A nuclear magnetic resonance study of the conformation and the interconversion between the enantiomeric conformers of bilirubin and mesobilirubin in solution

Abstract

¹ H N.m.r. spectra of bilirubin and mesobilirubin in chloroform solutions and their temperature dependence were studied. At room temperature the spectrum of the methylene protons of the two propionic acid residues appear as two partially overlapping ABCX multiplets. Specific assignments of the signals were obtained through the ¹H–¹H nuclear Overhauser enhancement and selectively protondecoupled ¹³C spectra. Approximate conformation of the propionic acid residues in terms of the dihedral angles within the CH₂CH₂ residue and its angles relative to the rest of the molecule was determined on the basis of the chemical shifts, geminal, and vicinal coupling constants. The results are consistent with previous determinations of the conformation of bilirubin and mesobilirubin in chloroform solutions and in the solid state. At elevated temperatures the spectral lineshape of the propionic methylene protons of bilirubin and mesobilirubin changes drastically. The changes were successfully simulated using the DNMR5 computer program, assuming an exchange of the type ABCX A′B′C′X′⇌XCBA X′C′B′A′, caused by inversion between the two enantiomeric conformations of these molecules. The activation parameters of the exchange are ΔH^‡ 74.1 ± 2.5 kJ mol^–1 and ΔS^‡– 8.8 ± 7.1 J K^–1 mol^–1 for bilirubin and ΔH^‡ 72.4 ± 1.2kJ mol^–1 and ΔS^‡– 12.5 ± 3.8J k ^–1 mol^–1 for mesobilirubin.