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Gas chromatographic analysis for quantification of plasma methanol requires laboratory equipment and personnel, and it is typically unavailable in short time notice, especially in low- and middle-income countries. Detection of formate with the enzyme formate oxidase (FOX) is a promising method that can make the diagnosis of methanol poisoning simple and fast. The aims of this study were to test the sensitivity and specificity of a modified FOX-enzyme and to test the specificity of a point-of-care (POC)-model containing FOX-enzyme with samples from patients with metabolic acidosis. The sensitivity and specificity of FOX-enzyme in aqueous solution were evaluated with a spectrometer and by visual detection for colour change. Formate concentrations between 1 and 20 mmol/L were used to test sensitivity, and 18 potentially interfering substances were tested for specificity. The sensitivity of the FOX-enzyme was 100% and the specificity 97%. When specificity of the POC-model was tested, no false positives were detected. As such, the sensitivity and specificity of this modified FOX-enzyme for detection of formate were high. The results with this enzyme confirm the potential for its use in formate analysis as a fast diagnosis of methanol poisoning.

Anaerobic digestion (AD) is an important technology for the effective conversion of waste and wastewater to methane. Here, syntrophic bacteria transfer molecular hydrogen (H

A growing body of work indicates that continental subsurface rocks host a substantial portion of the Earth's biosphere. However, the activities of microbial cells inhabiting pore spaces and microfractures in subsurface rocks remain underexplored. Here, we develop and optimize microcosm assays to detect organic acid transformation activities of cells residing in mafic to ultramafic igneous rocks. Application of this assay to gabbro core from the Stillwater Mine, Montana, USA, revealed maximal methane production from acetate at temperatures approximating that of the mine. Controls show that these activities are not due to contamination introduced during drilling, exhumation, or laboratory processing of the core. The assay was then applied to rocks cored from the Samail Ophiolite, Oman, which is undergoing low-temperature serpentinization. Production of (i) carbon dioxide from acetate and formate and (ii) methane from formate were detected in a dunite/harzburgite rock core interfacing pH 9.6 waters, and estimates of microbial activities were up to three orders of magnitude higher in the rock core pore space than in corresponding waters. The detection of endolithic microbial activities in igneous rocks has implications for life detection on other planetary bodies where similar rock types prevail, such as Mars, Europa and Enceladus.

A novel fermentation process was developed in which renewable electricity is indirectly used as an energy source in fermentation, synergistically decreasing both the consumption of sugar as a first generation carbon source and emission of the greenhouse gas CO2. As an illustration, a glucose-based process is co-fed with formic acid, which can be generated by capturing CO2 from fermentation offgas followed by electrochemical reduction with renewable electricity. This “closed carbon loop” concept is demonstrated by a case study in which cofeeding formic acid is shown to significantly increase the yield of biomass on glucose of the industrially relevant yeast species Yarrowia lipolytica. First, the optimal feed ratio of formic acid to glucose is established using chemostat cultivations. Subsequently, guided by a dynamic fermentation process model, a fed-batch protocol is developed and demonstrated on laboratory scale. Finally, the developed fed-batch process is tested and proven to be scalable at pilot scale. Extensions of the concept are discussed to apply the concept to anaerobic fermentations, and to recycle the O2 that is co-generated with the formic acid to aerobic fermentation processes for intensification purposes.

Unlike gaseous C1 feedstocks for acetogenic bacteria, there has been less attention on liquid C1 feedstocks, despite benefits in terms of energy efficiency, mass transfer and integration within existing fermentation infrastructure. Here, we present growth of Eubacterium limosum ATCC8486 using methanol and formate as substrates, finding evidence for the first time of native butanol production. We varied ratios of methanol-to-formate in batch serum bottle fermentations, showing butyrate is the major product (maximum specific rate 220 ± 23 mmol-C gDCW-1 day-1 ). Increasing this ratio showed methanol is the key feedstock driving the product spectrum towards more reduced products, such as butanol (maximum titre 2.0 ± 1.1 mM-C). However, both substrates are required for a high growth rate (maximum 0.19 ± 0.011 h-1 ) and cell density (maximum 1.2 ± 0.043 gDCW l-1 ), with formate being the preferred substrate. In fact, formate and methanol are consumed in two distinct growth phases - growth phase 1, on predominately formate and growth phase 2 on methanol, which must balance. Because the second growth varied according to the first growth on formate, this suggests butanol production is due to overflow metabolism, similar to 2,3-butanediol production in other acetogens. However, further research is required to confirm the butanol production pathway in E. limosum, particularly given, unlike other substrates, methanol likely results in mostly NADH generation, not reduced ferredoxin.

