Determination of glutamate decarboxylase by high-performance liquid chromatography.
An improved method for the determination of glutamate decarboxylase (GAD) activity is described. The enzyme was evaluated by incubation with glutamic acid (L-Glu) in the presence of pyridoxal 5'-phosphate (PLP): the gamma-aminobutyric acid (GABA) formed was derivatized to PTC-GABA; the latter was subsequently separated and assayed by isocratic HPLC (LiChrospher RP-18 column; isocratic elution with pH 5.8 acetate buffer in acetonitrile-water) with UV absorbance detection at 254 nm. The method described is a sensitive, reproducible and specific assay useful for following variations of GAD activity in vitro; this assay was subsequently used for the evaluation of GAD activity variations after irradiation with low doses of He-Ne laser radiation.
Rossetti
V
V
Dipartimento di Scienza e Tecnologia del Farmaco, University of Torino, Italy.
Lombard
A
A
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004307:Dose-Response Relationship, Radiation; D004790:Enzyme Induction / Q000528:radiation effects; D005968:Glutamate Decarboxylase / Q000032:analysis* / Q000096:biosynthesis / Q000528:radiation effects; D018698:Glutamic Acid / Q000378:metabolism; D007834:Lasers; D016014:Linear Models; D010670:Phenylthiourea / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet; D005680:gamma-Aminobutyric Acid / Q000032:analysis / Q000378:metabolism*
D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; D010670:Phenylthiourea; D005968:Glutamate Decarboxylase