Discovery and quantification of quality control chemical markers for Fomes Officinalis Kechuan oral liquid by UPLC-MS/MS combined with chemometrics.

Fomes Officinalis Kechuan oral liquid (FOKOL) is a distinctive ethnic medicine originating from the Uyghur medical system in the Xinjiang Uygur Autonomous Region of China. Clinically, it is primarily indicated for the treatment of asthma, as well as acute and chronic bronchitis, exerting notable antitussive, antiasthmatic, and expectorant effects. However, this formulation has not yet been included in the Chinese Pharmacopoeia, and the current quality control standard is limited to the quantification of a single component, glycyrrhizic acid. In this study, UHPLC-Orbitrap-MS/MS was employed to comprehensively characterize the chemical constituents of FOKOL, leading to the tentative identification of 62 compounds. 20 constituents (3 nucleosides, 1 furfural compound, 7 flavonoids, 6 caffeoylquinic acid derivatives, 1 organic acid, 1 phenylpropanoid, and 1 triterpenoid saponin) were unequivocally confirmed by comparison with reference standards. Subsequently, a UPLC-TQS-MS/MS method coupled with multiple reaction monitoring (MRM) mode was developed for the simultaneous quantification of these 20 components and successfully applied to 19 batches of FOKOL samples. The results demonstrated that the established method exhibited high sensitivity and satisfactory precision, making it well-suited for the concurrent quantitative analysis of the 20 analytes in FOKOL. Method validation revealed good linearity for all analytes within the tested concentration ranges, with correlation coefficients (r) no less than 0.990. The relative standard deviations (RSDs) for precision ranged from 0.67% to 4.84%, and the recoveries at three spiked concentration levels were between 91.48% and 109.9%, confirming acceptable accuracy of the method. Furthermore, hierarchical cluster analysis (HCA) was performed to classify the 19 batches of FOKOL samples. In combination with chemometric approaches, principal component analysis (PCA) and partial least squares-discriminant analysis (PLS-DA) were further applied to screen for core quality markers and stability indicators. As a result, glycyrrhizic acid, 1,3-dicaffeoylquinic acid, liquiritin, neoliquiritin, and liquiritin apioside were identified as the key quality markers for FOKOL. This study clarifies the chemical material basis of FOKOL, provides a scientific foundation for the development of its quality control methods, and facilitates its broader clinical application.