Low Quencher Concentrations Research Articles

Fluorescence quenching of poly(methylphenylsilane) in solution

The solution, electron-transfer fluorescence quenching of a typical aromatic polysilane[poly(methylphenylsilane)] by a series of electronic deficient aromatic monomers is described. The rate of fluorescence quenching is a function of the reduction potential of the quencher, and only very fast processes can be observed, due to the short polymer fluorescence lifetime. The measurement of quenching rate constants, which are considerably larger than diffusion control, suggests that extensive energy migration occurs in the polymer. Although the fluorescence quenching at low quencher concentrations follows Stern-Volmer kinetics, at high concentrations, contributions from static quenching are apparent. Strong fluorescence quenching can either accelerate or inhibit photodegradation, depending on the structure of the quencher.

Excimer formation of pyrene as a probe to investigate the recombination of geminate pairs: effect of multicomponent recombination on Stern-Volmer plot

Quenching of excimer and monomer flourescences induced by X-ray and UV-light as well as that of ODESR amplitude was observed with freon 113 as the quencher. The Stern-Volmer plots of these results show a bend at low quencher concentrations. Analysis of these plots give some precise information on the kinetic processes of the delayed geminate pairs.

Tryptophan fluorescence of terminal deoxynucleotidyl transferase: effects of quenchers on time-resolved emission spectra.

Terminal deoxynucleotidyl transferase (EC 2.7.7.31) is a eucaryotic DNA polymerase that does not require a template. The tryptophan environments in calf thymus terminal transferase were investigated by fluorescence. The heterogeneous emission from this multitryptophan enzyme was separated by time-resolved emission spectroscopy. Nanosecond fluorescence decays at 296-nm excitation and various emission wavelengths were deconvolved by global analysis, assuming that the lifetimes but not the relative weighting factors were independent of emission wavelength. The data were fit to three exponentials of lifetimes tau 1 = 1.4 ns, tau 2 = 4.5 ns, and tau 3 = 7.7 ns. The corresponding decay-associated emission spectra of the three components had maxima at about 328, 335, and 345 nm. The accessibility of individual tryptophan environments to polar and nonpolar fluorescence quenchers was examined in steady-state and time-resolved experiments. In the presence of iodide and acrylamide, the steady-state emission spectra shift to the blue. However, at low quencher concentrations, the emission from the 7.7-ns component (maximum 345 nm) is hardly affected, suggesting that this hydrophilic tryptophan environment is buried within the protein. On the other hand, the red shift in the steady-state emission spectrum in the presence of trichloroethanol indicates that the 1.4-ns component (maximum 328 nm) is an exposed hydrophobic tryptophan environment. The results are consistent with an inside-out model for terminal transferase protein, with the more hydrophobic tryptophan(s) near the surface and the most hydrophilic tryptophan(s) in the core.

Concentration-dependent fluorescence quenching and electron scavenging in liquids

We present a theory for the dependence of the fluorescence quenching lifetime τ on quencher concentration and the dependence of the electron-scavenger rate constant kf on scavenger concentration. At low quencher (scavenger) concentration these quantities are obtained by solution of the Smoluchowski–Debye diffusion equation. For nondilute quenchers we find positive deviations from the linear Stern–Volmer equation for 1/τ and for nondilute scavengers we find positive deviations from linear behavior in kf. With reactants that interact by a long range potential, such as charged fluorophores and quenchers, and electrons with charged scavengers, a length ã∼100–300 Å vs the collisional encounter length a ∼1–10 Å is the significant length. Corrections to dilute behavior are found to depend on the effective volume fraction of quenchers (or scavengers) φ̃=4π3ã3c/3 which can be much larger than the material volume fraction φ=4π3a3c/3 [c is the quencher (or scavenger) number density]. We consider both the initial value and steady state situations and find that, for our exact results, no transient behavior of the rate coefficient enters the initial value expression.

A model of dynamic quenching of fluorescence in globular proteins

A model is presented for the quenching of a fluorophore in a protein interior. At low quencher concentration the quenching process is determined by the acquisition rate of quencher by the protein, the migration rate of quencher in the protein interior, and the exit rate of quencher from the protein. In cases where the fluorescence emission observed in the absence of quencher could be described by a single exponential decay, the presence of quencher led to doubly exponential decay times, and the aforementioned exit rates of the quencher could be determined from experimental data. At high quencher concentration, the processes became more complex, and the deterministic rate equations used at low quencher concentration had to be modified to take into account the Poisson distribution of quencher molecules throughout the protein ensemble and also by using a migration rate for quencher in the protein interior that is a function of the quencher concentration. Simulations performed for typical fluorescent probes in proteins showed good agreement with experiments.

