EVALUATION OF A NEW HIGH PERFORMANCE LIQUID CHROMATOGRAPHY BASED METHOD FOR VORICONAZOLE ANALYSIS IN BLOOD

Abstract

Objective: To develop and evaluate a new high-performance liquid chromatography (HPLC) based method for Voriconazole (VRC) analysis in blood.

Study Design: Cross sectional study.

Place and Duration of Study: Department of Chemical Pathology and Endocrinology Armed Forces Institute of Pathology (AFIP), Rawalpindi, from May 2017 to Oct 2017.

Material and Methods: Trough venous blood samples of twenty-five patients on VRC therapy were collected, after five days of starting VRC. Samples were centrifuged. About 100 μl of Lorazepam as internal standard (5.0 μg/ml), was added to 0.5ml of each sample. Then extraction was done with 4.0 ml of mixture of hexane and ethyl acetate. Centrifugation was again done and supernatant layer was collected and dried under nitrogen. It was reconstituted with 200μl of deionized water and acetonitrile. Samples were analyzed on HPLC.

Results: Method was validated in terms of linearity, limit of detection, limit of quantification, precision, accuracy, recovery, analytical specificity and stability. Linearity calibration curve plot for assay obtained was linear over analytical measurement range of 1.0 to 8.0μg/ml and correlation coefficient was 0.99. Mean intra-assay accuracy for 1.0, 5.0 and 8.0 μg/ml was 96%, 98.6%, 99.5% respectively, and percentage Relative Standard Deviation (RSD) was 5.9%, 1.15%, 1.44% respectively. Mean inter-assay accuracy for 1.0, 5.0 and 8.0 μg/ml was 95%, 98.6%, 99.5% respectively, and % RSD was 5.68%, 1.03% and 1.01% respectively. Limit of Detection (LOD) was 0.25 μg/mL and Limit of Quantitation (LOQ) was 1.0 μg/ml.

Conclusion: HPLC based method for VRC determination is accurate and specific with good clinical correlation. This method may be used in routine for Therapeutic Drug Monitoring (TDM) of VRC.