Zbigniew Gasyna

Intermediate spin-states in one-electron reduction of oxygen-hemoprotein complexes at low temperature.

The oxy-heme unit of oxygen-hemoprotein complexes contains four oxidizing equivalents in excess of the ferrous state. Oneelectronequivalent reduction of the oxy-heme center would therefore convert it formally to the oxidation state of t5. This oxidation state is known to represent the catalytic intermediate compound I of peroxidase enzymes [ 1,2]. However, no such intermediate compound has been found in the case of the oxygen storage and transport hemoproteins such as myoglobin and hemoglobin. These results seem to be compatible with the difference in catalytic specificity and reactivity of oxy-heme complexes of heme proteins and enzymes. Various theoretical and spectroscopic studies [3-71 have been recently undertaken to define stereochemical origins of the oxy-heme reactivity. The potential application of low-temperature radiolysis method [8,9] to produce oneelectron-addition compounds of the oxygen-hemoprotein complexes in glassy matrices for spectroscopic analysis is demonstrated in the present work. The data show the differences in the pathway of oneelectron reduction of oxyperoxidase and oxymyoglobin, which can be correlated with basic functional properties of the hemoproteins.

Luminescence decay from copper(I) complexes of metallothionein

Abstract Luminescence in the 600 nm region from aqueous solutions of Zn 7 metallothionein (ZnMT) has been studied as a function of the extent of substitution of the ZnMT with Cu + . The steady state emission intensity shows a strong correlation with cluster formation. Analysis of the emission decay curves obtained by single photon-counting techniques using a multiexponential decay function, indicates that three components are responsible for the observed emission intensity. A short lived component exhibits a lifetime of 20 ns, while the two longer lived components exhibit lifetimes in the 1 μ range at room temperature. The longer-lived emission is assigned to the excited triplet states of the CuS chromophore, most probably the metal-centered 3d 9 4s 1 .

A luminescence probe for metallothionein in liver tissue: emission intensity measured directly from copper metallothionein induced in rat liver

We report the first use of an emission probe based on the Cu(I)‐thiolate chromophore, for the direct observation of copper metallothionein located in samples of rat liver. Elevated synthesis of Cu‐MT in the rat liver was induced by subcutaneous injections of a series of aqueous CuCl2 solutions containing increasing amounts of Cu(II). Luminescence intensity in the 600 nm region, detected from frozen solutions of Cu‐MT and from slices of the liver frozen at 77 K, following excitation in the 300 nm region, was dependent on the concentration of the Cu(II) used in the inducing solution. No such luminescence intensity was found for control samples obtained from the livers of rats not exposed to copper salts. It is suggested that this new method will allow direct visualization of Cu‐MT in tissue where genetic disorders impare copper metabolism.

One-electron photooxidation of porphyrins at low temperature

Abstract The π-cation radicals of the metalloporphyrins magnesium octaethylporphyrin (MgOEP), magnesium tetraphenylporphyrin (MgTPP), and zinc tetraphenylporphyrin (ZnTPP), as well as the free base porphyrins of tetratolylporphyrin (H 2 TTP) and tetraphenylporphyrin (H 2 TPP) have been formed at liquid nitrogen temperatures in a rigid matrix of alkyl chloride glasses containing CCl 4 or 1,1,2,2-tetrachloroethane (TCE), following photolysis of the porphyrins with visible light. The reaction proceeds via electron transfer from the photoexcited porphyrin to the solvent molecules; the efficiency of thie electron transfer may be qualitatively evaluated in terms of electron tunneling in the solid matrices. This is the first report of the photochemical formation of a free base porphyrin π-cation radical species.

Luminescent metallothioneins: Emission properties of copper, silver, gold and platinum complexes of MT

Abstract Observations of luminescence from complexes of the metals Cu(I), Ag(I), Au(I), Pt(II), and cisplatin and the protein metallothionein, are reported. In each case, emission intensity is observed in the 500—600 nm region. The luminescence properties of these metal complexes can be interpreted in terms of the presence of similar, lowe1st energy, d → s and d → p excited states, that are metal-centered states in origin. These data show that the emission spectrum provides a new probe to study metal binding to metallothionein.

Microcomputer-aided chemistry: 5. Interactive computing for instrument control and data analysis in photochemical studies

Gasyna, Z., Browett, W.R., Nyokong, T., Kitchenham, R. and Stillman, M.J., 1989. Microcomputer-aided chemistry. 5. Interactive computing for instrument control and data analysis in photochemical studies. Chemometrics and Intelligent Laboratory Systems, 5: 233–246. The implementation of an integrated package of scientific software for use in a spectroscopic research laboratory is described for the situation where several different experiments are carried out in each study, yet the data have to be assembled for analysis and presentation on a single computer. The software currently operates on a series of IBM 9001 computers and offers the user capabilities for data acquisition, data analysis, data management, and data presentation. Use of multitasking techniques to achieve real-time, interactive control of a spectrometer is illustrated by a description of the design of the program CDSCAN5. The program ELECTRA is used to acquire and display data from the PAR 273 electrochemical instrument. Two programs provide extensive data analysis capabilities. (i) SIMPFIT, which is based on a versatile Simplex algorithm, is used to perform deconvolution of spectral envelopes of both absorption and magnetic circular dichroism data. (ii) DECAYFIT, which employs fast Fourier transform procedures, is used to deconvolute the complex luminescence decay profiles obtained in the single-photon counting experiments and to analyse kinetic data from a variety of sources. The spectral data base management program, SPECTRA MANAGER, is used to produce publication quality diagrams.

