ORIGIN OF THE TEMPERATURE-DEPENDENT ISOTROPIC HYPERFINE COUPLING OF THE VINYLIC PROTON OF OXYPYRROLINYL NITROXYL SPIN-LABELS

Proton ENDOR spectra of the spin-label 2,2,5,5-tetramethyl-1-oxypyrrol ine-3-carboxylic acid in methanol, chloroform/toluene, and toluene exh ibited well resolved features, permitting accurate estimation of hyper fine coupling (hfc) components of the vinylic and methyl protons. The magnitude and relative sign of the isotropic hfc A(iso) for these two classes of protons were determined by electron nuclear double resonanc e (ENDOR) and TRIPLE spectroscopy. While the isotropic hfc of methyl p rotons was temperature independent, A(iso) of the vinylic proton was t emperature dependent in fluid and frozen solutions. The temperature de pendence was not influenced by solvent polarity, but at T less than or equal to 50 K, A(iso) of the vinylic proton became temperature indepe ndent. The temperature dependence is explained on the basis of the out -of-plane vibration of the vinylic proton. Above 50 K, the out-of-plan e vibration of the C-H-V bond causes the vinylic proton to become disp laced from the nodal plane. The ratio of the measured A(iso) values of the vinylic proton and deuteron indicated that the vibrational mechan ism is responsible for the temperature dependent isotropic hfc of the vinyl proton of the oxypyrrolinyl nitroxyl spin-labels.