Magnetic field dependence of photo-CIDNP MAS NMR on photosynthetic reaction centers of Rhodobacter sphaeroides WT

Photochemically induced dynamic nuclear polarization (photo-CIDNP) is observed in frozen and quinone depleted photosynthetic reaction centers of the purple bacteria Rhodobacter sphaeroides wild type (WT) by (13)C solid-state NMR at three different magnetic fields. All light-induced signals appear to be emissive at all three fields. At 4.7 T (200 MHz proton frequency), the strongest enhancement of NMR signals is observed, which is more than 10 000 above the Boltzmann polarization. At higher fields, the enhancement factor decreases. At 17.6 T, the enhancement factor is about 60. The field dependence of the enhancement appears to be the same for all nuclei. The observed field dependence is in line with simulations that assume two competing mechanisms of polarization transfer from electrons to nuclei, three-spin mixing (TSM) and differential decay (DD). These simulations indicate a ratio of the electron spin density on the special pair cofactors is 3:2 in favor of the L-BChl during the radical cation state. The good agreement of simulations with the experiments raises expectations that artificial solid reaction centers can be tuned to show photo-CIDNP in the near future.     

Multichannel flash spectroscopy of the reaction centers of wild-type and mutant Rhodobacter sphaeroides: bacteriochlorophyllB-mediated interaction between the carotenoid triplet and the special pair

Multichannel flash spectroscopy (with microsecond time resolution) has been applied to carotenoid (Car)-containing and Car-less reaction centers (RC) of Rhodobacter sphaeroides with a view to investigate the interaction between the Car and its neighboring pigments at room temperature. Under neutral redox potential conditions, where the primary quinone acceptor (QA) is oxidized, the light-induced spectral changes in the 350-1000 nm region are attributed to the photochemical oxidation of the special pair (denoted here as P870), the generation of P870(+)QA(-), and the attendant electrochromism of adjacent chromophores. A bathochromic shift of <1 nm in the visible absorption region of Car reveals the sensitivity of Car to the P870 photooxidation. Under low redox potential conditions, where QA is reduced, P870 triplets (P870(+)) are formed. The time-resolved triplet-minus-singlet (TmS) spectrum of Car-less RC shows a deep bleaching at 870 nm, which belongs to P870(+), and additional (but smaller) bleaching at 800 nm; the entire spectrum decays at the same rate (with a lifetime of about 50 micros). The bleaching at 800 nm arises from the pigment interaction between P870(+) and the accessory bacteriochlorophylls on A and B branches (BA,B). In Car-containing RC, the TmS spectra of Car are accompanied by two smaller, negative signals--a sharp peak at 809 +/- 2 nm and a broad band at 870 nm--which decay at the same rate as the TmS spectrum of Car (ca 10 micros). The former is ascribed to the perturbation, by Car(+), of the absorption spectrum of BB; the latter, to the TmS spectrum of P870(+), a species that appears to be in approximate thermal equilibrium with Car(+). These assignments are consistent with the absorption-detected magnetic resonance spectra obtained by other workers at low temperatures.     

The spin mixing process of a radical pair in low magnetic field observed by transient absorption detected nanosecond pulsed magnetic field effect

The spin mixing process of the radical pair in the sodium dodecyl sulfate (SDS) micelle is studied by using a novel technique nanosecond pulsed magnetic field effect on transient absorption. We have developed the equipment for a nanosecond pulsed magnetic field and observed its effect on the radical pair reaction. A decrease of the free radical yield by a reversely directed pulsed magnetic field that cancels static field is observed, and the dependence on its magnitude, which is called pulsed MARY (magnetic field effect on reaction yield) spectra, is studied. The observed spectra reflect the spin mixing in 50-200 ns and show clear time evolution. Theoretical simulation of pulsed MARY spectra based on a single site modified Liouville equation indicates that the fast spin dephasing processes induced by the modulation of electron-electron spin interaction by molecular reencounter affect to the coherent spin mixing by a hyperfine interaction in a low magnetic field.     

