Capillary gas chromatographic determination of proteins and biological amino acids as N(O)-tert.-butyldimethylsilyl derivatives.

Forty-seven biological amino acids containing all 22 protein amino acids were derivatized to N(O)-tert.-butyldimethylsilyl (tBDMSi) derivatives by a single-step reaction with N-methyl-N-(tert.-butyldimethylsilyl)trifluoroacetamide and successfully separated on an HP-1 capillary column. The relative standard deviations of the relative molar responses of most amino acids were < 5%. Cystine seems to be partially converted into cysteine during derivatization. An increase in carrier gas flow-rate towards the end of the analysis by inlet pressure programming with electron pressure control avoided the peak broadening and adsorption of the derivatives with high boiling points on the column and especially increased sensitivity of cystine to 5 pmol. Glutamine was converted almost completely into pyroglutamic acid during prolonged storage of a standard solution prepared in 0.01 M HCl but not during derivatization. These results compared with those for the phenylthiocarbamyl derivatives analysed by HPLC and the analytical results reported in the literature on soybean hydrolysate showed good agreement except for cysteine. The results for the amino acid composition of bovine serum albumin also showed good agreement with results in the literature except for cysteine. In human urine, seventeen free amino acids were detected as tBDMSi derivatives.