title: Quantification of γ-Aminobutyric Acid in Cerebrospinal Fluid Using Liquid Chromatography-Electrospray Tandem Mass Spectrometry.
abstract: We describe a simple stable isotope dilution method for accurate and precise measurement of γ-aminobutyric acid (GABA), a major inhibitory neurotransmitter in human cerebrospinal fluid (CSF) as a clinical diagnostic test. Determination of GABA in CSF (50 μL) was performed utilizing high performance liquid chromatography coupled with electrospray positive ionization tandem mass spectrometry (HPLC-ESI-MS/MS). Analysis of free and total GABA requires two individual sample preparations and mass spectrometry analyses. Free GABA in CSF is determined by a 1:2 dilution with internal standard (GABA-D2) and injected directly onto the HPLC-ESI-MS/MS system. Determination of total GABA in CSF requires additional sample preparation in order to hydrolyze all the bound GABA in the sample to the free form. This requires hydrolyzing the sample by boiling in acidic conditions (hydrochloric acid) for 4 h. The sample is then further diluted 1:10 with a 90 % acetonitrile/0.1 % formic acid solution and injected into the HPLC-ESI-MS/MS system. Each assay is quantified using a five-point standard curve and is linear from 6 nM to 1000 nM and 0.63 μM to 80 μM for free and total GABA, respectively.
authors.affiliation: Center of Metabolomics, Institute of Metabolic Disease, Baylor Research Institute, 3812 Elm Street, Dallas, TX, 75226, USA. erlanda@baylorhealth.edu.
authors.forename: Erland
authors.initials: E
authors.lastname: Arning
authors.affiliation: Center of Metabolomics, Institute of Metabolic Disease, Baylor Research Institute, 3812 Elm Street, Dallas, TX, 75226, USA.
authors.forename: Teodoro
authors.initials: T
authors.lastname: Bottiglieri
mesh_terms: D053000:Analytic Sample Preparation Methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D007554:Isotopes; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D013223:Statistics as Topic; D005680:gamma-Aminobutyric Acid / Q000134:cerebrospinal fluid*
chemical_list: D007554:Isotopes; D005680:gamma-Aminobutyric Acid
keywords: Cerebrospinal fluid; GABA; Mass spectrometry; Seizures