Studies on the rate-limiting reaction of photosynthetic oxygen evolution in spinach chloroplasts

The modulated oxygen polarograph has been used to study the rate-determining steps of photosynthetic oxygen evolution in spinach chloroplasts. The rate constant, k, of the reaction has a value of 218±10 (S.E.) s⁻¹ at 23 °C and an activation energy of 7±2 (S.E.) kcal · mol⁻¹. A kinetic isotope experiment indicated that this step is probably not the water-splitting reaction. These findings resemble previous results with the unicellular alga Chlorella (Sinclair, J. and Arnason, T. (1974) Biochim. Biophys. Acta 368, 393–400). In other experiments we changed the pH, O₂ concentration and osmolarity of the medium, and treated the chloroplasts with 1 mM NH₄Cl without detecting any significant change in k. These results suggest that the step is irreversible. However, a significantly lower value of k, 110±20 (S.E.) s⁻¹ was obtained when all salts except 1 mM MgCl₂ were removed from the medium bathing the chloroplasts.     

Mechanistic study on the reaction of boronic acids with H-resorcinol

Kinetics for the reaction of m-nitrophenyl- (m-NO₂Ph-), phenyl- (Ph-), methyl(Me-), and n-butyl- (n-Bu-) boronic acids with H-resorcinol has been studied at various temperatures and pressures. The pseudo first-order rate constant (k_obs) exhibited saturation in the plot of k_obs against the total boronic acid concentration (C_B) for the reactions of m-NO₂PhB(OH)₂ and PhB(OH)₂, while the plot was linear for the reactions of MeB(OH)₂ and n-BuB(OH)₂, which leads to the following equations, respectively: k_obs=k*KC_B/(1+KC_B) and k_obs=k*KC_B=k_fC_B. It was experimentally shown that the reaction of boronic acids with H-resorcinol proceeds with rate determining second chelate-ring closure.     

Biotransformation of glucose to gluconic acid by Aspergillus niger—study of mass transfer in an airlift bioreactor

A glucose–gluconic acid biotransformation system was suggested for the experimental study of oxygen transfer in bioreactors. This biosystem was used for the investigation of the effect of the flow rate and biomass concentration on the volumetric oxygen transfer coefficient k_La in a 10 dm³ internal-loop airlift bioreactor. For this purpose, the fermentation broth of the mycelial strain Aspergillus niger was employed, representing a three-phase system, where bubbles come into contact with dense rigid pellets. The results showed that the presented biotransformation system can be successfully utilised for the determination of the oxygen transfer rate in airlift bioreactors. The experiments showed a strong positive influence of the air flow rate on the rate (r_Glu), specific rate of gluconic acid production (k_Glu/X) as well as on the volumetric oxygen transfer coefficient (k_La). This confirmed an expected limitation of production rate by the oxygen transport from the gas to the liquid phase in the whole range of air flow rates applied. Moreover, consistent curves of the production rate r_Glu and k_La values vs. biomass concentration c_X (amount of enzymes) were observed. These exhibited a local maximum for c_X equal to 6.68 g dm⁻³. On the other hand, the specific production rate monotonously decreased with increasing biomass concentration. A decline of k_La values at higher c_X values was attributed to a bubble coalescence promoting effect of mycelial pellets.     

The migratory insertion of carbon monoxide in pentamethylcyclopentadienyliridium (III) complexes. Structural effects on reactivity and mechanism for rhodium and iridium systems

