Background: Literature reports innumerable methods for quantification of paclitaxel in biological matrices. Most of these involve complicated extraction procedures like solid phase extraction, separate procedure for elimination of interference of Cremophor El, advanced and expensive instruments. Objectives:The objective of the present research work is to develop and validate a simple, rapid, sensitive, economic and reproducible reverse phase -high performance liquid chromatography method for the estimation of paclitaxel concentration in human plasma that eliminates negative influence of Cremophor El on recovery of paclitaxel.Methods: Chromatographic separation of paclitaxel was carried out using C18 column (150 × 4.6 mm i.d., 4µm particle size, Waters, Australia) with 60% acetonitrile, 40% of 10mM ammonium acetate buffer solution and 0.1% formic acid as a mobile phase at a flow rate of 1.0mL/min at ambient temperature. Validation was performed as per ICH Q2 guidelines.Results: In this system the retention time was 3min. The detection limit was 5ng/mL and limit of quantification with reproducibility was 15ng/mL. Plasma samples were extracted using single solvent (tertiary -Butyl Methyl Ether) liquid-liquid extraction with a recovery of 96-99%. Robustness of the method was established with variation in flow rate, detection wave length and mobile phase composition. Stability of paclitaxel during the study period was studied and found to be stable. Conclusion:The developed method is easy to perform, quick, reproducible with good recovery without negative influence of Cremophor El and applicable to quantify paclitaxel in regular clinical practice for individualization of the therapies.