UPLC–PDA–ESI–MS/MS Analysis of Compounds Extracted by Cardiac h9c2 Cell from Polygonum orientale

Abstract

A flavonoid-enriched extract (FEE) of Polygonum orientale was reported to show cardioprotective effect but only very few compounds were reported to contribute to the effect. Identification of compounds interacting with the target cardiac cell is important for the understanding of active compounds. To develop an efficient method for the screening of potential active compounds directly acting on the target cardiac cell in FEE and to structurally characterise these compounds. Flavonoid-enriched extract was prepared by extraction of the plant with water, addition of ethanol to the solution to remove polysaccharides and proteins, and removal of tannins by a polyamide column chromatography. Cell extraction was conducted on a cardiac h9c2 cell and the solution containing compounds released from the cell were desalted by solid phase extraction. Compounds present in the cell extract were detected by ultra-performance liquid chromatography (UPLC) and targeted multi-reaction monitoring (MRM), while their structures were characterised by UPLC-photodiodide array (PDA)-electrospray ion source (ESI)-MS/MS investigations of the FEE. Twenty-three potentially active phenolics including ten flavonoid C-glycosides and six flavonoid O-glycosides have been identified from the 40 compounds screened in the cell extract. Among these compounds, three were new and nine were identified from this plant for the first time. Strategies for the structural characterisation of flavonoid glycosides were also discussed. The study has shown that FEE contains the flavonoid as its major principles and the coupling of UPLC-PDA-ESI-MS/MS and targeted UPLC-MRM with target cell extraction is an efficient method for the screening and structural characterisation of potential active compounds.