Small Hyperfine Coupling Research Articles

An orientation-selected ENDOR and HYSCORE study of the Ni-C active state of Desulfovibrio vulgaris Miyazaki F hydrogenase

Electron nuclear double resonance (ENDOR) and hyperfine sublevel correlation spectroscopy (HYSCORE) are applied to study the active site of catalytic [NiFe]-hydrogenase from Desulfovibrio vulgaris Miyazaki F in the reduced Ni-C state. These techniques offer a powerful tool for detecting nearby magnetic nuclei, including a metal-bound substrate hydrogen, and for mapping the spin density distribution of the unpaired electron at the active site. The observed hyperfine couplings are assigned via comparison with structural data from X-ray crystallography and knowledge of the complete g-tensor in the Ni-C state (Foerster et al. (2003) J Am Chem Soc 125:83-93). This is found to be in good agreement with density functional theory calculations. The two most strongly coupled protons (a(iso)=13.7, 11.8 MHz) are assigned to the beta-CH(2) protons of the nickel-coordinating cysteine 549, and a third proton (a(iso)=8.9 MHz) is assigned to a beta-CH(2) proton of cysteine 546. Using D(2)O exchange experiments, the presence of a hydride in the bridging position between the nickel and iron-recently been detected for a regulatory hydrogenase (Brecht et al. (2003) J Am Chem Soc 125:13075-13083)-is experimentally confirmed for the first time for catalytic hydrogenases. The hydride exhibits a small isotropic hyperfine coupling constant (a(iso)=-3.5 MHz) since it is bound to Ni in a direction perpendicular to the z-axis of the Ni (3d(z)(2)) orbital. Nitrogen signals that belong to the nitrogen N(epsilon) of His-88 have been identified. This residue forms a hydrogen bond with the spin-carrying Ni-coordinated sulfur of Cys-549. Comparison with other hydrogenases reveals that the active site is essentially the same in all proteins, including a regulatory hydrogenase.

Theoretical treatment of ion–molecular charge transfer reactions involving dimer radical ions

For the first time spin evolution of radical ions participating in ion–molecular charge transfer reactions involving dimer radical ion and neutral monomer molecule (dimer–monomer charge transfer reaction) has been studied quantitatively. The peculiarity of the problem is that only a fragment (monomer) of a radical ion is changed in the course of the reaction. For this case the kinetic equations of the encounter theory for spin density matrix of a radical ion have been obtained. Calculations of EPR spectra of radical ions in the presence of dimer–monomer charge transfer reaction have been performed. The cases of small and large hyperfine coupling constants, slow and fast charge transfer reactions have been studied in detail. The shifts and broadening of the EPR spectral lines in these cases have been calculated.

17O ENDOR detection of a solvent-derived Ni-(OH(x))-Fe bridge that is lost upon activation of the hydrogenase from Desulfovibrio gigas.

