Comprehensive Two-dimensional Liquid Chromatography Method for the Determination of Flavonoids in Complex Plant Extracts

Flavonoids are a large group of polyphenolic secondary metabolites of plants with high biologically active properties. Their determination in plant extracts (medicinal, edible, and other plants, as well as products made from them) with a complex chemical composition is a difficult task and requires effective separation of the components. In this study, a new approach to the determination of 18 flavonoids in plant extracts using reversed-phase comprehensive two-dimensional liquid chromatography with UV detection (LC×LC-UV) is proposed. The approach has been successfully tested on various plant raw materials: above-ground parts of bog rosemary, bog bilberry, bilberry, lingonberry, and cranberry, as well as aspen and apricot knots. It was found that aspen knots are a rich source of aromadendrin and naringenin and their derivatives, while apricot knots are a rich source of catechin and epicatechin, and bog rosemary is a rich source of quercetin glycosides-hyperoside, reinoutrin, and avicularin. The approach for isolation of pure hyperoside, reinoutrin, and avicularin from bog rosemary extract by preparative HPLC was proposed. Other phenolic compounds of the studied plant extracts were classified (by LC×LC-MS/MS) as hydroxycinnamic acid derivatives, iridoids, and flavonoids. The developed approaches are promising for bioprospecting of flavonoids and phenolic compounds in various plant materials. It can also be used for quality control of medicinal, edible, and other plants, as well as products made from them.