Ultra performance liquid chromatography coupled with principal component and cluster analysis of Swertia chirayita...

This article describes the study to standardize phytochemically and distinguish Swertia chirayita from that of possible substitution/adulteration using ultra performance liquid chromatography (UPLC) with photodiode array detector (PDA) and chemometric tools viz. principal component analysis (PCA) and hierarchical clustering analysis (HCA). Five ecotypes of Swertia chirayita and five possible substitutions, e.g.,Swertia bimaculata (SB), Swertia chordata (SCH), Swertia ciliata (SCL), Swertia paniculata (SP), and Halenia elliptica (HE) collected from different Indian Himalayan region. Samples evaluated for 04 marker compounds- swertiamarin (SM), mangiferin (MF), gentiopicroside (GP), and sweroside (SW). Reverse phase column (Waters Acquity BEH C₁₈, 50 mm × 2.1 mm , 1.7 μm) provided high resolution for all target analytes with binary gradient elution. The detector response was linear (concentration 2.5-125 μg/mL, R² > 0.999). The limit of detection (LOD) and quantification (LOQ) of targeted compounds was in the range of 1.40-2.06 and 4.57-6.27 μg/mL respectively. The combined relative standard deviation (%RSD) for intra-day and inter-day precision values were less than 2%. The recoveries study comply the method suitability. Chromatogram similarity analysis based on congruence coefficient was higher than 0.925 for the chirayita ecotypes while much lower than 0.629 for possible substitutes. HCA showed that the samples could be clustered (all 5 clusters in two-level) reasonably into different ecotypes and substitutes. HCA together with loading plots has indicated different chemical properties of all five groups. PCA results showed that the discrimination of chirayita ecotypes is because of the presence of SW while SM may have more influence on the targeted substitutes to discriminate from chirayita ecotypes. Therefore, UPLC fingerprint in association with chemometric tools provides a reliable and accurate quality assessment and detection of possible adulteration.