Crystallographic studies of the hydrogenases (Hases) from Desulfovibrio gigas (Dg) and Desulfovibrio vulgaris Miyazaki (DvM) have revealed heterodinuclear nickel-iron active centers in both enzymes. The structures, which represent the as-isolated (unready) Ni-A (S = (1)/(2)) enzyme state, disclose a nonprotein ligand (labeled as X) bridging the two metals. The bridging atom was suggested to be an oxygenic (O(2)(-) or OH(-)) species in Dg Hase and an inorganic sulfide in DvM Hase. To determine the nature and chemical characteristics of the Ni-X-Fe bridging ligand in Dg Hase, we have performed 35 GHz CW (17)O ENDOR measurements on the Ni-A form of the enzyme, exchanged into H(2)(17)O, on the active Ni-C (S = (1)/(2)) form prepared by H(2)-reduction of Ni-A in H(2)(17)O, and also on Ni-A formed by reoxidation of Ni-C in H(2)(17)O. In the native state of the protein (Ni-A), the bridging ligand does not exchange with the H(2)(17)O solvent. However, after a reduction/reoxidation cycle (Ni-A --> Ni-C --> Ni-A), an (17)O label is introduced at the active site, as seen by ENDOR. Detailed analysis of a 2-D field-frequency plot of ENDOR spectra taken across the EPR envelope of Ni-A((17)O) shows that the incorporated (17)O has a roughly axial hyperfine tensor, A((17)O) approximately [5, 7, 20] MHz, discloses its orientation relative to the g tensor, and also yields an estimate of the quadrupole tensor. The substantial isotropic component (a(iso)((17)O) approximately 11 MHz) of the hyperfine interaction indicates that a solvent-derived (17)O is indeed a ligand to Ni and thus that the bridging ligand X in the Ni-A state of Dg Hase is indeed an oxygenic (O(2)(-) or OH(-)) species; comparison with earlier EPR results by others indicates that the same holds for Ni-B. The small (57)Fe hyperfine coupling seen previously for Ni-A (A((57)Fe) approximately 0.9 MHz) is now shown to persist in Ni-C, A((57)Fe) approximately 0.8 MHz. However, the (17)O signal is lost upon reductive activation to the Ni-C state; reoxidation to Ni-A leads to the reappearance of the signal. Consideration of the electronic structure of the EPR-active states of the dinuclear center leads us to suggest that the oxygenic bridge in Ni-A(B) is lost in Ni-C and is re-formed from solvent upon reoxidation to Ni-A. This implies that the reductive activation to Ni-C opens Ni/Fe coordination sites which may play a central role in the enzyme's activity.

Dual-Mode EPR Detects the Initial Intermediate in Photoassembly of the Photosystem II Mn Cluster: The Influence of Amino Acid Residue 170 of the D1 Polypeptide on Mn Coordination

We report the first parallel polarization EPR signal from the Mn(III) ion formed by photooxidation of Mn(II) bound at the high affinity Mn-binding site of photosystem II (PSII). This species corresponds to the first photoactivation intermediate formed on the pathway to assembly of the water-splitting Mn cluster. The parallel mode EPR spectrum of the photooxidation product of 1.2/1 stoichiometry Mn(II)/Mn-depleted wild-type Synechocystis sp. PCC 6803 PSII particles consists of six well-resolved transitions split by a relatively small 55Mn hyperfine coupling (44 G). This spectral signature is absent in photooxidized Mn apoPSII complexes prepared from D1-Asp170Glu and D1-Asp170His mutants, providing direct spectral evidence for a role for this specific D1-Asp170 residue in the initial photoactivation chemistry. Temperature-dependence measurements and spectral simulations performed on the Mn(III) parallel mode EPR signal of the wild-type sample give an axial zero-field splitting value of D ≈ −2.5 cm-1 and a r...

Electronic Structure of Antiferromagnetically Coupled Dinuclear Manganese (MnIIIMnIV) Complexes Studied by Magnetic Resonance Techniques

