SPIN RELAXATION MEASUREMENTS USING FIRST-HARMONIC OUT-OF-PHASE ABSORPTION EPR SIGNALS

The dependence on spin-lattice (T-1) relaxation of the first-harmonic absorption EPR signal (V-1') detected in phase quadrature with the Zee man modulation has been investigated both theoretically and experiment ally for nitroxide spin labels. Spectral simulations were performed by iterative solution of the Bloch equations that contained explicitly b oth the modulation and microwave magnetic fields (T, Pali, V. A. Livsh its, and D. Marsh, 1996, J. Magn. Reson, B 113, 151-159), It was found that, of the various non-linear EPR displays, the first-harmonic out- of-phase V-1'-signal, recorded under conditions of partial saturation of the microwave absorption, is particularly favorable for determining spin-lattice relaxation enhancements because of its superior signal i ntensity and relative insensitivity to spin-spin (T-2) relaxation, By varying the Zeeman modulation frequency it is also possible to tune th e optimum sensitivity of the V-1'-signal to different ranges of the T- 1-relaxation time. A Zeeman modulation frequency of 25 kHz appears to be particularly suited to spin label applications. Calibrations are gi ven for the dependence on T-1-relaxation time of both the amplitude an d the second integral of the V-1'-signal recorded under standard condi tions. Experiments on different spin labels in solution and in membran es demonstrate the practical usable sensitivity of the V-1'-signal, ev en at modulation frequencies of 25 kHz, and these are used to investig ate the dependence on microwave field intensity, in comparison with th eoretical predictions. The practicable sensitivity to spin-lattice rel axation enhancements is demonstrated experimentally for a spin-labeled membrane system in the presence of paramagnetic ions. The first-harmo nic out-of-phase V-1'-signal appears to be the non-linear CW EPR metho d of choice for determining T-1-relaxation enhancements in spin-labele d systems. (C) 1998 Academic Press.