NY/T 2006-2011Determination of beta-glucan in cereal and its products (English PDF)

1 Scope

This standard specifies the enzymatic-colorimetric determination method for beta-glucan content in cereals and their products.

This standard applies to the determination of beta-glucan content in oats, barley, highland barley, rye and other cereals and their processed products (flour, bran, oatmeal, beverages, etc.).

2 Normative references

The following documents are essential to the application of this document. For the dated documents, only the versions with the dates indicated are applicable to this document;

for the undated documents, only the latest version (including all the amendments) is applicable to this standard.

GB 5009.3 National food safety standard - Determination of moisture in foods

GB/T 5491 Inspection of grain and oilseeds - Methods for sampling and sample reduction

GB/T 6682 Water for analytical laboratory use - Specification and test methods

3 Principle

This method uses the enzymatic hydrolysis of (1->3)(1->4)-beta-D-glucan (mixed beta-D- glucan, or beta-glucan for short) in the sample by lichenase (or glucanase) and beta- glucosidase, uses lichenase to specifically hydrolyze beta-glucan into oligosaccharides, while beta-glucosidase hydrolyzes oligosaccharides into glucose; glucose generates gluconic acid and hydrogen peroxide under the action of glucose oxidase; hydrogen peroxide is oxidized and condensed with 4-aminoantipyrine under the action of peroxidase to generate red quinone compounds. This compound absorbs at 510 nm, its absorbance value is proportional to the glucose content.

4 Reagents

Unless otherwise specified, only confirmed analytical reagents are used in the analysis;

the experimental water shall meet the requirements of grade 3 water in GB/T 6682.

4.6.1 200 mmol/L, pH 4.0 sodium acetate buffer solution: Take 7.6 mL of glacial acetic acid in 900 mL water; add 4.8 g of sodium acetate trihydrate; dissolve it; adjust pH to

4.0; dilute to 1000 mL.

4.6.2 50 mmol/L, pH 4.0 sodium acetate buffer solution: Take 250 mL of 200 mmol/L, pH 4.0 sodium acetate buffer solution; dilute to 1000 mL.

4.7 1.000 mg/mL glucose standard stock solution

Dry glucose powder (purity greater than 99%) at 100 °C under normal pressure for 2 h;

cool in a desiccator and store in a sealed container at room temperature. Accurately weigh 0.1000 g of dry glucose; dissolve it in 50 mmol/L, pH 4.0 sodium acetate buffer (4.6.2); make the volume reach to 100 mL. The standard solution can be stored at 4 °C for one month.

Note: The enzyme preparations involved in 4.1, 4.2, 4.3 can all be provided by Megazyme mixed beta-D-glucan assay kit. Megazyme is the trade name of the products of Megazyme

Company of Ireland. This information is given for the convenience of users of this standard. If there are other products with the same effect, these equivalent products can be used.

5 Instruments and equipment

5.1 Cyclone mill or pulverizer: The sample can be pulverized to pass through a 0.5 mm or 35 mesh sieve.

5.2 Centrifuge: Can accommodate glass test tubes of 15 mm × 100 mm or 10 mm × 75 mm, centrifugal force 1000 × g.

5.3 Water bath: Temperature stability ± 0.2 °C.

5.4 Vortex mixer.

5.5 pH meter: Accuracy 0.01.

5.6 Analytical balance: Sensitivity 0.0001 g.

5.7 Drying oven: Can maintain 105 °C ± 1 °C.

5.8 Spectrophotometer: Detection wavelength 510 nm.

5.9 Cuvette.

5.10 Pipette: Range 10 µL ~ 100 µL, 20 µL ~ 200 µL, 1000 µL ~ 5 000 µL, equipped with disposable pipette tips.

5.11 Volumetric flask: Volume 100 mL, 1000 mL.

Accurately weigh an appropriate amount of sample into a test tube; add 10 mL of 50% ethanol solution (4.4); pre-extract at 80 °C for 10 min; centrifuge (1000×g, 10 min);

discard the supernatant; repeat the alcohol wash of the precipitate once; centrifuge (1000×g, 10 min); retain the precipitate to prepare for use.