The reactions of complex (C₅Me₅)Ir(Cl) (CO) (Me) (1a) with cyclohexylisocyanide and phosphines (L=CyNC, PHPh₂, PMePh₂, PMe₂Ph) give the products of alkyl migratory insertion (C₅Me₅Ir(Cl) (COMe) (L), in toluence or tetrahydrofuran at 323 K or higher temperature. The phenyl analogue (C₅Me₅)Ir(Cl)(CO)(Ph) or the iodide complexes (C₅Me₅)Ir(I) (CO) (R) (R=Me, Ph_are not reactive under the same conditions. The reaction of (C₅Me₅)Ir(Cl)(CO)(Me) with PMePh₂ and PMe₂Ph in acetonitrile yields the chloride substitution product [(C₅Me₅)Ir(CO)(L)(Me)]⁺Cl⁻. Kinetic measurements for the reactions of (C₅Me₅)Ir(Cl)(CO)(Me) in toluene are first order in the iridium complex and exhibit a saturation dependence on the incoming donors L. Analysis of the data suggests a two-step process involving (i) rapid formation of a molecular complex [(C₅Me₅)Ir(Cl)(CO)(Me), (L)], in which the structure of 1a is unperturbed within the limits of spectroscopic analysis, and (ii) rate determining methyl migration. The reaction parameters are K for the pre-equilibrium step (K = 1.5 (CyNC), 7.3 (PHPh₂), 7.1 (PMePh₂) dm³ mol⁻¹ at 323 K) and k₂ for the slow carbon---carbon bond formation (k₂ (10⁵) = 6.9 (CyNC), 1.2 (PHPh₂), 1.0 (PMePh₂) s⁻¹ at 323 K). The activation parameters for the methyl migration step in the reaction with PMePh₂ obtained between 308 and 338 K, are ΔH^≠ = 106±16 kJ mol⁻¹ and ΔS^≠ = − 14±5 J K⁻¹ mol⁻¹. The reaction of 1a with PMePh₂ proceeds at similar rates in tetrahydrofuran (K = 3.7 dm³ mol⁻¹, k₂ (10⁵) = 1.2 s⁻¹, 323 K). The crystal structure of (C₅Me₅)Ir(Cl)(COMe) (PMe₂Ph) has been determined by X-ray diffraction. C₂₀H₂₉ClOPIr: M_r = 544.1, monoclinic, P2₁/n, A = 8.084 (2), B = 9.030(2), C = 28.715 (3) Å, β = 91.41 (3)°, Z = 4, D_c = 1.71 g cm⁻³, V = 2095.5 ų, room temperatyre, Mo K, γ = 0.71069, μ = 65.55 cm⁻¹, F(000) = 1044, R = 0.037 for 2453 independent observed reflections. The complex shows a deformed tetrahedral coordination assuming the η⁵-C₅Me₅ molecular fragment as a single coordination site. The iridium-chlorine bond is staggered with respect to two adjacent C(ring)-methyl bonds, while the Ir---P and the Ir---COMe bonds are eclipsed with respect to C(ring)-methyl bonds.     

Algorithm for application of Fourier analysis for biorhythmic baselines of pharmacodynamic indirect response models

The change of an indirect pharmacological response R(t) can be described by a periodic time-dependent production rate k_in (t) and a first-order loss constant k_out. If k_in(t) follows some biological rhythm (e.g., circadian), then the response R(t) also displays a periodic behavior. A new approach for describing the input function in indirect response models with biorhythmic baselines of physiologic substances is introduced. The present approach uses the baseline (placebo) response R_b(t) to recover the equation for k_in(t). Fourier analysis provides an approximate equation for R_b(t) that consists of terms (usually two or three) of the Fourier series (harmonics) that contribute most to the overall sum. The model differential equation is solved backward for k_in(t), yielding the equation involving R_b(t). A computer program was developed to perform the square L²-norm approximation technique. Fourier analysis was also performed based on nonlinear regression. Cortisol suppression after inhalation of fluticasone propionate (FP) was modeled based on the inhibition of the secretion rate k_in(t) using ADAPT II. The pharmacodynamic parameters k_out and IC₅₀ were estimated from the model equation with k_in(t) derived by the new approach. The proposed method of describing the input function needs no assumption about the behavior of k_in(t), is as efficient as methods used previously, and is more flexible in describing the baseline data than the nonlinear regression method. (Chronobiology International, 17(1), 77-93, 2000)     

Specificity and kinetic parameters of recombinant Microdochium nivale carbohydrate oxidase

A new carbohydrate oxidase from Microdochium nivale heterologously expressed in Aspergillus oryzae (rMnO) has been characterized. The carbohydrate oxidase is a flavoenzyme which oxidizes glucose and other mono- or oligosaccharides. It shows a broad substrate specificity towards carbohydrates reacting with aldoses in the 1-position. The rMnO oxidizes the β-form of -glucose, and the product of -glucose oxidation is -gluconic acid.

