A simplified and sensitive validated RP-HPLC method for determination of temozolomide in rat plasma and its application to a pharmacokinetic study

The present study describes the first fully validated RP-HPLC method for the quantification of temozolomide, an anti-cancer drug, in rat plasma. The method is simple, rapid, specific, sensitive and also environment-friendly. Further, the method can be translated for analysis with an LC-MS system with minor modifications. The procedure involved a simple step of protein precipitation for extraction of temozolomide and the internal standard, ganciclovir from the plasma. Efficient chromatographic separation was performed on reversed phase C-18 Oyster BDS Premium (5 μm, 4.6×150 mm) column using dual wavelength UV detector at 329 nm for temozolomide and 310 nm for the internal standard using a mobile phase consisting of a mixture of 0.5% aqueous acetic acid and methanol (98:2, v/v) in an isocratic mode at a flow rate of 1 mL/min. The method was sensitive with an LLOQ of 50 ng mL−1 and demonstrated good linearity from 50 to 10000 ng mL−1 with a regression coefficient of 0.998. The extraction efficiency was found to be >90.0% and all other validation parameters such as accuracy, precision and stability were well within limits. The method was successfully employed for the in-vivo pharmacokinetic study of temozolomide in rat.