A kinetic and thermodynamic study of procyanidin oligomer conformation by 1H NMR and DFT

Two procyanidin dimers, four trimers and single tetramer, pentamer and hexamer species were isolated by preparative liquid chromatography from fresh cocoa beans. The most abundant procyanidin dimer was identified as procyanidin B2 by means of UHPLC-ESITOF-MS and one- and two-dimensional hetero- and homonuclear NMR spectroscopy. The effect of aggregation was discounted based on DOSY NMR experiments in conjunction with the Einstein-Stokes equation. The hydrodynamic radius of the molecule (8.5 Å) is too small for aggragation. Line shape analysis of variable temperature 1H NMR procyanidin B2 spectra by a custom developed program (NMRfit) yielded first order kinetic reaction parameters for the conformational interchange. The calculated is 57.03 ± 3.92 kJ.mol-1 , is 53.36 ± 3.82 kJ.mol-1 , while is -0.3454 ± 14.76 J.mol-1 K -1 and is -4.398 ± 14. 39 J.mol-1 K -1 . The thermodynamic parameters for the conformational interchange reaction are: = 3.670 ± 0.217 kJ/mol and = 4.053 ± 1.57 J.mol-1 K -1 . The conformational geometries and standard reaction Gibbs free energies were calculated via DFT. Current conformational models reported in literature for procyanidin B2 were found to be insufficient. Four conformers were identified in this text, as opposed to only two conformers reported previously. The new conformations have been named: fully compact (FC), fully extended (FE), partially compact (PC) and partially extended (PE). Intramolecular interactions that include hydrogen bonds were shown to occur to varying degrees within each conformer. The resulting electronic stability hierarchy was found to be FC>FE>PC>PE with two of the three functionals used in the calculations. However, it was found that the entropy component of the Gibbs free energy plays a significant role in determining the thermodynamic distribution. The conformations with the most internal interactions were also found to be less available to solvent interactions due to a decrease in the polarization of their hydroxyl moieties and the reduced availability of the solvation sites, due to a closed conformation. The final Gibbs free energy hierarchy (PC>FE>PE>FC) indicates that the PC and FE conformers are the two conformers visible on NMR spectra. Finally, the distribution of conformers is shown to approach 1:1 in increasing solvent polarity.National Research Foundatio