Development and Validation of a RP-HPLC Method for Vemurafenib in Human Urine

Vemurafenib (propane-1-sulfonic acid{3-[5-(4-chlorophenyl)-1H-pyrrolo[2,3-b]pyridine-3-carbonyl]-2,4difluoro- phenyl}-amide) is a BRAF enzyme inhibitor approved by the U.S. Food and Drug Administration (FDA) for the treatment of late stage melanoma. Reversed phase high performance liquid chromatography (RP-HPLC) method was developed and validated for the determination of vemurafenib and erlotinib was used as an internal standard. Erlotinib (IS) and vemurafenib were separated by using a X-Terra RP-18 column (250 x 4.60 mm, ID 5 mu m) and detected by a DAD detector set at 249 nm. The mobile phase consisted of a mixture of acetonitrile: water 60:40 (v/v) at a flow rate of 1.0 mL/min. Isocratic separation was performed in less than 7 min. The calibration was found to be linear in the range 0.2-10 mg/L. Inter-day and intra-day precision was less than 0.72 %. This analytical method was successfully applied in spiked samples of human urine.