Optically detected magnetic resonance phosphorescence

J. Zuclich, D. Schweitzer, and A. H. Maki, Optically detected magnetic resonance of the tryptophan phosphorescent state in proteins, Photochem. Photobiol. 18, 161-168 (1973). 

K. L. Bell and H. C. Brenner, Phosphorescence and optically detected magnetic resonance study of the tryptophan residue in human serum albumin, Biochemistry 21, 799-804 (1982). 

W. C. Galley, Heterogeneity in protein emission spectra, in Concepts of Biochemical Fluorescence Vol. 2 (R. F. Chen and H. Edelhoch, eds.), pp. 409-439, Marcel Dekker, New York (1976).32. S.-Y. Mao and A. H. Maki, Comparative phosphorescence and optically detected magnetic resonance studies of fatty acid binding to serum albumin, Biochemistry 26, 3576-3582 (1987). 

M. V. Hershberger, A. H. Maki, and W. C. Galley, Phosphorescence and optically detected magnetic resonance studies of a class of anomalous tryptophan residues in globular proteins, Biochemistry 19, 2204—2209 (1980). 

In this section, results of three different experimental methods that have been apphed to Pd(2-thpy)2 are reported. Information from optically detected magnetic resonance (ODMR spectroscopy), microwave-recovery measurements, and phosphorescence microwave double resonance (PMDR spectroscopy) is presented. These methods complement each other to a large extent. The discussion presented here can be fimited to the basic impfications of the methods, since a comprehensive review by Max Glasbeek concerning these aspects is found in Volume 213 of this series [90] and a detailed report by Glasbeek, Yersin et al. [61] concerning Pd(2-thpy)2 has only recently been published. 

TTie porphyrin ZnP in a crystalline n-octane matrix at 1.2 K was investigated by optically detected magnetic resonance (ODM) and microwave induced delayed phosphorescence (MIDP) . ... 

Reports on chlorophyll fluorescence,404-408 phosphorescence,409 and photochemiluminescence 410 411 have been published. The properties of the triplet state of chlorophyll have been studied by e.s.r.412-416 and by optically detected magnetic resonance.416 417... 

Rousslang and co-workers (1978, 1979) and Ross and co-workers (1980) used optically detected magnetic resonance (ODMR) of tryptophan triplet states to detect conformational variations in proteins and polypeptides. This method used in combination with phosphorescence appears to be an excellent probe for detection of conformational variations which affect the environment of tryptophan. 

Double-resonance Spectroscopy.—A review has been given of double-resonance methods in spectroscopy.378 Attention will be focused here on optically (usually phosphorescence) detected magnetic resonance experiments (ODMR). Microwave-optical double-resonance experiments have been carried out on the spectrum of gaseous N02,379 permitting assignment of the rotational = 0—4 side-bands of the 493 nm band. 

The new techniques of phosphorescence-microwave multiplet resonance spectroscopy with optical detection have been reviewed by El-Sayed and Kwiram Such exciting experiments as the optical detection on electron-nuclear double resonance (ENDOR) and of electron-electron double resonance (EEDOR) in zero magnetic field have been achieved, and it is certain that much detailed knowledge concerning the phosphorescent states will evolve from this field. 

PMDR distinguishes this field from the general field of optical microwave double resonance (OMDR) by being confined to the studies of the magnetic, radiative, nonradiative, and structural properties of the lowest triplet state of molecules that exhibit phosphorescence radiation. Titles such as optical detection of ESR transitions are not correct since the interest in this field goes beyond the magnetic properties of the triplet state. 

UV spectra were obtained with a Varian spectrometer (Cary 15 and 17). Fluorescence, phosphorescence spectra, and the zero-field splitting parameters D and E of the triplet state were determined at 1.3K with an apparatus (31) for optical detection of magnetic resonance (ODMR) which was similar to the one described by Zuclich et al. (32). 

From magnetic resonance spectroscopy [49] it is well-known that IB effects are adequately circumvented by the tricks of a spin echo experiment. For instance, in a two-pulse echo experiment, IB effects are averaged out and one probes spin dephasing determined by time-dependent fluctuations characteristic of HB only (and not IB). More specifically, a nll-r-n microwave pulse sequence is applied, where the first nil pulse creates a coherent superposition state for which a la = 1 and the n pulse, applied at time r after the first pulse, generates a spin coherence (the echo) at time 2r after the initial pulse. The echo amplitude is traced with r. The echo amplitude decay time is characteristic of the pure dephasing dynamics. For phosphorescent triplet states it is possible to make the echo optically detectable by means of a final nil probe pulse applied at time f after the second pulse [44]. In Fig. 3b, the optically detected echo amplitude decay for the zero-field transition at 2320 MHz of... 

Slow-passage ODMR signals frequently are observed by the continuous wave method in which the optical effect is monitored using broadband detection. On the other hand, if the triplet state decay constants are sufficiently large, the microwave power may be amplitude modulated at an audio frequency which results in modulated phosphorescence when the microwave frequency is at resonance. The phosphorescence is then monitored with narrow-band phase-sensitive detection, for a great improvement in the signal/noise ratio. The latter detection method is frequently used to produce a magnetic resonance-induced phosphorescence spectrum by a technique referred to as phosphorescence-microwave double resonance (PMDR). The microwave frequency is fixed at resonance,... 

The differences in the population and depopulation rate constants and the phosphorescence probabilities of the three components of the triplet states form the basis of all the methods for Optical Detection of Magnetic Resonance in triplet states of jr-electron systems. These methods were developed after the discovery of optical spin polarisation and extended to inorganic solids. The essential physical difference from the optical double resonance in atoms developed by Alfred Kastler is to be found in the selection mechanism in optical double resonance, the polarisation of the resonant UV light, i.e. the symmetry of an applied field, is responsible for the selection. In optical spin polarisation, the selection is due to the spin-orbit coupling, and thus to an internal field. 

To our knowledge the first appheation of ODMR to a molecule of biological importance was made by Kwiram who in 1970 reported the optical detection of magnetic resonance signals from the tryptophan moiety of lysozyme. A more recent report dealt with frozen glassy solutions of tryptophan, tyrosine, and the protein, bovine serum albumin (BSA) loob) See Fig. 10. Discrepancies between the lifetimes involved in the phosphorescence decay and the fast-passage ODMR responses... 

If the separation among the sublevels is in the range of microwave frequency, sublevel properties can be obtained by observing the effect of microwave resonance on the emission from this state. The zero-field splitting is of the order of microwave frequency for most of rr/r states. Thus, the sublevel properties can be obtained by analyzing the effect of microwave resonance on the phosphorescence intensity. The method is called phosphorescence-microwave double resonance (PMDR) or optical detection of magnetic resonance (ODMR). 

Spin selective information on the lowest triplet state decay was obtained by optical detection of magnetic resonance transitions between the spin components of the T state of FBP in n-octane. Because of the absence of phosphorescence at these conditions, the ODMR signals were detected via changes in the 5i->5o fluorescent intensity [6, 29], Our calculations reproduce the fluorescent frequency and radiative constant rather well. In order to complete the interpretation of the microwave-induced fluorescent ODMR measurements [6, 29] one has to calculate the zero-field splitting in the T state and hyperflne coupling between electron and nuclear spins. 

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