GB/T 18415-2026Determination of benzoyl peroxide in wheat flour and its products (English PDF)

4.2.13 Benzoic acid standard working solution (100 µg/mL). Accurately pipette

10.00 mL of benzoic acid standard stock solution (4.2.12) into... Store in a refrigerator at 2°C~8°C. Shelf life is 3 months.

4.2.14 Capped centrifuge tubes. 50 mL.

4.2.15 Glass beads. 5mm~6mm in diameter.

4.3 Instruments and Equipment

4.3.1 Gas chromatograph. equipped with a flame ionization detector.

4.3.2 Analytical balance. sensitivity of 0.01g and 0.0001g.

4.3.3 Crusher. capable of crushing dry samples to the point that all of them can pass through a sieve with a mesh size of 2 mm.

4.3.4 Tissue homogenizer. capable of pulverizing wet samples to the point that all of them pass through a sieve with a sieve aperture of 2 mm.

4.3.5 Oscillator. reciprocating type, oscillation frequency.200 times/min.

4.3.6 High-speed centrifuge. speed >= 10000 r/min.

4.3.7 Vortex mixer.

4.4 Analysis Steps

4.4.1 Sample Preparation Take at least.200g of representative sample from all samples. Place the wheat flour sample directly into a clean container, seal it, and store it for later use. After being cut or chopped, the wheat flour products are ground in a grinder (4.3.3) or a tissue homogenizer (4.3.4) until all of them pass through a 2mm aperture. Sift, mix well, and store in a clean, sealed container for later use.

4.4.2 Sample Pretreatment Weigh 5g (accurate to 0.01g) of the sample into a 50mL capped centrifuge tube (4.2.14) pre-filled with 5 to 6 glass beads (4.2.15). Add 20 mL of sodium thiosulfate solution (4.2.7) to the centrifuge tube (4.2.14) and mix well. Place the capped centrifuge tube horizontally on a shaker (4.3.5) and centrifuge. Shake at a frequency of.200 times/min for 30 minutes, add 10 mL of sodium bicarbonate solution (4.2.8), repeat shaking for 5 minutes, and then place in a high-speed centrifuge. (4.3.6) Centrifuge at 10000 r/min for 5 min, take 15 mL of the supernatant aqueous solution and place it in another container with 5 g to 6 g of sodium chloride placed in it beforehand. (4.2.6) In a 50 mL capped centrifuge tube (4.2.14), shake thoroughly to dissolve sodium chloride until saturated, then add

0.5 mL of hydrochloric acid solution. (4.2.9) First, gently shake to release the gas, then shake thoroughly to release the gas. Next, add 5 mL of petroleum ether-diethyl ether mixture (4.2.10) and place it in a vortex mixer. Vortex in (4.3.7) for 1 min, then place in a high-speed centrifuge (4.3.6) and centrifuge at 10000 r/min for 5 min. Place the supernatant in a container... The sample bottle is ready for testing.

4.4.3 Creation of Standard Working Curves Weigh out 6 portions of 5g (accurate to 0.01g) blank samples of the same matrix that do not contain benzoic acid and benzoyl peroxide, and pre-fill 5 tablets. Add 20 mL of sodium thiosulfate solution (4.2.7) to a 50 mL capped centrifuge tube (4.2.14) containing 6 glass beads (4.2.15) and mix well. Place the capped centrifuge tubes (4.2.14) horizontally on a shaker (4.3.5) and shake at a frequency of.200 times/min for 30 minutes. After removing the tubes, add benzoic acid. The standard solutions (4.2.13) are 0 mL,

0.10 mL,

0.50 mL,

5.0 mL, followed by sodium bicarbonate solution (4.2.8). Add to a total volume of 10 mL, and proceed as described in section 4.4.2, "Place the capped centrifuge tube (4.2.14) horizontally in the shaker" during sample pretreatment. (4.3.5) Shake for 5 minutes, then place the supernatant in a sample vial for testing. The final mass concentrations of the standard working solutions are as follows: 0µg/mL, 1.0µg/mL, 5.0µg/mL, 10µg/mL, 25µg/mL and 50µg/mL. The standard series working solutions were injected sequentially into the gas chromatograph in order of increasing concentration (4.3.1) to obtain the standard solutions at each concentration. Chromatogram of the solution. A standard curve was plotted with the benzoic acid concentration (µg/mL) on the x-axis and the peak area on the y-axis. (Standard solution) The gas chromatogram is shown in Figure A.1 in Appendix A.

