High Formate Concentrations Research Articles

Coupling glycerol oxidation reaction using Ni-Co foam anodes to CO2 electroreduction in gas-phase for continuous co-valorization

Electrocatalytic reduction of CO2 is a promising alternative for storing energy and producing valuable products, such as formic acid/formate. Continuous gas-phase CO2 electroreduction has shown great potential in producing high concentrations of formic acid or formate at the cathode while allowing the oxygen evolution or the hydrogen oxidation reactions to occur at the anode. It is advantageous to use a more relevant oxidation reaction, such as glycerol which is a plentiful by-product of current biodiesel production process. This work successfully manages to couple the glycerol oxidation reaction with continuous gas-phase CO2 electroreduction to formate with the implementation of Ni-Co foam-based anodes. The MEA-electrolyzer developed can achieve significantly high formate concentrations of up to 359 g L-1 with high Faradaic efficiencies of up to 95%, while also producing dihydroxyacetone at a rate of 0.434 mmol m−2 s−1. In comparison with existing literature, this represents an excellent trade-off between relevant figures of merit and can remarkably contribute to a future implementation of this coupled electrochemical system approach at larger scales.

Investigating formate tolerance mechanisms in Saccharomyces cerevisiae and its application

Current global energy and environmental crisis have spurred efforts towards developing sustainable biotechnological solutions, such as utilizing CO2 and its derivatives as raw materials. Formate is an attractive one-carbon source due to its high solubility and low reduction potential. However, the regulatory mechanism of formate metabolism in yeast remains largely unexplored. This study employed adaptive laboratory evolution (ALE) to improve formate tolerance in Saccharomyces cerevisiae and characterized the underlying molecular mechanisms. The evolved strain was applied to produce free fatty acids (FFAs) under high concentration of formate with glucose addition. The results showed that the evolved strain achieved a FFAs titer of 250 mg/L. Overall, this study sheds light on the regulatory mechanism of formate tolerance and provides a platform for future studies under high concentrations of formate.

Core and auxiliary functions of one-carbon metabolism in Pseudomonas putida exposed by a systems-level analysis of transcriptional and physiological responses.

The soil bacterium Pseudomonas putida is a robust biomanufacturing host that assimilates a broad range of substrates while efficiently coping with adverse environmental conditions. P. putida is equipped with functions related to one-carbon (C1) compounds (e.g. methanol, formaldehyde, and formate) oxidation-yet pathways to assimilate these carbon sources are largely absent. In this work, we adopted a systems-level approach to study the genetic and molecular basis of C1 metabolism in P. putida. RNA sequencing identified two oxidoreductases, encoded by PP_0256 and PP_4596, transcriptionally active in the presence of formate. Quantitative physiology of deletion mutants revealed growth defects at high formate concentrations, pointing to an important role of these oxidoreductases in C1 tolerance. Moreover, we describe a concerted detoxification process for methanol and formaldehyde, the C1 intermediates upstream formate. Alcohol oxidation to highly-reactive formaldehyde by PedEH and other broad-substrate-range dehydrogenases underpinned the (apparent) suboptimal methanol tolerance of P. putida. Formaldehyde was mostly processed by a glutathione-dependent mechanism encoded in the frmAC operon, and thiol-independent FdhAB and AldB-II overtook detoxification at high aldehyde concentrations. Deletion strains were constructed and characterized towards unveiling these biochemical mechanisms, underscoring the worth of P. putida for emergent biotechnological applications-e.g. engineering synthetic formatotrophy and methylotrophy. IMPORTANCE C1 substrates continue to attract interest in biotechnology, as their use is both cost-effective and ultimately expected to mitigate the impact of greenhouse gas emissions. However, our current understanding of bacterial C1 metabolism remains relatively limited in species that cannot grow on (i.e., assimilate) these substrates. Pseudomonas putida, a model Gram-negative environmental bacterium, constitutes a prime example of this sort. The biochemical pathways active in response to methanol, formaldehyde, and formate have been largely overlooked-although the ability of P. putida to process C1 molecules has been previously alluded to in the literature. By using a systems-level strategy, this study bridges such knowledge gap through the identification and characterization of mechanisms underlying methanol, formaldehyde, and formate detoxification-including hitherto unknown enzymes that act on these substrates. The results reported herein both expand our understanding of microbial metabolism and lay a solid foundation for engineering efforts toward valorizing C1 feedstocks.

