GB 5009.124-2016National food safety standard - Determination of Amino Acid in Foods (English PDF)

3.1 Reagents

3.1.1 Hydrochloric acid (HCl). concentration>=36%, guarantee reagent.

3.2 Reagents preparation

3.2.1 Hydrochloric acid solution (6mol/L). take 500mL of hydrochloric acid, dilute with

water to 1000mL, mix evenly.

3.2.2 Refrigerant. mix the commercial salt and ice by the mass ratio of 1.3.

3.3 Standard substance

3.3.1 Mixed amino acid standard solution. standard solution approved by the state or

awarded the standard substance certificate by the state.

3.3.2 16 individual amino acid standard substance. solid with purity>=98%.

3.4 Preparation of standard solution

3.4.1 Mixed amino acid standard stock solution (1µmol/mL). separately and

accurately take individual amino acid standard substances (accurate to 0.00001g) into

the same 50mL beaker; use 8.3mL of 6mol/L hydrochloric acid solution to dissolve;

4.1 Laboratory used tissue crusher and grinder.

4.2 Homogenizer.

4.7 Electric blower thermostat or hydrolysis furnace.

4.8 Test tube concentrator or parallel evaporator (with supporting 15mL~25mL test

tube).

4.9 Amino acid analyzer. ninhydrin post-column derivatization ion exchange

chromatography.

5.1 Specimen preparation

Crush the solid or semi-solid specimen by tissue crusher or grinder; use the

homogenizer to homogenize the liquid specimen into homogenate, sealed and frozen

to store; when analyzing, thaw it to use.

5.2 Specimen weighing

For sample with good uniformity such as milk powder, etc., accurately weigh a certain

quantity of specimen (accurate to 0.0001g); so that the protein content in the specimen

is in the range of 10mg~20mg. For samples with unknown protein content, the protein

content in the specimen can be measured firstly. Then place the weighed specimen

into the hydrolysis tube.

5.3 Specimen hydrolysis

According to the protein content in the specimen, add 10mL~15mL of 6mol/L

hydrochloric acid solution into the hydrolysis tube. For the specimen with high water

content and low protein content, such as beverages, fruits, vegetables, etc., firstly add

the same volume of hydrochloric acid to mix evenly, after that supplement by 6mol/L

hydrochloric acid solution to about 10mL. Continue to add 3 drops ~ 4 drops of phenol

into the hydrolysis tube.

5.4 Determination

5.4.1 Instrument conditions

Inject the mixed amino acid standard working solution into the automatic amino acid

analyzer; refer to the test protocol and instrument manual of the amino acid analyzer

stipulated in JJG1064-2011; appropriately adjust the instrument operation procedures,

parameters, and reagent ratio of buffer solution; confirm the instrument operating

conditions.

5.4.2 Chromatographic reference conditions

6.1 Calculation of individual amino acids in mixed amino acid standard stock

solution

6.2 Calculation of amino acid content in the sample

The amino acid content in the sample assay solution can be calculated by Formula (2).