Exergonic Electron Transfer Research Articles

A study of electrochemiluminescence mechanism of Rubrene/TPrA system in an aqueous solution via stochastic collision electrochemistry in an emulsion reactor

BackgroundElectrochemiluminescence (ECL) is an electrochemically induced process in which radicals generated at the electrode surface undergo exergonic electron transfer reaction to form excited states and luminesce. ECL, with high sensitivity and superior spatiotemporal control, has been widely applied in bioanalysis and light-emitting devices. The ECL signal of rubrene (Rub) was observed in Rub/TPrA oil-in-water (o/w) emulsions, which was inconsistent with the theory of ion-transfer coupled electron-transfer in Rub emulsion droplets, and the conventional ECL mechanism in Rub/TPrA system couldn't explain this phenomenon. ResultsChoosing the toluene oil-in-water emulsion droplets as reactors, we put forward and verified a new ECL mechanism of Rub in aqueous solution on the basis of cyclic voltammograms, single-entity electrochemistry, ECL detection techniques and COMSOL finite element simulation. The new mechanism involved that Rub was reduced to Rub•- by the highly reducing species TPrA•, Rub•- was then oxidized to the excited state Rub∗ by the stable intermediate TPrA•+, and finally Rub∗ was inactivated by emitting photons. In simulation, the standard oxidation rate constant of TPrA in the emulsion droplets was deduced as 1.5 × 10−4 cm/s, and the deprotonation rate of TPrA•+ was about 200 s−1. Besides, emulsion droplets colliding on Au ultramicroelectrode (UME) were found to be able to amplify ECL signals. SignificanceThe newly proposed ECL mechanism of Rub/TPrA emulsion reactors broke the solubility problem of Rub in an aqueous, promoting the research and application of the organic luminophore Rub in bioanalysis. With the ECL signals amplification capability, the o/w emulsion reactors are promising to future design of ECL biosensors with higher sensitivity.

Anti-Dissipative Strategies toward More Efficient Solar Energy Conversion.

In natural and artificial photosynthesis, light absorption and catalysis are separate processes linked together by exergonic electron transfer. This leads to free energy losses between the initial excited state, formed after light absorption, and the active catalyst formed after the electron transfer cascade. Additional deleterious processes, such as internal conversion (IC) and vibrational relaxation (VR), also dissipate as much as 20-30% of the absorbed photon energy. Minimization of these energy losses, a holy grail in solar energy conversion and solar fuel production, is a challenging task because excited states are usually strongly coupled which results in negligible kinetic barriers and very fast dissipation. Here, we show that topological control of oligomeric {Ru(bpy)3} chromophores resulted in small excited-state electronic couplings, leading to activation barriers for IC by means of inter-ligand electron transfer of around 2000 cm-1 and effectively slowing down dissipation. Two types of excited states are populated upon visible light excitation, that is, a bridging-ligand centered metal-to-ligand charge transfer [MLCT(Lm)], and a 2,2'-bipyridine-centered MLCT [MLCT(bpy)], which lies 800-1400 cm-1 higher in energy. As a proof-of-concept, bimolecular electron transfer with tri-tolylamine (TTA) as electron donor was performed, which mimics catalyst activation by sacrificial electron donors in typical photocatalytic schemes. Both excited states were efficiently quenched by TTA. Hence, this novel strategy allows to trap higher energy excited states before IC and VR set in, saving between 100 and 170 meV. Furthermore, transient absorption spectroscopy suggests that electron transfer reactions with TTA produced the corresponding Lm•--centered and bpy•--centered reduced photosensitizers, which involve different reducing abilities, that is, -0.79 and -0.93 V versus NHE for Lm•- and bpy•-, respectively. Thus, this approach probably leads in fine to a 140 meV more potent reductant for energy conversion schemes and solar fuel production. These results lay the first stone for anti-dissipative energy conversion schemes which, in bimolecular electron transfer reactions, harness the excess energy saved by controlling dissipative conversion pathways.

