ICH and US-FDA validated HPTLC methods with greenness assessments for the assay of mixtures prescribed in stroke prophylaxis: application to pharmaceutical preparations and human plasma

Abstract

The importance of the binary mixtures of the novel oral anticoagulants (NOACs): apixaban (APX), edoxaban tosylate (EDX) and rivaroxaban (RIV) with the lipid-lowering statin, rosuvastatin calcium (ROS) is highly emerging to save lives of cardiovascular patients as these combinations are used in prophylaxis from stroke. A high-performance thin-layer chromatography (HPTLC) method was developed for the quantitative assay of these life-saving mixtures in tablets and human plasma. Two mobile phases were developed for the assay in bulk and tablets; the first one: toluene‒ethyl acetate‒methanol‒25% ammonia (3.5:4.5:2:0.2, V/V) (method I) used for the three mixtures, and the second one: methanol‒25% ammonia (9.95:0.05, V/V) (method II) used for EDX/ROS mixture only. For analysis in human plasma, APX was used as internal standard in RIV/ROS and EDX/ROS mixtures using methods I and II, respectively, while RIV was used as internal standard in APX/ROS mixture using method I; the methods were validated according to the Food and Drug Administration (FDA) regulation for analysis in biological fluids. The method selectivity was demonstrated by its ability to simultaneously analyze the drugs in the presence of dosage form excipients and in the presence of plasma interferences (analysis in biological fluid) at single wavelength (291 nm) by use of the internal standard.