Inhibition of sulfate-reducing bacteria with formate.

Despite hostile environmental conditions, microbial communities have been found in µL-sized water droplets enclosed in heavy oil of the Pitch Lake, Trinidad. Some droplets showed high sulfate concentrations and surprisingly low relative abundances of sulfate-reducing bacteria in a previous study. Hence, we investigated here whether sulfate reduction might be inhibited naturally. Ion chromatography revealed very high formate concentrations around 2.37 mM in 21 out of 43 examined droplets. Since these concentrations were unexpectedly high, we performed growth experiments with the three sulfate-reducing type strains Desulfovibrio vulgaris, Desulfobacter curvatus, and Desulfococcus multivorans, and tested the effects of 2.5, 8, or 10 mM formate on sulfate reduction. Experiments demonstrated that 8 or 10 mM formate slowed down the growth rate of D. vulgaris and D. curvatus and the sulfate reduction rate of D. curvatus and D. multivorans. Increasing formate concentrations delayed the onsets of growth and sulfate reduction of D. multivorans, which were even inhibited completely while formate was added constantly. Contrary to previous studies, D. multivorans was the only organism capable of formate consumption. Our study suggests that formate accumulates in the natural environment of the water droplets dispersed in oil and that such levels are very likely inhibiting sulfate-reducing microorganisms.

107 Roles of Amino Acids in One-carbon Metabolism and Nutition.

The critical importance of non-essential and conditionally-essential amino acids for physiological function is perhaps best exemplified in one-carbon metabolism (OCM), a complex metabolic process whereby chemical groups containing a single carbon atom are used as biosynthetic building blocks. OCM is required for the synthesis of three of the building blocks of nucleic acids (adenine, guanine, thymidine). In addition, OCM provides methyl groups for many methyltransferases. OMC involves two vitamins (folate, Vitamin B12) and amino acids such as serine and glycine which serve as sources of the one-carbon groups. As much as 70% of the whole-body labile methyl groups (S-adenosylmethionine) produced by OMC in piglets is utilized for creatine synthesis. Formate plays a critical role in OCM. Formate, produced in mitochondria from serine and other substrates is released to the cytosol where it is incorporated into 10-formyl-THF. This may be directly incorporated into purines or reduced to 5,10-methylene-THF which is used either for thymidine synthesis or for the production of methyl groups. We have measured formate in serum from a variety of animals (mice, rats, sheep, pigs, humans) and find substantial concentrations, (25–50 µM), in all cases. An important exception is in late pregnant fetal lambs and fetal rats, where we found very high formate concentrations (200–400 µM) in the fetal circulation. Serine concentrations in the fetal circulation are also very high almost 10-fold higher than in the pregnant ewe. Formate concentrations in the human umbilical cord are 70% higher than in the mother. These data suggest an important role for formate in fetal development. We examined this possibility by examining the fate of formate in fetal sheep. Label from 13C-formate infused into the fetal circulation is incorporated into methionine. This requires that formate serves as a substrate for OCM in fetal sheep. Formate is a new frontier in OCM.

Structural and metabolic responses of Staphylococcus aureus biofilms to hyperosmotic and antibiotic stress.

