GB/T 15400-2018Determination of tryptophan in feeds (English PDF)

3.1 Principle

The tryptophan in the sample is hydrolyzed by lithium hydroxide solution, separated by high performance liquid chromatography, detected by ultraviolet or fluorescence detector, external standard method Quantitative.

3.2 Reagents or materials

Unless otherwise specified, the reagents used in this standard are of analytical grade.

3.2.1 Water. Meet the requirements of the first grade water in GB/T 6682.

3.2.2 Methanol. chromatographically pure.

3.2.3 Potassium hydroxide solution (0.1mol/L). Weigh 0.56g of potassium hydroxide and dissolve it in 100mL of water.

3.2.4 Lithium hydroxide solution (4mol/L). Weigh 83.9g of lithium hydroxide and dissolve it in 500mL of water.

3.2.5 sodium acetate buffer solution (0.0085mol/LNa, pH=4.5). Weigh 0.70g of anhydrous sodium acetate and dissolve it in 1000mL of water. The pH was adjusted to 4.5 with glacial acetic acid.

3.2.6 Mobile phase. sodium acetate buffer solution (3.2.5) methanol (3.2.2) = 95 5.

3.2.7 L-tryptophan standard stock solution. accurately weigh 25mg (accurate to 0.0001g) tryptophan reference substance (purity greater than 99%), set In a 25 mL beaker, add potassium hydroxide solution (3.2.3) dropwise with a dropper to dissolve it, and quantitatively transfer it to a 250 mL brown volumetric flask. The volume was adjusted to a concentration of 100 µg/mL. Store at 2 ° C ~ 8 ° C, valid for 1 month.

3.2.8 L-tryptophan standard series solution. accurately remove L-tryptophan standard stock solution (3.2.7) 5.00mL, 7.50mL, 10.00 mL, 12.50 mL, 15.00 mL, and 17.50 mL were placed in a 25 mL brown volumetric flask, and the volume was adjusted to the mark with water and shaken. its The solution concentrations were 20 µg/mL, 30 µg/mL, 40 µg/mL, 50 µg/mL, 60 µg/mL, and 70 µg/mL, respectively. 2 ° C ~ 8 ° C Save, valid for 2 weeks. […]