Amino acid nitrogen

Determination of Amino Acid Nitrogen

Principle                                                     

A known quantify of KCI extract of soil is treated with citric acid and ninhydrin and then distilled with phosphate borate buffer and NaOH. The released ammonia is absorbed in boric acid and titrated against standard H2S04. From the volume of H2S04 consumed, the amino acid-N content of the soil is calculated.

Apparatus required

v  50 ml conical flask

v  Water bath

v  Distillation set

Reagents required

v  0.5 N NaOH

v  Ninhydrin

v  5N NaOH

v  Citric acid

v  Phosphate borate buffer (pH 11.2)

v  0.002 N H2S04

Procedure

v  Take about 5 ml of the hydrolysate in a 50 ml conical flask

v  Add one ml of 0.5 N NaOH and heat the contents in a water bath at 100°C until the volume of the same got reduced to 3 ml

v  After cooling, add 0.5 g of citric acid and 100 mg of ninhydrin and keep the flask immersed in water bath at 100°C

v  After one minute. swirl the flask for a few seconds without removing it from the water bath and then allow it to remain in the water bath for an additional nine minutes

v  Transfer the contents to Bremner's distillation flask by giving washings with distilled water

v  To this, add 1 0 ml of phosphate borate (pH 11.2) and 1 ml of 5 N NaOH and start distillation

v  Note the volume of H2S04 consumed and from this value calculate the amino acid N content in the soil

Note: Amonio acid N content of soil is nearly 40% of total nitrogen. Calculation

Calculation

Volume of hydrolysate prepared

=

10 g

Weight of soil taken

=

100 ml

Volume of hydrolysate taken for estimation

=

5 ml

Volume of 0.002N H2S04 consumed

=

X ml

% of amino acid N

=

=

‘X’ x 0.0056