A facile synthesis of 4H-benzo[d][1,3]oxazin-4-one derivatives by Pd-catalyzed carbonylative cross-coupling between N-(ortho-bromoaryl)amides and benzene-1,3,5-triyl triformate (TFBen) was developed. This procedure does not require the toxic and flammable gas CO as the carbonyl source and tolerates a wide scope of functional groups. Remarkably, 4H-benzo[d][1,3]oxazin-4-ones incorporated to natural products and drugs can be constructed by this method.

On-site hydrogen peroxide production through electrocatalytic and photocatalytic oxygen reduction reactions has recently attracted broad research interest. However, practical applications have thus far been plagued by the low activity and the requirement of complex equipment. Here, inspired by the process of biological hydrogen peroxide synthesis catalyzed by enzymes, we report a Pt-Au alloy to mimic the catalytic function of natural formate oxidase for hydrogen peroxide synthesis through aerobic oxidation of formic acid. The mass activity of the Pt-Au alloy is three times higher than that of formate oxidase. Density functional theory calculations revealed that the efficient dehydrogenation of formic acid and the high selectivity of the subsequent reduction of oxygen to hydrogen peroxide account for the high hydrogen peroxide productivity. In addition, the formic acid aqueous solution provides an acidic environment, which is conducive to the utilization of the in situ generated hydrogen peroxide for oxidation reactions, including C-H bond oxidation and sterilization.

Acknowledging the importance of medicinal compounds, flavonoids, and phenolic acids in plants and human health; understanding the best time to harvest plants to get the most potentially therapeutic biological activity; and predicting and controlling the quality of medicinal plants are very useful. The rich chemical composition of medicinal lavender, as well as its antioxidant activity, has led to its wide application in the pharmaceutical, cosmetic, and food industries. In the current research, seasonal differences in phenolic content; the main constituents of essential oil, essential oil yield, and antioxidant activity; and flavonoid content were evaluated. In this investigation, for all the investigated factors, three replications were considered as completely randomized designs. The predominant natural components were α-pinene, camphene, 1,8-cineole, limonene, camphor, borneol, cryptone, isobornyl formate, cumin aldehyde, carvone, caryophyllene oxide, and epi-α-cadinol. The highest rate of 1,8-cineole was seen in summer, while camphor and borneol showed their highest percentages in the cold season. Some of the compounds such as caryophyllene oxide, isobornyl formate, and cumin aldehyde showed their highest amounts in the spring with the activation of some important plant enzymes such as peroxidase. The total content of flavonoids and some phenolic compounds (coumarin and p-coumaric acid) increased when plants received sunlight for a longer time than in the colder to warmer seasons. On the other hand, increases in chlorogenic acid and vanillin were observed in spring. Also, the highest antioxidant activity was observed in lavender, which was accompanied by an increase in the important phenolic composition of rosmarinic acid in the cold season. Generally, this investigation provides beneficial evidence about the most appropriate harvest time for lavender to produce the most favorable constituents for optimization in food, pharmaceutical, and herbal products.

Photocatalysis is a promising technology for conversion of the glycerol into formic acid, but photocatalytic oxidation of C-C bonds in glycerol exhibits poor selectivity towards formic acid because the photogenerated radicals (e.g., hydroxyl radicals) further oxidize formic acid to CO

SHR0302, as a novel Janus kinase (JAK) inhibitor 1, is used for treatment of rheumatoid arthritis (RA) in humans. A novel and sensitive ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) has been developed and validated for determining the concentration of SHR0302 in human plasma. A precipitation deproteinization method was used for plasma pretreatment with methanol. Detection was carried out on an Agilent 1,260 UPLC coupled with a Triple Quad 4000 mass spectrometer operated in positive multiple reaction monitoring mode, and the analytes were separated on a Synergi Polar-RP C

