¹H–¹³C HSQC NMR Spectroscopy for Estimating Procyanidin/Prodelphinidin and cis/trans-Flavan-3-ol Ratios of Condensed Tannin Samples: Correlation with Thiolysis

Abstract

Studies with a diverse array of 22 purified condensed tannin (CT) samples from nine plant species demonstrated that procyanidin/prodelphinidin (PC/PD) and cis/trans-flavan-3-ol ratios can be appraised by (1)H-(13)C HSQC NMR spectroscopy. The method was developed from samples containing 44-∼100% CT, PC/PD ratios ranging from 0/100 to 99/1, and cis/trans ratios ranging from 58/42 to 95/5 as determined by thiolysis with benzyl mercaptan. Integration of cross-peak contours of H/C-6' signals from PC and of H/C-2',6' signals from PD yielded nuclei-adjusted estimates that were highly correlated with PC/PD ratios obtained by thiolysis (R(2) = 0.99). cis/trans-Flavan-3-ol ratios, obtained by integration of the respective H/C-4 cross-peak contours, were also related to determinations made by thiolysis (R(2) = 0.89). Overall, (1)H-(13)C HSQC NMR spectroscopy appears to be a viable alternative to thiolysis for estimating PC/PD and cis/trans ratios of CT if precautions are taken to avoid integration of cross-peak contours of contaminants.

Galloylated proanthocyanidins from shea (Vitellaria paradoxa) meal have potent anthelmintic activity against Ascaris suum

“…the percentage of galloylated subunits (Xg) ranged from 29, 46 and 38% in the F1-, F2-and F3-fractions. Given our previous observations we had expected that PA contents would increase in line with the acetone content (Zeller et al, 2015). However, shea meal appears to contain non-PA impurities (see Section 2.3.2) that were eluted by 50% aqueous acetone and this explains the relatively low PA content of the F2-fraction.…”

“…The F2-fraction had the highest PA content (45 g/100 fraction) and was subjected to NMR (HSQC) (Grabber et al, 2013;Zeller et al, 2015) in order to assess the nature of the impurities and to check for any other tannins that might not have been detected by thiolysis. This suggested that the major impurities consisted of lipids, sugars and other phenolic compounds (i.e.…”

“…1 H-13 C correlation 2D NMR (HSQC) spectra were recorded at 27°C on a Bruker Avance III 500 ( 1 H 500.13 MHz, 13 C 125.76 MHz) instrument equipped with TopSpin 2.4 software and a 5-mm BBI 1 H/ 13 C gradient probe (Bruker, Coventry, U.K.) (Zeller et al, 2015). Spectral resonances were referenced to the residual signals of DMSO-d 6 (2.49 ppm for 1 H and 39.5 ppm for 13 C spectra).…”

Activity, Purification, and Analysis of Condensed Tannins: Current State of Affairs and Future Endeavors

“…Examination of purified CT samples by two-dimensional 1 H-13 C HSQC NMR spectroscopy as a means to determine PC/PD and cis/trans ratios correlated strongly with thiolysis data (Zeller et al, 2015a). Efforts to extract and validate additional structural information from these data such as mDP, type, and quantitation of interflavan connectivity (A-type and C4-C6) are ongoing.…”

Benefits of Condensed Tannins in Forage Legumes Fed to Ruminants: Importance of Structure, Concentration, and Diet Composition

“…An important constraint in the quest for valid tannin data is the requirement for high purity standards for quantitation, which means that the CT concentration and purity of the standards needs to be assessed by CT-specific methods such as thiolysis (Gea et al, 2011;Grabber et al, 2013;Williams et al, 2014a) or nuclear magnetic resonance spectroscopy (Zeller et al, 2015a). As Zeller (2019) has addressed this topic in detail, only a few additional techniques are described below.…”