Influence of surface heterogeneity on the luminescence decay of probe molecules in heterogeneous systems Ru(bpy) 2+ 3 on clays

The decay of a luminescent probe [Ru(bpy)2+3] adsorbed on solids (clays) containing quenching impurities (Fe3+) has been examined. The time law is a multi-exponential function which stems from (i) the quasi-total translational immobility of the probe on the microsecond scale, and (ii) the heterogeneous nature of the surface, which is influenced by the local concentration of iron in the clay lattice. An elementary model has been proposed based on a randomly decorated 2-dimensional lattice for the probe and a second underlying and randomly decorated 2-dimensional lattice for the quencher ions. Assuming that the quenching probability for an excited probe is linearly related to the number of neighbouring quenchers, a decay function has been derived which, for very low quencher concentrations, reduces to the Infelta–Grätzel–Thomas equation for quenching in micellar solutions. The parameters of the decay function have been correlated to the chemical composition of the clays and to their swelling properties.

THE PHOTOREDUCTION OF METHYLENE BLUE BY ARYLAMINOMETHANE‐SULFONATES

Abstract— The photoreduction of methylene blue in the presence of arylaminomethanesulfonates (RAMS = RC6H4NHCH2SO3Na) was studied by laser and conventional flash photolysis. These compounds quenched the methylene blue triplet deviating from a normal Stern‐Volmer behaviour. For low quencher concentrations, a Rehm‐Weller relationship was found between the kq's and the DLG's obtained for the electron transfer reactions. The lack of further quenching at higher [RAMS] is ascribed to the formation of a ground state ion pair between the dye and the anionic quencher which, on excitation, forms a triplet state unable to under go electron transfer for steric reasons. A second order decay rate constant was found for the semireduced species (MB') (ca. 5 × 109M‐1 s‐1, independent of the RAMS used) and is attributed to a proton transfer from the radical zwitterion (RC6H4NH CH2SO3−) to MB. The overall dependence on the substituent of the bleaching observed by continuous irradiation follows the triplet behaviour.

Photochemical modifications of the tryptophan residues of wheat-germ agglutinin in the presence of trichloroethanol.

Trichloroethanol is an efficient quencher of indole fluorescence of model compounds and proteins [Eftink, M. R. and Ghiron, C. A. (1976) J. Phys. Chem. 80, 486--493]. At low quencher concentrations, the quenching follows the classical Stern-Volmer law. Bimolecular rate constants calculated from measured quenching constants and lifetimes are equal to 6 X 10(9) M-1s-1 and 1.2 X 10(9) M-1s-1 for N-acetyltrypotophanamide and wheat germ agglutinin, respectively. Upon ultraviolet irradiation in the presence of trichloroethanol, transformation of fluorescent tryptophan occurs, leading to a fluorescent photoproduct. This can be easily used as a method for the quantitative determination of fluorescent tryptophan residues in proteins. In good agreement with previous results, two fluorescent tryptophan residues per polypeptide chain are found in wheat germ agglutinin. Concomitantly with the photochemical reactions, the hemagglutinating protein activity and its affinity constant towards chitin oligomers are reduced. A probable location of tryptophan residues in the binding sites of wheat germ agglutinin is proposed.

Photochemistry of 6-substituted 2-methyl-2-alkoxycarbonylcyclohexanones

David S. Weiss Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109 (Received inUSd 17 November 1977; received inUK for publication 30 JanaarY x978) Ketone photochemistry, 1 the Norrish type I reaction of cyclic ketones in particular, has received considerable mechanistic attention2 and some synthetic application. 3 The primary photoproduct of this reaction, an acyl alkyl biradical, can and often does cascade to a multitude of products the variety apparently being determined mainly by structural features in the biradical. Therefore, a knowledge of these features is obviously of considerable relevance in terms of biradical chemistry as well as for the rational design of synthetic applications of ketone photochemistry. Recently, some of the effects of molecular structure on the disproportionation reactions resulting in alkenals and ketenes, 2 and the oxycarbene producing ring closure, 4 have been elucidated. Our interest is in the decarbonylation reaction, and although it has been known for some time that radical stabilizing substituents facilitate photochemical acleavage 1,2 as well as acyl radical decarbonylation, 5 studies were often carried out on molecules in which processes other than decarbonylation were structurally suppressed. We wish to report our results on the photochemistry of three 2-methyl-2-alkoxycarbonylcyclohexanones in which the competition between decarbonylation, disproportionation and reclosure of the acyl alkyl biradical intermediate has been quantitatively determined as a function of structure. Irradiation of compounds l-3 in cyclohexane-methanol(ethano1) resulted in -the products shown with the indicated quantum yields. 6 The reactions of 1 and 2 were quenched with trans-1,3-pentadiene resulting in nonlinear Stern-Volmer behavior and the formation of new, unidentified, products. However, by taking the Stern-Volmer slopes from the linear, low quencher concentration portion of the plots, 7 we estimate that T -1 =6x10gsec -1 for both 1 and 2. 8 Assuming tha-L the rate of a-cleavage is influenced only by the a-substituents and that this determines the triplet lifetime (-c -1 = 1.1 x 177 set -1 for cyclohexanone,' T-1 = 2.6 x lo* set -1 for 2-methylcyclohexanone,' and T -1 = 1.8 x 10' set -' for 2,2-dimethylcyclohexanonel' ),we predict that about lOO%, 96% and 77% of the cleavage should occur at I, 2 and 3, respectively the a-carbon bearing the alkoxycarbonyl substituent in The increased rate of cleavage occasioned by the