Poly(ferrocenylsilane) Hydrogels as a Foundry for Metal Nanoparticle Synthesis by Direct Reduction of Electrolytes via a Catalytic Route

Poly(ferrocenylsilane)s (PFSs) as redox-responsive organometallic polymers have been used in redox reactions in noble metal ion-containing electrolyte solutions and in hydrogels swollen by electrolytes for successful preparation of metal nanoparticles (NPs) by in situ direct reduction. We report a surprising imbalance of stoichiometric ratios in redox reactions with transition metals between metal cations and PFS observed in hydrogels. Due to the redox-active ferrocene center in PFS backbones, well-dispersed noble metal NPs are directly formed in situ in the hydrogel matrix depending on their redox potential. By careful analysis, an extra-stoichiometric amount of gold NPs was found in the reaction when following the principle of exergonic electron transfer from ferrocene to gold(III) with the reduction of AuIII to Au0, and the iron center’s oxidation state increases from II to III. This surprising result is targeted to account for a stoichiometric balance of the electrons in the redox reaction, that is, we tackle the question “where do the electrons come from?” First, HAuCl4 was allowed to react with an oxidized PFS hydrogel to determine the source of the unaccounted electrons. Surprisingly, essentially regardless of the redox state of PFS, similar amounts of gold NPs were formed. The formation of H+ ions in the reaction indicates that H2O could be a possible electron source. A comparison with the stoichiometric relations in PtNP formation further confirms this hypothesis.

Transient luminescence spectroscopy of dialkylRu(bpy)3 2+-citratoperoxotitanate-dialykylviologen triad LB film deposited on quartz slab waveguide

Amphiphilic Ru(bpy)3 2+ complex (DC17Ru)—citratoperoxotitanate (CPT) —ditetradecylviologen (DC14V) triad nano-film was fabricated on quartz slab optical waveguide by organic-inorganic hybrid Langmuir-Blodgett method. Transient phosphorescence of the DC17Ru monolayer at 610 nm excited by 532 nm pulsed laser was observed by SOWG spectroscopy. The phosphorescence was partly quenched by the CPT layer and the quenching was enhanced by the DC14V monolayer underneath the CPT layer in the DC17Ru-CPT nano-film. Energy levels of the components, DC17Ru, CPT and DC14V suggests an exergonic cascade electron transfer mechanism from the excited state of DC17Ru to DC14V via the CPT.

Generation of Catalytically Active Gold Nanocrystals in Water Induced with Ferrocene Carboxylate

AbstractThe fabrication and properties of anisotropic gold nanoparticles (AuNPs) are of great interest from fundamental and application standpoints. Whereas various neutral ferrocene derivatives have been shown to reduce AuIII to Au0NPs owing to slightly exergonic electron transfer in biphasic systems, the question remained open for water soluble ferrocenes containing electron withdrawing substituents increasing the FeIII/FeII oxidation potential. Here, it is shown that reaction of sodium ferrocene carboxylate, NaFcCO2, alone, does not reduce H[AuIIICl4] to stable AuNPs under ambient conditions in water. On the other hand, reaction between NaFcCO2 and H[AuIIICl4] in water in the presence of poly (vinyl pyrrolidone) (PVP), citrate or other 1,2,3‐triazole (trz)‐containing stabilizers is shown to produce catalytically active Au nanocrystals with various shapes. EDX results confirm the fixation of the ferrocenyl carboxylate ligand onto the AuNP surface. 4‐nitrophenol reduction to 4‐aminophenol by Na[BH4] in water proceeds in the presence of the various stabilizers, without induction time in most cases, which confirms the weakness of the ferrocenyl carboxylate ligand bonded to the AuNP surface and the favorable role of ferrocenyl carboxylate ligand for catalysis at the AuNP surface.

