GB 5009.149-2016Determination of crocin in foods (English PDF)

3.1 Reagents

3.1.1 Methanol (CH3OH): chromatographic pure.

3.1.2 Acetonitrile (CH3CH): chromatographic pure.

3.1.3 Glacial acetic acid (CH3COOH).

3.1.4 Ammonium acetate (CH3COONH4).

3.2 Preparation of reagents

3.2.1 Acetic acid-ammonium acetate solution (pH=4): accurately weigh 0.77 g of

ammonium acetate in a 1 L volumetric flask; add 900 mL of water to dissolve; use glacial acetic acid to adjust pH=4.0; add water to fix volume to 1 L; mix well. Use it after it is filtered through a 0.45 µm microporous membrane.

5.1.1 Liquid sample

Shake well and pack the samples such as beverage, alcohol and soy sauce; seal and store at room temperature or in a refrigerator.

5.1.2 Semi-solid sample

For samples such as jelly, take the edible portion to homogenate; stir evenly; package;

seal; refrigerate or freeze to store.

5.1.3 Solid sample

For samples of low-water content, such as biscuit, cake, cooked meat products and cocoa products, use the high-speed pulverizer to pulverize; package; seal and store at room temperature out of the sun.

5.2 Sample processing

5.2.1 Liquid sample

Weigh 2 g (accurate to 0.01 g) of uniform sample (carbon dioxide in the sample, if any, shall be removed by ultrasound firstly) in a beaker; use an appropriate amount of methanol to dissolve it; transfer it to a 25 mL volumetric flask; add methanol to fix volume; shake well. Pipette 1 mL of the solution; filter it through a 0.45 µm organic filter for testing.

5.2.2 Semi-solid and solid samples

Weigh 2 g (accurate to 0.01 g) of uniform sample in a 50 mL centrifuge tube; accurately measure 25 mL of methanol solution into it; use ultrasound for 20 min and vortex for 2 min; centrifuge at 4 000 r/min for 10 min; absorb 1 mL of supernatant to filter through a 0.45 µm organic filter membrane, which is for later test.

5.2.3 Soy sauce

Weigh 1 g (accurate to 0.01 g) of uniform sample in a beaker; use an appropriate amount of methanol to dissolve it; transfer it to a 50 mL volumetric flask; add methanol to fix volume; shake well. Pipette 1 mL of the solution; filter it through a 0.45 µm organic filter for testing.

Note: 5.2 shall be performed in an environment without strong light exposure.

5.3 Apparatus reference conditions

Apparatus reference conditions are listed as below:

a) Chromatographic column: C18 column with a column length of 250 mm, an inner diameter of 4.6 mm, a particle size of 5 µm, or columns of equivalent performance;

X -- the content of crocin or crocetin in the sample, in grams per kilogram (g/kg);

rho -- the concentration of crocin or crocetin in the sample solution that is obtained from the standard curve, in micrograms per milliliter (µg/mL);

V -- the volumetric volume of the sample solution, in milliliters (mL);

1 000 -- conversion coefficient;

m -- the sample mass of the final sample, in grams (g);

F -- the purity conversion factor of the standard.

The calculation result shall keep two significant figures.