIGH 2 g (accurate to 0.01 g) of sample into a 50 mL centrifuge tube; ADD 200 μL of internal standard working solution (3.4.4); and then ADD 9.8 mL of 40 g/L sodium hydroxide-methanol mixed solution (3.2.3); homogenize for 1 min.The homogenizer cutter head is washed twice using 5.0 mL of 40 g/L sodium hydroxide-methanol mixed solution (3.2.3).The washing liquid is combined into a sample extraction centrifuge tube.The sample is centrifuged at 10000 r/min for 10 min to be purified. 5.2.2 Purification. The solid-phase extraction column (3.5.2) is activated in advance subsequently using 2 mL of methanol and 3 mL of 4 g/L sodium hydroxide solution (3.2.2).MEASURE and TAKE 10 mL of extract into the solid- phase extraction column.After discarding the effluent, subsequently USE 3 mL of 4 g/L sodium hydroxide-methanol mixed solution (3.2.4) and 2 mL of n- hexane to rinse.After each rinse, the rinse solution in the column shall be drained.Finally, USE 1.5 mL of ethanol-dichloromethane solution (3.2.5) to elute.The elution flow rate shall be less than 1 mL/min.After the eluent is nitrogen-concentrated in a water bath at 35 °C to near dryness, 1.0 mL of acetic acid buffer solution-acetonitrile mixed solution (3.2.8) is added; vortex-mixed; and filtered through the microporous membrane (3.5.1) to the injection vial, for assay determination on the machine. 5.3 Determination 5.3.1 Liquid chromatographic conditions 5.3.1.1 Chromatographic column. C18 column (2.5 μm, 100 mm×2.1 mm) or equivalent. 5.3.1.2 Mobile phase. A is acetic acid-ammonium acetate buffer solution (3.2.7). B is acetonitrile.For the gradient elution procedures, see Appendix A, A.1. 5.3.1.3 Flow rate. 0.3 mL/min. 5.3.1.4 Column temperature. 40 °C. 5.3.1.5 Injection volume. 5 μL. ......
Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
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