SIMULTANEOUS HPLC METHOD DEVELOPMENT AND VALIDATION FOR ESTIMATION OF SALBUTAMOL IN COMBINATION WITH VASICINE

Papiya Bigoniya* and Smita Mishra

ABSTRACT

A sensitive high-performance liquid chromatography method has been developed for the simultaneous determination of salbutamol and vasicine isolated from Adhathoda vasica. Isolated vasicine was authenticated by TLC, FT-IR and HPLC analysis compared with reference standard. Salbutamol was obtained as pure compound and further authenticated for purity by FT-IR, UV and HPLC analysis with reference to Indian Pharmacopoeia. Simultaneous estimation was carried out on a reversed-phase C18 column (150 mm × 4.6 mm; 5 μm) using isocratic solvent system consisting of methanol and de-ionized water in ratio of 40:60 as mobile phase at detection web length of 298 nm. Quantification was performed by preparation of calibration curve using standardized vasicine and salbutamol as the internal standard. Calibration curves showed good linearity between concentration of 100-1000 μg/ml for vasicine and 1-10 μg/ml for salbutamol with correlation coefficients higher than 95%. The average recovery rates were between 98.35 to 99.34%. The intra- and inter-day relative standard deviations were below 2%. The lower limit of quantification was found to be 136.25 and 21.69 ng/ml for vasicine and salbutamol respectively. This validated method was sufficiently sensitive, accurate and precise with less than 2% relative standard deviation. The validated HPLC method developed was further optimized for co-estimation of vasicine and salbutamol from in vitro plasma. Specific identifiable peaks were obtained in the optimized HPLC condition with excellent selectivity and sensitivity for the co-analysis of vasicine and salbutamol in the in vitro blood samples.

Keywords: Authentication, Co-estimation, HPLC, Salbutamol, Validation, Vasicine.