A fast, uncomplicated, sensitive and fully validated high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method has been developed for estimating l-amino acids in the plasma of schizophrenic patients. The gradient-elution chromatographic method was implemented with the Luna® PFP column (50 × 2.0 mm, 5 μm), and a mobile phase of 0.1% formic acid in water and methanol was used. The intra- and interday variability of the l-amino acids was13.11%, their accuracy ranged from 85.14 to 116.75% at the quality control levels and the lower limit of quantification ranged from 2.5 to 15 nm. The extraction efficiency (apparent recovery) of amino acids from healthy plasma was employed by spiking the plasma with standard amino acids at the quality control levels. Their percentage recoveries ranged from 80.4 to 119.94%. Our method has a short run time and fast sample preparation compared with existing methods, which suffer from long preparative steps and/or time-consuming analysis, restricted reagents and the suboptimal performance characteristics of presently available technologies. Therefore, the proposed HPLC-MS/MS method was effectively applied for monitoring the l-amino acids in the plasma of schizophrenic patients and healthy volunteers.

Escherichia coli is able to utilize the mixture of carbon sources and produce molecular hydrogen (H2 ) via formate hydrogen lyase (FHL) complexes. In current work role of transcriptional activator of formate regulon FhlA in generation of fermentation end products and proton motive force, N'N'-dicyclohexylcarbodiimide (DCCD)-sensitive ATPase activity at 20 and 72 hr growth during utilization of mixture of glucose, glycerol, and formate were investigated. It was shown that in fhlA mutant specific growth rate was ~1.5 fold lower compared to wt, while addition of DCCD abolished the growth in fhlA but not in wt. Formate was not utilized in fhlA mutant but wt cells simultaneously utilized formate with glucose. Glycerol utilization started earlier (from 2 hr) in fhlA than in wt. The DCCD-sensitive ATPase activity in wt cells membrane vesicles increased ~2 fold at 72 hr and was decreased 70% in fhlA. Addition of formate in the assays increased proton ATPase activity in wt and mutant strain. FhlA absence mainly affected the ΔpH but not ΔΨ component of Δp in the cells grown at 72 hr but not in 24 hr. The Δp in wt cells decreased from 24 to 72 hr of growth ~40 mV while in fhlA mutant it was stable. Taken together, it is suggested that FhlA regulates the concentration of fermentation end products and via influencing FO F1 -ATPase activity contributes to the proton motive force generation.

Biotechnology and Bioengineering published by Wiley Periodicals LLC Autotrophic or mixotrophic use of one-carbon (C1) compounds is gaining importance for sustainable bioproduction. In an effort to integrate the reductive glycine pathway (rGP) as a highly promising pathway for the assimilation of CO2 and formate, genes coding for glycine synthase system from Gottschalkia acidurici were successfully introduced into Clostridium pasteurianum, a non-model host microorganism with industrial interests. The mutant harboring glycine synthase exhibited assimilation of exogenous formate and reduced CO2 formation. Further metabolic data clearly showed large impacts of expression of glycine synthase on the product metabolism of C. pasteurianum. In particular, 2-oxobutyrate (2-OB) was observed for the first time as a metabolic intermediate of C. pasteurianum and its secretion was solely triggered by the expression of glycine synthase. The perturbation of C1 metabolism is discussed regarding its interactions with pathways of the central metabolism, acidogenesis, solventogenesis, and amino acid metabolism. The secretion of 2-OB is considered as a consequence of metabolic and redox instabilities due to the activity of glycine synthase and may represent a common metabolic response of Clostridia in enhanced use of C1 compounds.

