Discovery of antitussive material basis and mechanisms in Citri Sarcodactylis Fructus by coupling UHPLC-Q/Orbitrap HRMS combined spectrum-effect relationship and metabolomics analyses.

ABSTRACT

Citri Sarcodactylis Fructus (CSF) is widely used as food raw material and traditional Chinese medicine. Fingerprints of different fractions of CSF were established for spectrum-effect relationship analysis, and the main compounds were identified by UHPLC Quadrupole Orbitrap high-resolution mass spectrometry (UHPLC-Q/Orbitrap HRMS). The antitussive effect was evaluated using a classical mouse model of cough induced by ammonia water. One-way ANOVA was used to determine differences in efficacy. The potential active compounds were screened by spectrum-effect relationship with grey relational degree analysis (GRA), Pearson bivariate correlation analysis (Pearson's), and partial least squares analysis (PLS) analyses. Differential metabolites associated with cough in serum were screened and identified using orthogonal partial least squares-discriminant analysis, HMDB database, and UHPLC-Q/Orbitrap HRMS. Metabolic pathway analysis was performed using MetaboAnalyst 5.0. Results indicate that 70 % ethanol elution fraction (70 % EF) is the major active fraction, and 8 components were identified to possess antitussive effects. Metabolomic analysis showed that 19 metabolites are potential biomarkers related to cough, and 70 % EF can remarkable restore 13 of them to normal levels (P < 0.05). These biomarkers are mainly involved in glycerophospholipid metabolism and sphingolipid metabolism. This study aims to reveal the main pharmacodynamic active sites and potential active ingredients of CSF's antitussive effect. In addition, metabolomics was used to preliminarily elucidate the in-vivo regulatory mechanism of the antitussive effect of the 70 % EF of CSF.