A new, simple, sensitive, accurate and precise RP-HPLC method was developed for the estimation of quetiapine fumarate in human plasma. Full factorial design was used for the optimization of an extraction method. The main effect of volume of deproteinating agent, speed of centrifugation, time of centrifugation and temperature of centrifugation was found to be significant at P<0.0001 on all the responses. After deproteinization, the drug was analyzed on a C18 (150 × 4.6mm, 5 µm) column using UV detector. The mobile phase consisting acetonitrile and phosphate buffer (pH 3) in the ratio of 50:50 (v/v) at a flow rate of 1.0 ml/min. The standard calibration curve was constructed in the concentration range of 5 µg/ml to 30 µg/ml and linearity was found to be 0.999. Irbesartan was used as the internal standard. The retention time of quetiapine fumarate and the internal standard was found to be 5.42 and 2.89 min, respectively. No interference peak was perceived. The high performance liquid chromatography method was successfully demonstrated as rapid and sensitive method which can be used as an alternative for the analysis of quetiapine fumarate in plasma samples.