Biofilms alter their metabolism in response to environmental stress. This study explores the effect of a hyperosmotic agent-antibiotic treatment on the metabolism of Staphylococcus aureus biofilms through the use of nuclear magnetic resonance (NMR) techniques. To determine the metabolic activity of S. aureus, we quantified the concentrations of metabolites in spent medium using high-resolution NMR spectroscopy. Biofilm porosity, thickness, biovolume, and relative diffusion coefficient depth profiles were obtained using NMR microimaging. Dissolved oxygen concentration was measured to determine the availability of oxygen within the biofilm. Under vancomycin-only treatment, the biofilm communities switched to fermentation under anaerobic condition, as evidenced by high concentrations of formate (7.4 ± 2.7 mM), acetate (13.1 ± 0.9 mM), and lactate (3.0 ± 0.8 mM), and there was no detectable dissolved oxygen in the biofilm. In addition, we observed the highest consumption of pyruvate (0.19 mM remaining from an initial 40 mM concentration), the sole carbon source, under the vancomycin-only treatment. On the other hand, relative effective diffusion coefficients increased from 0.73 ± 0.08 to 0.88 ± 0.08 under vancomycin-only treatment but decreased from 0.71 ± 0.04 to 0.60 ± 0.07 under maltodextrin-only and from 0.73 ± 0.06 to 0.56 ± 0.08 under combined treatments. There was an increase in biovolume, from 2.5 ± 1 mm3 to 7 ± 1 mm3 , under the vancomycin-only treatment, while the maltodextrin-only and combined treatments showed no significant change in biovolume over time. This indicated that physical biofilm growth was halted during maltodextrin-only and combined treatments.

Formate: The Neglected Member of One-Carbon Metabolism.

Formate, the only non-tetrahydrofolate (THF)-linked intermediate in one-carbon metabolism, is produced in mammals from a variety of metabolic sources. It occurs in serum of adults at a concentration of approximately 30 μM. Its principal function lies as a source of one-carbon groups for the synthesis of 10-formyl-THF and other one-carbon intermediates; these are primarily used for purine synthesis, thymidylate synthesis, and the provision of methyl groups for synthetic, regulatory, and epigenetic methylation reactions. Although formate is largely produced in mitochondria, these functions mostly occur in the cytoplasm and nucleus. Formate plays a significant role in embryonic development, as evidenced by the effectiveness of formate in the pregnant dam's drinking water on the incidence of neural tube defects in some genetic models. High formate concentrations in fetal lambs may indicate a role in fetal development and suggest that extracellular formate may play a role in the interorgan distribution of one-carbon groups.

Triple‐pulse Method for Monitoring Formate in CO2 Conversion Process

AbstractIn CO2‐to‐formic‐acid process, real‐time monitoring of formate is pivotal for the efficient control of the process. A stable sensing method that is suitable for measuring the high concentrations of formate produced during this process is required. Here, we developed a triple‐potential‐step method to monitor the formate in the range of 0.1 to 2.0 M and showed that a long‐term, continuous monitoring is possible. Among the Pd‐, Pt‐, and Au‐electrode materials tested, Au was the best option due to its low sensitivity and its stability in formate oxidation.

Engineering Hydrogen Gas Production from Formate in a Hyperthermophile by Heterologous Production of an 18-Subunit Membrane-bound Complex

Biohydrogen gas has enormous potential as a source of reductant for the microbial production of biofuels, but its low solubility and poor gas mass transfer rates are limiting factors. These limitations could be circumvented by engineering biofuel production in microorganisms that are also capable of generating H2 from highly soluble chemicals such as formate, which can function as an electron donor. Herein, the model hyperthermophile, Pyrococcus furiosus, which grows optimally near 100 °C by fermenting sugars to produce H2, has been engineered to also efficiently convert formate to H2. Using a bacterial artificial chromosome vector, the 16.9-kb 18-gene cluster encoding the membrane-bound, respiratory formate hydrogen lyase complex of Thermococcus onnurineus was inserted into the P. furiosus chromosome and expressed as a functional unit. This enabled P. furiosus to utilize formate as well as sugars as an H2 source and to do so at both 80° and 95 °C, near the optimum growth temperature of the donor (T. onnurineus) and engineered host (P. furiosus), respectively. This accomplishment also demonstrates the versatility of P. furiosus for metabolic engineering applications.

