Development and Validation of Stability-indicating HPLC-DAD method for simultaneous determination of Emtricitabine, Rilpivirine, and Tenofovir Alafenamide in bulk and their Pharmaceutical dosage forms

A simple and rapid high performance liquid chromatographic method was developed and validated for simultaneous estimation of Emtricitabine(EMT), Rilpivirine (RPV), and Tenofovir alafenamide fumarate(TAF)in bulk active pharmaceutical ingredients & its tablet formulation. The method was established using Agilent C18 (250 × 4.6 mm, i.d., 5 μm) column, mobile phase consisting of 0.1%Formic acid: Acetonitrile (65:35%, v/v) at a flow rate of 1 mL/min with isocratic elution, injecting 20 μL sample into the chromatographic system. The eluted compounds were detected by using PDA detector at detection wavelength of 250 nm and temperature was maintained at 30 °C. Retention times of Reference Standard Emtricitabine, Rilpivirine, and Tenofovir alafenamide was found to be 2.90, 4.34, 6.58mins respectively. The calibration curve was plotted over the concentration range 4-20 μg/mL for EMT, 2-10 μg/mL of RPV and 1-5 μg/mL of TAF.The recoveries for EMT, RPV and TAF were found to be 99.12, 99.38, and 99.18%, respectively. All the validation parameters results were obtained with in acceptance limit.Developed method was subjected to forced degradation studies under specified conditions, which meets the required criteria. The present method was specific, sensitive, reproducible, precise, rapid and simple. Keawords : RP-HPLC, Emtricitabine, Rilpivirine, and Tenofovir alafenamide, stability studies. Introduction Around 33.4 million people were living with HIV in year 2008 and around 2 million people have died in the same year. Highly active antiretroviral therapy (HAART) has brought new hope for those people who live with HIV/AIDS by decreasing the morbidity and mortality among people infected with HIV. Highly active antiretroviral therapy also has improved the quality of life among the people who live with HIV/AIDS. Saidulu. P et al /International Journal of ChemTech Research, 2018,11(09): 329-339 DOI= http://dx.doi.org/10.20902/IJCTR.2018.110939 International Journal of ChemTech Research CODEN (USA): IJCRGG, ISSN: 0974-4290, ISSN(Online):2455-9555 Vol.11 No.09, pp 329-339, 2018 Saidulu. P et al /International Journal of ChemTech Research, 2018,11(09): 329-339. 330 Combination therapy is preferred to be the gold standard for the treatment of AIDS so as to maximize potency, minimize toxicity, and diminish the risk for resistance development and reduction of pill burden to once-daily dosing so as to optimize the patient’s compliance and reduce the treatment costs. The nucleoside reverse transcriptase inhibitors and non-nucleoside reverse transcriptase inhibitors as multidrug combinations are effective in the therapy of human immunodeficiency virus (HIV) infection and are used as a part of highly active antiretroviral Therapy, for the treatment of HIV 1, 2 . Literature indicates spectrophotometry 5-11 , HPLC 12– 15 , HPTLC 16 and LC/MS/MS 17 methods for determination of TDF individually and in combination with other drugs in pharmaceutical formulations, drug substance, and biological matrices. Similarly for EMT individually and in combination with other drugs by UV 18, 19 , HPLC in pharmaceutical formulations, drug substance and biological matrices 20–25 , HPTLC, LC/MS/MS 26 and stability indicating liquid chromatographic methods 27 were reported. A detailed literature survey for RPV revealed that few analytical methods are available using spectrophotometric 28 , HPLC 29 and HPTLC 30 , individually. Literatures are available to show the existence of HPLC method for the triple drug combination of TDF, EMT, and RPV as well 3, 4 .