Objective: The present report embodies detailed diagnostic features of five important Red listed Swertia species for easy and authentic identification along with HPTLC fingerprinting using the c-glucoxanthone mangiferin as a biomarker. The key objectives of this study were to reduce indiscriminate harvesting of different Swertia species from its natural habitat and the development of a reliable kit based on HPTLC fingerprinting to reduce adulteration in commercial trade. Methods:Chromatography was performed on Silica gel 60 F254 TLC plates with ethyl acetate-glacial acetic acid-formic acid- water- 100:11.0:11.0:26 as mobile phase. Densitometric measurement was performed at λ=254 nm. The method was found to be simple, reliable, precise and convenient for routine analysis. Results: Although mangiferin was detected in substantial amount from all populations of Swertia chirata irrespective of their geographical locations and to a lesser degree in Swertia nervosa, the biomarker was totally absent in Swertia bimaculata, Swertia dilatata and Swertiapaniculata. Conclusions:Floral morphology can be effectively used for the construction of an easy identification tool that can be utilized at the field level to screen out Swertia chirata from other closely related species that flower at the same time along with Swertia chirata. High Performance TLC using mangiferin as a biomarker can be utilized for quality screening and checking adulteration among different species and populations of Swertia.