Document Type: Original Article

Authors

1 a Department of Pharmaceutical Analysis, College of Pharmacy, SRIPMS affiliated to Dr. M.G.R Medical University, Coimbatore, India.

2 Department of Pharmaceutical Analysis, College of Pharmacy, SRIPMS affiliated to Dr. M.G.R Medical University, Coimbatore, India.

Abstract

A simple, specific, sensitive and rapid stability-indicating high performance thin layer chromatographic method has been developed for the simultaneous estimation of ledipasvir and sofosbuvir in combined dosage form. In this method, the separation was achieved on silica gel 60F254 thin layer chromatography plates using mobile phase comprising of hexane: ethyl acetate: methanol (5:3:2,v/v/v) with 3 drops of ammonia at 288 nm as selected wavelength on a densitometer. The Rf value of sofosbuvir and ledipasvir were observed to be 0.21±0.02 and 0.43±0.02, respectively. The linearity, precision, accuracy, robustness, specificity, limit of detection and limit of quantitation of the method were validated according to the ICH guidelines. The linear regression analysis for calibration plots produced r2=0.9948±0.0005 and r2 = 0.9927±0.0003 for ledipasvir and sofosbuvir, respectively. Percent recovery of the drugs from tablet formulation was carried out by standard addition method and was found to be close to 100 and relative standard deviation was less than 2%, which indicated good accuracy and precision of the method. The factors evaluated in the robustness test were determined to have an insignificant effect on the selected responses. To make the method more specific stress studies were conducted for both drugs individually as well as in the combined form by subjected to acid, alkali and neutral hydrolysis, oxidation, thermal and photolytic degradation. Degradation studies indicated ledipasvir to be susceptible to acid and alkaline hydrolysis, thermal, photolytic and oxidative degradation whereas sofosbuvir was susceptible only to alkaline hydrolysis. The degradation product peaks were well resolved from the pure drug with significant differences in their Rf values. The results indicate that the method is suitable for the routine quality control testing of marketed tablet formulation.

Graphical Abstract

Keywords

Main Subjects

1. National Center for Biotechnology Information. PubChem Compound Database; CID=67505836,https://pubchem.ncbi.nlm.nih.gov/compound/67505836 (accessed Apr. 10, 2018). Ledipasvir structure

2. Pol, S., Vallet-Pichard, A., and Corouge, M. (2016) Daclatasvir-sofosbuvir combination therapy with or without ribavirin for hepatitis C virus infection: from the clinical trials to real life. Hepatic Medicine: Evidence and Research. 8: 21–26.

3. Fung, A., Jin, Z., Dyatkina, N., Wang, G., Beigelman, L., and Deval, J. (2014). Antimicrobial Agents and Chemotherapy. 58: 3636-3645.

4. National Center for Biotechnology Information. PubChem Compound Database;CID=45375808,https://pubchem.ncbi.nlm.nih.gov/compound/45375808 (accessed Apr. 10, 2018). Sofosbuvir structure

5. Afdhal, N., Zeuzem, S., Kwo, P., Chojkier, M., Gitlin, N., Puoti, M., Romero-Gomez, M., Zarski, J. P., Agarwal, K., Buggisch, P., Foster, G.R., Bräu, N., Buti, M., Jacobson, I. M., Subramanian, G.M., Ding, X., Mo, H., Yang, J.C., Pang, P.S., Symonds, W. T., McHutchison, J. G., Muir, A. J., Mangia, A., and Marcellin, P. (2014). N Engl J Med. 370: 1889–1898.

6. Mohan Vikas, P., Satyanarayana, T., Vinod Kumar, D., Mounika, E., Sri Latha, M., Anusha, R., and Sathish, Y. (2016) J. Global Trends Pharm. Sci. 7: 3013-3015.

7. Devilal, J., Durgaprasad, B., Pal, N., and Avanapu, S. R. (2016) WJPPS. 6: 1312-1321.

8. Rezk, M., Basalious, E., and Karim, A. I. (2015) J. Pharmaceut. Biomed. Anal. DOI: 10.1016/j.jpba.2015.05.006.

9. Nebsen, M., and Eman, S. E. (2016) J. Chromatogra. Sci. 54: 1631–1640.

10. Debasish, S., and Gananadhamu, S. (2017) J. Pharmaceut. Biomed. Anal. 138: 29-42.

11. Amira, S., Eldin, S. M., Azab, A. S., and Magda El-Maamly. (2017) Antiviral drugs. J. Pharm. Pharmacol. Res. 1: 028-042.

12. Nehal, F. F., and Nada, A.W.S. (2017). J of liquid chromatography and related technologies. 327-332.

13. Madhavi, S., and Prameela R. A. (2017). Int. J. Pharm. Pharmaceut. Sci. 9: 35-41.

14. Bakht, Z., Faisal, S., and Waseem, Hassan. (2016). Chromatographia. 79: 1605–1613.

15. Mahsa, K., Mahmoud, R. S., Vahid, M., and Nahid, T. Z. (2018). Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy. 194: 141-151.

16. Fathy, M. S., Khalid, A. A., Ahmed, A. A., Ahmed El-Olemy., and Ebrahim, A. (2017). J. Adv. Pharmacy Res. 1: 185-192.

17. Fathy, M. S., Khalid, A. A., Ahmed, A. A., Ahmed El-Olemy., and Ebrahim, A. (2017). Anal. Che. Lett. 7: 241-247.

18. Elkady, E. F., and Aboelwafa, A. A. (2016). Int. J. AOAC. 99: 1252-1259.

19. Sherif Abdel‐Naby Abdel‐Gawad. (2017). Eur. J. Chem. 8: 8‐12.

20. Guidance for Industry, Analytical Procedures and Methods Validation, Chemistry, Manufacturing, and Controls Documentation, Draft Guidance, U.S. Department of Health and Human Services, Food and Drug Administration, Center for Drug Evaluation and Research (CDER), Center for Biologics Evaluation and Research (CBER), July 2000.

21. ICH Harmonised Tripartite Guideline Stability Testing of New Drug Substances and Products Q1A(R2), Current Step 4 version, dated 6 February 2003.

22. Monika. B., and Singh, S. J., (2002). Biomed. Anal. 28: 1011–1040.

23. Sethi, P. D., 1997. HPTLC Quantitative Analysis of drug in pharmaceutical

formulation, third ed. New Delhi: CBS Publishers and Distributors, pp. 1-56.

24. ICH Harmonised Tripartite Guideline, Validation Of Analytical Procedures: Text and Methodology, Q2(R1), Current Step 4 version, Parent Guideline dated 27 October 1994, (Complementary Guideline on Methodology dated 6 November 1996 incorporated in November 2005).