The absolute yield of bacteriochlorophyll fluorescence in vivo

Abstract— –The method of Weber and Teale for determining absolute fluorescence quantum yield of dyes in solution was modified for determination of the yield of bacteriochlorophyll fluorescence from chromatophores and whole cells of photosynthetic bacteria. Measured yields ranged from about 1–6 per cent. The yield depended on intensity and wavelength of the exciting light. The higher yield at higher light intensity was interpreted as due to saturation of photosynthesis. The lower yield in some strains when excited at 810 nm was attributed to preferential excitation of the reaction center pigment P800. From this study and the lifetime measurements of others, the relation τ=Q.τ₀ was substantiated for the fluorescence of bacteriochlorophyll in vivo, τ being the actual lifetime, τ₀ the intrinsic lifetime as estimated from the absorption band area, and Q the quantum yield of fluorescence.     

THE EFFECT OF TREATMENT OF CHLOROPLAST MEMBRANES WITH GUANIDINE HC1 AND AQUEOUS ACETONE ON THE FLUORESCENCE OF BOUND ANS AND CHLOROPHYLL-a†

Abstract The fluorescence intensity of the extrinsic chromophore 1-anilino-naphthalene-8-sulfonate (ANS) bound to pea chloroplast fragments shows a sigmoidal rise as the pH of the suspending medium is decreased by the addition of HC1. The abrupt increase occurs at pH – 4.5. A 70% decrease in the maximal fluorescence intensity (pH range 3.5-4.5) of bound ANS was observed when soluble chloroplast proteins were removed by washing with water. Extraction of chloroplast membranes with 6 M guanidine-HC1 abolishes the acid–induced enhancement of ANS fluorescence. However, the subsequent removal of lipids (by 80% acetone extraction) from the guanidine-HC1-extracted naked membranes restores the acid-induced fluorescence increase. These results suggest that ANS binds mainly to the surface of the chloroplast membrane and the fluorescence changes of ANS by acidification mainly reflect the changes in the associated proteins. The lack of enhancement of the fluorescence of ANS by acidification of the guanidine-HCl treated membranes and the recovery of the acid-induced fluorescence rise after extraction of the lipids from the guanidine-HCl treated membranes suggest that the boundary lipids somehow prevent the entry of the ANS molecules into the hydrophobic interior of the naked membrane. The lipid-depleted, guanidine-HCl extracted naked membrane fragments do not show any shift in the position of the peak of emission of ANS (λ= 470 nm) upon acidification as the lipid-depleted preparations without guanidine-HCl treatment do (shift from 460 to 470 nm). Divalent cations (Mn²⁺, Ca²⁺, Mg²⁺) also increased ANS fluorescence intensities when added to both types of lipid-depleted chloroplast preparations. A comparative analysis of ANS fluorescence bound to the lipid-depleted and guanidine-HCl treated chloroplast fragments with that of just lipid-depleted fragments shows that the acidification of the latter brings about a greater change in the value of the relative binding sites (n) and the dissociation constant k_d of ANS than the protonation of the former. The role of chloroplast protein and lipid components in the structural changes of the thylakoid membrane imposed by external perturbations is discussed.     

Positron annihilation in amino acids

Positron lifetime measurements were performed on eight different amino acids, τ₁, τ₂ and I₂ values were observed at around 0.35 nsec, 0.7–1.2 nsec and 20–30%, respectively. The long-lived component seems to be produced by positronium states trapped in the crystalline lattice or by positronium atoms bound to molecules.     

Rheological properties of ionic and nonionic polyacrylamide solutions

Non-Newtonian shear viscosities were measured over six decades of strain rate k for 13 solutions of both the ionic and nonionic forms of polyacrylamide. By using the Weissenberg rheogoniometer with both the cone-and-plate and the parallel-plate attachments, the normal stress functions σ₁ (k²) and σ₂(k²) were obtained for four of the solutions. From the measurements of the shear viscosity and the normal stresses at low rates of strain, characteristic times τ and τ_N, respectively, were determined for each solution. The quantity τ was then used to nondimensionalize the strain rate τk, and when plotted versus the reduced shear viscosity, found successfully to correlate the experimental data for all the polyelectrolyte solutions over the entire range of τk and the data for the concentrated solutions of the nonionic polymer over a smaller range of τk. However, in order to correlate the normal stress data for the polyelectrolyte solutions, a second reduced strain rate (τ_Nk) was used. Thus, two different times were required to correlate all the observed data. The shear viscosity data for the dilute solutions of the nonionic polymer were well represented by the two-parameter, non-Newtonian intrinsic viscosity function that has been computed by Fixman.     