Effects of active site arginine residue on redox behavior of flavins in bifurcating electron transfer flavoprotein

Flavin based electron bifurcation (FBEB) is observed in bifurcating electron transfer flavoproteins (BfEtfs). Electron bifurcation couples endergonic and exergonic electron transfer reactions to exploit lower potential electron transfer reaction step therefore this is considered as a third mode of energy conservation mechanism in biological systems. We study about the bifurcating EtfAB system from Rhodoseudomonas palustris (Rpal). This is a heterodimer containing two flavin adenine dinucleotides (FADs), one in each domain. One flavin accepts two electrons from NADH and bifurcates (Bf-FAD) each electron to higher potential electron transfer FAD (ET-FAD) and lower potential ferredoxin or flavodoxin. ET-FAD likely rests in its anionic semiquinone (ASQ) state and upon accepting an electron from the Bf-FAD it becomes a hydroquinone (HQ). BF-FAD accepts a pair of electrons from NADH to form HQ and distributes the electrons to separate acceptors without accumulating a SQ intermediate. This contrasting redox behavior of the two flavins in this system in turn points a spotlight on whether the conserved active site arginine residues have different effects in the two binding sites. R273 favors the ASQ of ET-FAD, whereas R165 near the Bf-FAD appears not to, possibly due to neutralization of its positive charge by nearby C174. R273 forms a pi- pi stacking interaction with ET-FAD whereas R165 appears to form hydrogen bond interactions with Bf-FAD. We performed site directed mutagenesis to change the size of the positively charged residue by replacing with lysine, neutralize the charge by inserting alanine and make the charge tunable via modulation of pH by introducing a histidine. Amino acid substitutions were made in each of the two flavin-binding sites separately. Based on the Spectro-electrochemical redox titrations, all the variants retained the WT pattern of single electron reactivity at the ET site and pairwise reactivity of the Bf-FAD. Reduction midpoint potential titrations showed 50% less accumulation of ASQ in R273H and R273A, whereas R273K produced similar an amount similar to that observed in WT. This confirms that positive charge helps in stabilization of ASQ. Redox titrations performed at pH 7 and 8 suggest that the substituting H273 residue remains neutral at these pHs. Amino acid replacement in the bifurcating site, R165K did not affect the formation of ASQ in ET-FAD, but removal of residue 165's positive charge in R165H resulted in diminished stability prevented FAD binding in the Bf site at pH8. Therefore, R273 plays a vital role in Bf-ETF by stabilizing the ASQ of the ET-FAD. On the other hand, R165 stabilizes the Bf-FAD that is essential for electron bifurcation in RpalEtf.

Spectroscopic, thermodynamic and computational evidence of the locations of the FADs in the nitrogen fixation-associated electron transfer flavoprotein.

Flavin-based electron bifurcation allows enzymes to redistribute energy among electrons by coupling endergonic and exergonic electron transfer reactions. Diverse bifurcating enzymes employ a two-flavin electron transfer flavoprotein (ETF) that accepts hydride from NADH at a flavin (the so-called bifurcating FAD, Bf-FAD). The Bf-FAD passes one electron exergonically to a second flavin thereby assuming a reactive semiquinone state able to reduce ferredoxin or flavodoxin semiquinone. The flavin that accepts one electron and passes it on via exergonic electron transfer is known as the electron transfer FAD (ET-FAD) and is believed to correspond to the single FAD present in canonical ETFs, in domain II. The Bf-FAD is believed to be the one that is unique to bifurcating ETFs, bound between domains I and III. This very reasonable model has yet to be challenged experimentally. Herein we used site-directed mutagenesis to disrupt FAD binding to the presumed Bf site between domains I and III, in the Bf-ETF from Rhodopseudomonas palustris (RpaETF). The resulting protein contained only 0.80 ± 0.05 FAD, plus 1.21 ± 0.04 bound AMP as in canonical ETFs. The flavin was not subject to reduction by NADH, confirming absence of Bf-FAD. The retained FAD displayed visible circular dichroism (CD) similar to that of the ET-FAD of RpaETF. Likewise, the mutant underwent two sequential one-electron reductions forming and then consuming anionic semiquinone, reproducing the reactivity of the ET-FAD. These data confirm that the retained FAD in domain II corresponds the ET-FAD. Quantum chemical calculations of the absorbance and CD spectra of each of WT RpaETF's two flavins reproduced the observed differences between their CD and absorbance signatures. The calculations for the flavin bound in domain II agreed better with the spectra of the ET-flavin, and those calculated based on the flavin between domains I and III agreed better with spectra of the Bf-flavin. Thus calculations independently confirm the locations of each flavin. We conclude that the site in domain II harbours the ET-FAD whereas the mutated site between domains I and III is the Bf-FAD site, confirming the accepted model by two different tests.

