The versatile coordination chemistry of tricyanoethenolate, [(NC)2C=C(CN)O]

The reaction of meso-tetrakis (4-dimethoxyphenyl) porphinatomanganese(II), MnTP_OMeP, with TCNE (TCNE = tetracyanoethylene) leads to the formation of [MnTP_OMeP]⁺ [TCNE]⁻ and [MnTP_OMeP]⁺[OC(CN)C(CN)₂]⁻. The single-crystal X-ray structures of the latter as well as [Cu(bipy)₂Cl]⁺ [OC(CN)C(CN)₂]⁻ were determined. The former has a disordered [OC(CN)C(CN)₂]⁻ bridging via C and O between a pair of Mn^III sites, whereas the latter has an isolated [OC(CN)C(CN)₂]⁻ unbound to Cu^II. The IR characterization for μ₂-C,O bound [OC(CN)C(CN)₂]⁻ is at 2219m and 2196s (νCN) cm⁻¹ and at 1558s (νCO) cm⁻¹ while for unbound [OC(CN)C(CN)₂]⁻ it is at 2210m, 2203m, 2181m (νCN) cm⁻¹ and at 1583s (νCO) cm⁻¹.     

Pigment content and molar extinction coefficients of photochemical reaction centers from Rhodopseudomonas spheroides

Reaction center particles isolated from carotenoidless mutant Rhodopseudomonas spheroides were studied with the aim of determining the pigment composition and the molar extinction coefficients.

Two independent sets of measurements using a variety of methods show that a sample with A_{800 nm} = 1.00 contains 20.8 ± 0.8 μM tetrapyrrole and that the ratio of bacteriochlorophyll to bacteriopheophytin is 2:1.

Measurements were made of the absorption changes attending the oxidation of cytochrome c coupled to reduction of the photooxidized primary electron donor in reaction centers, using laser flash excitation. The ratio of the absorption change at 865 nm (due to the bleaching of P870) to that at 550 nm (oxidation of cytochrome) was found to be 5.77.

These results, combined with other data, yield a pigment composition of 4 bacteriochlorophyll and 2 bacteriopheophytin molecules in a reaction center. Based on this choice, extinction coefficients are determined for the 802- and 865-nm bands: _{802 nm} = 288 (± 14) mM⁻¹ · cm⁻¹ and _{865 nm} = 128 (± 6) mM⁻¹ · cm⁻¹. For reversible bleaching of the 865-nm band, Δ^{red - ox}_865nm = 112 (± 6) mM⁻¹ · cm⁻¹ (referred to the molarity of reaction centers). Earlier reported values of photochemical quantum efficiency are recomputed, and the revised values are shown to be compatible with those obtained from measurements of fluorescence transients.     

Light-induced absorbance changes in chloroplasts mediated by Photosystem I and Photosystem II at low temperature

Stable light-induced absorbance changes in chloroplasts at −196 °C were measured across the visible spectrum from 370 to 730 nm in an effort to find previously undiscovered absorbance changes that could be related to the primary photochemical activity of Photosystem I or Photosystem II. A Photosystem I mediated absorbance increase of a band at 690 nm and a Photosystem II mediated absorbance increase of a band at 683 nm were found. The 690-nm change accompanied the oxidation of P₇₀₀ and the 683-nm increase accompanied the reduction of C-550. No Soret band was detected for P₇₀₀.

A specific effort was made to measure the difference spectrum for the photooxidation of P₆₈₀ under conditions (chloroplasts frozen to −196 °C in the presence of ferricyanide) where a stable, Photosystem II mediated EPR signal, attributed to P₆₈₀⁺ has been reported. The difference spectra, however, did not show that P₆₈₀⁺ was stable at −196 °C under any conditions tested. Absorbance measurements induced by saturating flashes at −196 °C (in the presence or absence of ferricyanide) indicated that all of the P₆₈₀⁺ formed by the flash was reduced in the dark either by a secondary electron donor or by a backreaction with the primary electron acceptor. We conclude that P₆₈₀⁺ is not stable in the dark at −196 °C: if the normal secondary donor at −196 °C is oxidized by ferricyanide prior to freezing, P₆₈₀⁺ will oxidize other substances.     