The mechanism of carbohydrate oxidation by oxygen and artificial electron acceptors has been described by a ping-pong scheme. Compared to Aspergillus niger glucose oxidase (GOx) the reactivity of rMnO at pH 7.0 is significantly lower; k_cat is 20, k_ox 11 and k_red 22 times less, using oxygen as electron acceptor. Also with other two electron acceptors, like DPIP, the activity is low. However, compared to oxygen the rMnO shows 2–10 times higher activity towards some artificial single electron acceptors (AAs). The enzyme activity increases at higher ionic strength of the solution, if positively-charged AAs are used.

The high activity towards AAs and low rate for oxygen as well as broad specificity to carbohydrates indicates that rMnO may have some advantages compared to the most used GOx in connection with enzyme use for analytical devices and for biotechnological purposes.     

The respiratory chain of Azotobacter vinelandii II. The effect of cyanide on cytochrome d

1. Cyanide causes a slow disappearance of the oxidized band (648 nm) of cytochrome d in particles of Azotobacter vinelandii and inhibits the appearance of the reduced band (631 nm). No effect of cyanide is found on the reduced band of cytochrome d.

2. The kinetics of the disappearance of the 648-nm band of cytochrome d with excess cyanide deviates from first-order kinetics at lower temperatures (22 °C) indicating that at least two conformations of the enzyme are involved. At higher temperatures (32 °C) the observed kinetics of the cyanide reaction are first order with a k_on = 0.7 M⁻¹·s⁻¹ and with an estimated k_off of approximately 5·10⁻⁵ s⁻¹.

3. The value of the k_off (7·10⁻⁴−14·10⁻⁴ s⁻¹ at 32 °C) determined from the rate of reduction of cyanocytochrome d by Na₂S₂O₄ or NADH is one order of magnitude larger than the k_off value found when the enzyme is in its oxidized state.

4. No effect of cyanide is found on the spectrum of cytochrome a₁.     

Kinetics of ligand substitution in bis(N-alkylsalicylaldiminato)nickel(II) complexes: A search for kinetically-effective aromatic ring stacking

The square-planar bis chelate complexes Ni(R-sal)₂ (= bis(N-alkyl)salicylaldiminato)nickel(II)) with R = (CH₂)₂Ph (I; Ph = phenyl), (CH₂)₃Ph (II), (CH₂)₄Ph (III) and (CH₂)₂(4-hydroxyphenyl) (IV) were prepared and characterized. ComplexesII and III meet the steric requirements for intramolecular aromatic ring stacking. Stopped-flow spectrophotometry was used to study the kinetics of ligand substitution in complexesI–IV by H₂salen (=N,N′-disalicylidene-ethylenediamine) in acetone. For the substitution of the two bidentate ligands in Ni(R-sal)₂ only one step is kinetically observed which follows a second-order rate law, rate =k[H₂salen] [Ni(R-sal)₂], with k = 43.4 (I), 64.0 (II), 87.0 (III) and 49.5 (IV) M⁻¹ s⁻¹ at 298 K. It is found, therefore, that the size of k does not change significantly upon lengthening of the alkane chain in Ni(Ph(CH₂)_nsal)₂ from n = 2 to 4 and that there is no kinetic evidence for intramolecular stacking interactions. The equilibrium constants and thermodynamic parameters for the formation of the bis adductsIII·(py)₂ and III·(MeOH)₂ in acetone are reported.     