On the nature of organic and inorganic centers that bifurcate electrons, coupling exergonic and endergonic oxidation-reduction reactions.

Bifurcating electrons to couple endergonic and exergonic electron-transfer reactions has been shown to have a key role in energy conserving redox enzymes. Bifurcating enzymes require a redox center that is capable of directing electron transport along two spatially separate pathways. Research into the nature of electron bifurcating sites indicates that one of the keys is the formation of a low potential oxidation state to satisfy the energetics required of the endergonic half reaction, indicating that any redox center (organic or inorganic) that can exist in multiple oxidation states with sufficiently separated redox potentials should be capable of electron bifurcation. In this Feature Article, we explore a paradigm for bifurcating electrons down independent high and low potential pathways, and describe redox cofactors that have been demonstrated or implicated in driving this unique biochemistry.

Structural Changes and Proton Transfer in Cytochrome c Oxidase

In cytochrome c oxidase electron transfer from cytochrome c to O2 is linked to transmembrane proton pumping, which contributes to maintaining a proton electrochemical gradient across the membrane. The mechanism by which cytochrome c oxidase couples the exergonic electron transfer to the endergonic proton translocation is not known, but it presumably involves local structural changes that control the alternating proton access to the two sides of the membrane. Such redox-induced structural changes have been observed in X-ray crystallographic studies at residues 423–425 (in the R. sphaeroides oxidase), located near heme a. The aim of the present study is to investigate the functional effects of these structural changes on reaction steps associated with proton pumping. Residue Ser425 was modified using site-directed mutagenesis and time-resolved spectroscopy was used to investigate coupled electron-proton transfer upon reaction of the oxidase with O2. The data indicate that the structural change at position 425 propagates to the D proton pathway, which suggests a link between redox changes at heme a and modulation of intramolecular proton-transfer rates.

Structural Changes and Proton Transfer in Cytochrome c Oxidase.

In cytochrome c oxidase electron transfer from cytochrome c to O2 is linked to transmembrane proton pumping, which contributes to maintaining a proton electrochemical gradient across the membrane. The mechanism by which cytochrome c oxidase couples the exergonic electron transfer to the endergonic proton translocation is not known, but it presumably involves local structural changes that control the alternating proton access to the two sides of the membrane. Such redox-induced structural changes have been observed in X-ray crystallographic studies at residues 423–425 (in the R. sphaeroides oxidase), located near heme a. The aim of the present study is to investigate the functional effects of these structural changes on reaction steps associated with proton pumping. Residue Ser425 was modified using site-directed mutagenesis and time-resolved spectroscopy was used to investigate coupled electron-proton transfer upon reaction of the oxidase with O2. The data indicate that the structural change at position 425 propagates to the D proton pathway, which suggests a link between redox changes at heme a and modulation of intramolecular proton-transfer rates.

Electron-transfer acceleration investigated by time resolved infrared spectroscopy.