Phosphorescence of adenosine and poly(riboadenylic acid). Evidence for dual emissions and n pi states

Phosphorescence from the 9-adenylyl group in the fom of microcrystalline powders of adenosine films of poly(riboadenylic acid) (poly(rA)) in hyaluronic acid has been studied at 77 K. For adenosine, clearly resolved vibronic structure consists of two progressions, A and B, with _A ‡ 1363 cm⁻¹ and _B ‡ 1575 cm ⁻¹, correlated with in-plane C₅-N₇ and in-plane C₄-C₅ stretch, respectively. The relative strength of the progressions varies with excitation wavelength and this, together with the absence of a common origin, indicates the existence of two independent emitting states with 0-0' levels separated by either 300 or 1000 cm⁻¹. Two different excitation spectra are observed lying below the normal (ππ*) adsorption and one is assigned as a previously undetected ¹(nπ*) transition. For poly(rA) films the emission band envelope is identical with that of adenosine but the vibronic structure is lost. Only one excitation peak is observed at 32.9×10³ cm⁻¹, identical with one of the adenosine spectra. The second adenosine excitation spectrum probably represents an intermolecular charge transfer transition. Comparison is made with the predictions of six semi-empirical MO calculations.     

High resolution spectroscopy of the lowest excited state in Re tetra-carbonyl complexes

The luminescence and absorption properties of [Re(bpy)(CO)₄](PF₆) and [Re(phen)(CO)₄](PF₆) are consistent with representation of the lowest excited states as nominally ³LC with an admixture of ¹CT character. Using high resolution spectroscopic techniques at cryogenic temperatures, such as luminescence line narrowing spectroscopy or spectroscopy in single crystals, the vibrational sideband information which is normally lost in the ‘natural’ solution environment can be observed in the luminescence and absorption spectra. Mixing between the ³LC and ¹CT excitation (3%) has previously been reported in [Re(bpy)(CO)₄](PF₆), resulting in metal-ligand sidebands at 184 and 198 cm⁻¹ in the absorption spectrum and a short luminescence lifetime (33.0 μs). In the luminescence spectra (line narrowed) the metal-ligand sidebands are observed at 194 cm⁻¹. Weak mixing ( 1%) of the ¹CT excitation (32 100 cm⁻¹) with the ³LC excitation (22 100 cm⁻¹) in [Re(phen)(CO)₄](PF₆) gives rise to the observation of metal-ligand vibrational sidebands in the luminescence spectrum (204 cm⁻¹) and a luminescence lifetime of τ= 295±5 μs at 20 K. A spin-orbit mixing matrix element of ³LC|H_so|¹CT for [Re(phen)(CO)₄](PF₆) of 65 cm⁻¹ is calculated, compared to 261 cm⁻¹ in [Re(bpy)(CO)₄](PF₆).     

A convenient electrochemical preparation of reduced methyl viologen and a kinetic study of the reaction with oxygen using an anaerobic stopped-flow apparatus

1. Single reduced methyl viologen (MV^.+) acts as an electron donor in a number of enzyme systems. The large changes in extinction coefficient upon oxidation (λ_max 600 nm; MV^.+, = 1.3 · 10⁴ M⁻¹ · cm⁻¹; oxidised form of methyl viologen (MV²⁺), = 0.0) make it ideally suited to kinetic studies of electron transfer reactions using stopped-flow and standard spectrophotometric techniques.

2. A convenient electrochemical preparation of large amounts of MV^.+ has been developed.

3. A commercial stopped-flow apparatus was modified in order to obtain a high degree of anaerobicity.

4. The reaction of MV^.+ with O₂ produced H₂O₂ (k > 5 · 10⁶ M⁻¹ · s⁻¹, pH 7.5, 25 °C). H₂O₂ subsequently reacted with excess MV^.+ (k = 2.3 · 10³ M⁻¹ · s⁻¹, pH 7.5, 25 °C) to produce water. The kinetics of this reaction were complex and have only been interpreted over a limited range of concentrations.

