RP-HPLC Method Development and Validation for Simultaneous Estimation of Hydrocortisone and Acyclovir in Pharmaceutical dosage forms.

ABSTRACTThe Chromatographic condition were successfully developed for the separation of Acyclovir and     Hydrocortisone by using Agilent C8 column (25 cm x 4.6 mm i.d., 5 ?). A simple, selective, rapid, precise and economical reverse phase high performance liquid chromatographic method has been developed for the simultaneous estimation of Hydrocortisone and Acyclovir from pharmaceutical formulation. The method was carried out on a C8 (25 cm x 4.6 mm i.d., 5 ?) column with a mobile phase consisting of Methanol: water (adjusted to pH 3.0 using o-phosphoric acid) in the ratio of 80:20 v/v. The retention time of  Hydricortusone and Acyclovir was 3.50 min and 6.00 min respectively with the flow rate of 1mL/ min. Eluents were detected at 254 nm.  The linear regression analysis data for the linearity plot showed good linear relationship with correlation coefficient value for Hydrocortisone and Acyclovir were R2=0.9995 and R2=0.9996 in the concentration range of 10-40µg. mL-1 , 20-80 µg. mL-1 respectively. The relative standard deviation for intra-day precision was lower than 2.0 %. The method was validated according to the ICH guidelines. The method was also found to be robust. The developed method was validated in terms of accuracy, precision, linearity, limit of detection, limit of quantification and solution stability. Keywords: RP-HPLC, Hydrocortisone, Acyclovir, Agilent , Mobile Phase, Retention time.