Background: The rapid growth of COVID-19 infections may result in second wave of infection and an overwhelmed health care providing systems. Ledipasvir and sofosbuvir can be a good choice for management of COVID-19 patients. Development of simple, sensitive, and rapid assay for simultaneous determination of ledipasvir / sofosbuvir to investigate their pharmacokinetic parameters in human plasma, and aid in therapeutic drug moitoring in COVID-19 patients seems to be essential. Besides, its application in bioequivalence study of ledipasvir 90mg / sofosbuvir 400mg film coated tablets generic and reference products to ensure bioanalytical method reliability. Methods: After extraction of ledipasvir and sofosbuvir from human plasma, it was chromatographed with mobile phase consisting of ammonium formate pH 2.8 : acetonitrile (10 : 90 V/V) at flow rate 0.55ml/min, ESI positive mode, and m/z 889.8130.1, 530.3243.1, 739.4565.3 for ledipasvir, sofosbuvir and daclatasvir (internal Standard) respectively. The bioequivalence study was conducted in a partial replicated crossover design invovlving 36 volunteers. The criteria used to assess bioequivalence of the two products were AUC 0-t, AUC 0-inf, Cmax, and Tmax for sofosbuvir, and AUC 0-72, Cmax, and Tmax for ledipasvir. Results: The described method of analysis showed that the average recovery of Ledipasvir and Sofosbuvir from human plasma was 95.180%, and 94.721%. The limit of quantitation was 0.1ng/ml for both drugs, and the correlation coefficient (r2) was equal to 0.999 for ledpasvir and sofosbuvir. Statistical analysis (ANOVA) of the measured parameters showed that there was no significant difference between the two products. Conclusion: The LC/MS/MS method presented is direct, simple, reproducible, sensitive, and linear for determination of ledipasvir / sofosbuvir in plasma, and is adequate for its clinical pharmacokinetic studies, and use in therapeutic drug monitoring. Besides the generic product was found to be biologically equivalent to the reference product regarding their kinetic behavior.