4.4.4 Measurement Under the same conditions as the standard curve determination, the sample solution to be tested was injected into the gas chromatograph (4.3.1), and the retention time was used for qualitative analysis to determine the peak surface. The concentration of benzoic acid in the sample solution is calculated based on the standard curve.

4.4.5 Instrument Reference Conditions The instrument reference conditions are as follows:

a) Chromatographic column. Nitro-terephthalic acid modified polyethylene glycol highly polar capillary column, 30m × 0.53mm × 0.50µm, or Comparable in performance;

b) Inlet temperature. 250°C;

c) Detector temperature. 250°C;

d) Column temperature. 190°C for 6 min, then increased to 240°C at a rate of 20°C/min and held for

e) Nitrogen. 8 mL/min;

4.5 Calculation and Presentation of Results

4.5.1 Result Calculation The content (X) of benzoyl peroxide in the sample is expressed as a mass fraction and calculated according to formula (1).

4.5.2 Results Presentation The results are expressed as the arithmetic mean of two independent measurements obtained under repeatability conditions, and are retained to two significant figures.

4.6 Precision Under repeatability conditions, the absolute difference between two independent measurements should not exceed 10% of the arithmetic mean; under reproducibility conditions... The absolute difference between two independent measurements should not exceed 15% of the arithmetic mean.

4.7 Other When the sample size is 5g, the detection limit of this method is 0.0005g/kg and the quantitation limit is 0.0015g/kg.

5.2.6 Ammonium acetate solution (20 mmol/L). Weigh

1.54 g of ammonium acetate (5.2.2), dissolve and dilute to 1000 mL with water, mix well, and then... Use after filtration with a 0.45µm water-type filter membrane (5.2.14).

5.2.7 Ammonium acetate and formic acid solution (20 mmol/L ammonium acetate and 2 mmol/L formic acid). Weigh

1.54 g of ammonium acetate (5.2.2) and dissolve it in water to dilute it. Dilute to 1000 mL, add 75.2 µL of formic acid (5.2.3), mix well, and filter through a 0.45 µm aqueous filter membrane (5.2.14) before use.

5.2.8 Benzoic acid (C6H5COOH, CAS No.. 65-85-0). Purity >= 99.0%, or a standard sample certified by the state and granted a certificate. Standard/Standard Substances.

5.2.9 Benzoic acid standard stock solution (1 mg/mL). Accurately weigh 100 mg of benzoic acid (5.2.8) (accurate to

0.1 mg) into 100 mL of caustic soda. In a glass, dissolve the contents in methanol (5.2.4), then transfer to a 100 mL volumetric flask and dilute to the mark. Mix well. Transfer the solution to a brown reagent bottle and seal tightly. Store in a refrigerator at -18°C or below. Shelf life is 6 months.

5.2.10 Benzoic acid standard intermediate solution (100 µg/mL). Accurately pipette

2.5 mL of benzoic acid standard stock solution (5.2.9) into a 25 mL container. Dilute with methanol (5.2.4) in a volumetric flask and bring to volume to the mark. Mix well and prepare fresh before use.

5.2.11 Benzoic acid standard working solution. Accurately pipette 0 mL,

0.20 mL,

0.80 mL, and

0.90 mL of the intermediate benzoic acid standard solution (5.2.10) respectively.

10.0 mL were added to

25.0 mL volumetric flasks and diluted to volume with methanol (5.2.4). The standard curve uses solutions with different mass concentrations. The concentrations are 0 µg/mL, 0.80 µg/mL, 3.2 µg/mL, 8.0 µg/mL, 32 µg/mL, and 40 µg/mL, and should be prepared fresh before use.

5.2.12 Syringe. 2mL.

5.2.13 Needle-type microporous filter membrane. pore size 0.22µm, organic type.

5.2.14 Filter membrane. pore size 0.45µm, water type.

5.2.15 Stoppered colorimetric tubes. 50 mL.

5.3 Instruments and Equipment

5.3.1 High-performance liquid chromatograph. equipped with an ultraviolet detector or a diode array detector.

5.3.2 Analytical balance. sensitivity of 0.01g and 0.0001g.

5.3.3 Crusher. capable of crushing dry samples to the point that all of them pass through a sieve with a sieve aperture of 2 mm.