Ultrafast electron transfer (ET) processes are important primary steps in natural and artificial photosynthesis, as well as in molecular electronic/photonic devices. In biological systems, ET often occurs surprisingly fast over long distances of several tens of angströms. Laser-pulse irradiation is conveniently used to generate strongly oxidizing (or reducing) excited states whose reactions are then studied by time-resolved spectroscopic techniques. While photoluminescence decay and UV-vis absorption supply precise kinetics data, time-resolved infrared absorption (TRIR) and Raman-based spectroscopies have the advantage of providing additional structural information and monitoring vibrational energy flows and dissipation, as well as medium relaxation, that accompany ultrafast ET. We will discuss three cases of photoinduced ET involving the Re(I)(CO)3(N,N) moiety (N,N = polypyridine) that occur much faster than would be expected from ET theories. [Re(4-N-methylpyridinium-pyridine)(CO)3(N,N)](2+) represents a case of excited-state picosecond ET between two different ligands that remains ultrafast even in slow-relaxing solvents, beating the adiabatic limit. This is caused by vibrational/solvational excitation of the precursor state and participation of high-frequency quantum modes in barrier crossing. The case of Re-tryptophan assemblies demonstrates that excited-state Trp → *Re(II) ET is accelerated from nanoseconds to picoseconds when the Re(I)(CO)3(N,N) chromophore is appended to a protein, close to a tryptophan residue. TRIR in combination with DFT calculations and structural studies reveals an interaction between the N,N ligand and the tryptophan indole. It results in partial electronic delocalization in the precursor excited state and likely contributes to the ultrafast ET rate. Long-lived vibrational/solvational excitation of the protein Re(I)(CO)3(N,N)···Trp moiety, documented by dynamic IR band shifts, could be another accelerating factor. The last discussed process, back-ET in a porphyrin-Re(I)(CO)3(N,N) dyad, demonstrates that formation of a hot product accelerates highly exergonic ET in the Marcus inverted region. Overall, it follows that ET can be accelerated by enhancing the electronic interaction and by vibrational excitation of the reacting system and its medium, stressing the importance of quantum nuclear dynamics in ET reactivity. These effects are experimentally accessible by time-resolved vibrational spectroscopies (IR, Raman) in combination with quantum chemical calculations. It is suggested that structural dynamics play different mechanistic roles in light-triggered ET involving electronically excited donors or acceptors than in ground-state processes. While TRIR spectroscopy is well suitable to elucidate ET processes on a molecular-level, transient 2D-IR techniques combining optical and two IR (or terahertz) laser pulses present future opportunities for investigating, driving, and controlling ET.

S-glutathionylation reactions in mitochondrial function and disease.

Mitochondria are highly efficient energy-transforming organelles that convert energy stored in nutrients into ATP. The production of ATP by mitochondria is dependent on oxidation of nutrients and coupling of exergonic electron transfer reactions to the genesis of transmembrane electrochemical potential of protons. Electrons can also prematurely “spin-off” from prosthetic groups in Krebs cycle enzymes and respiratory complexes and univalently reduce di-oxygen to generate reactive oxygen species (ROS) superoxide (O2•−) and hydrogen peroxide (H2O2), important signaling molecules that can be toxic at high concentrations. Production of ATP and ROS are intimately linked by the respiratory chain and the genesis of one or the other inherently depends on the metabolic state of mitochondria. Various control mechanisms converge on mitochondria to adjust ATP and ROS output in response to changing cellular demands. One control mechanism that has gained a high amount of attention recently is S-glutathionylation, a redox sensitive covalent modification that involves formation of a disulfide bridge between glutathione and an available protein cysteine thiol. A number of S-glutathionylation targets have been identified in mitochondria. It has also been established that S-glutathionylation reactions in mitochondria are mediated by the thiol oxidoreductase glutaredoxin-2 (Grx2). In the following review, emerging knowledge on S-glutathionylation reactions and its importance in modulating mitochondrial ATP and ROS production will be discussed. Major focus will be placed on Complex I of the respiratory chain since (1) it is a target for reversible S-glutathionylation by Grx2 and (2) deregulation of Complex I S-glutathionylation is associated with development of various disease states particularly heart disease. Other mitochondrial enzymes and how their S-glutathionylation profile is affected in different disease states will also be discussed.

Exciplexes versus Loose Ion Pairs: How Does the Driving Force Impact the Initial Product Ratio of Photoinduced Charge Separation Reactions?