Improving d-mannitol productivity of Escherichia coli: Impact of NAD, CO 2 and expression of a putative sugar permease from Leuconostoc pseudomesenteroides

The highly productive whole-cell biotransformation of d-fructose to d-mannitol with recombinant, resting cells of Escherichia coli BL21(DE3) requires the combined expression of mdh, fdh and glf which encode mannitol and formate dehydrogenases and a sugar facilitator, respectively. However, long-term stability of the system was restricted, possibly due to loss of the cofactor NAD, high concentrations of formate, formation of CO 2 affecting the internal pH of the cells, accumulation of high intracellular concentrations of d-mannitol, and export of d-mannitol. Downstream of the mdh gene of Leuconostoc pseudomesenteroides, we identified an open reading frame encoding for a putative mannitol permease. The gene was cloned and expressed in E. coli. Biochemical analyses revealed an activity as secondary carrier for d-fructose. Therefore, the carrier was named FupL and participation in d-mannitol transport was excluded. In biotransformation experiments, the productivity of d-mannitol formation obtained with the strain expressing the additional fupL gene was enhanced by 20%.

Positional Ordering of Reacting Groups Contributes Significantly to the Efficiency of Proton Transfer at an Antibody Active Site

Catalytic antibody 34E4 accelerates the conversion of benzisoxazoles to salicylonitriles with surprising efficiency, exploiting a carboxylate base with an elevated pKa for proton abstraction. Mutagenesis of this antibody, produced as a chimeric Fab, confirms the prediction of a homology model that GluH50 is the essential catalytic residue. Replacement of this residue by glutamine, alanine, or glycine reduces catalytic activity by more than 2.6 x 10(4)-fold. By comparing the chemical proficiencies of the parent antibody with the chemical proficiencies of acetate and the mutants, the effective concentration of the catalytic side chain was estimated to be >51 000 M. The 2.1 kcal/mol destabilization of the transition state observed when GluH50 is replaced by aspartate suggests that positional ordering imposed by the antibody active site contributes significantly to the efficiency of proton transfer. The observation that the GluH50Ala and GluH50Gly variants could not be chemically rescued by exogenous addition of high concentrations of formate or acetate further underscores the advantage the antibody derives from covalently fixing its base at the active site. Although medium effects also play an important role in 34E4, for example in enhancing the reactivity of the carboxylate side chain through desolvation, comparison of 34E4 with less proficient antibodies shows that positioning a carboxylate in a hydrophobic binding pocket alone is insufficient for efficient general base catalysis. Our results demonstrate that structural complementarity between the antibody and its substrate in the transition state is an important and necessary component of 34E4's high activity. By harnessing an additional catalytic group that could serve as a general acid to stabilize developing negative charge in the leaving group, overall efficiencies rivaling those of highly evolved enzymes should be accessible.

Experimental constraints on the hydrothermal reactivity of organic acids and acid anions: I. Formic acid and formate

A series of hydrothermal experiments covering a range of temperatures from 175 to 260°C examined the decomposition of formic acid and formate and also investigated the production of formate from reduction of CO 2. Decomposition rates measured in this study, which were conducted in gold-TiO 2 reactors, were several orders of magnitude slower than those reported in previous studies conducted in steel and Ti-metal reactors, indicating the previous studies substantially overestimated the rate of the reaction owing to reactor catalysis. Although experiments were conducted with several different minerals present (hematite, magnetite, serpentinized olivine, NiFe-alloy), the decomposition rates were similar in each experiment once the effects of fluid pH were accounted for, suggesting that the minerals had no effect on the stability of formic acid or formate. At higher temperatures (>225°C), the rates of both the decomposition of formate and the reduction of CO 2 to formate were sufficiently rapid that reactions between dissolved CO 2 and formate rapidly attained a state of metastable thermodynamic equilibrium. The results suggest that the amount of formate in many subsurface and hydrothermal fluids is likely to be controlled by equilibrium with dissolved CO 2 at the prevailing oxidation state and pH of the fluid. This may account for the high concentrations of formate observed in strongly reducing environments such as serpentinites, as well as the low concentrations relative to other organic acid anions in mildly reducing environments such as oil-filed brines and formation waters in sedimentary basins. Although formate has been suggested to be a reaction intermediate in the formation of abiotic hydrocarbons from reduction of aqueous CO 2, production of hydrocarbons was not observed in any of the experiments, except for trace amounts of methane, despite high concentrations of formate and strongly reducing conditions.