The following dinuclear exchange-coupled manganese complexes are investigated: [dtneMnIIIMnIV(μ-O)2μ-OAc](BPh4)2 (dtne = 1,2-bis(1,4,7-triazacyclonon-1-yl)ethane), [(CH3)4dtneMnIIIMnIV(μ-O)2μ-OAc](BPh4)2 ((CH3)4dtne = 1,2-bis(4,7-dimethyl-1,4,7-triazacyclonon-1-yl)ethane), [(CH3)4dtneMnIVMnIV(μ-O)2μ-OAc](ClO4)3, [(tacn)2MnIIIMnIV(μ-O)2μ-OAc](BPh4)2 (tacn = 1,4,7-triazacyclononane), [bpy4MnIIIMnIV(μ-O)2](ClO4)3 (bpy = 2,2‘-bipyridyl), and [phen4MnIIIMnIV(μ-O)2](ClO4)3 (phen = 1,10-phenanthroline). For three of these complexes, X-ray structural data obtained on single crystals are reported here. All complexes are strongly antiferromagnetically coupled, with exchange coupling constants ranging from J = −110 cm-1 (bis-μ-oxo-μ-acetato-bridged) to −150 cm-1 (bis-μ-oxo-bridged). EPR investigations at X- and Q-band frequencies are reported for all five mixed-valence MnIIIMnIV complexes. G tensors and 55Mn hyperfine coupling constants (hfc's) were obtained by simultaneous simulation of the EPR spectra at both frequency bands. By using the vector model of exchange-coupled systems, tensor axes could be related to the molecular structure of the complexes. Hyperfine coupling constants from 55Mn cw-electron−nuclear double-resonance (ENDOR) spectra were in agreement with those obtained from the simulation of the EPR spectra. Ligand hyperfine couplings (1H and 14N) were also measured using cw-ENDOR spectroscopy. Electron spin−echo envelope modulation spectroscopy (ESEEM) spectra yielded information about small 14N hyperfine and quadrupole coupling constants that could not be resolved in the ENDOR spectra. On the basis of specifically deuterated complexes and results from orientation-selection ENDOR spectra, some proton hfc's could be assigned to positions within the complexes. Using an extended point-dipole model and the coordinates provided by the X-ray structure analysis, all dipolar hfc's of the complexes were calculated. Comparison of these hfc's with experimentally obtained values led to a consistent assignment of most hf tensors to molecular positions. The electronic structures of the investigated complexes are compared with each other, and the relevance of the results for metalloenzymes containing at least a dinuclear manganese core is discussed.

In Situ Radiolysis Steady-State ESR Study of Carboxyalkyl Radical Trapping by 5,5-Dimethyl-1-pyrroline-N-oxide: Spin Adduct Structure and Stability

Short-lived carboxyalkyl radicals formed in the reaction of three mono- and two dicarboxylic acids with radiolytically produced hydroxyl radicals or hydrated electrons were trapped successfully with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) in dilute aqueous solution. The in situ radiolysis steady-state ESR spectra of the spin adducts were analyzed to determine accurate ESR parameters for these spin adducts in a uniform environment. DMPO spin adducts of the five carboxyalkyl radicals were identified for the first time. Parent radicals include carboxymethyl, dicarboxymethyl, 1,2-dicarboxyethyl, 1-amino-1-carboxymethyl (glycine radical), and 2-carboxyethyl radicals. ESR parameters for DMPO spin adducts of carboxyalkyl radicals are rather similar with nitrogen hyperfine coupling constants of approximately 16 G and with g values around 2.0054 except for glycine radical. Proton hyperfine coupling constants due to C2−H vary between 21 and 25 G. Nitrogen and C2-proton hyperfine coupling constants are smaller than those of alkyl radical adducts with larger g values. These changes are ascribed to the spread of the π-conjugation system to include the carboxyl groups. Moreover, the ESR spectrum of both the carboxymethyl and the 1,2-dicarboxyethyl radical is a superposition of spectra from adduct radicals with protonated and deprotonated carboxyl groups in the spin addend. The protonated radical adducts have small additional proton hyperfine couplings due to these carboxyl protons. These carboxyalkyl radicals exhibit intramolecular hydrogen bonding between the hydroxyl function and the aminoxyl oxygen in aqueous solution, which has a profound effect upon the stability of their DMPO spin adducts against spin adduct rearrangement and fragmentation. From carboxymethyl and dicarboxymethyl−DMPO adducts, (CH3)2C(CH2CHCH2)N(O·)CH2CO2H and (CH3)2C(CH2CHCH2)N(O·)CH(CO2-)CO2H are proposed to form following the opening of the DMPO ring. A similar fragmentation reaction is proposed to occur from DMPO−dicarboxyhydroxymethyl adduct formed in γ-irradiated tartronic acid (hydroxymalonic acid) solution. DMPO spin adducts of glycine and 2-carboxyethyl radicals also show the small proton hyperfine splitting, revealing the intramolecular hydrogen bonding.

