title: Free amino acid quantification by LC-MS/MS using derivatization generated isotope-labelled standards.
abstract: The further development of derivatizing reagents for plasma amino acid quantification by tandem mass spectrometry is described. The succinimide ester of 4-methylpiperazineacetic acid (MPAS), the iTRAQ reagent, was systematically modified to improve tandem mass spectrometer (MS/MS) product ion intensity. 4-Methylpiperazinebutyryl succinimide (MPBS) and dimethylaminobutyryl succinimide (DMABS) afforded one to two orders of magnitude greater MS/MS product ion signal intensity than the MPAS derivative for simple amino acids. CD(3) analogues of the modified derivatizing reagents were evaluated for preparation of amino acid isotope-labelled quantifying standards. Acceptable accuracy and precision was obtained with d(3)-DMABS as the amino acid standards derivatizing reagent. The product ion spectra of the DMABS amino acid derivatives are diagnostic for structural isomers including valine/norvaline, alanine/sarcosine and leucine/isoleucine. Improved analytical sensitivity and specificity afforded by these derivatives may help to establish liquid chromatography tandem mass spectrometry (LC-MS/MS) with derivatization generated isotope-labelled standards a viable alternative to amino acids analysers.
authors.affiliation: SA Pathology, Dept of Genetic Medicine, Women's and Children's Hospital, 72 King William Road, North Adelaide, South Australia 5006, Australia. david.johnson@adelaide.edu.au
authors.forename: David W
authors.initials: DW
authors.lastname: Johnson
mesh_terms: D000596:Amino Acids / Q000032:analysis* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D007202:Indicators and Reagents / Q000737:chemistry; D007553:Isotope Labeling / Q000379:methods; D010949:Plasma / Q000737:chemistry*; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D013388:Succinimides / Q000737:chemistry; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D000596:Amino Acids; D007202:Indicators and Reagents; D013388:Succinimides