Mechanisms and parameters of transients and oscillations of delayed chlorophyll fluorescence in the thylakoid membrane of the intact maize leaf

Standard induction processes of delayed fluorescence (DF) of chlorophyll (induction signals) occur when an intact leaf segment of maize inbreds and hybrids is initially kept in the phosphoroscope darkroom for more than 15 min (τ > 15 min), and then the leaf is illuminated with the intermittent white light and measured. Resolved induction processes of DF chlorophyll into transients: A, B, C, D, and E occur when the intact leaf segment of maize inbreds and hybrids is kept in the phosphoroscope darkroom for a significantly shorter period (30 s ≤ τ ≤ 240 s), with the time rate τ of 30 s, prior to its illumination with the intermittent white light. Induction transients: A, B, C, D, and E are characterised with their temporal parameters: t _A, t _B, t _C, t _D, and t _E, dynamics of changes in transients intensities and mechanisms of their generation. The induction processes of chlorophyll DF of the intact leaf of maize inbreds and hybrids resolved into transients: A, B, C, D, and E are accompanied by the occurrence and different levels of activation energy (E _a, kJ mol⁻¹) that correspond to different critical temperatures. The generation mechanisms of induction transients: A, B, C, D, and E are classified into two groups. Transients A and B are of a physical character, while the transients: C, D, and E are of a chemical character. It is shown that the generation of the induction transients: B, C, D, and E simultaneously follows establishing of the oscillations of induction processes of the DF chlorophyll. Oscillating of induction processes of DF chlorophyll is explained by the ion (K⁺, Na⁺, H⁺, Cl⁻) transport mechanism across the thylakoid membrane of the intact leaf of maize inbreds and hybrids grown under conditions of air drought, increased temperatures and water deficiency in the medium. The article is published in the original.     

Picosecond laser photolysis study of the photocycloreversion of heterocoerdianthrone endoperoxide

The formation of heterocoerdianthrone (HCD) following laser pulse photolysis of its endoperoxide (HCDPO) was observed by laser-induced fluorescence. The rise time of HCD was determined to be τ_r = 40±10 ps. The lifetime of the S₃(ππ^*) state of HCDPO, from which cycloreversion occurs, was estimated by steady-state fluorescence measurements to be τ < 3 ps. These results indicate photocycloreversion of HCDPO as a two-step reaction.     

Chain fragmentation under liquid-state and solid-state conditions

Thermally Activated fragmentation of copolycarbonates PC(T_xA_1-x) of bisphenol A (unit CA) and the Heat-sensitive diol 1,1,2,2-tetraethyl-1,2-di-(p-hydroxy) phenylethane (unit CT) was studied in the bulk, i.e., in the pure copolymers and in their blends with the polycarbonates of bisphenol A (PCA) or tetramethyl bisphenol A (TMPC). Fragmentation proceeds via dissociation followed by disproportionation at the central C? C bond of the unit CT. The reaction has rates that are convenient to study near the glass transition temperature. The “chemical” time constants τ for the entire reaction and τ₂ for the disproportionation step compete with the “physical” time constants δ_α for segmental motion and δ_q for fragment diffusion. A cage effect is observed below τ₂ = δ_α and effects of delayed matrix response below τ = δ_α and τ = δ_q. Owing to the two latter effects, parameters such as the glass transition temperature and the structure factor of concentration fluctuations do not respond primarily to the fragmentation, but rather to subsequent relaxation and diffusion processes in the polymer matrix.     