EPR and ENDOR Studies of X-Irradiated Single Crystals of Deoxycytidine 5‘-Phosphate Monohydrate at 10 and 77 K

The EPR spectra of single crystals of deoxycytidine 5‘-phosphate monohydrate (5‘dCMP), X-irradiated at 10 K, exhibit signals from several distinct radical species. Analysis of the ENDOR spectra from two of these radicals indicates that these result from oxidation and reduction of the cytosine base. The reduced species exhibits hyperfine coupling to the C6−Hα proton, and an additional small exchangeable hyperfine coupling from the N3−H proton. No additional couplings that may be associated with protonation of the amino group have been observed. Since the native molecule is protonated at N3, it appears that reduction of the cytosine base does not result in any further protonation. The oxidized species exhibits hyperfine couplings to C5−Hα and C1‘−Hβ, and two small exchangeable couplings from the C4−NH2 protons. Since no hyperfine coupling to N3−H was observed, the oxidized species is believed to be the N3−H deprotonated cation. At high X-ray dose there is also evidence for both C5 and C6 H-addition radicals...

Spectral, magnetic and electrochemical properties of metal oxa- and oxathia-porphyrins

Metal derivatives (CuII, NiII) of monooxa-, dioxa- and oxathia-tetraphenylporphyrins and their one-electron oxidised and reduced species have been studied. Electronic spectra of the monooxa and oxathia derivatives exhibit split Soret bands and a complicated pattern of Q-bands revealing their lower symmetry. The spectra of one-electron reduced species show only marginal shifts while the one-electron oxidised product of copper monooxaporphyrins show broad, red-shifted bands. Cyclic voltammetric studies indicated one-electron metal-centered reduction at fairly low potentials forming copper(I) and nickel(I) porphyrins. The ring-oxidised product exhibits weak antiferromagnetic interaction between the unpaired electrons of copper and the porphyrin ring. The ESR spectra of the copper dioxa- and oxathia-porphyrins exhibit rhombic symmetry with unusually small metal hyperfine couplings. A comparison of ACu values and the E½ values for metal reduction suggests that distortion towards tetrahedral symmetry and the presence of a soft donor atom like sulfur in porphyrins are required to generate spectral and electrochemical properties like those observed for the type I copper center in proteins.

Conformation of a Geminal Dicyclopropyl-Substituted Trimethylenemethane Triplet [2-(Dicyclopropylmethylidene)cyclopentane-1,3-diyl

Triplet 2-(dicyclopropylmethylidene)cyclopentane-1,3-diyl, CP2TMM, has a Deltam(s) = 2 ESR transition revealing hyperfine splitting consistent with coupling to two equivalent alpha hydrogens and five equivalent beta hydrogens and one other hydrogen with a small hyperfine coupling constant. This is consistent only with a conformation in which one cyclopropane ring conjugates with the pi triplet so that the other cyclopropane is twisted allowing the tertiary cyclopropyl hydrogen to hyperconjugate. Geometry optimization of the triplet state of CP2TMM was carried out using the 3-21G basis set with a UHF wave function and while the singlet state was optimized using a GVB perfect pairing wave function. The most stable conformation of each spin state is a C(2)-symmetric structure providing a 10 kcal/mol singlet-triplet energy gap favoring the triplet. This gap is reproduced at the 6-31G level using the 3-21G geometries. However, this structure cannot reproduce the hyperfine splitting observed; 1 kcal/mol higher in energy is an asymmetrical conformation of triplet CP2TMM which can account for the hyperfine interactions.