Optical absorption and magnetic circular dichroism studies of hydrogen, copper(II), zinc(II), nickel(II), and cobalt(II) crown ether-substituted monomeric and dimeric phthalocyanines

Monomeric and dimeric species of phthalocyanines with four, 15-crown-5-ether (1,4,7,10,13-pentaoxacyclopentadecane) voids at the 2,3 positions (ML)(where M = H2, Cu, Zn, Ni, or Co), have been studied using optical absorption and magnetic circular dichroism (m.c.d.) spectroscopies. The ready solubility of these crown ether-substituted complexes in chloroform allows the measurement of spectral data over a wide wavelength range. In particular, absorption and m.c.d. data are reported for H2L between 250 and 750 nm. The contribution by individual bands to the absorption and m.c.d. spectral envelopes measured for the monomeric and dimeric species of H2L and NiL, has been determined using deconvolution calculations carried out on pairs of related absorption and m.c.d. spectra. M.c.d. A terms have been used to identify the O, B1, B2, and Nπ→π* degenerate transitions in the spectra of the monomer of NiL, at 14 946 cm–1(669 nm) in the visible region, and at 24 398 (410), 31 414 (318), and 36 286 cm–1(276 nm) in the u.v. region. The deconvolution calculations were also used to find the pairs of bands that arise from the exciton splittings in the Q and B bands of the cofacial dimers. In the spectra of the dimer of NiL, two degenerate bands are required to fit the observed envelope in the Q-band region [at 14 887 (672) and 15 832 cm–1(632 nm)], which indicates that the splitting is ca. 950 cm–1, while the pair of B bands are separated by ca. 2 180 cm–1. The deconvolution of the spectra of the H2L complex gives a value of 1 925 cm–1 for the separation of the two components in the B band region.

Luminescent Ag12-metallothionein: Dependence of emission intensity on silver-thiolate cluster formation

We report the observation of emission intensity at 77 K that is a function of Ag(I)‐thiolate bonds formation within the protein metallothionein. The emission characteristics (a large, 250 nm, Stokes shift and long emission lifetime) suggests that the transition occurs from the excited triplet state. The emission intensity and circular dichroism both indicate that silver(I) clusters form with stoichiometric ratios of 12 Ag(I) to the 20 thiolate sulfur groups that are present in the protein. These data are the first to show that Ag(I)‐metallothionein complexes are luminescent and that a specific Ag12‐MT species forms.

Primary electron-accepting sites and electron transport reaction in human ceruloplasmin: Low-temperature radiolysis study

The reaction of ceruloplasmin with a reducing substrate is an important primary step in the fourelectron reduction of molecular oxygen to water catalysed by the enzyme. The functional properties and the interaction of different copper sites of ceruloplasmin in this reaction have been intensively investigated and the mechanism of electron transfer in the enzyme~atalysed reactions has been rationalized to some extent [l--5]. Specifically, the pathway of reduction equivalent to Type I Cu2+ in ceruloplasmin has been delineated for reduction in solution by hydrated electron using the method of pulse radiolysis combined with a fast recording spectrophotometric system 161. In the present work, the reaction of ‘dry’ electrons, which are the simplest reagent in redox reactions, has been studied at low temperature. The electrons can be produced by high-energy irradiation of aqueous organic solution frozen to a glassy solid at 77 K. It has been shown [7,8] that solutes in such a matrix capture electrons produced upon irradiation yielding the primary electron addition compounds. The spectral properties of the low-temperature products of ceruloplasmin and their thermal stability investigated by absorption and EPR spectroscopies have been used to identify the primary electron accepting sites and intramolecular electron transport reactions. 2. Materials and methods

The temperature dependence of the MCD spectrum of horseradish peroxidase compound I

The magnetic circular dichroism spectrum of the compound I species of horseradish peroxidase, which contains an iron (IV) porphyrin pi-cation radical complex, has been measured between 273 K and 4.2 K. The spectrum is temperature independent between 273 K and 30 K. However, very strong temperature dependence is observed below 30 K. These data do not appear to fit the temperature dependence expected for the presence of a simple MCD C term, or combination of C terms, but suggest that an increase in the coupling between the S = 1 iron (IV), and the S = 1/2 porphyrin pi-cation radical occurs forming a degenerate ground state. This increase in coupling below 30 K may be the result of a phase change in the protein which in turn affects the electronic structure of the heme group.