SIMPLE AND RAPID GAS-CHROMATOGRAPHIC METHOD FOR QUANTITATION OF TOTAL AND FREE VALPROIC ACID IN HUMAN SERUM

Abstract

Valproic acid (VPA) is one of the mostly used antiepileptic drugs that may have some side effects so, it is highly recommended to evaluate its serum concentrations. The aim of this study was to develop a simple, fast and economic method using gas-chromatography equipped with flame ionization detector (GC/FID) and VPA analysis. To do this, 200 µl of serum was mixed with an aliquot of caproic acid (200 µL, methanolic solution) as internal standard and extracted by stepwise addition of hydrochloric acid and chloroform with slight agitation between each step. After centrifugation, 1.0 µl of the bottom layer was injected into a wide-bore nonpolar capillary column. Injectable samples for analysis of unbonded VPA were prepared by ultra filtration followed by solid phase extraction (SPE). Caproic acid and VPA were eluted after 1.5 min and 3.0 min, respectively (total GC run time about 3.2 minutes). This GC/FID method was linear over a range of 2.5-6400 µg/ml with the mean recovery of 92%. The intra- and inter-assay precision in the range of 25-100 µg/ml was 1.50-, 2.95, and 2.35- 3.22%, respectively. The simplicity of sample preparation with no derivatization, short run-time and high sensitivity sufficient to detect low concentrations of the drug makes this method suitable for research as well as routine use.