Identification of O-phosphoamino acids in urine hydrolysate by gas chromatography-mass spectrometry

The occurrence of O-phosphoserine (P-Ser), O-phosphothreonine (P-Thr) and O-phosphotyrosine (P-Tyr) in the hydrolysate of human urine was demonstrated. Urine samples were separated into non-adsorbed and adsorbed fractions by DEAE-cellulose column chromatography. After acid and base hydrolyses of these fractions, the O-phosphoamino acids in the hydrolysates were converted into their N-isobutoxycarbonyl methyl ester derivatives and identified by gas chromatography-mass spectrometry. By gas chromatography with flame photometric detection, the urinary contents of P-Ser, P-Thr and P-Tyr were estimated to be 945 +/- 3, 109 +/- 3 and 2.9 +/- 0.2 ng/ml, respectively.