Absorbance-detected electron spin echo spectroscopy of non-radiative triplet states in zero field : An application to the primary donor of the photosynthetic bacterium rhodobacter sphaeroides R-26

Electron spin echoes of triplet states in zero field were optically detected using the singlet ground-state absorbance. The technique has been applied to reaction centers of the photosynthetic bacterium Rhodobacter sphaeroides R-26. The spin-spin relaxation time T₂ of the triplet state of the primary electron donor P (a bacteriochlorophyll dimer) was 1.16±0.05 μs, independent of temperature in the range 1.2–2.1 K.     

Electron spin polarization in the photoexcited triplet state of porphyrins

Electron spin polarization in the photoexcited triplet state of tetraphenyl porphyrin was detected at 100°K using EPR technique. The zero field splitting parameters |D| and |E| the free base porphyrin were found to be 0.0369 ± 0.0005 and 0.0082 ± 0.0005 cm^?1, respectively.     

Spin-selective depopulation of triplet sublevels in rapidly rotating triplet exciplexes detected by a heavy-atom-induced magnetic field effect

A mechanism is presented explaining a reported heavy-atom-induced magnetic field effect as a consequence of non-equilibrium triplet sublevel population in an intermediate exciplex. The triplet exciplex spin polarization is induced by sub-level-selective intersystem crossing from the exciplex triplet to its singlet ground state and is decreased by an external magnetic field. The theory accounts almost quantitatively for the observed influence of magnetic field strength and heavy-atom substituents.     

Configuration of spheroidene in the photosynthetic reaction center of Rhodobacter sphaeroides : a comparison of wild-type and reconstituted R26

We compare the resonance Raman spectra acquired at two excitation wavelengths, 496.5 and 514.5 nm, of spheroidene in the wild-type reaction center of Rhodobacter sphaeroides and reconstituted into the reaction center of the Rhodobacter sphaeroides mutant R26. Our earlier work showed that the reconstituted R26 reaction center holds spheroidene in two configurations: 15,15'-cis and another configuration. Here we show that in the wild-type reaction center only 15,15'-cis spheroidene is present. In the resonance Raman spectra of the reconstituted R26 reaction centers, a transition is identified that arises exclusively from the second configuration. According to density-functional-theory calculations, this transition is specific for the 13,14-cis configuration.     

Photo-CIDNP MAS NMR in intact cells of Rhodobacter sphaeroides R26: molecular and atomic resolution at nanomolar concentration

Photochemically induced dynamic nuclear polarization (photo-CIDNP) is observed in photosynthetic reaction centers of the carotenoid-less strain R26 of the purple bacterium Rhodobacter sphaeroides by (13)C solid-state NMR at three different magnetic fields (4.7, 9.4, and 17.6 T). The signals of the donor appear enhanced absorptive (positive) and of the acceptor emissive (negative). This spectral feature is in contrast to photo-CIDNP data of reactions centers of Rhodobacter sphaeroides wildtype reported previously (Prakash, S.; Alia; Gast, P.; de Groot, H. J. M.; Jeschke, G.; Matysik, J. J. Am. Chem. Soc. 2005, 127, 14290-14298) in which all signals appear emissive. The difference is due to an additional mechanism occurring in RCs of R26 in the long-living triplet state of the donor, allowing for spectral editing by different enhancement mechanisms. The overall shape of the spectra remains independent of the magnetic field. The strongest enhancement is observed at 4.7 T, enabling the observation of photo-CIDNP enhanced NMR signals from reaction center cofactors in entire bacterial cells allowing for detection of subtle changes in the electronic structure at nanomolar concentration of the donor cofactor. Therefore, we establish in this paper photo-CIDNP MAS NMR as a method to study the electronic structure of photosynthetic cofactors at the molecular and atomic resolution as well as at cellular concentrations.     