Many donor–acceptor systems can undergo a photoinduced charge separation reaction, yielding loose ion pairs (LIPs). LIPs can be formed either directly via (distant) electron transfer (ET) or indirectly via the dissociation of an initially formed exciplex or tight ion pair. Establishing the prevalence of one of the reaction pathways is challenging because differentiating initially formed exciplexes from LIPs is difficult due to similar spectroscopic footprints. Hence, no comprehensive reaction model has been established for moderately polar solvents. Here, we employ an approach based on the time-resolved magnetic field effect (MFE) of the delayed exciplex luminescence to distinguish the two reaction channels. We focus on the effects of the driving force of ET and the solvent permittivity. We show that, surprisingly, the exciplex channel is significant even for an exergonic ET system with a free energy of ET of −0.58 eV and for the most polar solutions studied (butyronitrile). Our findings demonstrate that exciplexes play a crucial role even in polar solvents and at moderate driving forces, contrary to what is usually assumed.

Bimolecular Electron Transfers That Deviate from the Sandros–Boltzmann Dependence on Free Energy: Steric Effect

As we reported recently, endergonic to mildly exergonic electron transfer between neutral aromatics (benzenes and biphenyls) and their radical cations in acetonitrile follows a Sandros-Boltzmann (SB) dependency on the reaction free energy (ΔG); i.e., the rate constant is proportional to 1/[1 + exp(ΔG/RT)]. We now report deviations from this dependency when one reactant is sterically crowded: 1,4-di-tert-butylbenzene (C1), 1,3,5-tri-tert-butylbenzene (C2), or hexaethylbenzene (C3). Obvious deviation from SB behavior is observed with C1. Stronger deviation is observed with the more crowded C2 and C3, where steric hindrance increases the interplanar separation at contact by ~1 Å, significantly decreasing the π orbital overlap. Consequently, electron transfer (k(et)) within the contact pair becomes slower than diffusional separation (k(-d)), causing deviation from the SB dependency, especially near ΔG = 0. Fitting the data to a standard electron-transfer theory gives small matrix elements (~5-7 meV) and reasonable reorganization energies. A small systematic difference between reactions of C3 with benzenes vs biphenyls is rationalized in terms of small differences in the electron-transfer parameters that are consistent with previous data. The influence of solvent viscosity on the competition between k(et) and k(-d) was investigated by comparing reactions in acetonitrile and propylene carbonate.

The 2-benzoyl xanthone/triethylamine system as a type II photoinitiator: A laser flash photolysis and computational study

2-Benzoylxanthone ( BzX) was synthesized, characterized and used as type II photoinitiator ( PI) in combination with triethylamine for the polymerization of methylmethacrylate ( MMA). The photophysical/photochemical behaviour of the photoinitiator, the involved excited state and the reaction with the amine co-initiator was studied by means of absorption and nanosecond time-resolved absorption spectroscopy. Upon irradiation with 266 or 355 nm laser light, the triplet state 3 BzX* ( λ max = 355 nm and 530 nm) was generated as the only transient (lifetime of 22.7 μs) in nitrogen saturated MeCN solution. 3 BzX* was confirmed through quenching experiments with oxygen, 2-methylbutadiene, perylene and MMA and spectral similarity to benzophenone triplet ( 3 BP*). The quantum yield of its formation ( Φ T = 0.8) and the molar absorption coefficient ( ɛ = 7500 L mol −1 cm −1) was measured in MeCN. The triplet was photoreduced by triethylamine (TEA) via photoinduced electron/proton transfer giving the corresponding ketyl and α-amino ethyl radical ( CHMe–NEt 2). From the reduction potential of BzX measured via cyclic voltametry (two cathodic peaks at −1.63 V and −1.91 V vs. Ag/AgCl), an exergonic electron transfer reaction results. The ketyl radical resembles the well-known benzophenone ketyl. In conclusion, the triplet 3 BzX* corresponds to an n → π* transition localized on the benzoyl substituent and resembles the benzophenone triplet 3 BP* but deviates from the triplet state of xanthone ( 3 X*). This is supported through DFT/B3LYP calculations, viz., (i) fully ground and triplet state geometry optimizations show that charge and spin densities are localized on the benzoyl group, and (ii) calculation of the electronic transitions via TD-DFT at B3LYP/631G+(d) and PB1BPE/631G+(d) levels of theory shows the local character. Agreement with experiment is better by applying the conductor like polarized continuum model (CPCM) to consider the solvent effect (MeCN). The effectiveness of BzX as photoinitiator for the polymerization of MMA is found to be double that of the unsubstituted xanthone ( X). The photopolymerization rates ( R p ) were found to be 7.06 × 10 −4 mol L −1 s −1 in the case of BzX and 3.04 × 10 −4 mol L −1 s −1 in the case of BX. This is attributed to the fact that triplet 3 BzX* behaves like the benzophenone triplet 3 BP* and deviates from that of 3 X*, i.e., it is the localization of the triplet excitation on the benzoyl subunit which renders BzX a good type- II photoinitiator.