5.3.4 Tissue homogenizer. capable of pulverizing wet samples to the point that all of them pass through a sieve with a sieve aperture of 2 mm.

5.3.5 Vortex mixer.

5.4 Analysis Steps

5.4.1 Sample Preparation Same as 4.4.1.

5.4.2 Sample Pretreatment Weigh 5 g (accurate to

0.01

g) of the sample into a 50 mL stoppered colorimetric tube (5.2.15), and add 10 mL of methanol (5.2.4). Vortex mix. Mix in a vortex mixer (5.3.5) for 1 min, let stand for 5 min, add

5.0 mL of potassium iodide solution (5.2.5), and mix in a vortex mixer (5.3.5). 1 min, let stand for 10 min, add water to 50 mL, mix well, let stand, and then pass the supernatant through a needle-type microporous membrane (5.2.13). Place the filtrate in the injection port. The sample is to be tested in the bottle.

5.4.3 Creation of Standard Working Curves Six 5g (accurate to 0.01g) blank samples of the same matrix, free of benzoic acid and benzoyl peroxide, were weighed into 50mL stoppered containers. Add 10 mL of benzoic acid standard working solution (5.2.11) to each color tube (5.2.15), and then proceed as described in 5.4.2 "in a vortex mixer". Mix thoroughly in (5.3.5) for 1 min, let stand for 5 min, and place the filtrate in a sample vial for testing. The final mass concentrations of the standard working solutions are respectively... It is 0µg/mL, 0.16µg/mL, 0.64µg/mL, 1.6µg/mL, 6.4µg/mL, and 8.0µg/mL. The standard series working solutions were injected sequentially into the high-performance liquid chromatograph (5.3.1) in order from low to high concentration to obtain the concentrations. Chromatogram of the standard solution. A standard curve was plotted with the benzoic acid concentration (µg/mL) on the x-axis and the peak area on the y-axis. The solution chromatograms are shown in Figures A.2 and A.3.

5.4.4 Measurement Under the same conditions as the standard curve determination, the sample solution to be tested was injected into the high-performance liquid chromatograph (5.3.1), and the retention time was used for qualitative analysis to determine the result. The peak area is used to calculate the concentration of benzoic acid in the sample solution based on the standard curve.

5.4.5 Instrument Reference Conditions The instrument reference conditions are as follows:

a) Chromatographic column. C18 column (250 mm length,

4.6 mm inner diameter, 5 µm packing particle size), or a similar column, to extend column length. For extended battery life, a C18 protective post should be added.

b) Mobile phase. Methanol (5.2.4) + Ammonium acetate solution (5.2.6) = 10.90;

d) Detection wavelength. 230nm;

e) Column temperature. 30°C;

f) Injection volume. 50 µL.

Note. When interfering peaks are present or auxiliary qualitative analysis is required, a mobile phase containing formic acid (5.2.3) should be used for determination, such as a mobile phase containing methanol (5.2.4) or ammonium acetate formic acid. Solution (5.2.7) = 10 90, see Figure A.3 for reference chromatogram.

5.5 Calculation and Presentation of Results

5.5.1 Result Calculation The content (X) of benzoyl peroxide in the sample is expressed as a mass fraction and is calculated according to formula (2).

5.5.2 Results Presentation The results are expressed as the arithmetic mean of two independent measurements obtained under repeatability conditions, and are retained to two significant figures.

5.6 Precision Under repeatability conditions, the absolute difference between two independent measurements should not exceed 10% of the arithmetic mean; under reproducibility conditions... The absolute difference between two independent measurements should not exceed 15% of the arithmetic mean.

5.7 Other When the sample size is 5g, the detection limit of this method is 0.0005g/kg and the quantitation limit is 0.0015g/kg. GB/T 18415-2026. Determination of benzoyl peroxide in wheat flour and its products ICS

04 National Standards of the People's Republic of China Replaces GB/T 18415-2001 and GB/T 22325-2008 Determination of benzoyl peroxide in wheat flour and its products Published on 2026-03-

31 Implemented on October 1, 2026 State Administration for Market Regulation The State Administration for Standardization issued a statement.