ESR in zero field of the photoinduced triplet state in isolated reaction centers of rhodopseudomonas sphaeroides R-26 detected by the singlet ground-state absorbance

We have measured zero-field resonance transitions of the triplet state of the primary donor monitoring the transmittance at 890 nm at 1.2 K in isolated reaction centers of Rhodopseudomonas sphaeroides R-26. The transitions correspond to a decrease in transmittance, confirming the energy transfer model for the transitions detected via the antenna fluorescence in whole cells.     

Electron hopping and magnetic field dependent spin dynamics of radical ions in solution

The role of hopping on the geminate recombination of radical ions (N,N-dimethylaniline cation and anthracene anion) in acetonitrile is studied via the nanosecond time-resolved magnetic field effect on the triplet yield and the influence of donor concentration thereon. Increasing donor concentration leads to lifetime broadening of the magnetic field dependence of the triplet yield. Responsible for this effect is the perturbation of the coherent spin motion caused by hopping of the electron spin between donor sites of different nuclear spin configuration. Comparison of experimental results with calculations based on the semiclassical theory of spin motion yields an estimate of the hopping rates. Deuteration of both radicals influences the halfwidth of the magnetic field effect: at long probing times and low donor concentrations the halfwidth measured for protonated radical ions exceeds the one for the deuterated species: at short delay times and large donor concentrations, i.e. high hopping rates, this isotopic effect is reversed.     

Anisotropic spin lattice relaxation of triplet quinoxaline in a naphthalene host crystal at high magnetic field

The spin lattice relaxation in the excited triplet state of quinoxaline molecules in a perdeutero-naphthalene host crystal was investigated at high magnetic field (B = 5 T) between T = 1.7 K and T = 4.2 K. Relaxation rates were deduced from the non-monoexponential phosphorescence decay of the individual Zeeman sublevels. Besides the so far investigated Δm = 1 transition the Δm = 2 transition was observed and attributed to a purely direct process. Its anisotropy is different to that of the Δm = 1 direct process at 1.7 K. The temperature effect upon Δm = 1 relaxation anisotropy is discussed.     

Spin dynamics of the radical pair in a low magnetic field studied by the transient absorption detected magnetic field effect on the reaction yield and switched external magnetic field

The spin dynamics of the radical pair generated from the photocleavage reaction of (2,4,6-trimethylbenzoyl)diphenylphosphine oxide (TMDPO) in micellar solutions was studied by the time-resolved magnetic field effect (MFE) on the transient absorption (TA) and by a novel technique, absorption detected switched external magnetic field (AD-SEMF). Thanks to the large hyperfine coupling constant (A = 38 mT), a characteristic negative MFE on the radical yield was observed at a magnetic field lower than 60 mT whereas a positive effect due to the conventional hyperfine (HFM) and relaxation mechanisms (RM) was observed at higher magnetic field. The negative effect can be assigned to the mechanism "so-called" low field effect (LFE) mechanism and has been analyzed thoroughly using a model calculation incorporating a fast spin dephasing process. The time scale of the spin mixing process of LFE studied by AD-SEMF is shorter than the lifetime of the recombination kinetics of the radical pair. These results indicate that the LFE originates from the coherent spin motion. This can be interfered from the fast spin dephasing caused by electron spin interaction fluctuations.     

Electron spin density distribution of a series of nitroxide radicals with five-member ring as studied by solid-state 1H, 2H and 13C NMR

Stable nitroxide radicals are useful to construct molecular magnetic systems. Particularly, radicals substituted by –COOH and –CONH₂ can be coordinated to magnetic metal ions and may be used as cladding reagents of gold nano-particles for modifying magnetism. Nitroxide molecules with unsaturated five-member ring have almost planner structure and electron spin delocalization may be expected. We determined the hyperfine coupling constants (hfcc) of ¹H, ²H and ¹³C of a series of nitroxide radicals with five-member ring. Experimental values of hfcc were compared with those deduced from calculations based on density functional theory. The electron spin density distribution at β position of ring was sensitive to the ring structure, although the electron spin density at β position is small compared with N–O site. Magnetic susceptibility and UV–Vis absorption spectra were also measured and discussed.     