Operational calixarene-based fluorescent sensing systems for choline and acetylcholine and their application to enzymatic reactions

Electron-rich anionic calixarenes and resorcinarenes are known receptors for trimethylammonium-containing neurotransmitters, but the development of practical sensor applications has been impeded by the lack of suitable supramolecular sensing ensembles as well as the low selectivity and sensitivity of the macrocyclic cation-receptor hosts. The host–guest complexes between p-sulfonatocalix[n]arenes (n = 4–5) and the cationic aromatic fluorescent dye lucigenin (LCG) have been characterised by optical spectroscopic techniques, NMR, cyclic voltammetry, isothermal titration calorimetry, and X-ray crystallography. The dye is complexed with binding constants of the order of 107 M−1 and undergoes a strong static fluorescence quenching (factor 140) upon complexation as a consequence of exergonic electron transfer within the complex. LCG has been utilised in combination with p-sulfonatocalix[4]arene to set-up a refined reporter pair for label-free continuous real-time enzyme assays according to the supramolecular tandem assay principle. This affords product-selective tandem assays for amino acid decarboxylases with a one order of magnitude higher sensitivity and a 3 orders of magnitude lower host/dye concentration range, a convenient substrate-selective tandem assay for direct monitoring of choline oxidase, and a conceptually novel substrate-selective enzyme-coupled tandem assay for acetylcholinesterase. The applicability of the method to the measurement of enzyme-kinetic parameters, the screening for inhibitors of acetylcholinesterase, and the highly selective determination of absolute, low micromolar concentrations of both choline and acetylcholine by simple fluorescence measurements has been demonstrated. A domino tandem assay can be employed to measure the two analytes in the same sample. The described applications bypass problems related to the unselective binding of the macrocycle by coupling the signalling event with highly specific enzymatic transformations.

ChemInform Abstract: Photochemical Generation, Isomerization, and Oxygenation of Stilbene Cation Radicals.

The cation radicals of cis- and trans-stilbene and several of their ring-substituted derivatives have been generated in solution directly by means of pulsed-laser-induced electron transfer to singlet cyanoanthracenes or indirectly via electron transfer from biphenyl to the singlet cyanoanthracene followed by secondary electron transfer from the stilbenes to the biphenyl cation radical. Transient absorption spectra of the cis- and trans-stilbene cation radicals generated by secondary electron transfer are similar to those previously obtained in 77 K matrices. Quantum yields for radical ion-pair cage escape have been measured for direct electron transfer from the stilbenes to three neutral and one charged singlet acceptor. These values increase as the ion-pair energy increases due to decreased rate constants for radical ion-pair return electron transfer, in accord with the predictions of Marcus theory for highly exergonic electron transfer. Cage-escape efficiencies are larger for trans- vs cis-stilbene cation radicals, possibly due to the greater extent of charge delocalization in the planar trans vs nonpolar cis cation radicals. Cage-escape stilbene cation radicals can initiate a concentration-dependent one way cis- {yields} trans-stilbene isomerization reaction.

Does Bimolecular Charge Recombination in Highly Exergonic Electron Transfer Afford the Triplet Excited State or the Ground State of a Photosensitizer?