Pre´paration chimique et proprie´te´s optiques de CeP5O14 triclinique

Crystals of a new cerium(III)-ultraphosphate form, CeP₅O₁₄, have been grown from CeCl₃ · 7H₂O and NH₄H₂PO₄. Synthesis and structural characterization by X-ray diffraction and ir absorption spectroscopy are given. The new CeP₅O₁₄ crystallizes in a triclinic unit cell, P1, with parameters: a = 9.229(2), b = 8.879(1), c = 7.201(1) (Å), α = 110.27(1), β = 102.75(1), γ = 82.13(1)°, Z = 2, and D_x = 3.20. This compound is piezoelectric and has no known structural analog. The excitation and emission spectrum of this Ce-ultraphosphate variety are reported. This material emits strongly in the near-uv. The emission band peaks at 322 nm and decays, at first, with τ₁ = 14 nsec, then, with τ₂ = 60 nsec.     

Luminescence and nonradiative relaxation of Pb2+, Sn2+, Sb3+, and Bi3+ in oxide glasses

Absorption and fluorescence spectra of Sn²⁺ and Sb³⁺ in borax, phosphate, and germanate glasses were measured in the temperature range 87–295°K. Fluorescence decay times of these ions in borax glass at 87°K was a single exponent with τ ≈ 6–11 μsec. At 293°K, two decay times were resolved in the range of 50–2000 nsec. The nonexponential behavior is interpreted by the repopulation of the ³P₁ level from the ³P₀ level. The temperature dependence of fluorescence and the low values of quantum efficiencies of fluorescence are explained by means of the configurational coordinate diagram model.     

A theoretical approach to the kinetics of polymer degradation in solution

After main-chain scission in a polymer, the frequency of encounter between segments in the different fragments is related to the separating process between the fragments. The relationship obtained shows that the separating time is proportional to M ^½, where M is the molecular weight, when the excluded volume disappears. When good solvent is used, the half-time for the separation is obtained as τ_½ = const. M ^0.16–0.22, which is approximated to the experimental data obtained previously (τ_½ = const. M ^0.34 and τ_½ = const. M ^0.22) for the degradations of polyisobutene and poly(phenyl vinyl ketone), respectively. The increase of the half-time with increasing coil density can be explained by the excluded volume. The inverse proportionality of the diffusion of segments to solvent viscosity explains the proportionality of the half-time to microviscosity. The above separating process reverses the reaction between polymer radicals. From their dependence on the chain length, τ_½/k_D = const. M ^½ (where k_D is the specific rate for the reaction), is estimated. Such an approximation holds, regardless of the type of solvent.     

Investigation of Structure and Mobility of Silica Bonded Stationary Acridine Phases by Means of Fluorescence and NMR Spectroscopy

Ligand mobility of silica-based HPLC stationary phases modified by various surface coverages of acridine-9-carboxy(N-aminoethylaminopropyl)amide ligands was investigated by fluorescence spectroscopy, time-resolved fluorescence anisotropy measurements, as well as solid-state ¹³C-CP/MAS- and ¹H-MAS-NMR spectroscopy. Rotational correlation times, τ_R, of the bound acridine fluorophore obtained from fluorescence anisotropy measurements are significantly longer in the bound phase, than in solution. Also, in time-resolved experiments anisotropies do not decay to zero. These results are interpreted in terms of wobble-in-cone ligand motion. The mobility of the fluorophore in the presence of liquid phase correlates strongly with the solubility of the model compound acridine-9-carboxy-n-butylamide in the same solvent. In the good solvent acetonitrile τ_R = 3.2 ns is found, whereas in methanol, τ_R > 80 ns is obtained. NMR measurements of the dry phase yield large linewidths, cross polarization constants, T_CH, and spin-lattice relaxation times, T_1ρH, shifting around the minimum in the correlation time curve. Both fluorescence and NMR data indicate medium to low ligand mobility. No difference in the mobilities of alkyl spacer and aromatic group is observed, probably due to the rigidity of the amide group.     

Conformation of random polypeptide chains

The characteristic ratios C_of and C_∞ are calculated for infinitely long poly(L-amino acid) chains having a? CH₂? R′ substitution at the C^α atom, as functions of the valence angle τ at the C^α atom. The value of C_of is found to increase from 1.6 to 2.3 as τ varies from 105 to 115°. On the other hand, C_∞ decreases with increasing τ in the range 105–111°, passes through a minimum at 111°, and then increases slowly. Since, C_∞ is much less dependent on τ in the most commonly observed range (110–115°), any variation in τ in this range due to solvent would not appreciably affect C_∞. The calculated temperature coefficient of C_∞ is negative and increases significantly in absolute magnitude even for small deviations in τ from 110°.     