Contributions of spin-spin interactions to the magnetic field dependence of the triplet quantum yield in photosynthetic reaction centers

Calculations are presented which show that dipolar coupling in the primary radical ion pair of quinone-depleted photosynthetic reaction centers substanually affects the magnetic field dependence of the triplet quantum yield, as does exchange coupling to the semiquinone-Fe(II) complex, when quinone is present. Inclusion of these interactions resolves significant discrepancies between theory and experiment.     

Electron transfer in the reaction center of the Rb. sphaeroides R-26 studied by transient absorption

Electron transfer at the reaction center of the purple photosynthetic bacterium Rb. sphaeroides R-26 was measured at room temperature by the time-resolved transient absorption spectroscopy technique with 200 fs temporal resolution. The absorbance changes characteristic of the excited state of the primary donor and extending over the whole spectral range investigated from 350 nm up to 720 nm appeared after excitation with a laser pulse of about 100 fs duration at 800 nm. The time evolution of the spectra reflected the excitation of bacteriochlorophylls (BChl) M and L and the subsequent transfer of this excitation to the primary electron donor (P), with the time constant shorter than 1 ps. The decay time constant of the excited primary donor P was determined as about 3 ps, and it was faster than the rise of the reduced intermediary acceptor bacteriopheophytin (BPhe(L)). Photoreduction of BPhe(L) and its further reoxidation was clearly observed as an increase in its bleaching band intensity at around 540 nm in about 4 ps and its decrease in about 200 ps. Our findings support the theoretical model assuming the involvement of the intermediate state P(+)BChl- in the so-called "two-step" model. In this model an electron is transferred in a sequence from the excited special pair P* to bacteriochlorophyll, BChl(L), then to bacteriopheophytin, BPhe(L), and further on to quinone, Q(A). The branched charge separation, partially via P and partially via BChl(L), was also observed.     

Electron spin dynamics as a probe of molecular dynamics: temperature-dependent magnetic field effects on charge recombination within a covalent radical ion pair

The electron spin-spin exchange interaction, 2J, in radical pairs (RPs) is exquisitely sensitive to the details of molecular structure and can thus serve as an important probe of structural dynamics in RPs of potential interest to photonic and electronic devices. Photoinitiated ultrafast two-step charge separation produces (1)(MeOAn(+)(*)-6ANI-NI(-)(*)), where MeOAn = p-methoxyaniline, 6ANI = 4-(N-piperidinyl)naphthalene-1,8-dicarboximide, and NI = naphthalene-1,8:4,5-bis(dicarboximide). Radical pair intersystem crossing subsequently produces (3)(MeOAn(+)(*)-6ANI-NI(-)(*)), and the total RP population decays with approximately 10 ns lifetime at 140 K, which increases to nearly 30 ns at 300 K in toluene. The activation energy observed for this process is negative and can be explained by a mechanism involving a conformational preequilibrium of the RP followed by charge recombination. Over the same temperature range, the magnetic field effect (MFE) on yield of the triplet recombination product, MeOAn-6ANI-(3)()NI, yields the magnitude of 2J, which directly monitors the superexchange electronic coupling for charge recombination. A single resonance in the MFE plot is observed at 300 K, which splits into two resonances at temperatures below 230 K, suggesting that there are two distinct groups of RP conformations at low temperature. The magnitude of 2J for the lower field resonance (10 mT) at 140 K is 5 times smaller than that of the high field resonance. At 300 K the equilibrium is shifted almost entirely to the set of conformers with the stronger electronic coupling. The motion that couples these two groups of conformations is the motion that most effectively gates the donor-acceptor electronic coupling.     

