摘要： A highly sensitive and specific fluorescence dependent approach was created for quantitation of a recently approved anti-HCV drug (ledipasvir). This approach relies on the innovative enhancement in fluorescence intensity of ledipasvir upon exposing the cited drug to direct UV irradiation as a photo-physical fluorescence enhancer. The fluorescence of the resultant solution was measured at an emission peak of 375 nm (321 nm excitation). The photoluminescence properties of the resultant product were carefully examined and the quantitative rectilinear concentration plot for the method was 5.0–150.0 ng mL?1 with a detection limit of 0.9 ng mL?1 and a quantitation limit of 2.7 ng mL?1. The excellent analytical features of the proposed method allow to the specific and sensitive estimation of ledipasvir either in plasma samples or in tablet dosage form without any interference from pharmaceutical excipients or other co-formulated anti-HCV drugs (sofosbuvir). The developed analytical and bio-analytical procedures were created and validated according to ICH guidelines and FDA guidelines, respectively. Since the main elimination route for ledipasvir is via faecal excretion, the studied drug was determined for the first time in faecal samples by the method with adequate recovery. Moreover, the pharmacokinetic parameters (Cmax, tmax, t1/2, AUC0–t, and AUC0–N) for ledipasvir were determined by the proposed method. Additionally, the proposed method was successfully applied for supervising the content uniformity for ledipasvir in pharmaceutical tablets.