Trapping, loss and annihilation of excitations in a photosynthetic system: II. Experiments with the purple bacteria Rhodospirillum rubrum and Rhodopseudomonas capsulata

In this paper a number of experiments with the purple bacteria Rhodospirillum rubrum and Rhodopseudomonas capsulata is described in which the total fluorescence yield and/or the total fraction of reaction centers closed after a picosecond laser pulse were measured as a function of the pulse intensity. The conditions were such that the reaction centers were either all in the open or all in the closed state before the pulse arrived. These experiments are analysed using the theoretical formalism discussed in the preceding paper (Den Hollander, W.T.F., Bakker J.G.C., and Van Grondelle, R., Biochim. Biophys. Acta 725, 492–507). From the experimental results the number of connected photosynthetic units, λ, the rate of energy transfer between neighboring antenna molecules, k_h, and the rate of trapping by an open reaction center, k^o_t, can be estimated. For R. rubrum it is found that λ = 14−17, k_h = (1−2)·10¹² s⁻¹ and k^o_t = (4−6)·10¹¹ s⁻¹, for Rps. capsulata λ ≈ 30, k_h ≈ 4·10¹¹ s⁻¹ and k^o_t ≈ 3·10¹¹ s⁻¹. The findings are discussed in terms of current models for the structure of the antenna and the kinetic properties of the decay processes occurring in these purple bacteria.     

One-electron reduction of selenomethionine oxide

Experimental evidence is provided that selenomethionine oxide (MetSeO) is more readily reducible than its sulfur analogue, methionine sulfoxide (MetSO). Pulse radiolysis experiments reveal an efficient reaction of MetSeO with one-electron reductants, such as e^-_aq (k = 1.2 × 10¹⁰M^-1s^-1), CO^·-₂ (k = 5.9 × 10⁸ M^-1s^-1) and (CH₃)₂) C^·OH (k = 3.5 × 10⁷M^-1s^-1), forming an intermediate selenium-nitrogen coupled zwitterionic radical with the positive charge at an intramolecularly formed Se_∴ N 2σ/1σ* three-electron bond, which is characterized by an optical absorption with λ_max at 375 nm, and a half-life of about 70 μs. The same transient is generated upon HO^· radical-induced one-electron oxidation of selenomethionine (MetSe). This radical thus constitutes the redox intermediate between the two oxidation states, MetSeO and MetSe. Time-resolved optical data further indicate sulfur-selenium interactions between the Se_∴ N transient and GSH. The Se_∴ N transient appears to play a key role in the reduction of selenomethionine oxide by glutathione.     

Enantioselective cleavage of activated amino acid esters promoted by chiral palladacycles

The ester cleavage of R- and S-isomers N-CBZ-leucine p-nitrophenyl ester intermolecularly catalyzed by R- (a) and S-stereoisomers (b) of the Pd(II) metallacycle [Pd(C₆H₄C^*HMeNMe₂)Cl(py)] (3) follows the rate expression k_obs = k_o + k_cat [3], where the rate constants k_cat equal 25.8 ± 0.4 and 7.6 ± 0.5 dm³ mol⁻¹ s⁻¹ for the S- and R-ester, respectively, in the case of 3a, but are 5.7 ± 0.6 and 26.7 ± 0.5 dm³ mol⁻¹ s⁻¹ for the S- and R-ester, respectively, in the case of 3b (pH 6.23 and 25°C). Thus, the best catalysis occurs when the asymmetric carbons of the leucine ester and Pd(II) complex are R and S, or S and R configured, respectively. Molecular modeling suggests that the stereoselection results from the spatial interaction between the CH₂CHMe₂ radical of the ester and the -methyl group of 3. A hydrophobic/stacking contact between the leaving 4-nitrophenolate and the coordinated pyridine also seems to play a role. Less efficient intramolecular enantioselection was observed for the hydrolysis of N-t-BOC-S-metthionine p-nitrophenyl ester with R- and S-enantiomers of [Pd(C₆H₄C*HMeNMe₂)Cl] coordinated to sulfur.     