5. The results support the theory that the herbicidal action of methyl viologen (Paraquat, Gramoxone) is due to H₂O₂ (or radicals derived from H₂O₂) induced damage of plant cell membrane.     

Structure, physical, chemical and photophysical properties of Pt(bph) (CO)2, where bph is the biphenyl dianion

The complex Pt(bph) (CO)₂ crystallizes in the space group Cmcm with a = 18.647(6), B = 9.566(2) and C = 6.4060(5) Å. The geometry of the molecule is slightly distorted from square planar with a Pt---C(CO) bond distance of 1.98(2) Å and a Pt---C(bph) bond distance of 2.04(2) Å. The Pt(bph)(CO)₂ complex serves as a precursor for the preparation of a wide variety of Pt(bph)X₂ complexes, where X = monodentate ligands such as acetonitrile, pyridine, etc., and X₂ = bidentate ligands such as bypyridine, 1,10-phenanthroline, etc. In the solid state, the complex exhibits a green color, but when ground with an alkali metal salt turns deep blue to purple. In CH₂Cl₂, the color disappears but optical transitions are observed at 271 nm (2.7 × 10⁴ M⁻¹ cm⁻¹), 303 nm (1.1 × 10⁴ M⁻¹ cm⁻¹) and 330 nm (5.5 × 10³ M⁻¹ cm⁻¹). The complex is a weak emitter exhibiting a structured spectrum in CH₂Cl₂ at r.t. with maxima located at 562 and 594 nm and an emission lifetime of 3.1 μs when excited at 337 nm.     

Copper(II) complexes with (2-hydroxybenzyl-2-pyridylmethyl)amine–Hbpa: syntheses, characterization and crystal structures of the ligand and [Cu(II)(Hbpa)2](ClO4)2·2H2O

Copper(II) complexes were synthesized and characterized by means of elemental analysis, IR and visible spectroscopies, EPR and electrochemistry, as well as X-ray structure crystallography. The group consists of discrete mononuclear units with the general formula [Cu(II)(Hbpa)₂](A)₂·nH₂O, where Hbpa=(2-hydroxybenzyl-2-pyridylmethyl)amine and A=ClO₄ ⁻, n=2 (1), CH₃COO⁻, n=3 (2), NO₃ ⁻, n=2 (3) and SO₄ ²⁻, n=3 (4). The structures of the ligand Hbpa and complex 1 have been determined by X-ray crystallography. Complexes 1–4 have had their UV–Vis spectra measured in both MeCN and DMF. It was observed that the compounds interact with basic solvents, such that molecules coordinate to the metal in axial positions in which phenol oxygen atoms are coordinated in the protonated forms. The values were all less than 1000 M⁻¹ cm⁻¹. EPR measurements on powdered samples of 1–3 gave g/A values between 105 and 135 cm⁻¹, typical for square planar coordination environments. Complex 4·3H₂O exhibits a behaviour typical for tetrahedral coordination. The electrochemical behaviour for complexes 1 and 2 was studied showing irreversible redox waves for both compounds.     

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.     

Resonance raman spectroscopy of chemically modified and isotopically labelled purple membranes. II. Kinetic studies

Kinetic resonance Raman spectra of native and isotopically labelled purple membranes are compared. Using these data and the assignments of the previous paper in this sequence, we have confirmed that the Schiff base is deprotonated at times that are short in comparison to M₄₁₂ evolution. In addition, by monitoring the kinetic resonance Raman spectra in ²H₂O with 488.0 nm excitation we have been able to characterize in more detail the vibrational features associated with this unprotonated intermediate that precedes M₄₁₂. Furthermore, the kinetic spectra of fully deuterated purple membranes in H₂O have allowed us to assign the 1465 cm⁻¹ band in these spectra to the C=C stretching frequency of BR₅₇₀ and the 1512 cm⁻¹ band to the C=C stretching frequency of M₄₁₂. These spectra have also provided an indication of a Raman spectral feature associated with O₆₄₀ and, finally, our kinetic spectra have provided evidence that there is a significant alteration in the rate constants for the evolution of the various intermediates when the non-exchangeable protons on the membrane are replaced by deuterons.     