Aqueous-phase oxidation by H

Geobacter sulfurreducens is a model bacterium to study the degradation of organic compounds coupled to the reduction of Fe(III). The response of G. sulfurreducens to the electron donors acetate, formate, hydrogen and a mixture of all three with Fe(III) citrate as electron acceptor was studied using comparative physiological and proteomic approaches. Variations in the supplied electron donors resulted in differential abundance of proteins involved in the citric acid cycle (CAC), gluconeogenesis, electron transport, and hydrogenases and formate dehydrogenase. Our results provided new insights into the electron donor metabolism of G. sulfurreducens. Remarkably, formate was the preferred electron donor compared to acetate, hydrogen, or acetate plus hydrogen. When hydrogen was the electron donor, formate was formed, which was associated with a high abundance of formate dehydrogenase. Notably, abundant proteins of two CO2 fixation pathways (acetyl-CoA pathway and the reversed oxidative CAC) corroborated chemolithoautotrophic growth of G. sulfurreducens with formate or hydrogen and CO2, and provided novel insight into chemolithoautotrophic growth of G. sulfurreducens. This article is protected by copyright. All rights reserved., This work was performed in the TTIW-cooperation framework of Wetsus, European Centre of Excellence for Sustain able Water Technology (www.wetsus.nl). Wetsus is funded by the Dutch Ministry of Economic Affairs, the European Union Regional Development Fund, the Province of Fryslân, the City of Leeuwarden and the EZ/Kompas program of the “Samenwerkingsverband Noord-Nederland”. The authors like to thank the participants of the research theme ‘Resource Recovery’ for fruitful discussions and their financial support. Research of AJMS is financed by an advanced grant of the European Research Council under the European Union’s Seventh Framework Programme (FP/2007e2013)/ERC Grant Agreement (project 323009). Research of AJMS and PHAT is supported by a Gravitation grant (project 024.002.002) of the Netherlands Ministry of Education, Culture and Science., info:eu-repo/semantics/publishedVersion

In this study we focused on the two acetogenic bacteria Acetobacterium woodii and Thermoanaerobacter kivui and demonstrated their potential as whole cell biocatalysts for the conversion of CO into the bulk chemicals acetate and formate. Notably, no external electron donor such as H2 was required for the conversion of CO. We could further shed light into the dark of CO utilization in A. woodii and T. kivui by using mutants with deficiencies in key enzyme activities of the central metabolism., Summary Acetogenic bacteria have gained much attraction in recent years as they can produce different biofuels and biochemicals from H2 plus CO2 or even CO alone, therefore opening a promising alternative route for the production of biofuels from renewable sources compared to existing sugar‐based routes. However, CO metabolism still raises questions concerning the biochemistry and bioenergetics in many acetogens. In this study, we focused on the two acetogenic bacteria Acetobacterium woodii and Thermoanaerobacter kivui which, so far, are the only identified acetogens harbouring a H2‐dependent CO2 reductase and furthermore belong to different classes of ‘Rnf’‐ and ‘Ech‐acetogens’. Both strains catalysed the conversion of CO into the bulk chemical acetate and formate. Formate production was stimulated by uncoupling the energy metabolism from the Wood–Ljungdahl pathway, and specific rates of 1.44 and 1.34 mmol g−1 h−1 for A. woodii ∆rnf and T. kivui wild type were reached. The demonstrated CO‐based formate production rates are, to the best of our knowledge, among the highest rates ever reported. Using mutants of ∆hdcr, ∆cooS, ∆hydBA, ∆rnf and ∆ech2 with deficiencies in key enzyme activities of the central metabolism enabled us to postulate two different CO utilization pathways in these two model organisms.

Separation selectivity and detection sensitivity of reversed-phase high-performance liquid chromatography with tandem mass spectrometry analyses were compared for formic (0.1%) and formic/heptafluorobutyric (0.1%/0.005%) acid based eluents using a proteomic data set of ∼12 000 paired peptides. The addition of a small amount of hydrophobic heptafluorobutyric acid ion-pairing modifier increased peptide retention by up to 10% acetonitrile depending on peptide charge, size, and hydrophobicity. Retention increase was greatest for peptides that were short, highly charged, and hydrophilic. There was an ∼3.75-fold reduction in MS signal observed across the whole population of peptides following the addition of heptafluorobutyric acid. This resulted in ∼36% and ∼21% reduction of detected proteins and unique peptides for the whole cell lysate digests, respectively. We also confirmed that the separation selectivity of the formic/heptafluorobutyric acid system was very similar to the commonly used conditions of 0.1% trifluoroacetic acid, and developed a new version of the Sequence-Specific Retention calculator model for the formic/heptafluorobutyric acid system showing the same ∼0.98 R2 -value accuracy as the Sequence-Specific Retention calculator formic acid model. In silico simulation of peptide distribution in separation space showed that the addition of 0.005% heptafluorobutyric acid to the 0.1% formic acid system increased potential proteome coverage by ∼11% of detectable species (tryptic peptides ≥ four amino acids).

