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GB 5009.240-2023 English PDF (GB5009.240-2023)

GB 5009.240-2023 English PDF (GB5009.240-2023)

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GB 5009.240-2023: National food safety standard - Determination of fumonisin in foods
GB 5009.240-2023
GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard - Determination of
Fumonisin in Foods
ISSUED ON. SEPTEMBER 6, 2023
IMPLEMENTED ON. MARCH 6, 2024
Issued by. National Health Commission of the People’s Republic of China;
State Administration for Market Regulation.
Table of Contents
Foreword... 3
1 Scope... 4
Method I - Immunoaffinity Column Purification - Post-column Derivatization High-
performance Liquid Chromatography... 4
2 Principle... 4
3 Reagents and Materials... 4
4 Instruments and Equipment... 7
5 Analytical Procedures... 7
6 Expression of Analysis Results... 9
7 Precision... 10
8 Others... 10
Method II - High-performance Liquid Chromatography - Tandem Mass Spectrometry
... 10
9 Principle... 10
10 Reagents and Materials... 11
11 Instruments and Equipment... 13
12 Analytical Procedures... 14
13 Expression of Analysis Results... 18
14 Precision... 19
15 Others... 19
Method III - Immunoaffinity Column Purification - Pre-column Derivatization High-
performance Liquid Chromatography... 19
16 Principle... 19
17 Reagents and Materials... 19
18 Instruments and Equipment... 22
19 Analytical Procedures... 22
20 Expression of Analysis Results... 25
21 Precision... 25
22 Others... 25
Appendix A Post-column Derivatization - High-performance Liquid Chromatogram 27
Appendix B MRM Mass Chromatogram... 28
Appendix C Pre-column Derivatization - High-performance Liquid Chromatogram. 31
Appendix D Column Capacity and Column Recovery Verification Methods... 32
National Food Safety Standard - Determination of
Fumonisin in Foods
1 Scope
This Standard specifies the immunoaffinity column purification - post-column derivatization
high-performance liquid chromatography, high-performance liquid chromatography - tandem
mass spectrometry and immunoaffinity column purification - pre-column derivatization high-
performance liquid chromatography for the determination of fumonisin in foods.
This Standard is applicable to the determination of fumonisin B1, fumonisin B2 and fumonisin
B3 (hereinafter referred to as FB1, FB2 and FB3) in cereals and products, cereal supplementary
foods for infants and young children, and vegetable oils.
Method I - Immunoaffinity Column Purification - Post-
column Derivatization High-performance Liquid
Chromatography
2 Principle
The specimen is extracted, purified by an immunoaffinity column, separated by C18 reversed-
phase chromatography column, derivatized by o-phthalaldehyde, detected by fluorescence
detector, and quantified by the external standard method.
3 Reagents and Materials
Unless it is otherwise specified, the reagents used in this Method are all analytically pure, and
the water is Grade-1 water specified in GB/T 6682.
3.1 Reagents
3.1.1 Methanol (CH3OH). chromatographically pure.
3.1.2 Acetonitrile (CH3CN). chromatographically pure.
3.1.3 Formic acid (HCOOH). chromatographically pure.
3.1.4 Acetic acid (CH3COOH).
3.1.5 Sodium hydroxide (NaOH).
3.1.6 Sodium chloride (NaCl).
3.1.7 Disodium hydrogen phosphate (Na2HPO4).
3.1.8 Potassium dihydrogen phosphate (KH2PO4).
3.1.9 Potassium chloride (KCl).
3.1.10 Borax (Na2B4O7  10H2O).
3.1.11 2-mercaptoethanol (C2H6OS).
3.1.12 o-phthalaldehyde (C8H6O, OPA).
3.1.13 Tween-20 (C58H114O26).
3.1.14 Hydrochloric acid (HCl).
3.2 Preparation of Reagents
3.2.1 Formic acid - water solution (0.1%). accurately transfer-take 1 mL of formic acid, use
water to dilute to 1,000 mL and evenly mix it.
3.2.2 Acetonitrile - water solution (50 + 50). respectively measure-take 500 mL of acetonitrile
and 500 mL of water, and evenly mix them.
3.2.3 Acetonitrile - water solution (20 + 80). respectively measure-take 20 mL of acetonitrile
and 80 mL of water, and evenly mix them.