Mixed anion lanthanide(III) crown ether complexes: crystal structures of [LaCl2(NO3)(12-crown-4)]2, [La(NO3)(OH2)4-(12-crown-4)]Cl2·CH3CN and [LaCl2(NO3)(18-crown-6)]

Reaction of LaCl₃·7H₂O containing small amounts of La(NO₃)₃·7H₂O as an impurity with 12-crown-4 or 18-crown-6 in 3:1 CH₃CN:CH₃OH resulted in the isolation of the mixed anion complexes [LaCl₂(NO₃)(12-crown-4)]₂, [La(NO₃)(OH₂)₄(12-crown-4)]Cl₂·CH₃CN and [LaCl₂(NO₃)(18-crown-6)]. The nine-coordinate dimer, [LaCl₂(NO₃)(12-crown-4)]₂, has all of the anions in the inner coordination sphere and La³⁺ has a capped square antiprismatic geometry. It crystallizes in the orthorhombic space group Pbca with (at −150 °C) a = 12.938(6), B = 15.704(3), C = 13.962(2) Å, and D_calc = 2.08 g cm⁻³ for Z = 4. The second complex isolated from the same reaction, [La(NO₃)(OH₂)₄(12-crown-4)]Cl₂·CH₃CN, has the bidentate nitrate anion in the inner coordination sphere but the two chloride anions are in a hydrogen bonded outer sphere. This complex is ten-coordinate 4A,6B-expanded dodecahedral and crystallizes in the monoclinic space group P2₁ with (at 20 °C) A = 7.651(2), B = 11.704(7), C = 11.608(4) Å, β = 95.11(2)°, and D_calc = 1.80 g cm⁻³ for Z = 2. The 18-crown-6 complex, [LaCl₂(NO₃)(18-crown-6)], has all inner sphere anions and has ten-coordinate 4A,6B-expanded dodecahedral La³⁺ centers. It crystallizes in the orthorhombic space group Pbca with (at 20 °C) a = 14.122(7), B = 13.563(5), C = 19.311(9) Å, and D_calc = 1.89 g cm⁻³ for Z = 8.     

Syntheses, structures and magnetic properties of mono- and di-manganese inclusion compounds

Mono- and di-manganese inclusion compounds 1 and 2 are reported. Two mono-manganese molecules Mn(bpy)₂(NO₃)₂ (bpy=2,2′-bipyridine) and [Mn(bpy)₂(NO₃)(H₂O)]·NO₃ coexist in the mole ratio of 1:1 in the structure of 1, while two di-manganese molecules [Mn₂O(bpy)₂(phtha)₂(H₂O)₂]·(NO₃)₂ (phtha=phthalate) and [Mn₂O(bpy)₂(phtha)₂(NO₃)(H₂O)]·NO₃ in the structure of 2. Refluxing Mn(NO₃)₂/bpy/phthalic acid reaction mixtures in CH₃CN leads to the isolation of 1, further concentration of the reaction solution in raising temperature results in 2. The Mn₁ and Mn₂ units in the inclusion compounds 1 and 2 are similar to other reported Mn₁ and Mn₂ analogs, respectively. The Jahn–Teller distortion was observed to give rise to the elongation along the O_terminal---Mn---O_carboxyl axes for all the four Mn(III) sites in 2, leading to unexpected longer Mn(III)---O_aqua than Mn(II)---O_aqua in 1. Extensive hydrogen bonding interactions among H₂O, NO₃ ⁻ and COOH were observed in the two inclusion compounds. Cyclic voltammetry of 2 in DMF displays two quasi-reversible redox couples at +0.10/+0.22 and −0.43/−0.36 V assigned to the Mn(III)Mn(IV)/2Mn(III) and 2Mn(III)/Mn(III)Mn(II), respectively. Variable temperature magnetic susceptibilities of 1 and 2 were measured. The data were fit to a model including axial zero-field splitting term and a good fit was found with D=1.77 cm⁻¹, g=1.98 and F=1.48×10⁻⁵ for 1. For 2, the least-squares fitting of the experimental data led to J=2.37 cm⁻¹, g=2.02 and D=0.75 cm⁻¹ with R=1.45×10⁻³.     

