GB 5009.305-2025 PDF English
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National food safety standard - Determination of bisphenol A, bisphenol F and bisphenol S in foods
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GB 5009.305-2025: National food safety standard - Determination of bisphenol A, bisphenol F and bisphenol S in foods
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GB
NATIONAL STANDARD OF THE
PEOPLE’S REPUBLIC OF CHINA
National Food Safety Standard - Determination of Bisphenol
A, Bisphenol F and Bisphenol S in Foods
Issued on: MARCH 16, 2025
Implemented on: SEPTEMBER 16, 2025
Issued by. National Technical Commission of the People’s Republic of China;
State Administration for Market Regulation.
Table of Contents
1 Scope... 3
2 Principle... 3
3 Reagents and Materials... 3
4 Instruments and Equipment... 5
5 Analysis Procedures... 6
6 Expression of Analysis Results... 10
7 Precision... 10
8 Others... 11
Appendix A Immunoaffinity Column Verification Method... 12
Appendix B Liquid Chromatography-Mass Spectrometry/Mass Spectrometry of
Bisphenol A, Bisphenol F and Bisphenol S Standard Solutions... 13
National Food Safety Standard - Determination of Bisphenol
A, Bisphenol F and Bisphenol S in Foods
1 Scope
This Standard specifies the liquid chromatography-tandem mass spectrometry method for the
determination of bisphenol A, bisphenol F and bisphenol S in foods.
This Standard is applicable to the determination of bisphenol A, bisphenol F and bisphenol S
in meat and meat products, milk and dairy products, aquatic products, cereals, eggs, vegetables,
fruits, beverages, infant formula foods and infant cereal complementary foods.
2 Principle
Bisphenol A, bisphenol F and bisphenol S in the specimen were extracted with acetonitrile or
methanol-water solution, purified by immunoaffinity column, determined by liquid
chromatography-tandem mass spectrometry and quantified by internal standard method.
3 Reagents and Materials
Unless otherwise specified, all reagents used in this method were analytically pure and water
was Grade-1 water as specified in GB/T 6682.
3.1 Reagents
3.1.1 Acetonitrile (CH3CN). Chromatographically pure.
3.1.2 Methanol (CH3OH). Chromatographically pure.
3.1.3 Sodium chloride (NaCl).
3.1.4 Disodium hydrogen phosphate (Na2HPO4).
3.1.5 Potassium dihydrogen phosphate (KH2PO4).
3.1.6 Potassium chloride (KCl).
3.1.7 Hydrochloric acid (HCl). Guaranteed reagent.
3.2 Preparation of reagents
3.2.1 Hydrochloric acid solution (10+90). Pipette 10 mL of hydrochloric acid; add to 90 mL of
water; and mix well.
3.2.2 Phosphate buffer solution (hereinafter referred to as PBS). Weigh 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; dissolve in 900 mL of water; adjust the pH to 7.4±0.1 with hydrochloric
acid solution (10+90); and then dilute to 1,000 mL with water.
3.2.3 Methanol-water solution (80+20). Pipette 80 mL of methanol; add 20 mL of water; and
mix well.
3.2.4 Methanol-water solution (40+60). Pipette 40 mL of methanol; add 60 mL of water; and
mix well.
3.3 Standard sample
3.3.1 Bisphenol A standard sample (C15H16O2, CAS. 80-05-7). Purity ≥ 99.0%; or a standard
sample certified by the state and granted a standard substance certificate.
3.3.2 Bisphenol F standard sample (C13H12O2, CAS. 620-92-8). Purity ≥ 99.0%, or a standard
sample certified by the state and granted a standard substance certificate.
3.3.3 Bisphenol S standard sample (C12H10O4S, CAS. 80-09-1). purity ≥ 99.0%, or a standard
sample certified by the state and awarded a standard substance certificate.
3.3.4 13C12-Bisphenol A isotope internal standard (C313C12H16O2, CAS. 263261-65-0). 100 mg/L,
or a solid standard sample with a purity of ≥ 98.0%.
3.3.5 13C12-Bisphenol F isotope internal standard (C13C12H12O2). 100 mg/L, or a solid standard
sample with a purity of ≥ 98.0%.
3.3.6 13C12-bisphenol S isotope internal standard (13C12H10O4S). 100 mg/L, or solid standard
sample with a purity of ≥98.0%.
3.4 Preparation of standard solutions
3.4.1 Standard stock solution (100 mg/L). Accurately weigh 10 mg (accurate to 0.1 mg) of
bisphenol A, bisphenol F and bisphenol S standard samples; dissolve them in methanol and
dilute to 100 mL, respectively; and mix well. Transfer the solution to a brown glass reagent
bottle and store it at -18 ℃ away from light. The validity period is 12 months.
3.4.2 Standard intermediate solution (1 mg/L). Accurately pipette 0.10 mL of the standard stock
solution of bisphenol A, bisphenol F and bisphenol S, respectively, into 10 mL volumetric flasks;
make constant volume to the scale with methanol; and mix well. Transfer the solution to a
brown glass reagent bottle and store it at -18 ℃ away from light. The validity period is 6 months.
3.4.3 Standard mixed solution (bisphenol A and bisphenol F. 100 μg/L; bisphenol S. 10 μg/L).
Accurately pipette 1.00 mL of each standard intermediate solution of bisphenol A and bisphenol
F, respectively, and 0.10 mL of the standard intermediate solution of bisphenol S; place them
in a 10 mL volumetric flask; make constant volume to the scale with methanol-water solution
(40+60); and mix well. Prepare immediately before use.