Muonium Adducts of the Nitro Group: Muoxynitroxides

Implantation of positive muons in liquid samples of nitroalkanes leads to radicals, detectable by the transverse field muon spin rotation (TF-μSR) technique, with small hyperfine couplings (ca. 26 MHz). The results indicate that muonium has added to one of the oxygen atoms of the nitro group and that the bound muon remains close to the ‘nodal’ region of the nitrogen 2pz orbital, which contributes to the SOMO. This is a consequence of π-bonding in the O—N—O(Mu) unit, which is maximally preserved. An abrupt change is found for the nitrobenzene adduct, in which the coupling is raised to 37.14 MHz. Semi-empirical (PM3) calculations provide an explanation for this in terms of the different geometries of RNO2H· and PhNO2H· radicals, the former being bent at the nitrogen centre while PhNO2H· is planar. When PhNO2H· is located in NaX zeolite, its coupling is increased; this is thought to be due to the electrostatic field from Na+ cations. © 1997 John Wiley & Sons, Ltd.

Local fields and nuclear spin-lattice relaxation of 27Al in the heavy-fermion system YbNiAl

We report measurements of the Knight-shift, the linewidth and the spin-lattice (T1−1 and spin-spin (T2−1 relaxation rates of 27Al in the heavy fermion system YbNiAl at temperatures 1.3–70 K and in external fields between 0.3 and 8 T. From T2−1 the endpoints of the critical line of the transition to the magnetically ordered state are TN = 2:6K and Bcr = 3:0T. In the paramagnetic regime we find a linear dependence of the Knight-shift and the linewidth on the susceptibility χ with a small mean hyperfine coupling (0:011mT/μB) but a large linewidth indicating the cancellation of stronger couplings. T1−1 is independent of T above 10 K, below it becomes strongly field and temperature dependent due to the approaching phase transition. While the high field behaviour can be described consistently with χ within a simple two band model this model predicts a strong field dependence which is incompatible with the experiment. This indicates that many body effects in the Kondo-lattice can be identified much better from the field than from the temperature dependence of the physical properties.

Low and zero field stimulated nuclear polarization in cyclic ketones

13C stimulated nuclear polarization (SNP) in acyl-alkyl biradicals has been investigated at a low magnetic field B 0. For carbons in the carbonyl position, which have a strong hyperfine interaction (hfi), a significant SNP effect is observed at low fields B 0 < hfi, in contrast to carbons in the α-position with smaller hyperfine coupling. A qualitative analysis using a two-state biradical model shows that this effect has a strong dependence onthe non-spin selective channels of the biradical decay.

Formate dehydrogenase from Desulfovibrio desulfuricans ATCC 27774: isolation and spectroscopic characterization of the active sites (heme, iron-sulfur centers and molybdenum)

An air-stable formate dehydrogenase, an enzyme that catalyzes the oxidation of formate to CO2, was purified from a sulfate-reducing organism, Desulfovibrio desulfuricans ATCC 27774. The enzyme has a molecular mass of approximately 150 kDa (three different subunits: 88, 29 and 16 kDa) and contains three types of redox-active centers: four c-type hemes, nonheme iron arranged as two [4Fe-4S]2+/1+ centers and a molybdenum-pterin site. Selenium was also chemically detected. The enzyme specific activity is 78 units per mg of protein. Mo(V) EPR signals were observed in the native, reduced and formate-reacted states. EPR signals related to the presence of multiple low-spin hemes were also observed in the oxidized state. Upon reduction, an examination of the EPR data under appropriate conditions distinguishes two types of iron-sulfur centers, an [Fe-S] center I (g max=2.050, g med=1.947, g min=1.896) and an [Fe-S] center II (g max=2.071, g med=1.926, g min=1.865). Mossbauer spectroscopy confirmed the presence of four hemes in the low-spin state. The presence of two [4Fe-4S]2+/1+ centers was confirmed, one of these displaying very small hyperfine coupling constants in the +1 oxidation state. The midpoint redox potentials of the enzyme metal centers were also estimated.

