Development, Validation and Forced Degradation Study of Emtricitabine and Tenofovir Alafenamide in its Pharmaceutical Dosage Form Using RP-HPLC

Aims: The present research was aimed to develop and validate a reverse phase high performance liquid chromatographic (RP-HPLC) method for the quantification of Emtricitabine (EMT) and Tenofovir Alafenamide (TEN) in combination.

Methodology: Separation was achieved under optimized chromatographic condition on an Inertsil C18, 250 x 4.6 mm, 5μm column. Various composition of mobile phase was tried. Separation of EMT and TEN was started with Methanol: Buffer and Methanol finally using solvent system of Buffer (pH 3.5) and Methanol in ratio of (30:70) and flow rate adjust at 1.0 ml/min was used as solvent system, the detection was carried out at 262nm using Shimazdu UV-visible detector. The mobile phase run time for the developed analytical method was 10 minutes.

Results: The standard curve was found linear in the concentration range of 20-60 μg/ ml (r2- 0.9994) and 2.5-7.5 μg/ ml (r2-0.9992) for EMT and TEN respectively. The %RSD was found to be 0.80-0.95% and 0.63-1.09 for EMT and TEN respectively. Percentage (%) recoveries for EMT and TEN to be in range of 100%-100.6% and 99.32%-100.83% respectively. The limit of detection and the limit of quantification were found to be 4.80 μg/ ml and 14.7 μg/ ml respectively for EMT and 0.11 μg/ ml and 0.33μg/ ml respectively for TEN. Results of forced degradation study showed EMT degradation in acid and base medium while TEN was showed degradation in oxidative stress. The proposed developed RP-HPLC method was validated statistically and the values were found to be within the acceptable limits.

Conclusion: In conclusion, the developed RP-HPLC method was found to be simple, specific, and rugged for simultaneous estimation of EMT and TEN. Validation results of method was found within the acceptable limits. Hence it can be used for analysis of EMT and TEN.