Comprehensive Analysis of Bile‐derived Medicinal Materials Based on Ultra‐High‐Performance Liquid Chromatography Coupled With Charged Aerosol Detection and Chemometrics

ABSTRACT The medicinal use of bile‐derived materials dates back thousands of years. However, due to their complex origins and morphological similarities, adulteration of expensive bile types with cheaper alternatives remains prevalent in the market, significantly compromising quality control and regulatory oversight. To address this critical issue, this study employed ultra‐high‐performance liquid chromatography coupled with charged aerosol detection (UHPLC‐CAD) technology combined with chemometric approaches, including chromatographic fingerprint similarity analysis, hierarchical cluster analysis, principal component analysis, and orthogonal partial least squares‐discriminant analysis to systematically analyze 10 bile‐derived medicinal materials: biotransformed bear bile powder, bear bile powder, pig bile powder, ox bile powder, sheep bile powder, chicken bile powder, duck bile powder, goose bile powder, rabbit bile powder, and snake bile powder. Chromatographic fingerprint similarity analysis revealed significant differences among the medicinal materials from different sources. Hierarchical cluster analysis and principal component analysis successfully achieved accurate classification of the 10 types of bile‐derived medicinal materials, while orthogonal partial least squares‐discriminant analysis identified nine characteristic differential components with variable importance in projection >1: taurocholic acid, taurochenodeoxycholic acid, glycodeoxycholic acid, tauroursodeoxycholic acid, glycocholic acid, taurodeoxycholic acid, glycohyodeoxycholic acid, glycochenodeoxycholic acid, and chenodeoxycholic acid. This study established a UHPLC‐CAD method capable of simultaneously separating 17 bile acids, combined with multidimensional chemometric approaches, to conduct a comprehensive analysis of 10 bile‐derived medicinal materials. The research successfully identified characteristic compounds for each bile powder type, providing a reliable methodology for both the identification and quality control of bile‐derived medicinal materials.