Escherichia coli is able to ferment not only single but also mixtures of carbon sources. The formate metabolism and effect of formate on various enzymes have been extensively studied during sole glucose but not mixed carbon sources utilization. It was revealed that in membrane vesicles (MV) of wild type cells grown at pH 7.5 during fermentation of the mixture of glucose (2 g/L), glycerol (10 g/L), and formate (0.68 g/L), in the assays, the addition of formate (10 mM) increased the N,N'-dicyclohexylcarbodiimide (DCCD)-inhibited ATPase activity on ~30% but no effect of potassium ions (100 mM) had been detected. In selC (coding formate dehydrogenases) and fdhF (coding formate dehydrogenase H) single mutants, formate increased DCCD-inhibited ATPase activity on ~40 and ~70%, respectively. At pH 5.5, in wild type cells MV, formate decreased the DCCD-inhibited ATPase activity ~60% but unexpectedly in the presence of potassium ions, it was stimulated ~5.8 fold. The accessible SH or thiol groups number in fdhF mutant was less by 28% compared with wild type. In formate assays, the available SH groups number was less ~10% in wild type but not in fdhF mutant. Taken together, the data suggest that proton ATPase activity depends on externally added formate in the presence of potassium ions at low pH. This effect might be regulated by the changes in the number of redox-active thiol groups via formate dehydrogenase H, which might be directly related to proton ATPase FO subunit.

The abundance and evolving pathogenic behavior of bacterial microorganisms give rise to antibiotic tolerance and resistance which pose a danger to global public health. New therapeutic strategies are needed to keep pace with this growing threat. We propose a novel approach for targeting bacteria by harnessing formate, a cell metabolite found only in particular bacterial species, to activate an antibacterial prodrug and selectively inhibit their growth. This strategy is premised on transfer hydrogenation reaction on a biorthogonal substrate utilizing native formate as the hydride source as a means of uncaging an antibacterial prodrug. Using coordination-directed 3-component assembly to prepare a library of 768 unique Ru-Arene Schiff-base complexes, we identified several candidates that efficiently reduced sulfonyl azide functional group in the presence of formate. This strategy paves the way for a new approach of targeted antibacterial therapy by exploiting unique bacterial metabolites.

Successful application of microorganisms to heavy metal remediation depends on their resistance to toxic metals. This study contrasted the differences of tolerant mechanisms between Pb2+ and Cd2+ in Enterobacter sp. Microbial respiration and production of formic acid showed that Enterobacter sp. had a higher tolerant concentration of Pb (>1000 mg l-1 ) than Cd (about 200 mg l-1 ). Additionally, SEM confirmed that most of Pb and Cd nanoparticles (NPs) were adsorbed onto cell membrane. The Cd stress, even at low concentration (50 mg l-1 ), significantly enlarged the sizes of cells. The cellular size raised from 0.4 × 1.0 to 0.9 × 1.6 μm on average, inducing a platelet-like shape. In contrast, Pb cations did not stimulate such enlargement even up to 1000 mg l-1 . Moreover, Cd NPs were adsorbed homogeneously by almost all the bacterial cells under TEM. However, only a few cells work as 'hot spots' on the sorption of Pb NPs. The heterogeneous sorption might result from a 'self-sacrifice' mechanism, i.e., some cells at a special life stage contributed mostly to Pb sorption. This mechanism, together with the lower mobility of Pb cations, caused higher microbial tolerance and removal efficiency towards Pb2+ . This study sheds evident contrasts of bacterial resistance to the two most common heavy metals.

