VALIDATION OF WARFARIN ANALYSIS METHOD IN HUMAN BLOOD PLASMA USING HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY WITH FLUORESCENCE DETECTION

Objective: Validation of analysis method is important, especially in analyzing narrow-index drugs such as warfarin (WF). This study aimed to obtain a validated human plasma according European Medicine Agency guidelines.
 Methods: The optimum conditions for the using fluorescence detector HPLC with Chiralcel OD-RH column (4.6 x 150 mm i.d., 5μm); guard (4.0 10 mm, 5μm), temperature 45 °C. mobile phase used was acetonitrile: phosphate buffer pH 2 (40:60), an isocratic flow rate 1 ml/min and injection volume 20 μl. Excitation emission wavelengths were 310 350 nm (warfarin) 300 400 (griseofulvin). retention time griseofulvin 6-7.5 min; R-warfarin 10-11.5 S-warfarin 14-16 min.
 Results: result this validation obtained condition. yielded LOD values 0.0674 ppm (R-warfarin) 0.0897 (S-warfarin). LOQ 0.225 0.298 Linearity at concentrations 0.2-3 line equation y = 0.0705x+0.0704 R2 0.978 0.0513x+0.0297 R² 0.9924 S-warfarin. 75% seven met reverse concentration requirements, which below±15%. requirements accuracy precision within between runs, selectivity carryover where %RSD %diff below±15%
 Conclusion: analytical can be declared valid sample measurement pharmacokinetic studies.