The investigations were made on photoinduced electron transfer (ET) from the singlet excited state of rubrene (1RU*) to p-benzoquinone derivatives (duroquinone, 2,5-dimethyl-p-benzoquinone, p-benzoquinone, 2,5-dichloro-p-benzoquinone, and p-chloranil) in benzonitrile (PhCN) by using the steady state and time-resolved spectroscopies. The photoinduced ET produces solvent-separated type charge-separated (CS) species and the charge-recombination (CR) process between RU radical cation and semiquinone radical anions obeys second-order kinetics. Not only the CS species but also the triplet excited state of RU (3RU*) is seen in the transient absorption spectra upon laser excitation of a PhCN solution of RU and p-benzoquinone derivatives. The comparison of their time profiles clearly suggests that the CR process between RU radical cation and semiquinone radical anions to the ground state is independent from the deactivation of 3RU*. This indicates that the CR in a highly exergonic ET occurs at a longer distance with a large solvent reorganization energy, which results in faster ET to the ground state than to the triplet excited state that is lower in energy than the CS state. Photoinduced ET from 3RU* in addition from 1RU* also occurs when p-benzoquinone derivatives with electron-withdrawing substituents were employed as electron acceptors.

The Protonation State of a Heme Propionate Controls Electron Transfer in CytochromecOxidase

In cytochrome c oxidase (CcO), exergonic electron transfer reactions from cytochrome c to oxygen drive proton pumping across the membrane. Elucidation of the proton pumping mechanism requires identification of the molecular components involved in the proton transfer reactions and investigation of the coupling between internal electron and proton transfer reactions in CcO. While the proton-input trajectory in CcO is relatively well characterized, the components of the output pathway have not been identified in detail. In this study, we have investigated the pH dependence of electron transfer reactions that are linked to proton translocation in a structural variant of CcO in which Arg481, which interacts with the heme D-ring propionates in a proposed proton output pathway, was replaced with Lys (RK481 CcO). The results show that in RK481 CcO the midpoint potentials of hemes a and a(3) were lowered by approximately 40 and approximately 15 mV, respectively, which stabilizes the reduced state of Cu(A) during reaction of the reduced CcO with O(2). In addition, while the pH dependence of the F --> O rate in wild-type CcO is determined by the protonation state of two protonatable groups with pK(a) values of 6.3 and 9.4, only the high-pK(a) group influences this rate in RK481 CcO. The results indicate that the protonation state of the Arg481 heme a(3) D-ring propionate cluster having a pK(a) of approximately 6.3 modulates the rate of internal electron transfer and may act as an acceptor of pumped protons.

Formation and Anomalous Behavior of Aminonaphthalene−Cinnamonitrile Exciplexes

The solvent dependence of the electronic spectra of several 1-aminonaphthalenes and 2-aminonaphthalenes in the absence and presence of two styrene derivatives, cinnamonitrile and 1-phenylpropene, has been investigated. The absorption maxima of the aminonaphthalenes are dependent upon both the polarity and hydrogen-bond donor and acceptor properties of the solvent and display both red and blue solvent shifts. The fluorescence maxima are more sensitive to solvent polarity than are the absorption maxima and display red shifts in both non-hydroxylic and hydroxylic solvents. The excited state dipole moments of the 1-aminonaphthalenes are larger than those of the 2-aminonaphthalenes. Quenching of the aminonaphthalene fluorescence by cinnamonitrile occurs with diffusion-controlled rates in all solvents, in accord with the occurrence of exergonic electron transfer. Exciplex fluorescence is observed only for the tertiary aminonaphthalenes in nonpolar solvents. The exciplex fluorescence displays anomalous temperature dependence in hexane solution, an increase in temperature resulting in blue-shifted exciplex fluorescence and an increase in the exciplex decay time. Quenching of the aminonaphthalene fluorescence by 1-phenylpropene is slower than diffusion-controlled and is not accompanied by exciplex fluorescence.