Formate hydrogenlyase (FHL) is the main hydrogen‐producing enzyme complex in enterobacteria. It converts formate to CO 2 and H2 via a formate dehydrogenase and a [NiFe]‐hydrogenase. FHL and complex I are evolutionarily related and share a common core architecture. However, complex I catalyses the fundamentally different electron transfer from NADH to quinone and pumps protons. The catalytic FHL subunit, HycE, resembles NuoCD of Escherichia coli complex I; a fusion of NuoC and NuoD present in other organisms. The C‐terminal domain of HycE harbours the [NiFe]‐active site and is similar to other hydrogenases, while this domain in NuoCD is involved in quinone binding. The N‐terminal domains of these proteins do not bind cofactors and are not involved in electron transfer. As these N‐terminal domains are separate proteins in some organisms, we removed them in E. coli and observed that both FHL and complex I activities were essentially absent. This was due to either a disturbed assembly or to complex instability. Replacing the N‐terminal domain of HycE with a 180 amino acid E. coli NuoC protein fusion did not restore activity, indicating that the domains have complex‐specific functions. A FHL complex in which the N‐ and C‐terminal domains of HycE were physically separated still retained most of its FHL activity, while the separation of NuoCD abolished complex I activity completely. Only the FHL complex tolerates physical separation of the HycE domains. Together, the findings strongly suggest that the N‐terminal domains of these proteins are key determinants in complex assembly., Complex I and the formate hydrogenlyase (FHL) complex from Escherichia coli contain either the fusion protein NuoCD or HycE, respectively. We deleted the N‐terminal, cofactor‐free domains from both proteins or genetically separated them into two individual proteins and monitored enzyme activity. The deletion caused instability of both complexes, while after separation, FHL retained activity but complex I was inactive.

Dimethyl sulfide, a major byproduct of the Kraft pulping process, was used as an inexpensive and sustainable catalyst/co-reagent (methyl donor) for various methylations with dimethyl carbonate (as both reagent and solvent), which afforded excellent yields of O-methylated phenols and benzoic acids, and mono-C-methylated arylacetonitriles. Furthermore, these products could be isolated using a remarkably straightforward workup and purification procedure, realized by dimethyl sulfide's neutral and distillable nature and the absence of residual salts. The likely mechanisms of these methylations were elucidated using experimental and theoretical methods, which revealed that the key step involves the generation of a highly reactive trimethylsulfonium methylcarbonate intermediate. The phenol methylation process represents a rare example of a Williamson-type reaction that occurs without the addition of a Brønsted base.

Methanol poisoning kills thousands of people every year and remains a diagnostic challenge, especially where the resources are scarce, but also in high-income countries worldwide. We are in the course of developing a bedside strip to detect formate - the toxic metabolite of methanol. We hereby present the first clinical methanol case where formate was detected bedside from a drop of blood: The patient, a 61-year-old male, was admitted with a suspect methanol poisoning and severe metabolic acidosis. The test strip was positive after 3 minutes. Sodium bicarbonate (500 mmol/L), fomepizole, dialysis and folinic acid were given based on the positive test. The diagnosis was some hours later confirmed by GC-MS, showing a methanol concentration of 62 mmol/L (200 mg/dL) and a formate concentration of 19 mmol/L. Implementation of this technology into routine clinical use can potentially offer an opportunity for a step change in the management of methanol poisoning.

We report a stereoselective conversion of terminal alkynes to α-chiral carboxylic acids using a nickel-catalyzed domino hydrocarboxylation-transfer hydrogenation reaction. A simple nickel/BenzP* catalyst displayed high activity in both steps of regioselective hydrocarboxylation of alkynes and subsequent asymmetric transfer hydrogenation. The reaction was successfully applied in enantioselective preparation of three nonsteroidal anti-inflammatory profens (>90 % ees) and the chiral fragment of AZD2716.

Summary Pectobacterium atrosepticum SCRI1043 is a phytopathogenic Gram‐negative enterobacterium. Genomic analysis has identified that genes required for both respiration and fermentation are expressed under anaerobic conditions. One set of anaerobically expressed genes is predicted to encode an important but poorly understood membrane‐bound enzyme termed formate hydrogenlyase‐2 (FHL‐2), which has fascinating evolutionary links to the mitochondrial NADH dehydrogenase (Complex I). In this work, molecular genetic and biochemical approaches were taken to establish that FHL‐2 is fully functional in P. atrosepticum and is the major source of molecular hydrogen gas generated by this bacterium. The FHL‐2 complex was shown to comprise a rare example of an active [NiFe]‐hydrogenase‐4 (Hyd‐4) isoenzyme, itself linked to an unusual selenium‐free formate dehydrogenase in the final complex. In addition, further genetic dissection of the genes encoding the predicted membrane arm of FHL‐2 established surprisingly that the majority of genes encoding this domain are not required for physiological hydrogen production activity. Overall, this study presents P. atrosepticum as a new model bacterial system for understanding anaerobic formate and hydrogen metabolism in general, and FHL‐2 function and structure in particular., Pectobacterium atrospecticum contains the genes for formate hydrogenlyase‐2, considered the ancient progenitor of Complex I. Here, P. atrosepticum was harnessed as a new model system for advancing new knowledge in FHL‐2. The complex was found to contain an unusual selenium‐free formate dehydrogenase and a [NiFe]‐hydrogenase‐4 with a large membrane arm. FHL‐2 was established as the major source of hydrogen gas; however, some components of the membrane arm were surprisingly not essential for this activity.

