A new simple, accurate, rapid, selective and robust high pressure liquid chromatography (HPLC) method was developed and validated for estimation of itraconazole in bulk and marketed formulation. Acetonitrile and double distilled water was used as a mobile phase for chromatographic separation and estimation on HiQSil C18- HS (250 × 4.6 mm) in the ratio of 90:10 v/v at flow rate of 1.0 ml/min. The detection was carried out with UV detector set at 263 nm. The retention time for itraconazole was found to be 7.75 minutes. The linearity range for itraconazole was found to be 5-60 μg/ml with coefficient of linear regression 0.991. The method was validated in accordance with the requirements of International Conference on Harmonization (ICHQ2 (R1) 2005) guidelines for accuracy, precision, LOD & LOQ, linearity and robustness.