Discrimination of Two Species of Tibetan Medicine “Zachi” Based on Chromatographic Approach and Multivariate Statistical Analysis

ABSTRACT The Tibetan medicine “Zachi” derived from Saussurea and Ilex , is widely used for its therapeutic properties. However, the morphological and chemical similarities between these two species often lead to challenges in their identification and quality control. This study presents a comprehensive approach to discriminate between the two species using chromatographic techniques coupled with multivariate statistical analysis. ultra‐high‐performance liquid chromatography‐quadrupole time‐of‐flight mass spectrometry‐mass spectrometry (UHPLC/Q‐TOF‐MS/MS) was employed to profile the chemical compositions of both species, focusing on key bioactive compounds. Multivariate statistical tools, including principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS‐DA), were applied to identify distinct chemical markers and classify the samples. The results revealed significant differences in the chemical fingerprints of Saussurea and Ilex , with characteristic compounds such as luteolin and chicoric acid emerging as potential markers for species discrimination. PCA and OPLS‐DA models demonstrated robust classification accuracy, providing a reliable framework for distinguishing the two species. This integrated chromatographic and statistical methodology offers a practical solution for ensuring the authenticity and quality of “Zachi”, which also contributes to the standardization and modernization of Tibetan medicine by establishing a scientific basis for species identification, supporting its safe and effective clinical application.