EXCITATION TRANSFER BY CHLOROPHYLL a IN MONOLAYERS AND THE INTERACTION WITH CHLOROPLAST GLYCOLIPIDS*

In mixed monolayers with purified chloroplast glycolipids and other colorless lipids, chlorophyll a fluorescence exhibits a decrease in quantum efficiency with increasing chlorophyll concentration. The fluorescence, which is strongly polarized in dilute films, becomes progressively depolarized as the area fraction of chlorophyll increases, and it is completely depolarized in a pure chlorophyll a monolayer. The observed behavior is consistent with an inductive resonance mechanism of energy transfer among the chlorophyll molecules with a critical transfer distance of 20–90 Å, depending on the model chosen for the energy transfer mechanism. The purified glycolipids–mono-and digalactosyl diglycerides and sulfoquinovodiglyceride–separately form stable, compressible monolayers of the liquid-expanded type on an aqueous subphase and in an atompshere of nitrogen. At maximum compression the three glycolipids occupy areas of 55, 80 and 47 A²-molecule^-1, respectively, in the monolayer. Mixed monolayers of chlorophyll a with, separately, the monogalactolipid and the sulfolipid behave upon compression as two-dimensional solutions. The fluorescence polarization at high chlorophyll concentrations in mixed monolayers indicates that several of the lipid diluents facilitate local ordering of the pigment molecules.     

Excited state decay of tetrahalomanganese(ii) complexes

The excitation and emission spectra and decay times of several MnX^2-₄ (X = Cl^?, Br^?, 1^?) complexes of various tetraalkylammonium, -phosphonium, and -arsonium salts have been measured for the pure solids at 298°K and 77°K. High luminescence quantum yields (0.3-1.0) reveal that lifetimes fairly accurately reflect radiative decay rates. An impressive correlation exists between the lifetime, τ, of the ⁴T₁ (G) →⁶ A₁ emission and the ligand, X: for X = Cl^?, τ = 1.2 - 3,5 x 10^?3 sec; X = Br^?, τ = 0.35 - 0.43 X 10^?3 sec; X =l^?, τ = 0.036 – 0.055 X 10^?3 sec. We attribute this decreasing lifetime largely to the enhanced spin-orbital coupling associated with the heavier halide. We find that direct population of high energy charge-transfer (CT) states gives smaller emission yields than excitations in the ligand-field (LF) region.     

Period-four Modulation of Photosystem II Primary Quinone Acceptor (QA) Reduction/Oxidation Kinetics in Thylakoid Membranes

Photosystem II (PSII), a multiprotein complex mainly coded by the chloroplast genome in higher plants and algae, contains the oxygen-evolving complex with four manganese atoms responsible for the oxidation of water. After each absorption of a light quantum by pigment molecules in the light harvesting complexes of PSII, the Mn cluster advances in its oxidation states denoted from S₀ to S₄. The S₄ state decays to S₀ in the dark with the concurrent release of molecular oxygen. Therefore, the oxygen production in PSII exposed to successive single turnover excitations follows a period-four oscillation pattern. The intensity of chlorophyll a fluorescence of PSII is also known to be influenced by the oxidation state of the Mn cluster. In the present work, fluorescence induction kinetics was measured in isolated thylakoids with various initial S-state populations settled by preflashes. The shape of the fluorescence induction traces was strongly affected by preflashes. O-J and J-I phases of the induction followed a period-four oscillation pattern. The results indicate that these changes reflect the influence of the oxidation rate of the Mn cluster on the reduction/oxidation kinetics of the primary quinone acceptor (Q_A) of PSII.     