Coupling of nuclear wavepacket motion and charge separation in bacterial reaction centers

The mechanism of the charge separation and stabilization of separated charges was studied using the femtosecond absorption spectroscopy. It was found that nuclear wavepacket motions on potential energy surface of the excited state of the primary electron donor P* leads to a coherent formation of the charge separated states P⁺B_A⁻, P⁺H_A⁻ and P⁺H_B⁻ (where B_A, H_B and H_A are the primary and secondary electron acceptors, respectively) in native, pheophytin-modified and mutant reaction centers (RCs) of Rhodobacter sphaeroides R-26 and in Chloroflexus aurantiacus RCs. The processes were studied by measurements of coherent oscillations in kinetics at 890 and 935 nm (the stimulated emission bands of P*), at 800 nm (the absorption band of B_A) and at 1020 nm (the absorption band of B_A⁻) as well as at 760 nm (the absorption band of H_A) and at 750 nm (the absorption band of H_B). It was found that wavepacket motion on the 130–150 cm⁻¹ potential surface of P* is accompanied by approaches to the intercrossing region between P* and P⁺B_A⁻ surfaces at 120 and 380 fs delays emitting light at 935 nm (P*) and absorbing light at 1020 nm (P⁺B_A⁻). In the presence of Tyr M210 (Rb. sphaeroides) or M195 (C. aurantiacus) the stabilization of P⁺B_A⁻ is observed within a few picosseconds in contrast to YM210W. At even earlier delay (40 fs) the emission at 895 nm and bleaching at 748 nm are observed in C. aurantiacus RCs showing the wavepacket approach to the intercrossing between the P* and P⁺H_B⁻ surfaces at that time. The 32 cm⁻¹ rotation mode of HOH was found to modulate the electron transfer rate probably due to including of this molecule in polar chain connecting P_B and B_A and participating in the charge separation. The mechanism of the charge separation and stabilization of separated charges is discussed in terms of the role of nuclear motions, of polar groups connecting P and acceptors and of proton of OH group of TyrM210.     

Manganese(II) complexes of an acyclic phenol-based dinucleating ligand with four methoxyethyl chelating arms: synthesis, structure, magnetism and electrochemistry

Dinuclear manganese(II) complexes [Mn₂(bomp)(PhCO₂)₂]BPh₄ (1), [Mn₂(bomp)(MeCO₂)₂]BPh₄ (2), and [Mn₂(bomp)(PhCO₂)₂]PF₆ (3) were synthesized with a dinucleating ligand 2,6-bis[bis(2-methoxyethyl)aminomethyl]-4-methylphenol [H(bomp)]. Dinuclear zinc complex [Zn₂(bomp)(PhCO₂)₂]PF₆ (4) was also synthesized for the purpose of comparison. X-ray analysis revealed that the complex 1·CHCl₃ contains two manganese ions bridged by the phenolic oxygen and two benzoate groups, forming a μ-phenoxo-bis(μ-benzoato)dimanganese(II) core. Magnetic susceptibility measurements of 1–3 over the temperature range 1.8–300 K indicated antiferromagnetic interaction (J=−4 to −6 cm⁻¹). Cyclic voltammograms of 3 showed a quasi-reversible oxidation process at +0.9 V versus a saturated sodium chloride calomel reference electrode, assigned to Mn^IIMn^II/Mn^IIMn^III.     