Bone containing tissues such as osteochondral joint are resistant to routine tissue processing, therefore require decalcification. This technique causes removal of mineral salts, but in the process may macerate the organic tissue, hence the need for tissue fixation. Such severe processing demands careful antigen retrieval to necessitate optimal staining. The aim of our study was to compare five different antigen retrieval protocols (heat retrieval and protein digestion) following decalcification of rabbit knee joints using two different techniques (20% formic acid and 10% ethylenediamine-tetra acetic acid: EDTA). Osteochondral sections were compared based on time required for decalcification, ease of sectioning, morphological integrity using HE staining and antigen preservation (Collagen type II) using immunohistochemistry. The two decalcification solutions did not impair the tissue morphology and ease of sectioning. Joints processed with formic acid decalcified four times faster than EDTA. Among the five antigen retrieval approaches, maximal collagen II uptake with minimal nonspecific staining was found with protein digestion (pronase and hyaluronidase) in both formic acid and EDTA sections. For osteo-chondral sections, we recommend using 10% EDTA for decalcification and pronase plus hyaluronidase for antigen retrieval if maintaining tissue morphology is crucial, whereas if time is of the essence, 20% FA with pronase plus hyaluronidase is the faster option while still preserving structural integrity. Clin. Anat. 33:343-349, 2020. © 2019 Wiley Periodicals, Inc.

The integration of enzymes with synthetic materials allows efficient electrocatalysis and production of solar fuels. Here, we couple formate dehydrogenase (FDH) from Desulfovibrio vulgaris Hildenborough (DvH) to metal oxides for catalytic CO 2 reduction and report an in-depth study of the resulting enzyme–material interface. Protein film voltammetry (PFV) demonstrates the stable binding of FDH on metal-oxide electrodes and reveals the reversible and selective reduction of CO 2 to formate. Quartz crystal microbalance (QCM) and attenuated total reflection infrared (ATR-IR) spectroscopy confirm a high binding affinity for FDH to the TiO 2 surface. Adsorption of FDH on dye-sensitized TiO 2 allows for visible-light-driven CO 2 reduction to formate in the absence of a soluble redox mediator with a turnover frequency (TOF) of 11±1 s −1 . The strong coupling of the enzyme to the semiconductor gives rise to a new benchmark in the selective photoreduction of aqueous CO 2 to formate. authorsversion published

We report an approach to conducting the hydroaminomethylation of diverse α-olefins with a wide range of alkyl, aryl, and heteroarylamines at low temperatures (70–80 °C) and pressures (1.0–3.4 bar) of synthesis gas. This approach is based on simultaneously using two distinct catalysts that are mutually compatible. The hydroformylation step is catalyzed by a rhodium diphosphine complex, and the reductive amination step, which is conducted as a transfer hydrogenation with aqueous, buffered sodium formate as the reducing agent, is catalyzed by a cyclometallated iridium complex. By adjusting the ratio of CO to H(2), we conducted the reaction at one atmosphere of gas with little change in yield. A diverse array of olefins and amines, including hetreroarylamines that do not react under more conventional conditions with a single catalyst, underwent hydroaminomethylation with this new system, and the pharmaceutical ibutilide was prepared in higher yield and under milder conditions than those reported with a single catalyst.

