Asian Journal Of Research in Chemistry

A stereospecific RP-HPLC method for the separation and estimation of S-isomer content in tenofovir disoproxil fumarate drug substance was developed and validated on a reverse-phase amylose derivativesed chiral column. The effect of organic modifiers, namely, methanol, acetonitrile and triethylamine in mobile phase was optimized as 0.1% triethyl amine in mixture of water, methanol and acetonitrile (10:75:15, v/v/v) to obtain the best enantiomeric separation. UV detection was performed at 260nm. The flow rate was kept at 0.8 mL/min and the column temperature was set at 25°C. The retention times of tenofovir and its S-isomer were observed to be 5.137 min and 8.768 min respectively. The linear regression analysis data for calibration plots showed a good linear relationship over a concentration range of 0.0005mg/mL - 0.01mg/mL for S-isomer. The values of the correlation coefficient were 0.999 for S-isomer. The limit of detection (LOD) was 0.0001 mg/mL and the limit of quantification (LOQ) was 0.0005mg/mL. The precision of S-isomer at LOQ level was evaluated through six replicate injections and the % RSD of the peak response achieved is 3.07. The percentage recoveries of S-isomer from tenofovir disoproxil fumarate were ranged from 96.4% to 102.2%. The proposed method was found to be accurate, precise and suitable for the separation and enantiomer excess determination of unwanted S-isomer in active pharmaceutical ingredients (API). The analytical results were supported by statistical parameters. The proposed method could be successfully applied to the enantiomeric purity analysis of tenofovir disoproxil fumarate in drug substance. This method was validated in as per ICH Q2 (R1) and USP validation of compendial methods <1225>.