Journal
ANALYTICAL BIOCHEMISTRY
Volume 516, Issue -, Pages 75-85Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2016.10.017
Keywords
Aminobutyric acid isomers; LC-MS/MS; Chromatographic deconvolution; Marfey's derivatization
Funding
- National Institutes of Health
- NIAMS [P01-AG039355]
- NIAID [R21-121903]
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Isomeric molecules present a challenge for analytical resolution and quantification, even with MS-based detection. The eight aminobutyric acid (ABA) isomers are of interest for their various biological activities, particularly gamma-aminobutyric acid (GABA) and the D- and L-isomers of beta-aminoisobutyric acid (beta-AIBA; BAIBA). This study aimed to investigate LC-MS/MS-based resolution of these ABA isomers as their Marfey's (Mar) reagent derivatives. HPLC was able to separate three Mar-ABA isomers L-beta-ABA (L-BABA), and L- and D-alpha-ABA (AABA) completely, with three isomers (GABA, and D/L-BAIBA) in one chromatographic cluster, and two isomers (alpha-AIBA (AAIBA) and D-BABA) in a second cluster. Partially separated cluster components were deconvoluted using Gaussian peak fitting except for GABA and D-BAIBA. MS/MS detection of Marfey's derivatized ABA isomers provided six MS/MS fragments, with substantially different intensity profiles between structural isomers. This allowed linear deconvolution of ABA isomer peaks. Combining HPLC separation with linear and Gaussian deconvolution allowed resolution of all eight ABA isomers. Application to human serum found a substantial level of L-AABA (13 mu m), an intermediate level of L-BAIBA (0.8 mu M), and low but detectable levels (<0.2 mu M) of GABA, L-BABA, AAIBA, D-BAIBA, and D-AABA. This approach should be useful for LC-MS/MS deconvolution of other challenging groups of isomeric molecules. (C) 2016 Elsevier Inc. All rights reserved.
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