Activation by reduction of the resting form of cytochrome c oxidase: Tests of different models and evidence for the involvement of CuB

(1) The reaction of the resting form of oxidised cytochrome c oxidase from ox heart with dithionite has been studied in the presence and absence of cyanide. In both cases, cytochrome a reduction in 0.1 M phosphate (pH 7) occurs at a rate of 8.2 · 10⁴ M⁻¹ · s⁻¹. In the absence of cyanide, ferrocytochrome a₃ appears at a rate (k_obs) of 0.016 s⁻¹. Ferricytochrome a₃ maintains its 418 nm Soret maximum until reduced. The rate of a₃ reduction is independent of dithionite concentration over a range 0.9 mM–131 mM. In the presence or cyanide, visible and EPR spectral changes indicate the formation of a ferric a₃/cyanide complex occurs at the same rate as a₃ reduction in the absence of cyanide. A g = 3.6 signal appears at the same time as the decay of a g = 6 signal. No EPR signals which could be attributed to copper in any significant amounts could be detected after dithionite addition, either in the presence or absence of cyanide. (2) Addition of dithionite to cytochrome oxidase at various times following induction of turnover with ascorbate/TMPD, results in a biphasic reduction of cytochrome a₃ with an increasing proportion of the fast phase of reduction occurring after longer turnover times. At the same time, the predominant steady state species of ferri-cytochrome a₃ shifts from high to low spin and the steady-state level of reduction of cytochrome a drops indicating a shift in population of the enzyme molecules to a species with fast turnover. In the final activated form, oxygen is not required for fast internal electron transfer to cytochrome a₃. In addition, oxygen does not induce further electron uptake in samples of resting cytochrome oxidase reduced under anaerobic conditions in the presence of cyanide. Both findings are contrary to predictions of certain O-loop types of mechanism for proton translocation. (3) A measurement of electron entry into the resting form of cytochrome oxidase in the presence of cyanide, using TMPD or cytochrome c under anaerobic conditions, shows that three electrons per oxidase enter below a redox potential of around +200 mV. An initial fast entry of two electrons is followed by a slow (k_obs ≈ 0.02 s) entry of a third electron. Above +200 mV, the number of electrons taken up in the initial fast phase drops as a redox center (presumably Cu_A) titrates with an apparent mid-point potential of +240 mV. The slow phase of reduction remains at the more positive redox values. (4) The results are interpreted in terms of an initial fast reduction of cytochrome a (and Cu_A at redox values more negative than +240 mV) followed by a slow reduction of Cu_B. Cu_B reduction is proposed to spin-uncouple cytochrome a₃ to form a cyanide sensitive center, and trigger a conformational change to an activated form of the enzyme with faster intramolecular electron transfer.     

Kinetics and equilibria of Ga(III)---thiocyanate complex formation. Mechanism of ligand substitution reactions of Ga(III) in aqueous solution

The kinetics and equilibria of complex formation by Ga(III) with NCS⁻ in aqueous solution have been measured over a range of acidities and temperatures, the contributing paths to the reaction resolved, and their rate constants and activation parameters determined. The hydrolysis equilibria required to carry out this resolution of kinetic behaviour have also been measured.

