NY/T 1975-2010Water-soluble fertilizers - Determination of amino-acids contents (English PDF)

3.1 Principle

After precipitating the protein from the specimen by the use of sulfosalicylic acid, the EDTA complexes metal elements are used to release amino acids. The amino acids are separated by a separation column, to develop color with ninhydrin, to determine the contents of totally 17 amino acids: aspartic acid, threonine, serine, glutamic acid, Proline, glycine, alanine, cystine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine. The sum of these amino acids is the content of free amino acids.

3.2 Reagents and materials

The preparation of reagents, water and solutions used in this standard shall comply with the provisions of HG/T 2843 unless otherwise specified for the specifications and preparation methods.

3.2.1 Sodium citrate (Na3C6H5O7 · 2H2O).

3.2.2 Hydrochloric acid.

3.2.3 Hydrochloric acid solution: c(HCl) = 0.06 mol/L.

3.2.4 Sodium hydroxide solution: rho (NaOH) = 500 g/L.

3.2.5 Sulfosalicylic acid solution: rho (C6H6O6S · 2H2O) = 50 g/L.

3.2.6 Disodium edetate solution: rho (EDTA-Na) = 10 g/L.

3.2.7 Sodium citrate buffer: pH 2.2. Weigh 19.6 g of sodium citrate (3.2.1).

Dissolve it. Pipette it to a 1000 mL volumetric flask. Add 16.5 mL of hydrochloric acid (3.2.2). Add water to the mark. Mix it uniformly. If necessary, it may use hydrochloric acid (3.2.2) and sodium hydroxide solution (3.2.4) to adjust the pH to 2.2.

3.2.8 Mixed amino acid standard solution, chromatographically pure. It contains

total 17 amino acids: aspartic acid, threonine, serine, glutamic acid, Proline, glycine, alanine, cystine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine.

3.3 Instruments

3.3.1 Commonly used laboratory instruments.

3.3.2 Automatic amino acid analyzer.

3.3.3 Centrifuge (above 10000 r/min) or 0.45 µm microporous membrane, filter

and syringe.

3.3.4 Steam drying device: Test tube concentrator or other concentration device.

3.4 Analytical procedures

3.4.1 Preparation of specimens

After the solid sample has been reduced several times, take out about 100 g.

Grind it quickly to make all pass through a 0.50 mm aperture sieve (if the sample is wet, it may pass a 1.00 mm sieve). Mix it evenly. Put it in a clean, dry container.

After multiple shaking of the liquid sample, quickly take about 200 mL and place

After precipitating the protein of specimen by sulfosalicylic acid, use the EDTA to make the metal element release the amino acid. The amino acid is chemically derivatized and separated by liquid chromatography to determine the contents of totally 17 amino acids: aspartic acid, threonine, serine, glutamic acid, Proline, glycine, alanine, cystine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine. The sum of these amino acids is the content of free amino acids in the fertilizer.

4.2 Reagents and materials

The preparation of reagents, water and solutions used in this standard shall comply with the provisions of HG/T 2843 unless otherwise the specifications and preparation methods are indicated.

4.2.1 Sulfosalicylic acid solution: rho (C7H6O6S · 2H2O) = 50 g/L.

4.2.2 Disodium edetate solution: rho (EDTA - Na) = 10 g/L

4.2.3 Mixed amino acid standard solution, chromatographically pure. It contains

total 17 amino acids: aspartic acid, threonine, serine, glutamic acid, Proline, glycine, alanine, cystine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine.

4.2.4 Reagents required by the selected derivatization method.

4.3 Instruments

4.3.1 Commonly used laboratory instruments.

4.3.2 Centrifuge (above 10000 r/ min) or 0.45 µm microporous membrane and

filter, syringe.

4.3.3 Liquid chromatograph, equipped with UV detector.

4.3.4 The equipment required for the selected derivative method.

4.4 Analytical procedures

4.4.1 Preparation of specimen

After the solid sample has been reduced several times, take about 100 g and quickly grind it to pass through a 0.50 mm aperture sieve (if the sample is wet, it can pass through a 1.00 mm sieve). Mix it evenly. Place it in a clean, dry container. After shaking the liquid sample many times, quickly take about 100 mL and place it in a clean, dry container.

4.4.2 Preparation of sample solution

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