UHPLC-PDA-ESI/HRMSⁿ Profiling Method To Identify and Quantify Oligomeric Proanthocyanidins in Plant Products

Abstract

Oligomeric proanthocyanidins were successfully identified by UHPLC-PDA-HRMSn in a selection of plant-derived materials (jujube fruit, Fuji apple, fruit pericarps of litchi and mangosteen, dark chocolate, and grape seed and cranberry extracts). The identities of 247 proanthocyanidins were theoretically predicted by computing high-accuracy masses based on the degree of polymerization, flavan-3-ol components, and the number of A type linkages and galloyls. MSn fragments allowed characterization on flavan-3-ol based on the monomer, connectivity, and location of A-type bonds. Identification of doubly or triply charged ions of 50 PAs was made on the basis of theoretical calculations. A single catechin standard and molar relative response factors (MRRFs) were used to quantify the well-separated PAs. The ratios of the SIM peak counts were used to quantify each of the unseparated isomers. This is the first report of direct determination of each of the proanthocyanidins in plant-derived foods and proanthocyanidins containing an epifisetinidol unit in grape seeds.

Characterization of Polyphenolics in Grape Pomace Extracts Using ESI Q-TOF MS/MS

“…Their main fragments are aligned with -(progressively loss (epi)cat units) or loss water molecules. A portion of these precursor ions and possible isomers of tetramers and pentamers with one and two A-type linkages are mentioned previously in other foods such as peanuts and cranberry [5,24,35,36].…”

“…They displayed 2 amu difference from the corresponding B-type PACs at m/z 577, 865, 1153, 1441 and 1729 [5,7,24,31,35,36]. Further, the observed [M+H] + ions for A-type PAC dimers to hexamers at m/z 577*, 865*, 1153*, 1441* and 1729* also present 2 amu difference from the corresponding B-types at m/z 579*, 867*, 1155*, 1443* and 1731* [6,20,21] (Figure 1,2 and Table 2).…”

A computational tool for accelerated analysis of oligomeric proanthocyanidins in plants

“…Proanthocyanidins have a high structural diversity that is affected by the two chiral centers in each flavan-3-ol unit, existence of substitutions (e.g., galloyl), degrees of polymerization (DP), orders and types of linkage between the flavan-3-ol units, and different monomer units, e.g., epicatechin/catechin catechin/epicatechin (EC), epiafzelechin/afzelechin (EA), epigallocatechin/gallocatechin (EG), epifisetinidol/fisetinidol (EF), and epirobinetinidol/robinetinidol (ER)(Figure S1) (Hümmer and Schreier, 2008; Lin et al, 2014). These structural differences are often associated with the various nutritional outcomes of proanthocyanidins.…”

“…Analytical methods for the identification and quantification of PA oligomers in food to identify the role of PAs in health and nutrition is very important (Feliciano et al, 2012; Lin et al, 2014), but PA analysis presents a great challenge. Both normal and reverse-phase high-performance liquid chromatography (HPLC) has been applied to the analysis of PAs, but the selection of a chromatographic method mainly depended on the PAs expected to be present in the matrix and the research focus (Hümmer and Schreier, 2008).…”

“…Moreover, the multiply charged ions under ESI of proanthocyanidins with high degree polymerization usually stack on the single charged ions of monomers in the MS spectra. Thus, identification and quantitation of proanthocyanidins manually are very difficult, due to the presence of multiply charged ions, co-eluting peaks, and isomers (Appeldoorn et al, 2009b; Hokkanen et al, 2009; Irungu et al, 2008; Lin et al, 2014; Nunez et al, 2006; Sarnoski et al, 2012). …”