Chromatographic analysis of diverse fruit components using HPLC and UPLC

Abstract

The nutritional properties of fruits are due to the combined action of many categories of molecules. Studies on molecular nutrition, therefore, requires analytical methods that can cope with diverse molecular groups, including metabolites that may be currently unknown. Fractionation and identification protocols are crucial to validate identified compounds, and can be adapted from methodologies developed for metabolome studies. Pectin-related problems during fractionation of samples from ripe fruit products, both undigested and after digestion, as well as samples from in vitro cell culture bioassay were addressed, and chromatographic separations of target compounds (carotenoids, phenolics, sugars, and organic acids) were developed based on RP chromatography using HPLC and UPLC systems. The final method comprised of a chloroform–methanol–water (2 : 1 : 3) extraction to simultaneously extract as many polar and nonpolar compounds as possible, followed by chromatographic analysis using (a) a HPLC-ELSD-UV system with a Prevail Carbohydrate column for sugar analysis, (b) Prevail Organic Acid column for organic acid analysis, (c) LC-MS ESI of eluants from a XTerra C-18 column for phenolics and (d) UPLC-PDA systems in a C-18-Bridged Ethane in Hybrid matrix column for carotenoids and other hydrophobic components. Acetonitrile-based mobile phases can be used to run all chromatographic systems developed.