63,65Cu NMR investigation ofCuGeO3ssingle crystals:mThe uniform and the dimerized spin-Peierls phase

We report a Cu NMR study performed under high magnetic field on single crystals of the inorganic spin-Peierls ${\mathrm{CuGeO}}_{3}$ compound, in its uniform and dimerized phases. The temperature (T) dependence of the magnetic hyperfine shift, i.e., the local static spin susceptibility, is found to scale the macroscopic susceptibility. This allows the determination of the hyperfine coupling tensor, which can be well accounted for by the on-site coupling associated with the localized electronic spin of a ${\mathrm{Cu}}^{++}$ ion in an approximately axially symmetric environment. The nuclear spin-lattice relaxation rate (${\mathrm{T}}_{1}^{\mathrm{\ensuremath{-}}1g}$) in the dimerized phase is found to be activated for temperature below \ensuremath{\sim}${\mathrm{T}}_{\mathrm{SP}}$ /2. The magnetic-field dependence of the activation energy is in good agreement with that of the energy gap determined by neutron inelastic scattering. In the uniform phase, neither the magnitude of ${\mathrm{T}}_{1}^{\mathrm{\ensuremath{-}}1g}$ (an order of magnitude smaller) nor its T dependence (approximately linear in T instead of being nearly constant) correspond to the theoretical predictions for a simple S=1/2, one-dimensional Heisenberg antiferromagnet, unless we admit for a filtering of the antiferromagnetic fluctuations, due to a small supertransferred hyperfine coupling. As expected, we found no magnetic-field dependence of ${\mathrm{T}}_{1}$ (T\ensuremath{\gtrsim}1.5 ${\mathrm{T}}_{\mathrm{SP}}$ ) in the range 8.9--14.9 T.

A Combined EPR and Quantum Chemical Approach to the Structure of Surface Fs+(H) Centers on MgO

Fs+(H) color centers at the surface of MgO have been studied using a combined EPR and quantum chemical approach. Fs+(H) are paramagnetic excess electrons centers where the unpaired electron is trapped in a surface anion vacancy. They are formed at the surface of thoroughly dehydrated MgO (1073K) upon UV irradiation under hydrogen in parallel with the formation of minor fractions of different color centers. The whole EPR spectrum resulting from irradiation has been analyzed by simulation of the experimental profile. Fs+(H) centers are characterized by an axial g tensor and display a hyperfine interaction with a hydrogen nucleus (belonging to an hydroxyl group stabilized nearby the vacancy) and two distinct families of 25Mg nuclei characterized by a large (10.5 G) and a small (0.7 G) hyperfine coupling constant, respectively. Both EPR and ab initio calculations on clusters of ions converge in indicating that the features of the centers are due to the polarization of the electron density by the positive charge of the hydrogen in the OH group toward two (or possibly three) equivalent magnesium ions of the vacancy close to the OH group itself. The formation mechanism of the centers is strictly analogous to that occurring upon contact of low ionization energy metals with the surface of MgO leading to the formation of another type of color centers. Partially hydrated MgO samples also give rise to another family of paramagnetic center based on electrons trapped in anion vacancies. This finding indicates that the structure of the partially hydroxylated oxide and the mechanism of its dehydration are still open questions.

14N Electron Spin Echo Envelope Modulation (ESEEM) Spectroscopy of the Cation Radical P840+, the Primary Electron Donor of the Chlorobium limicola Reaction Center

AbstractThe electronic structure of the oxidized primary chlorophyll electron donor, P840+., of the green sulfur bacterium Chlorobium limicola has been investigated using electron spin echo envelope modulation (ESEEM) spectroscopy. This ESEEM investigation of the electron spin density distribution in the radical cation P840+. in membranes isolated from C. limicola confirms that the electron spin is shared eqully between the two bacteriochlorophyll a molecules. Observation of the small hyperfine couplings to the ring nitrogens by ESEEM gives results that are in agreement with those obtained from ENDOR measurements (S. E. J. Rigby, R. Thapar, M. C. W. Evans and P. Heathcote, FEBS Lett. 350,24–28, 1994) of the large hyperfine couplings to the methyl group protons. These results in combination with the Raman spectroscopy of P840 (U. Feiler, D. Albouy, B. Robert and T. A. Mattioli, Biochemistry 34,11099–11105, 1995) all indicate that the reaction center of green sulfur photosynthetic bacteria is functionally a protein homodimer providing a symmetrical protein environment for the primary electron donor.