Unlike the other reported complexation reactions of Ga(III) in aqueous solution, the separate reaction pathways can be assigned with no ambiguity. At 25 °C and ionic strength 0.5 M, the observed forward rate constant for the complex formation is described by {k₁ + k₂K_1h/[H⁺] + k₃K_1hK_2h/[H⁺]²} M⁻¹ s⁻¹. For these conditions, the first and second successive hydrolysis constants of Ga(H₂O)₆³⁺ are given by pK_1h = 3.69 ± 0.01 and pK_2h = 3.74 ± 0.04. The rate constants corresponding to the reactions of the species Ga(H₂O)₆³⁺, Ga(H₂O)₅(OH)²⁺ and Ga(H₂O)₄(OH)₂⁺ with NCS⁻ are k₁ = 57 ± 4 M^{−1 −1}, k₂ = (1.08 ± 0.01) × 10⁵ M⁻¹ s⁻¹ and k₃ = 3 × 10⁶ M⁻¹ s⁻¹ respectively. The complexation equilibrium quotient [GaNCS²⁺]/([Ga³⁺][NCS⁻]) has been independently determined by spectrophotometric titration to be 20.8 ± 0.3 M⁻¹ at 25 °C and ionic strength 0.5 M.

These kinetic results lead to an interpretation of the data, and a reinterpretation of other data for aquo-Ga(III) complex formation kinetics from the literature which support the assignment of a dissociative interchange mechanism for these reactions rather than the associative activation mode sometimes proposed.     

PATHWAYS OF INCORPORATION OF FATTY ACID INTO GLYCEROLIPIDS OF THE MURINE PERIPHERAL NERVOUS SYSTEM IN VIVO: ALTERATIONS IN THE DYSMYELINATING MUTANT TREMBLER MOUSE

In vivo glycerolipid metabolism was studied in sciatic nerves of normal and Trembler mice. The results showed that two kinetically independent pathways were implicated in the labeling of diacylglycerophospholipids from [³H]palmitate: the Kennedy pathway and a ‘direct acylation’ pathway. In normal nerves, 45% of the glycerophospholipids were labeled, with a rate constant k₃ = 3.9 × 10⁻³ min⁻¹, from phosphatidic acid and diacylglycerol intermediates, themselves formed with a rate constant of k₁ = 0.24 min⁻¹ from a free ³H-fatty acid pool, FFA₁, that represents 45% of the total injected label. The remaining 55% of the glycerophospholipids were labeled from a kinetically distinct free ³H-fatty acid pool, FFA₂, with a rate constant of k₄ = 9.8 × 10⁻², via a process that does not implicate a detectably labeled metabolic intermediate (‘direct acylation’). Glycerophospholipid labeling via the Kennedy pathway in the Trembler mouse sciatic nerves was reduced to 75% of the normal level, while labeling via the ‘direct acylation’ pathway was increased 1.4-fold. The values of the rate constants for free ³H-fatty acid utilisation (k₁ and k₄) were both increased about 2.5-fold, while that of glycerophospholipid formation from diacylglycerol (k₃) was close to normal. Copyright © 1996 Elsevier Science Ltd     

Study of the thermal stability of D-amino acid oxidase from Trigonopsis variabilis reveals enzyme inactivation via multiple steps

The thermal stability of a highly purified preparation of D-amino acid oxidase from Trigonopsis variabilis (TvDAO), which does not show microheterogeneity due to the partial oxidation of Cys-108, was studied based on dependence of temperature (20-60°C) and protein concentration (5-100 µmol L^-1). The time courses of loss of enzyme activity in 100 mmol L^-1 potassium phosphate buffer, pH 8.0, are well described by a formal kinetic mechanism in which two parallel denaturation processes, partial thermal unfolding and dissociation of the FAD cofactor, combine to yield the overall inactivation rate. Estimates from global fitting of the data revealed that the first-order rate constant of the unfolding reaction (k _a) increased 10⁴-fold in response to an increase in temperature from 20 to 60°C. The rate constants of FAD release (k _b) and binding (k _-b) as well as the irreversible aggregation of the apo-enzyme (k _agg) were less sensitive to changes in temperature, their activation energy (E _a) being about 52 kJ mol^-1 in comparison with an E _a value of 185 kJ mol^-1 for k _a. The rate-determining step of TvDAO inactivation switched from FAD dissociation to unfolding at high temperatures. The model adequately described the effect of protein concentration on inactivation kinetics. Its predictions regarding the extent of FAD release and aggregation during thermal denaturation were confirmed by experiments. TvDAO is shown to contain two highly reactive cysteines per protein subunit whose modification with 5,5'-dithio-bis (2-nitrobenzoic acid) was accompanied by inactivation. Dithiothreitol (1 mmol L^-1) enhanced up to 10-fold the recovery of enzyme activity during ion exchange chromatography of technical-grade TvDAO. However, it did not stabilize TvDAO at all temperatures and protein concentrations, suggesting that deactivation of cysteines was not responsible for thermal denaturation.     