Structures and magnetism of rubidium and cesium salts of dimeric oxovanadium(IV) tartrate(4−) complexes

Complexes of type A₄[VO(tart)]₂·nH₂O, where A = Rb or Cs and tart =d,l-tartrate(4−) (n = 2) or d,d-tartrate(4−) (n = 2 for Rb and n = 3 for Cs), were prepared from an aqueous mixture of V₂O₅, AOH and H₄tart. These complexes were studied by single-crystal X-ray diffraction methods: Rb₄[VO(d,l-tart)]₂·2H₂O, space group P1 with a = 8.156(1),b = 8.246(1),c = 8.719(1)Å, = 66.09(1)°, β = 65.07(1)°, γ = 82.40(1)°,Z = 2, 1917 observed reflections, and final R_w = 0.035; Cs₄[VO(d,l-tart)]₂·2H₂O, space group P₂₁/c with a = 9.350(1),b = 13.728(2),c = 8.479(1)Å, β = 106.77(1)°,Z = 4, 2235 observed reflections, and final R_w = 0.054; Rb₄[VO(d,d-tart)]₂·2H₂O, space group P₄₁₂₂ with a = 8.072(1),c = 32.006(3)Å,Z = 8, 1014 observed reflections and final R_w = 0.038; Cs₄[VO(d,d-tart)]₂·3H₂O, space group P₁₂₂ with a = 8.184(1),c = 33.680(5)Å,Z = 8, 1310 observed reflections, and final R_w = 0.063. Bulk magnetic susceptibility data (1.5–300 K) for these compounds and A₄[VOl,l-tart)]₂·nH₂O (A = Rb, Cs) were obtained on polycrystalline samples. These data were analyzed in terms of a Van Vleck exchange coupled S = 1/2 model which was modified to include an interdimer exchange parameters Θ. Analysis of the low-temperature (1.5–20 K) susceptibility data gave 2J = +1.30 cm⁻¹ and Θ = −1.86 K for Rb₄[VO(d,l-tart)]₂·2H₂O, 2J = +1.16 cm⁻¹ and Θ = −1.69 K for Cs₄[VO(d,l-tart)]₂·2H₂O, 2J = +1.90 cm⁻¹ and Θ = −0.82 K for Rb₄[VO(d,d-tart)]₂·2H₂O, 2J = +2.04 cm⁻¹ and Θ = −0.80 K for Rb₄[VO(l,l-tart)]₂·2H₂O, 2J = +1.52 cm⁻¹ and Θ = −0.25 K for Cs₄[VO(d,d-tart)]₂·3H₂O, and 2J = +1.64 cm⁻¹ and Θ = −0.31 K for Cs₄[VO(l,l-tart)]₂·3H₂O. These results suggest the magnitudes of intradimer (ferromagnetic and interdimer (antiferromagnetic) exchange interactions are similar in these complexes, as observed for the analogous Na salts.     

Circular dichroism of cytochrome oxidase, cytochrome b566, and cytochrome c in beef heart mitochondrial membrane fragments

1. Circular dichroism spectra of the cytochromes in membrane fragments derived from sonicated beef heart mitochondria have been obtained in the wavelength region 400–480 nm in which the major absorbance maxima of the heme prosthetic groups are found.

2. 2. Cytochrome oxidase in the mitochondrial membrane fragments has a band of positive ellipticity at 426 nm in the oxidized form and a pronounced band of positive ellipticity at 445 nm in the reduced form. The reduced-minus-oxidized difference molar ellipticity at 445 nm, Δ[θ]₄₄₅ is 3.0·10⁵ degree·cm⁻²·dmole⁻¹ heme a for membrane-bound oxidase compared to 1.6·10⁵ degree·cm⁻²·dmole⁻¹ heme a for the purified oxidase. The membrane-bound oxidase in the reduced form also appears to have a band of negative ellipticity at 426 nm not found in the purified oxidase.

3. 3. When reduced with succinate in the presence of cyanide and oxygen, cytochrome oxidase in the membrane fragments has a positive band at 442 nm very similar to that observed with the purified oxidase.

