HPLC-LC/MS/MS of secoiridoid glycoside and flavonoids in Enicostemma littorale

Abstract

Enicostemma littorale Blume. a small herb of family Gentianaceae is commonly used as an antidiabetic agent. Its anti-inflammatory, antioxidant, hypoglycemic and anticancer activities were reported [2, 3]. Swertiamarin, a bitter secoiridoid glycoside, alkaloids, flavonoids, phenolic acids and xanthones were isolated [3]. In the present study, new HPLC-LC/MS/MS method was developed for the analysis of Secoiridoid Glycoside and Flavonoids in the ethyl acetate fraction of the Enicostemma Littorale. HPLC-UV analysis was carried out on a Perkin-Elmer HPLC system with diode array detector using a RP-18 Column (250×4.6mm; 5µM Kromasil). Mobile phases used included 0.2% formic acid and pH 3.5 with Ammonia in water (solvent A), and Acetonitrile (solvent B). A gradient was used as 0–10min, 25% B; 10–20min, 80% B; 20–33min, 80% B; 33–34min, 10% B; and 34–40min, 10% B. HPLC-LC/MS/MS analysis was performed with an Acquity UPLC and Xevo Quards MS system. The operational conditions used were: capillary voltage at 3.5 V, cone 35 V, dissolveation temperature at 600°C, MS2 scan from 100 to 900 amu. Mass spectra of Swertiamarin, Secoiridoid Glycoside was m/z 374 and Swertisin m/z 446, prenylated flavanoids m/z 356 and m/z 358.This method is suitable for the routine analysis, as well as for the separation and identification of known and novel Secoiridoid Glycoside and Flavonoids.