Thermogravimetric study of starch derivatives with amine/ammonium ion-exchanging groups in oxidative environment

The amine/ammonium materials were prepared by cross-linking of starch (S) with epichlorohydrin (E→SE) in the presence of ammonia (A→SEA) or choline (C→SEC–HO⁻) or with 1,3-bis-(3-chloro-2-hydroxypropyl)imidazolium hydrogensulphate (BCHIHS→SHI–HO⁻) and transfered into the acid/salt forms with HCl (SEA–HCl, SEC–Cl⁻, or SHI–Cl⁻), H₂SO₄ (SEA–H₂SO₄, SEC–HSO₄⁻, or SHI–HSO₄⁻), and H₃PO₄ (SEA–H₃PO₄, SEC–H₂PO₄⁻, or SHI–H₂PO₄⁻) and analyzed with thermogravimetry (TG) under dynamic and isothermal conditions in nitrogen or oxygen environment. According to the values of thermooxidation maxima (TM) calculated from the maximal difference of measured residues on the dynamic TG curves run in nitrogen and oxygen environments the order of decreasing thermooxidation resistance is: S>SEA–H₃PO₄>SHI–H₂PO₄⁻>SHI–HSO₄⁻>SEA–H₂SO₄>SEC–HSO₄⁻>SEC–HO⁻>SEA–HCl>HCl–Cl⁻> SEC–Cl⁻>SHI–HO⁻>SEA>SEC–H₂PO₄⁻>SE. The first-order rate constants calculated by the linear regression method (regression coefficient R>0.95) represented the initial rate constants for residue formation (k_r's) and gasification (k_g's). All the derivatives had greater values of rate constants than S and the k_g's were about 1000 times greater than k_r's. The values obtained in nitrogen were smaller than those calculated from runs in oxygen environment with the exception of S. Most of the salt forms had greater values of k_g's in oxygen environment. The activation energies (E's) were usually greater in nitrogen than in oxygen as well as for residue formation than for gasification. The SHI–HO⁻ sample had high k_g's and low E_g's in oxygen environment while for SHI–H₂SO₄⁻ the opposite was true. This we consider as two extremes for labile and resistant samples for gasification.     

The Ene Reaction of Singlet Oxygen with Olefins

The reactions of singlet oxygen (¹O₂) with cis and trans butenes-1,1,1-d₃, at—80°C in Freon-11, show a product isotope effect (k_H/k_D) of 1.38 and 1.25 respectively. Isomerization of the starting materials or formation of dioxetanes were not observed during the course of the photooxygenation. Together with the isotope effects on the reactions of tetramethylethylene-d₆ isomers with singlet oxygen, these results require the reversible formation of a perepoxide or charge transfer intermediate.     

Radical-scavenging activity of butylated hydroxytoluene (BHT) and its metabolites