Protein-cofactor interactions in bacterial reaction centers from Rhodobacter sphaeroides R-26: effect of hydrogen bonding on the electronic and geometric structure of the primary quinone. A density functional theory study

The effect of hydrogen bonding to the primary quinone (Q(A) and Q(*)(-)(A)) in bacterial reaction centers was studied using density functional theory (DFT) calculations. The charge neutral state Q(A) was investigated by optimizing the hydrogen atom positions of model systems extracted from 15 different X-ray structures. From this analysis, mean values of the H-bond lengths and directions were derived. It was found that the N(delta)-H of His M219 forms a shorter H-bond to Q(A) than the N-H of Ala M260. The H-bond of His M219 is linear and more twisted out of the quinone plane. The radical anion Q(*)(-)(A) in the protein environment was investigated by using a mixed quantum mechanics/molecular mechanics (QM/MM) approach. Two geometry optimizations with a different number of flexible atoms were performed. H-bond lengths were obtained and spectroscopic parameters calculated, i.e. the hyperfine and nuclear quadrupole couplings of magnetic nuclei coupled to the radical. Good agreement was found with the results provided by EPR/ENDOR spectroscopy. This implies that the calculated lengths and directions of the H-bonds to Q(*)(-)(A) are reliable values. From a comparison of the neutral and reduced state of Q(A) it was concluded that the H-bond distances are shortened by approximately 0.17 Angstroms (His M219) and approximately 0.13 Angstroms (Ala M260) upon single reduction of the quinone. It is shown that the point-dipole approximation can not be used for an estimation of H-bond lengths from measured hyperfine couplings in a system with out-of-plane H-bonding. In contrast, the evaluation of the nuclear quadrupole couplings of (2)H nuclei substituted in the hydrogen bonds yields H-bond lengths close to the values that were deduced from DFT geometry optimizations. The significance of hydrogen bonding to the quinone cofactors in biological systems is discussed.     

Local electrostatic field induced by the carotenoid bound to the reaction center of the purple photosynthetic bacterium Rhodobacter sphaeroides

Electroabsorption (EA) spectra were recorded in the region of the reaction center (RC) Qy absorption bands of bacteriochlorophyll (Bchl) and bacteriopheophytin, to investigate the effect of carotenoid (Car) on the electrostatic environment of the RCs of the purple bacterium Rhodobacter (Rb.) sphaeroides. Two different RCs were prepared from Rb. sphaeroides strain R26.1 (R26.1-RC); R26.1 RC lacking Car and a reconstituted RC (R26.1-RC+ Car) prepared by incorporating a synthetic Car (3,4-dihydrospheroidene). Although there were no detectable differences between these two RCs in their near infrared (NIR) absorption spectra at 79 and 293 K, or in their EA spectra at 79 K, significant differences were detected in their EA spectra at 293 K. Three nonlinear optical parameters of each RC were determined in order to evaluate quantitatively these differences; transition dipole-moment polarizability and hyperpolarizability (D factor), the change in polarizability upon photoexcitation (Deltaalpha), and the change in dipole-moment upon photoexcitation (Deltamu). The value of D or Deltaalpha determined for each absorption band of the two RC samples showed similar values at 77 or 293 K. However, the Deltamu values of the special pair Bchls (P) and the monomer Bchls absorption bands showed significant differences between the two RCs at 293 K. X-ray crystallography of the two RCs has revealed that a single molecule of the solubilizing detergent LDAO occupies part of the carotenoid binding site in the absence of a carotenoid. The difference in the value of Deltamu therefore represents the differential effect of the detergent LDAO and the carotenoid on P. The change of electrostatic field around P induced by the presence of Car was determined to be 1.7 x 10(5) [V/cm], corresponding to a approximately 10% change in the electrostatic field around P.     

The effect of a magnetic field on the singlet/triplet ratio in geminate ion recombination

Excited state formation by ion recombination in solutions of fluorene in squalene has been studied by pulse radiolysis at the center of a large electromagnet. The products of the slower part of the ion recombination are affected by an applied magnetic field. The fluorescence yield increases by a factor of about 1.5 whereas the triplet yield decreases.     

Unexpected sense dependence of the effect of a magnetic field on delayed fluorescence

Delayed fluorescence intensity from pyrene and methylene phenanthrene crystals alters when an applied, steady magnetic field is reversed. This sense dependence vanishes at high fields. It is maximum along the pyrene two-fold axis and zero in the mirror plane.