3.2.4 Methanol - acetic acid solution (98 + 2). accurately transfer-take 2 mL of acetic acid, use
methanol to dilute to 100 mL and evenly mix it.
3.2.5 Sodium hydroxide solution (2 mol/L). accurately weigh-take 8.0 g of sodium hydroxide,
add 50 mL of water to dissolve it; after cooling, use water to dilute it to 100 mL and evenly mix
it.
3.2.6 Phosphate buffer solution (PBS). weigh-take 8.0 g of sodium chloride, 1.2 g of disodium
hydrogen phosphate, 0.2 g of potassium dihydrogen phosphate and 0.2 g of potassium chloride;
use 980 mL of water to dissolve it, use hydrochloric acid to adjust pH to 7.4, use water to dilute
to 1,000 mL and evenly mix it.
3.2.7 Tween-20/PBS solution (0.1%). weigh-take 1.0 g of Tween-20, add phosphate buffer
solution, dilute to 1,000 mL and evenly mix it.
3.2.8 Borax solution (0.05 mol/L, pH 10.5). weigh-take 19.1 g of borax, dissolve it in 980 mL
of water, use sodium hydroxide solution to adjust pH to 10.5, use water to dilute to 1,000 mL
and evenly mix it.
3.2.9 Derivatization solution. weigh-take 0.5 g of o-phthalaldehyde, dissolve it in 20 mL of
methanol, use borax solution (0.05 mol/L, pH 10.5) to dilute to 500 mL, add 500 L of 2-
mercaptoethanol and evenly mix it. After filtration, put it into a brown bottle; store it at room
temperature away from light. It shall remain valid for 1 week.
3.3 Reference Materials
3.3.1 Fumonisin B1 (C34H59NO15, FB1, CAS No.. 116355-83-0), purity  95%, or a standard
substance certified by the state and awarded a standard substance certificate.
3.3.2 Fumonisin B2 (C34H59NO14, FB2, CAS No.. 116355-84-1), purity  95%, or a standard
substance certified by the state and awarded a standard substance certificate.
3.3.3 Fumonisin B3 (C34H59NO14, FB3, CAS No.. 136379-59-4), purity  95%, or a standard
substance certified by the state and awarded a standard substance certificate.
3.4 Preparation of Standard Solutions
3.4.1 Standard stock solutions (100 g/mL). respectively and accurately weigh-take 10 mg
(accurate to 0.01 mg) of FB1, FB2 and FB3 into small beakers, use acetonitrile - water solution
(50 + 50) to dissolve them, and respectively transfer to 100 mL volumetric flasks. Use
acetonitrile - water solution (50 + 50) to reach a constant volume to the scale. Store them at 18
C away from light. They shall remain valid for 6 months.
3.4.2 Mixed standard stock solution. accurately transfer-take 1.0 mL of FB1 standard stock
solution, 0.5 mL of FB2 standard stock solution and 0.5 mL of FB3 standard stock solution into
the same 10 mL volumetric flask; add acetonitrile - water solution (50 + 50) to dilute to the
scale. The mass concentration of FB1 is 10 g/mL, and the mass concentration of FB2 and FB3
is 5 g/mL. Store it at 18 C away from light. It shall remain valid for 6 months.
3.4.3 Mixed standard working solution. accurately transfer-take 1.0 mL of the mixed standard
stock solution into a 10.0 mL volumetric flask, add acetonitrile - water solution (50 + 50) to
dilute it and reach a constant volume to the scale. The mass concentration of FB1 is 1 g/mL,
and the mass concentration of FB2 and FB3 is 0.5 g/mL. Store it at 4 C away from light. It
shall remain valid for 6 months.
3.4.4 Mixed standard series of working solutions. accurately transfer-take the mixed standard
working solution, use acetonitrile - water solution (20 + 80) to dilute it, and prepare mixed
standard series of working solutions respectively with a FB1 mass concentration of 20.0 ng/mL,
80.0 ng/mL, 160 ng/mL, 240 ng/mL, 320 ng/mL, 400 ng/mL and 480 ng/mL, and a FB2 and
FB3 mass concentration of 10.0 ng/mL, 40.0 ng/mL, 80....
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