Selective and sensitive determination of protein and non-protein amino acids by capillary gas chromatography with nitrogen-phosphorus selective detection.

A selective and sensitive method for the determination of protein and non-protein amino acids by capillary gas chromatography (GC) has been developed. The amino acids were converted into their N(O,S)-isobutoxycarbonyl methyl ester derivatives and measured by GC with nitrogen-phosphorus selective detection using a DB-17ht capillary column. Using this method, the derivatives of the 21 protein amino acids and the 33 non-protein amino acids provided excellent NPD responses, and were quantitatively and reproducibly resolved within 28 min. The detection limits of these amino acids were 6-150 pg per injection. The calibration curves were linear in the range 0.02-2 micrograms for each amino acid, the correlation coefficients being above 0.990. This method was successfully applied to small urine samples without prior clean-up, and analysed without any influence from coexisting substances. Overall recoveries of amino acids added to urine sample were 83-108%. The analytical results of free amino acid contents in urine samples of normal subjects are presented.