GB 5009.85-2016National food safety standard - Determination of vitamin B2 in foods (English PDF)

3.1 Reagents

3.1.1 Hydrochloric acid (HCl).

3.1.7 Taka-amylase. active units >= 100 U/mg, or similar performance.

3.2 Reagent preparation

3.2.1 Hydrochloric acid solution (0.1 mol/L). PIPETTE 9 mL of hydrochloric acid;

USE water to dilute it to 1000mL.

3.2.4 Sodium acetate solution (0.1 mol/L). Accurately WEIGH 13.60 g of sodium

acetate trihydrate; ADD 900 mL of water to dissolve it; USE water to dilute it to

3.2.5 Sodium acetate solution (0.05 mol/L). Accurately WEIGH 6.80 g of sodium

acetate trihydrate; ADD 900 mL of water to dissolve it; ADJUST the pH to 4.0 ~

5.0 with glacial acetic acid; USE water to dilute it to 1000 mL.

3.2.6 Mixed enzyme solution. Accurately WEIGH 2.345 g of papain and 1.175g

of taka-amylase; ADD water to dissolve it; DILUTE it to 50 mL. Prepare it before

use.

3.3 Standards

Vitamin B2 (C17H20N4O6, CAS number. 83-88-5). Purity>=98%.

3.4 Standard solution preparation

3.4.1 Vitamin B2 standard stock solution (100 µg/mL). PLACE the vitamin B2

standard in a vacuum desiccator or dryer with phosphorus pentoxide; after

drying for 24h, accurately WEIGH 10 mg (accurate to 0.1 mg) Vitamin B2

standard; ADD 2 mL of hydrochloric acid solution (1+1) to dissolve it in

ultrasound;

3.4.2 Vitamin B2 standard intermediate solution (2.00 µg/mL). Accurately

PIPETTE 2.00 mL of Vitamin B2 standard stock solution; USE water to dilute it

to 100 mL. Prepare before use.

4.1 High performance liquid chromatograph. equipped with fluorescence

detector.

4.2 Balance. sensitivities are 1 mg and 0.01 mg.

4.6 Tissue-mash-machine.

4.7 Thermostatic water bath.

4.8 Dryer.

4.9 Spectrophotometer.

5.1 Sample preparation

TAKE out 500 g of sample; USE tissue-mash-machine to smash it uniformly;

respectively PLACE it into clean brown grinding mouth bottle; MAKE a mark

after sealing; STORE it for spare-use in dark place.

5.2 Instrument reference conditions

5.3 Drawing of standard curve

LEAD the standard series working solution into high performance liquid

chromatograph respectively; MEASURE the corresponding peak area. With the

concentration of standard working solution as abscissa AND the peak area as

vertical axis, DRAW a standard curve.

5.4 Determination of sample solution

LEAD the sample solution into high performance liquid chromatograph to obtain

the corresponding peak area; OBTAIN the vitamin B2 concentration of the

solution under test according to the standard curve.

10.1 Reagents

10.1.1 Hydrochloric acid (HCl).

10.1.10 Hydrogen peroxide (H2O2). 30%.

10.1.11 Sodium dithionite (Na2S2O4).

10.2 Reagent preparation

10.2.1 Hydrochloric acid solution (0.1 mol/L). PIPETTE 9 mL of hydrochloric

acid; USE water to dilute it to 1000 mL.

10.2.5 Mixed enzyme solution. Accurately WEIGH 2.345 g of papain and 1.175

g of taka-amylase; ADD water to dissolve it; USE water to dilute it to 50 mL.

Prepare it before use.

10.2.6 Eluent. acetone-glacial acetic acid-water (5+2+9, volume ratio).

10.2.7 Potassium permanganate solution (30 g/L). Accurately WEIGH 3 g of

potassium permanganate; USE water to dissolve it and dilute it to 100 mL.

10.3 Standard products

Vitamin B2 (C17H20N4O6, CAS number. 83-88-5). Purity>=98%.

10.4 Standard solution preparation

10.4.1 Vitamin B2 standard stock solution (100 µg/mL). PLACE the vitamin B2

standard in a vacuum desiccator or dryer with phosphorus pentoxide; after

drying for 24h, accurately WEIGH 10 mg (accurate to 0.1 mg) Vitamin B2

standard; ADD 2 mL of hydrochloric acid solution (1+1) to dissolve it in

ultrasound; immediately USE water to transfer it and dilute it to 100 mL.

11.1 Fluorescence Spectrophotometer.

11.2 Balance. sensitivities are 1 mg and 0.01 mg.

11.6 Tissue-mash-machine.

11.7 Thermostat water bath.

11.8 Dryer.

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