GC-MS analysis of nonprotein amino acids in Gymnocladus dioicus as N(O,S)-isobutyloxycarbonyl silyl derivatives.

Abstract

Two different derivatives of novel nonprotein amino acids in Gymnocladus dioicus seeds are prepared for gas chromatographic (GC) and mass spectrometric (MS) analysis. The interfering components in a water extract of the seeds are removed by picric acid treatment and multistep washing with organic solvents followed by N(O,S)-isobutyloxycarbonylation combined with solid-phase extraction. The resulting N(O,S)-isobutyloxycarbonyl (isoBOC) amino acids are then converted to either tert-butyldimethylsilyl (TBDMS) or trimethylsilyl (TMS) derivatives, which are analyzed by dual capillary column GC and GC-MS. The characteristic ions in the mass spectra of N(O,S)-isoBOC TBDMS and N(O,S)-isoBOC TMS derivatives enable confirmation of molecular weight and existence of functional groups containing active protons. Temperature programmed retention index sets measured on DB-5 and DB-17 capillary columns (J&W Scientific; Folsom, CA) are useful for distinguishing among structural isomers. gamma-Methylglutamic acid, diastereoisomers of 3-hydroxy-4-methylglutamic acid, cis and trans isomers of 5-hydroxypipecolic acid, and some free protein amino acids are identified in Gymnocladus dioicus seeds.