ESR study on the structure of the ethylene radical cation stabilized in frozen SF6 and some fluoroalkanes: An evidence for a twisted structure

ESR spectra of the ethylene radical cation were detected at cryogenic temperatures in SF6, C2F6 and C3F8. From the unusually small hyperfine couplings estimated for1H and13C, it has been shown that the ethylene radical cation has a non-planar structure with a torsional angle in the range of 8°–23°. Upon annealing the sample at a temperature above 93 K, the ethylene radical cation in SF6 changed into a monofluoroethyl radical through charge recombination with fluoride anion or SF 6 − .

Electronic Structure of Neutral Tryptophan Radicals in Ribonucleotide Reductase Studied by EPR and ENDOR Spectroscopy

Two different tryptophan radicals (Wa• and Wb•) with lifetimes of several minutes at room temperature are formed during the reconstitution of the diiron center in the Escherichia coli ribonucleotide reductase mutant protein R2 Y122F. Detailed hyperfine parameters are for the first time determined for protein-linked oxidized neutral tryptophan radicals. Wa• is freeze-trapped and investigated by EPR and ENDOR in protonated and selectively deuterated proteins at 20 K. Two hyperfine couplings from the β-methylene protons, hyperfine tensors of two α-protons, and the complete nitrogen hyperfine tensor are determined. Based on the absence of a large hyperfine coupling from the N−H proton, which would be expected for a cation radical, and on comparison of the experimental data with theoretical spin densities from density functional calculations, Wa• is assigned to an oxidized neutral tryptophan radical. A small anisotropic hyperfine coupling detected in selectively deuterated Wa• is tentatively assigned to a prot...

Coherence Transfer by Selective Pulses in Photo-CIDNP Experiments

Identification of the products of a photoreaction by measurements of chemically induced dynamic nuclear polarization can be difficult, since often many of their allowed NMR transitions are unobservable with this technique due to small hyperfine coupling constants of the respective nuclei in the intermediate radicals. In this work, coherence-transfer methods are described that solve this problem for J-coupled spin systems containing at least one polarized nucleus. The limitations of the traditional two-dimensional NMR methods (a large number of acquisitions, which is not feasible in reacting systems) are overcome by the use of selective excitation pulses and pseudo-steady-state measurements. In this way, transfer spectra could be obtained with the minimum number of scans determined by the respective phase cycle (8 or 16). Correlation spectroscopy (1D COSY) and relayed coherence-transfer (ID RCT) experiments were performed with the photosystem 9,1 0-anthraquinone and triallyl amine. With the methods described, it was possible fully to determine the structures of the two unstable products formed by the reaction and to obtain valuable information about the mechanism and intermediates.

Metal atoms and clusters in fullerene cages

Fullerenes containing metal atoms and clusters can be formed by the arc-vaporization method. The electronic structure of these metallofullerenes can be probed using magnetic resonance techniques. Electron paramagnetic resonance (EPR) spectra of LaC82, YC82, ScC82 and Sc3C82 have been obtained. Metallofullerenes containing a single metal atom (MC82 with M = La, Y, or Sc), have small hyperfine couplings and g-values close to 2, implying that they can be described as + 3 metal cations within — 3 fullerene radical anion cages. In the La and Y cases, additional EPR active MC82 species have been found. The EPR spectrum of Sc3C82 shows that the metal atoms are equivalent, suggesting that they may form a triangular molecule. No EPR spectrum is found for Y2C82 or for Sc2C2n species (with 2n = 82,84,86), suggesting that they are diamagnetic. Sc NMR spectra of a solution containing Sc2C2n species have been obtained which confirm the diamagnetism of the discandium metallofullerenes.