Kinetics of Coil-to-Globule Transition of Dansyl-Labeled Poly(N-isopropyl-acrylamide) Chains in Aqueous Solution

The coil-to-globule transition of thermally sensitive linear poly(N-isopropylacrylamide) (PNIPAM) labeled with dansyl group is induced by 1.54 μm laser pulses (width≈10 ns). The dansyl group is used to follow the transition kinetics because its fluorescence intensity is very sensitive to its micro-environment. As the molar ratio of NIPAM monomer to dansyl group increases from 110 to 300, the effect of covalently attached dansyl fluorophores on the transition decreases. In agreement with our previous study in which we used 8-anilino-1-naphthalensulfonic acid ammonium salt free in water as a fluorescent probe, the current study reveals that the transition has two distinct stages with two characteristic times, namely, τ_fast≈0.1 ms, which can be attributed to the nucleation and formation of some "pearls" (lo-cally contracting segments) on the chain, and τ_slow≈0.5 ms, which is related to the merging and coarsening of the "pearls". τ_fast is independent of the PNIPAM chain length over a wide range (M_w=2.8×10⁶-4.2×10⁷ g/mol). On the other hand, τ_slow only slightly increases with the chain length.     

Relaxation times in a system with exponential transition probabilities

Thermal vibrational relaxation times (τ) are calculated for two properly normalzed exponential transition probability models. If both threshold for reaction (E₀ and a constant related to average energy lost in a collision (γ) are assumed to be temperature independent, τ increases sharply with temperature. A simple analytic formula is derived for τ, valid for γ/kT < 1 and for E₀/kT ? 15.     

Correction for instrument time constant and baseline in determination of reaction kinetics

Rates of reactions can be expressed as dn/dt = kf(n), where n is moles of reaction, k is a rate constant, and f(n) is a function of the properties of the sample. Instrumental measurement of rates requires c(dn/dt) = ckf(n), where c is the proportionality constant between the measured variable and the rate of reaction. When the product of instrument time constant, τ, and k is ? 1, the reaction is much slower than the time response of the instrument and measured rates are unaffected by instrument response. When τ k < 1, = 1, or >1, the reaction rate and instrument response rate are sufficiently comparable that measured rates are significantly affected by instrument response and correction for instrument response must be done to obtain accurate reaction kinetics. This paper describes a method for simultaneous determination of τ, k, c, and instrument baseline by fitting equations describing the combined instrument response and rate law to rates observed as a function of time. When τ cannot be neglected, correction for instrument response has previously been done by truncating early data or by use of the Tian equation. Both methods can lead to significant errors that increase as τk increases. Inclusion of instrument baseline as a fitting parameter significantly reduced variability in k and c compared with use of measured instrument baselines. The method was tested with data on the heat rate from acid‐catalyzed hydrolysis of sucrose collected with three types of calorimeters. In addition, to demonstrate the generality of this method of data analysis, equations including τ, k, c, and instrument baseline are derived for the relation between the reaction rate and the observed rate for first order, second order (first in each reactant), nth order in one reactant, autocatalytic, Michaelis–Menten kinetics, and the Ng equation. © 2010 Wiley Periodicals, Inc. Int J Chem Kinet 43: 53–61, 2011     

Permeability,permselectivity, and penetrant‐induced plasticization in fluorinated polyimides studied by positron lifetime measurements

The ortho‐positronium (o‐Ps) lifetime τ₃ and its intensity I₃ in various fluorinated polyimides were determined by the positron annihilation technique and were studied with the spin–lattice relaxation time T₁ and the propylene permeability, solubility, diffusivity, and permselectivity for propylene/propane in them. τ₃, I₃, and the distribution of τ₃ changed when the bulky moieties in the polyimides were changed. The polyimides, having both large τ₃ and I₃ values, exhibited a short T₁ and a high permeability with a low permselectivity. The propylene permeability and diffusivity were exponentially correlated with the product of I₃ and the average free‐volume hole size estimated from τ₃. In highly plasticized states induced by the sorption of propylene, the permeability increased with the propylene pressure in excellent agreement with the change in the free‐volume hole properties probed by o‐Ps. The large and broad distribution of the free‐volume holes and increased local chain mobility for the 2,2‐bis(3,4‐decarboxyphenyl) hexafluoropropane dianhydride‐based polyimides are thought to be important physical properties for promoting penetrant‐induced plasticization. These results suggest that o‐Ps is a powerful probe of not only the free‐volume holes but also the corresponding permeation mechanism and penetrant‐induced plasticization phenomenon. © 2003 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 41: 308–318, 2003