Overexpression of formate dehydrogenase in Arabidopsis thaliana resulted in plants tolerant to high concentrations of formate

The potential role played by formate dehydrogenase (FDH) in formate metabolism has been examined by the overexpression of FDH in Arabidopsis thaliana . Three independent transgenic lines were selected and shown to produce elevated amounts of FDH protein with a corresponding elevated FDH activity (2.5-5 fold) over wild-type (WT) plants. Under normal growth conditions, no altered phenotype was observed in these transgenic plants; in growth media supplied with formate, however, significant differences in shoot and root growth, compared to that of WT plants, were observed. WT plants were severely injured if grown in the presence of 16 mmol/L formate, while the transgenic plants were able to grow well. Formate delayed germination of both WT and transgenic seeds at concentrations above 4 mmol/L, but both types of seeds were eventually able to complete more than 95 % germination even at 32 mmol/L formate. Formate markedly inhibited primary root elongation, and its inhibitory action on WT was much stronger than on transgenic plants. Different formate salts affected root elongation similarly, indicating that the formate ion was the major factor inhibiting root growth. Sodium acetate (NaAc), an analogue of formate, also inhibited root elongation, but its action on WT and transgenic plants was the same, indicating that tolerance of transgenic plants to formate toxicity was specific. Transgenic plants showed no significant tolerance to the toxicity of two other one-carbon metabolites, methanol and formaldehyde. A role for FDH in detoxifying formate is proposed.

Singlet dioxygen formation in ozone reactions in aqueous solution

In ozone reactions, singlet dioxygen [O2(1Δg)] is formed when ozone reacts by O-atom transfer. O2(1Δg) yields were determined for more than 50 compounds using as reference the reaction of hydrogen peroxide with hypochlorite. Close to 100% yields were found in the reaction of O3 with sulfides, disulfides, methanesulfinic acid, and nitrite. In accordance with this, the only products are: methionine sulfoxide, methanesulfonic acid, and nitrate for the reaction of O3 with methionine, methanesulfinic acid, and nitrite, respectively. In the case of aliphatic tertiary amines (trimethylamine, triethylamine, and DABCO), the O2(1Δg) yields range between 70 and 90%, the aminoxide being the other major product. With EDTA and nitrilotriacetic acid (NTA), the O2(1Δg) yield is around 20%. The interpretation of the data with DABCO required the determination of the quenching constant of O2(1Δg) by this amine, kq = 1.8 × 109 dm3 mol−1 s−1 in H2O and D2O, two orders of magnitude lower than previously reported. Aromatic tertiary amines give even lower O2(1Δg) yields [N,N-dimethylaniline (7%), N,N,N′,N′-tetramethylphenylenediamine (9%)]. Substantial amounts of O2(1Δg) are also formed with the DNA model compounds, 2′-deoxyguanosine (40%) and 2′-deoxyadenosine (15%, in the presence of tert-butyl alcohol as ˙OH scavenger). The pyrimidine nucleobases only yield O2(1Δg) when deprotonated at N(1). O2(1Δg) formation is also observed with hydrogen sulfide (15%), azide (17%), bromide (56%), iodide (12%), and cyanide ions (20%). The O2(1Δg) yield from the reaction of O3 with phenols and phenolates is typically around 20%, but may rise closer to 50% in the case of pentachloro- and pentabromophenolate. Low O2(1Δg) yields are found with unsaturated acids such as dihydroxyfumarate (6%), muconate (2%), and acetylenedicarboxylate (15%). With saturated compounds, also, no O2(1Δg) (e.g. with propan-2-ol, acetaldehyde, acetaldehyde dimethylacetal and glyoxal) or very little O2(1Δg) (formic acid; 6%, at high formate concentrations) was detected. As shown with some examples, knowledge of the O2(1Δg) yield (in combination with that of other products) is a prerequisite for the elucidation of the mechanisms of O3 reactions in aqueous solutions.

