GB 5009.28-2016National food safety standard - Determination of Benzoic Acid, Sorbic Acid and Saccharin Sodium in Foods (English PDF)

3.1 Reagents

3.1.1 Ammonia (NH3·H2O).

3.1.6 Methanol (CH3OH). chromatographic purity.

3.1.7 Ammonium acetate (CH3COONH4). chromatographic purity.

3.1.8 Formic acid (HCOOH). chromatographic purity.

3.2 Preparation for reagents

3.2.1 Ammonia solution (1+99). take 1mL of ammonia; add it into 99mL water; mix it

evenly.

3.2.2 Potassium ferrocyanide solution (92g/L). weigh and take 106g of potassium

ferrocyanide; add appropriate amount of water to dissolve it; use water to dilute it to

1000mL.

3.2.5 Formic acid-ammonium acetate solution (2mmol/L formic acid + 20 mmol/L

ammonium acetate). weigh and take 1.54 g of ammonium acetate; add appropriate

amount of water to dissolve it; add 75.2µL of formic acid; use water to dilute it to

1000mL. After it is filtered by 0.22µm aqueous microfiltration membrane, it shall be set

for standby application.

3.3 Standard products

3.3.1 Sodium benzoate (C6H5COONa, CAS No.. 532-32-1). The purity shall be >=

99.0%; OR benzoic acid (C6H5COOH, CAS No.. 65-85-0). The purity shall be >= 99.0%;

OR the standard substance which is certified by the country AND granted for standard

substance certificate.

3.4 Preparation of standard solution

3.4.1 Benzoic acid, sorbic acid AND saccharin sodium (calculated as saccharine)

standard stock solution (1000mg/L).

3.4.2 Mixed standard intermediate solution (200mg/L) of benzoic acid, sorbic acid AND

saccharin sodium (calculated as saccharin). respectively and accurately pipette

10.0mL of the mixed standard stock solution of benzoic acid, sorbic acid AND

saccharin sodium into a 50mL volumetric flask; use water to set the constant volume.

At 4°C, store it. The preservation period shall be 3 months.

3.5 Materials

3.5.1 Aqueous microporous membrane. 0.22µm.

3.5.2 Plastic centrifuge tube. 50mL.

4.1 High performance liquid chromatography. be equipped with UV detector.

4.5 Homogenizer.

4.6 Constant temperature water bath.

4.7 Ultrasonic generator.

5.1 Preparation of sample

Take a plurality of prepackaged homogeneous samples, such as beverages and liquid

milk, etc.; mix them directly;

5.2 Sample's extraction

5.2.1 General Samples

Accurately weigh and take about 2g (accurate to 0.001g) of sample in a 50mL

stoppered centrifuge tube; add about 25mL of water; vortex-mix it; in 50°C water bath,

sonicate it for 20min;

5.2.2 Samples of gum-base jelly and candy, etc.

Accurately weigh and take about 2g (accurate to 0.001g) of sample in a 50mL

stoppered centrifuge tube; add about 25mL of water; vortex-mix it; in 70°C water bath,

heat and dissolve the sample; in 50°C water bath, sonicate it for 20min. Subsequent

operations shall be the same as those in 5.2.1.

5.3 Reference conditions of instrument

5.3.1 Chromatographic column. C18 column; column length - 250mm; inner diameter

- 4.6mm; particle size - 5µm; OR the chromatographic column with equivalent

performance.

5.3.2 Mobile phase. methanol + ammonium acetate solution = 5 + 95.

5.3.3 Flow rate. 1 mL/min.

5.3.4 Detection wavelength. 230nm.

5.5 Determination of sample solution

Inject the sample solution into the liquid chromatograph; obtain the peak area;

according to the standard curve, obtain the mass concentration of benzoic acid, sorbic

acid AND saccharin sodium (calculated as saccharine) in test solution.

10.1.1 Ether (C2H5OC2H5).

10.1.5 Hydrochloric acid (HCl).

10.1.6 Sodium chloride (NaCl).

10.1.7 Anhydrous sodium sulfate (Na2SO4). at 500°C, dry for 8 hours; in desiccator,

after it is cooled to room temperature, it shall be set for standby application.

10.2 Preparation of reagent

10.2.1 Hydrochloric acid solution (1+1). take 50mL of hydrochloric acid; at the time of

stirring it, slowly add it into 50mL water; mix it evenly.

10.3 Standard products

10.3.1 Benzoic acid (C6H5COOH, CAS No.. 65-85-0). the purity shall be >= 99.0%; OR

the standard substance which is certified by the country AND granted for standard

substance certificate.

10.4 Preparation of standard solution

10.4.1 Benzoic acid AND sorbic acid standard stock solution (1000mg/L). respectively

and accurately weigh and take 0.1g of benzoic acid AND sorbic acid (accurate to

0.0001g); use methanol to dissolve them AND dilute them to 100 mL respectively;

Transfer them into a sealed container; at -18°C, store it. The preservation period shall

be 6 months.

10.5 Materials

Plastic centrifuge tube. 50mL.

11.1 Gas chromatograph. Flame Ionization Detector (FID) with hydrogen.

11.5 Homogenizer.

11.6 Nitrogen blowing instrument.

......