To clarify the radical-scavenging activity of butylated hydroxytoluene (BHT), a food additive, stoichiometric factors (n) and inhibition rate constants (k_inh) were determined for 2,6-di-tert-butyl-4-methylphenol (BHT) and its metabolites 2,6-di-tert-butyl-p-benzoquinone (BHT-Q), 3,5-di-tert-butyl-4-hydroxybenzaldehyde (BHA-CHO) and 3,5-di-tert-butyl-4-hydroperoxy-4-methyl-2,5-cyclohexadiene-1-one (BHT-OOH). Values of n and k_inh were determined from differential scanning calorimetry (DSC) monitoring of the polymerization of methyl methacrylate (MMA) initiated by 2,2′-azobis(isobutyronitrile) (AIBN) or benzoyl peroxide (BPO) at 70 °C in the presence or absence of antioxidants (BHT-related compounds). The n values declined in the order BHT (1–2) > BHT-CHO, BHT-OOH (0.1–0.3) > BHT-Q (0). The n value for BHT with AIBN was approximately 1.0, suggesting dimerization of BHT. The k_inh values declined in the order BHT-Q ((3.5–4.6)×10⁴ M⁻¹ s⁻¹) > BHT-OOH (0.7–1.9×10⁴ M⁻¹ s⁻¹) > BHT-CHO ((0.4–1.7)×10⁴ M⁻¹ s⁻¹) > BHT ((0.1–0.2)×10⁴ M⁻¹ s⁻¹). The k_inh for metabolites was greater than that for the parent BHT. Growing MMA radicals initiated by BPO were suppressed much more efficiently by BHT or BHT-Q compared with those initiated by AIBN. BHT was effective as a chain-breaking antioxidant.     

Photogeneration of singlet oxygen (1O2) and free radicals (Sen·-, O·-2) by tetra-brominated hypocrellin B derivative

To improve photodynamic activity of the parent hypocrellin B (HB), a tetra-brominated HB derivative (compound 1) was synthesized in high yield. Compared with HB, compound 1 has enhanced red absorption and high molar extinction coefficients. The photodynamic action of compound 1, especially the generation mechanism and efficiencies of active species (Sen^·-, O^·-₂ and ¹O₂) were studied using electron paramagnetic resonance (EPR) and spectrophotometric methods. In the deoxygenated DMSO solution of compound 1, the semiquinone anion radical of compound 1 is photogenerated via the self-electron transfer between the excited and ground state species. The presence of electron donor significantly promotes the reduction of compound 1. When oxygen is present, superoxide anion radical (O^·-₂) is formed via the electron transfer from Sens^·- to the ground state molecular oxygen. The efficiencies of Sens^·- and O^·-₂ generation by compound 1 are about three and two times as much as that of HB, respectively. Singlet oxygen (¹O₂) can be produced via the energy transfer from triplet compound 1 to ground state oxygen molecules. The quantum yield of singlet oxygen (¹O₂) is 0.54 in CHCl₃ similar to that of HB. Furthermore, it was found that the accumulation of Sens^·- would replace that of O^·-₂ or ¹O₂ with the depletion of oxygen in the sealed system.     

Enantioselectivity of resolved Δ and Λ orthoruthenated 2-phenylpyridine complexes [Ru(o-C6H4-2-py)(LL)2]PF6 (LL = bpy and phen) toward glucose oxidase

Cyclometalated 2-phenylpyridine complexes [Ru^II(o-C₆H₄-2-py)(LL)₂]PF₆, LL = 2,2′-bipyridine (1) and 1,10-phenanthroline (2) were resolved into Δ and Λ enantiomers using column chromatography on SP Sephadex C-25 in the presence of (+)-2,3-dibenzoyl-D-tartrate. The absolute configuration of enantiomers was established using circular dichroism spectroscopy. The rate constants k_et for the electron transfer from reduced glucose oxidase (GO from Aspergillus niger) and PQQ-dependent glucose dehydrogenase (GDH) at the generated Ru^III species were measured by cyclic voltammetry and UV–vis spectroscopy. The electron transfer shows enantioselectivity. In the case of GO, the bell-shaped pH profile for the ratio k_Λ/k_Δ has a maximum at pH 7 (k_Λ/k_Δ equals 3.4 and 3.9 for 1 and 2, respectively), but its inversion is observed at pH around 5 and 9. The k_Λ/k_Δ ratio equals 2.0 for 2 and GDH at pH 7. The results of theoretical modeling of biological electron transfer for GO using functional docking Monte-Carlo simulations are presented and analyzed together with the experimental observations.