Preparation and crystal structures of formato complexes of the [MIV3O4]4+ and [MIV3S4]4+ (M = Mo, W) clusters. Convenient precursors to the corresponding aqua complexes.

In the aqueous chemistry of molybdenum(IV) and tungsten(IV), trinuclear, incomplete cubane-like, oxo and sulfido clusters of the type [M3E4]4+ (M = Mo, W; E = O, S) play a central role. We here describe how formato complexes of all these cluster cores can be prepared in high yields by crystallization from methanol-water or ethanol-water mixtures. Since potassium and ammonium formate are very soluble in these alcohol-water mixtures, high formate concentrations could be accomplished in the solutions from which the corresponding salts of cluster formato complexes crystallized. The [Mo3O4]4+ compounds could be synthesized without requiring the use of noncomplexing acids in the process. Some [M3E4]4+ compounds were characterized by single-crystal structure determinations. [NH4]3.20[K]0.80[H3O][Mo3O4(HCO2)8][HCO2].H2O was triclinic, space group P1 (No. 2) with a = 11.011(2) A, b = 13.310(2) A, c = 9.993(1) A, alpha = 106.817(7) degrees, beta = 91.651(9) degrees, gamma = 88.340(9) degrees, and two formula units per cell. [K]6[W3S4(HCO2)9][HCO2].2.27H2O.0.73CH3OH was monoclinic, space group C2/m (No. 12) with a = 19.605(6) A, b = 14.458(7) A, c = 13.627(5) A, beta = 118.94(2) degrees, and four formula units per cell. Generally, the nine coordination sites of [M3E4]4+ were occupied either by a mixture of monodentate and mu 2-bridging formato ligands or by monodentate formato ligands only. By dissolution in noncomplexing strong acid, all the formato complexes immediately hydrolyzed to form [M3E4(H2O)9]4+ aqua complexes. This allows, for example, high concentrations of [Mo3S4(H2O)9]4+ in CF3SO3H to be obtained and these solutions to be used for the synthesis of bimetallic clusters containing the cubane-like motif Mo3M'S4.

The influence of chemical structure on the anaerobic catabolism of refractory compounds: a case study of instant coffee wastes

Due to concerns about meeting the strict legislation currently in force, anaerobic treatment is being investigated for the treatment of a variety of waste streams. Instant coffee wastes are one type of industrial effluent that appears to be amenable to anaerobic digestion, and this paper presents some results on the biodegradation of this effluent. The method used was a batch bioassay technique known as the Biochemical Methane Potential assay (BMP), which gave 84% degradation of a composite coffee waste sample. Various fractions of the coffee waste revealed mixed degradation results mostly attributed to structural differences, with the liquid fractions being almost 60% degradable, and the solids filter cake fraction, containing most of the lignocellulosic material being only 9% degradable. Volatile fatty acid (VFA) analysis revealed high concentrations of formate being formed and subsequently degraded in the bioconversion process, and a possible role for formate production was postulated. Variations in structure did not appear to affect the route by which VFAs were produced. Bioconversion of 5-6 major classes of pure organic compounds thought to be potentially recalcitrant in coffee effluent was investigated to determine possible mechanisms of degradation, and the extent to which structural variation affected degradability. The phenolics and chlorogenic acids gave the highest degradation of 70% and 60% respectively, and the cyclic volatiles the least degradation at 40%.

