ABSTRACT:The fluorescence and absorption properties of a series of reduced flavoproteins have been measured and compared with the properties of suitable model compounds. Contrary to common belief, a number of reduced flavoproteins have been found to exhibit appreciable fluorescence emission with maxima in the range 500-530 nm. In keeping with common observation, the reduced model flavines are devoid of fluorescence in solution at room temperature, but show marked fluorescence emission in the range 476-512 nm at 77 OK in rigid glasses. The fluorescence quantum yield of reduced lactate oxidase (emission x , , , 507 nm) is increased 4.7 times upon formation of covalent N5 adducts and the emission maximum is shifted to 476 nm. In the case of the nonfluores-T he electronic properties of the isoalloxazine chromophore system, which constitutes the redox active moiety of the flavine coenzymes, have been the object of several thorough theoretical and experimental investigations (Sun et al., 1972 ;Song, 1971, and literature cited therein). These studies have dealt mainly with the oxidized form of the free and proteinbound flavine as well as with model compounds. A distinctive characteristic of oxidized free flavines is their relatively strong fluorescence (quantum yield -0.3; Sun et al., 1972), with an emission maximum typically around 520 nm. The energy and intensity of this emission are dependent on solvent polarity and temperature (Koziol, 1969; Sun el al., 1972;Kotaki et al., 1967), on formation of complexes with a variety of molecules (Weber, 1950; Slikin, 1971), and on the position and properties of substituents Walker et al., 1972; Sun et al., 1972). Similar effects have been observed with riboflavine, FMN, or FAD1 bound to various apoproteins.Unfortunately the spectroscopy of reduced flavines has received far less attention than that of the oxidized flavoquinones. A reason for this could be the absence of wellresolved structure in the near-ultraviolet absorption spectrum and of fluorescence at ambient temperature in any solvent.On the other hand, the nonplanarity of the flavine ring in its reduced forms introduces difficulties in theoretical calculations

Disulfide bond reduction by the CO2.- radical was investigated in aponeocarzinostatin, aporiboflavin-binding protein, and bovine immunoglobulin. Protein-bound cysteine free thiols were formed under gamma-ray irradiation in the course of a pH-dependent and protein concentration dependent chain reaction. The chain efficiency increased upon acidification of the medium, with an apparent pKa around 5, and decreased abruptly below pH 3.6. It decreased also at neutral pH as cysteine accumulated. From pulse radiolysis analysis, CO2.- proved able to induce rapid one-electron oxidation of thiols and of tyrosine phenolic groups in addition to one-electron donation to exposed disulfide bonds. The bulk rate constant of CO2.- uptake by the native proteins was 5- to 10-fold faster at pH 3 than at pH 8, and the protonated form of the disulfide radical anion, [symbol: see text], appeared to be the major protein radical species formed under acidic conditions. The main decay path of [symbol: see text] consisted of the rapid formation of a thiyl radical intermediate [symbol: see text] in equilibrium with the closed, cyclic form. The thiyl radical was subsequently reduced to the sulfhydryl level [symbol: see text] on reaction with formate, generating 1 mol of the CO2.- radical, thus propagating the chain reaction. The disulfide radical anion [symbol: see text] at pH 8 decayed through competing intramolecular and/or intermolecular routes including disproportionation, protein-protein cross-linking, electron transfer with tyrosine residues, and reaction with sulfhydryl groups in prereduced systems. Disproportionation and cross-linking were observed with the riboflavin-binding protein solely. Formation of the disulfide radical cation [symbol: see text], phenoxyl radical Tyr-O. disproportionation, and phenoxyl radical induced oxidation of preformed thiol groups should also be taken into consideration to explain the fate of the oxygen-centered phenoxyl radical.

The temperature dependence of the paramagnetic shifts in square planar ferrous complexes has been investigated for a number of porphyrin derivatives. Large anomalies in the Curie law have been observed in newly synthesized substituted tetraphenylporphins in which the environment of both faces of the tetrapyrrolic ring is strictly controlled. The pseudocontact and contact contributions to the measured hyperfine shifts as well as their temperature dependence have been interpreted in terms of a model with two electronic states. Following this model, the nature of the ground state of the ferrous ion depends critically upon the energy of the dz2 orbital relative to that of the degenerate dxz and dyz orbitals. For most of the porphyrin derivatives, the ground state is 3A2g, strongly perturbed by the closely lying 3Eg excited state. This strong mixing by spin–orbit coupling explains the large orbital contribution to the magnetic susceptibility of these complexes. A small axial perturbation can induce a reversal of the ground state with corresponding inversion of the magnetic anisotropy. Such a behavior, predicted by the model, is observed at low temperature by proton NMR spectroscopy of some complexes in solution.

Zopiclone and flunitrazepam had residual effects in the first part of the morning, whereas zolpidem had no residual effects. The hierarchical character of the effects of the molecules differed according to the test administered. This is probably linked more to drug-induced specific alterations than to different sensitivities of the tests.