Abstract
Simple and rapid reverse phase high-performance liquid chromatography (RP-HPLC) method was developed and validated using Phenomenex Gemini c18 (4.6 x 250 mm, 5 µ) reverse phase column for the determination of LAF in human plasma, Solid Phase Extraction (SPE) technique was used for the extraction of analyte, detection was carried out by Photo Diode Array detector at 216 nm. Chromatographic resolution of the LAF was achieved within 4.6 min by using mobile phase Methanol and 5 mM Di-Potassium Hydrogen Phosphate Buffer (pH 9.5) (80:20, v/v), flow rate was 1.0 mL/min. Calibration curve was linear with correlation coefficient of 0.9996 in the range of 50-1000 ng/mL, Limit of Detection (LOD) and Limit of Quantitation (LOQ) were 10 ng/mL and 30 ng/mL respectively, intra and inter-day deviations were lower than 3.92% and 3.98% respectively. The overall mean recovery of LAF was 94.57%. No any endogenous constituents were found to interfere at retention time of the analyte. This new RP-HPLC method was successfully validated and may be applied to conduct bioavailability & bioequivalence studies of LAF.
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