A novel photosynthetic mimic reaction catalysed by K[Ru(H-EDTA)Cl] · 2H 2O; reduction of carbon dioxide to formate and formaldehyde in the presence of an aqueous suspension of Pt—CdS—RuO 2

An efficient photocatalytic reduction of carbon dioxide to HCOOH and HCHO is reported using K[Ru(H-EDTA)Cl] · 2H 2O ( 1) as homogeneous catalyst and particulate Pt—CdS—RuO 2 as photon absorber at 505 nm. This system produces 0.22 M of HCOOH and 0.10 M of HCHO in 6 h of photolysis at rates of 3.05 × 10 −2 M h −1 and 2.0 × 10 −2 M h −1 respectively. Trace amounts of CH 3OH, CH 4 and CO are detected in the reaction vessel. The rates of formation of HCOOH and HCHO exhibit a first-order dependence on the catalyst and dissolved CO 2 concentrations. The reaction shows deuterium isotope effects ( k H/ k D) of 1.5 and 2.00 for the formation of HCOOH and HCHO respectively. Under identical experimental conditions, the rate of decomposition of formate was studied. The rate of decomposition of formate is slower (by two orders of magnitude compared with the formation of formate) even at high formate concentrations. A mechanism for the formation of HCOOH and HCHO is proposed.

Formate transport, growth inhibition and the membrane protonmotive force in two methylotrophs (T15 and L3)

Formate transport and the effect of formate on growth and the membrane protonmotive force were studied in two ribulose-monophosphate-type obligate methylotrophs (bacterial strains T15 and L3). Formate was accumulated very fast by the membrane ΔpH according to the general transport mechanism of short-chain organic acids. Formate accumulation was reduced or abolished by a number of factors (protonophores, high extracellular pH, cell-starvation conditions) that reduced or abolished the ΔpH. Formate transport was accompanied by removal of protons from the medium by the cells. Since protons are released by the cells upon substrate oxidation, the stoichiometry of proton uptake upon formate transport could not be directly determined, although data suggest that formate is cotransported with one proton. The net rate of proton removal from the medium by the cells due to formate transport and oxidation increased with increasing formate concentrations or decreasing medium-pH values. The membrane protonmotive force of strain T15 was also studied as a function of the pH. High formate concentrations (of 100 to 400 mM) reduced the membrane ΔpH by ca. 20 to 60% and the growth rate by ca. 20 to 100% for both strains but to a different extent. For example, 400 mM formate inhibited growth by ca. 60% in strain T15 and by 100% in strain L3. The nature of growth inhibition by formate is discussed in some detail.

Enzymatic synthesis of formic acid from H2 and CO2 and production of hydrogen from formic acid

Whole cells of Alcaligenes eutrophus (as well as isolated P. oxalaticus formate dehydrogenase and A. eutrophus hydrogenase coupled via NAD(+) or methyl viologen) have been shown to produce H(2) from formic acid. Immobilization of the cells in kappa-carrageenan gel greatly enhances their stability at room temperature. The rate of hydrogen production catalyzed by immobilized A. eutrophus has been studied as a function of the concentrations of the cells and formate and also pH. An inhibition by high concentrations of formate has been found. Immobilized cells were also capable of synthesizing formate from H(2) and bicarbonate. Yields of formate up to 30% have been obtained. The catalytic efficiency of immobilized A. eutrophus cells was compared with that of palladium adsorbed on activated carbon.

Do copper ions influence the reduction of ferricytochrome [formula omitted] by O −2?

Recently, it was suggested that the measured rate of reduction of ferricyto chrome C by O − 2 below pH 8, was too high in the presence of high concentrations of formate (Koppenol, W.H., Van Buuren, K.J.H., Butler J. and Braams, R. (1976) Biochim. Biophys. Acta 449, 157–168). The high values were attributed to the presence of impurities of copper, which compete for O − 2. This assumption is consistent with either a decrease in the reduction yield of ferricytochrome C in the presence of copper, or with a very fast reaction of Cu(I) with ferricytochrome C. It was previously shown by us and by others that the reduction yield of ferricytochrome C by O − 2 is 100%. We measured the rate of reduction of ferricytochrome C by Cu(I), and found that this reaction is slow: k = (1.5±0.5) · 10 3 M −1) · s −1 . Therefore, our results rule out the possibility that below pH 8 copper impurities affect the measured rate constant of the reduction of ferricytochrome C by O − 2.