4. 4. Cytochrome c, which has a positive band at 426 nm in the purified form when reduced, appears to have a negative band at this wavelength in the mito-chondrial membrane fragments which contributes to the pronounced negative band at 426 nm observed in the membrane fragments reduced with succinate in anaerobiosis. There is no evidence for a contribution to the CD spectra of the membrane fragments from cytochrome c₁ or from cytochrome b₅₆₁ in either the oxidized or the reduced form.

5. 5. Cytochrome b₅₆₆ in the mitochondrial membrane fragments has no detectable CD spectrum in the oxidized form, but has a small positive band at 427 nm and a small negative band at 436 nm in the reduced form. The same CD spectrum is observed with cytochrome b₅₆₆ reduced with succinate in the presence of antimycin A or 2-heptyl-4-hydroxyquinoline-N-oxide. The same increase in positive ellipticity is observed at 427 nm in the mitochondrial membrane fragments, treated with oligomycin to restore energy coupling, when cytochrome b₅₆₆ is reduced with succinate in the energized membrane, as is observed in the inhibitor-treated membrane fragments. The absence of a pronounced conformational change in cytochrome b₅₆₆ on energization, as revealed by its CD spectrum, favors the concept that its reduction by succinate in the energized state is due to reversed electron transport rather than an intrinsic shift in the cytochrome's midpoint redox potential.

Monovalent copper as a potential catalyst for formation of acetaldehyde via the migration of methyl radicals to the coordinated carbonyl in the complex (CO)Cu-CH3

Cu_aq⁺ forms stable complexes with carbon monoxide in aqueous solutions. Furthermore it reacts very fast with aliphatic radicals. The reaction of Cu(CO)_maq⁺ with methyl radicals, ^•CH₃ was studied using the pulse-radiolysis technique. The results point out that methyl radicals react with Cu(CO)_aq⁺ to form an unstable intermediate with a Cu^II-C σ bond identified as (CO)Cu^II-CH₃⁺, k = (1.1±0.2) × 10⁹ M⁻¹ s⁻¹. This intermediate has a strong LMCT charge transfer band (λ_max = 385 nm, _max = 2500 M⁻¹ cm⁻¹) which is similar to the absorption bands of other transient complexes with Cu^II-alkyl σ bonds. The coordinated carbon monoxide in (CO)Cu^II-CH₃⁺ inserts into the copper—carbon bond (or rather the coordinated methyl migrates to the coordinated carbon monoxide ligand) at a rate of (3.0±0.8) × 10² s⁻¹ to form the copperacetyl complex (CO)_mCu^II-C(CH₃)=O⁺ (λ_max = 480 nm, _max = 2100 M⁻¹ cm⁻¹). The rate of formation of (CO)Cu^II-CH₃⁺ and of the insertion reaction are pH independent. The complex (CO)_mCu^II-C(CH₃)=O⁺ is also unstable and decomposes heterolytically to yield acetaldehyde and Cu_aq²⁺ as the final stable products. This reaction is slightly pH dependent. The same reactivity pattern has been observed for the Cu(CO_naq⁺ complexes (n = 2 or 3). The results clearly point out that CO remains coordinated to transient complexes of the type Cu^II-alkyl.     

Studies on the kinetics and mechanism of anation reactions of some six-coordinate complexes of chromium(III) in aqueous solution

Rates of stepwise anation of cis-Cr(ox)₂(H₂O₂)⁻ with SCN⁻/N₃⁻, Cr(acac)₂(H₂O)₂⁺ with SCN⁻ and Cr(atda)(H₂O)₂ with SCN⁻ have been investigated in weakly acidic aqueous solutions. Rate constants, k_I and k_II for the two steps in each system, are composite as k_x = k_x⁰+k_x^X[X⁻] (x = I, II; X⁻ = SCN⁻, N₃⁻). These rate constants have been evaluated also as the corresponding ΔH^≠ and ΔS^≠ values. The results obtained and the plausible I_d mechanism seem to suggest Cr---OOC bond dissociation (hence a strongly negative ΔS^≠) generating the transition state in each system with outer-sphere association forming the precursor complex in the X⁻ dependent paths.     

