Discovery and identification of quality markers of Sparganii Rhizoma based on zebrafish thrombosis model

OBJECTIVE: The aim of the present study was to determine the quality marker (Q-Markers) of Sparganii Rhizoma against thrombus through an integration of investigations on its antithrombotic effect, content determination and spectrum-effect correlation analysis. METHODS: Based on the concept of Q-Marker, Sparganii Rhizoma was investigated for the identification of chemical component. The pharmacological effects on arachidonic acid-induced thrombosis in zebrafish were also investigated. The material basis in ethanol extract was determined by HPLC-UV. Furthermore, the potential Q-Markers were analyzed and predicted according to the effect-chemical correlation analysis. Finally, the anti-thrombotic Q-Markers were verified through the anti-thrombotic test of monomer components. RESULTS: The model of thrombosis zebrafish was established with larvae exposed to 100 µmol/L arachidonic acid for 1 h. Nine ingredients in Sparganii Rhizoma were identified as 5-hydroxymethylfurfural, vanillic acid, ferulic acid, p-hydroxybenzaldehyde, p-hydroxybenzoic acid, vanillin, protocatechuic acid, p-coumaric acid and isoferulic acid. According to the determination effect of zebrafish thrombosis model and HPLC content analysis results, all the other contents present positive correlation except 5-hydroxymethylfurfural, and the P values of three representative potential Q-Markers (ferulic acid, protocatechuic acid and p-coumaric acid) were 0.002, 0.001 and 0.026, respectively. CONCLUSION: Sparganii Rhizoma showed a dose-dependent effect on the recovery of reducing cardiac red blood cell on zebrafish model. Three phenolic acids (ferulic acid, protocatechuic acid and p-coumaric acid) were proved to possess the anti-thrombotic effects which could be regarded as the potential Q-Markers for quality assessment of Sparganii Rhizoma.