3.4.4 Isotope internal standard intermediate solution (1 mg/L). Accurately pipette 0.10 mL of
each 13C12-bisphenol A standard solution, 13C12-bisphenol F standard solution, and 13C12-
bisphenol S standard solution; respectively place them in 10 mL volumetric flasks; make
constant volume to the scale with methanol; and mix well. Transfer the solution to a brown
glass reagent bottle and store it at -18 ℃ away from light. The validity period is 6 months.
3.4.5 Mixed isotope internal standard solution (13C12-bisphenol A and 13C12-bisphenol F. 100
μg/L; 13C12-bisphenol S. 10 μg/L). Accurately pipette 1.00 mL of each 13C12-bisphenol A and
13C12-bisphenol F intermediate solution and 0.10 mL of 13C12-bisphenol S intermediate solution,
respectively; place them in a 10 mL volumetric flask; make constant volume to the scale with
methanol-water solution (40+60); and mix well. Prepare immediately before use.
3.4.6 Standard series working solution. Accurately and respectively pipette 0.10 mL, 0.20 mL,
0.50 mL, 0.80 mL, 1.00 mL, and 2.00 mL of the standard mixed working solution into a 10 mL
volumetric flask; add 0.50 mL of isotope internal standard mixed working solution; make
constant volume to the scale with methanol-water solution (40+60); and mix well. The mass
concentrations of bisphenol A and bisphenol F in the standard series working solutions are 1.00
μg/L, 2.00 μg/L, 5.00 μg/L, 8.00 μg/L, 10.0 μg/L, and 20.0 μg/L, respectively; the mass
concentrations of bisphenol S are 0.100 μg/L, 0.200 μg/L, 0.500 μg/L, 0.800 μg/L, 1.00 μg/L,
and 2.00 μg/L, respectively; the mass concentrations of 13C12-bisphenol A and 13C12-bisphenol
F are 5.00 μg/L; and the mass concentration of 13C12-bisphenol S is 0.500 μg/L. Prepare
immediately before use.
3.5 Materials
3.5.1 Bisphenol A, bisphenol F and bisphenol S composite immunoaffinity column. column
capacity of each compound ≥200 ng, column recovery ≥90% (see Appendix A for verification
method).
NOTE. Different batches of immunoaffinity columns need to be quality verified before use.
3.5.2 Polypropylene centrifuge tubes. 2 mL and 50 mL.
3.5.3 Glass nitrogen blowpipe. 15 mL.
4 Instruments and Equipment
4.1 Liquid chromatography-tandem quadrupole mass spectrometer. Equipped with electrospray
ion source.
5.2.2 Eggs, milk and liquid dairy products
Weigh 1 g of specimen (accurate to 0.01g); place it in a 50 mL polypropylene centrifuge tube;
add 50 μL of isotope internal standard mixed solution; shake and mix; and stand for 30 min.
Add 5 mL of acetonitrile; vortex and mix; then ultrasonically extract for 15 min; and centrifuge
at 10,000 r/min and at 4 ℃ for 10 min. Take the supernatant and place it in a glass nitrogen
blowpipe; slowly blow it with nitrogen at 40 ℃ to about 2 mL; add 8 mL of PBS and mix well;
and wait for purification.
5.2.3 Cereals and infant cereal complementary foods
Weigh 1 g of specimen (accurate to 0.01 g); place in a 50 mL polypropylene centrifuge tube;
add 50 μL of isotope internal standard mixed solution; shake and mix; and stand for 30 min.
Add 10 mL of methanol-water solution (80 +20); vortex for 30 s; ultrasonically extract for 15
min; and centrifuge at 10,000 r/min and at 4 ℃ for 10 min. Take the supernatant and place it in
a glass nitrogen blowpipe; slowly blow it with nitrogen at 40 ℃ to about 2 mL; add 8 mL of
PBS and mix well; and wait for purification.
5.2.4 Vegetables and fruits
Weigh 1 g of specimen (accurate to 0.01 g); place in a 50 mL polypropylene centrifuge tube;
add 100 μL of hydrochloric acid solution (10+90) and mix well. Then add 50 μL of isotope
internal standard mixed solution; shake and mix; and stand for 30 min. Add 5 mL of acetonitrile;
vortex for 30 s; ultrasonically extract for 15 min; and centrifuge at 10,000 r/min and at 4 °C for
10 min. Take the supernatant and place it in a glass nitrogen blowpipe; slowly blow it with
nitrogen at 40 °C to about 2 mL; add 8 mL of PBS and mix well; and wait for purification.
5.2.5 Beverages
Weigh 1 g of specimen (accurate to 0.01 g); place it in a 2 mL polypropylene centrifuge tube;
add 50 μL of isotope internal standard mixed solution; shake and mix; and stand for 30 min.
Centrifuge at 10,000 r/min for 10 min. Take the supernatant to a 50 mL polypropylene
centrifuge tube; add 10 mL of PBS and mix well; and wait for purification.
NOTE. Clarified specimens (such as purified water, mineral water, etc.) can be directly added to PBS
and mixed well without centrifugation.
5.3 Purification of specimen
Restore the immunoaffinity column stored at low temperature to room temperature; and then
let the original liquid in the column flow out naturally. Take all the specimen extracts in 5.2 and
pass them through the column; let them flow out naturally; discard all the effluent; and then
rinse the immunoaffinity column with 10 mL of water. After the water drips out, use a vacuum
pump to dry the immunoaffinity column. Add 1 mL of methanol for elution; collect the eluate
in a glass nitrogen blowpipe; and slowly blow dry with nitrogen at 40 °C. Accurately add 0.40
mL of methanol; vortex and mix for 10 s; then accurately add 0.60 mL of water; vortex and mix
......
Source: Above contents are excerpted from the full-copy PDF -- translated/reviewed by: www.ChineseStandard.net / Wayne Zheng et al.
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