Protein-based complex medium design for recombinant serine alkaline protease production

This work reports on the design of a complex medium based on simple and complex carbon sources, i.e. glucose, sucrose, molasses, and defatted-soybean, and simple and complex nitrogen sources, i.e. (NH₄)₂HPO₄, casein, and defatted-soybean, for serine alkaline protease (SAP) production by recombinant Bacillus subtilis carrying pHV1431::subC gene. SAP activity was obtained as 3050 U cm⁻³ with the initial defatted-soybean concentration C_soybean^o=20 kg m⁻³ and initial glucose concentration C_G^o=8 kg m⁻³; whereas, addition of the inorganic nitrogen source (NH₄)₂HPO₄ decreased SAP production considerably. Further increase in SAP production (3850 U cm⁻³) was obtained when sucrose was replaced with glucose at C_sucrose^o=15 kg m⁻³ and C_soybean^o=20 kg m⁻³. Nevertheless, when molasses was replaced with sucrose, the maximum activity was obtained with molasses having 10 kg m⁻³ initial sucrose concentration and C_soybean^o=15 kg m⁻³as 2130 U cm⁻³; moreover, when casein was replaced with defatted-soybean SAP production decreased considerably (ca. 250 U cm⁻³). Thereafter, the effects of inorganic ionic compounds were investigated; and except phosphate, inorganic compounds supplied from defatted-soybean were found to be sufficient for the bioprocess. The highest SAP activity was obtained as 5350 U cm⁻³ in the medium that contained (kg m⁻³): C_soybean^o=20, C_sucrose^o=15, C_Na2HPO4^o=0.021, and C_NaH2PO4^o=2.82, that was 6.5-fold higher than that of the SAP produced in the defined medium. By using the designed complex medium, oxygen transfer characteristics of the bioprocess were investigated; and, Damköhler number that is the oxygen transfer limitation increases with the cultivation time until t=14 h; and, at t>20 h both mass transfer and biochemical reaction resistances were effective. Overall oxygen transfer coefficient varied between 0.010 and 0.044 s⁻¹; volumetric oxygen uptake rate varied between 0.001 and 0.006 mol m⁻³ s⁻¹; and specific oxygen uptake rate varied between 0.0001 and 0.0022 mol kg⁻¹ DW s⁻¹ throughout the bioprocess.     

New herbicide-binding site in the photosynthetic electron-transport chain: Competitive herbicide binding at the photosystem I phylloquinone-(vitamin K1)-binding site

The binding of herbicides to the phylloquinone-(primary electron acceptor A₁)-binding site in green plant photosystem (PS) I reaction centers is shown. Dissociation constants (K_d) of various herbicides to the phylloquinone-binding site were estimated by analyzing their competitive inhibition of the reconstitution of the phylloquinone analogue, menadione (vitamin K₃), to the phylloquinone-extracted spinach PS I particles. The phylloquinone-binding site was found to bind o-phenanthroline (K_d = 1.2 × 10⁻⁴ M), but only weak binding was observed with atrazine (K_d > 10⁻² M), although both are known to bind specifically to the quinone-(Q_B)-binding site in reaction centers of purple photosynthetic bacteria or PS II. The inhibitors of the cytochrome b/c₁(ƒ) complex, myxothiazol (K_d=9.5 × 10⁻⁶ M) or antimycin A (K_d = 2.8 × 10⁻⁶ M), also strongly bound to the phylloquinone site. This is the first report showing that the PS I reaction center complex also has a herbicide-binding site, although the site is probably not sensitive in vivo to these herbicides due to its higher affinity for phylloquinone than herbicides. The inhibitor specificity of the PS I phylloquinone site is different from that of the other quinone-functioning sites in the photosynthetic or respiratory electron-transfer chain, suggesting it to have a unique structure.