Rationale Whole-bone proteomic analyses rely on lengthy sample preparation including demineralization and digestion to break bone down into peptides to recover using mass spectrometry. However, microwave-assisted acid hydrolysis, a technique used in proteomic analyses of other soft tissues and cells, will combine both demineralization and digestion and only take minutes. Methods To test microwave-assisted hydrolysis on whole moose bone, we microwaved five concentrations of acetic and formic acids (15%, 12.5%, 10%, 7.5% and 5%) for three times (10, 20 and 30 min) at 140°C using an ETHOS UP high performance microwave digestion system. Peptides were injected and separated using Thermo BioBasic C18 columns and detected with an LTQ Orbitrap Velos mass spectrometer. We searched the raw data on PEAKS 8.5 against the white-tailed deer database. Results Formic acid hydrolysis led to the most complete digestion, and therefore the highest number of peptide spectrum matches, more protein groups and better sequence coverage for collagenous proteins. However, for the formic acid samples there is a tradeoff with digestion completeness and a higher incidence of in vitro modifications (i.e. formylation) that are not induced using acetic acid. Acetic acid has greater cleavage specificity and higher sequence coverage for non-collagenous proteins. Conclusions Depending on the goals of analysis, there are benefits and drawbacks to using both acetic acid and formic acid. Overall, microwave-assisted acid hydrolysis was successful in demineralizing and digesting bone fragments to considerably speed up the preparation for bottom-up proteomics analysis.

We investigated the effects of moisture, rain damage, and additives on fermentation quality and vitamins change in alfalfa (Medicago sativa L.) silage. Two moisture content alfalfa (53 and 72% moisture content) were received artificial rain damage. Then, the formic acid and L. plantarum (LP) + sucrose were added. After 30 days, silos were opened. The significant interaction between rain damage and moisture content were found for nutrients and fermentation characteristics in alfalfa silage, which showed more rain damage effects occurred in 53% moisture content rather than in 72% moisture content material. Treatment with LP + sucrose had lower pH value, ammonia-N/total N, and higher lactic acid content compared to the other additives. The concentrations of α-tocopherol and phylloquinone increased, but thiamine, riboflavin, and ascorbic acid decreased in alfalfa silage treated with LP + sucrose, especially when the alfalfa was subjected to rain damage. The highly significant correlations were found between pantothenic acid (VB5) and pH, and between VB5 and lactic acid. The improvements in fermentation quality by adding of LP + sucrose was observed after rain damage for either 53% or 72% moisture alfalfa, which can be a guideline for forage processor who ensiles alfalfa after rain damage.

The human gut microbiota is a crucial factor for the host's physiology with respect to health and disease. Metagenomic shotgun sequencing of microbial gut communities revealed that Prevotella copri is one of the most important players in the gastrointestinal tract of many individuals. Because of the importance of this bacterium we analyzed the growth behavior and the central metabolic pathways of P. copri. Bioinformatic data, transcriptome profiling and enzyme activity measurements indicated that the major pathways are based on glycolysis and succinate production from fumarate. In addition, pyruvate can be degraded to acetate and formate. Electron transport phosphorylation depends on fumarate respiration with NADH and reduced ferredoxin as electron donors. In contrast to Bacteroides vulgatus, P. copri showed a more pronounced dependency on the addition of CO2 or bicarbonate for biomass formation, which is a remarkable difference between P. copri and Bacteroides spp. with important implication in the context of gut microbial competition. The analysis of substrate consumption and product concentrations from many P. copri cultures with different optical densities allowed a prediction of the carbon and electron flow in the central metabolism and a detailed calculation of growth yields as well as carbon and redox balances.

Methanol mass poisoning is a global problem with high fatality rates and often severe sequelae in survivors. Patients typically present late to the hospital with severe metabolic acidosis followed by a rapid deterioration in their clinical status. The hypothesis 'Circulus hypoxicus' describes the metabolic acidosis following methanol poisoning as a self-enhancing hypoxic circle responsible for methanol toxicity. We wanted to test the validity of this hypothesis by an observational study based on 35 patients from the methanol outbreaks in Norway (2004) and the Czech Republic (2012). Comprehensive laboratory values, including S(serum)-methanol, S-formate, S-lactate, arterial blood gases, anion and osmolal gaps, were used in the calculations. Laboratory values and calculated gaps were compared to each other using linear regression. S-lactate and S-formate correlated better with the increased base deficit and anion gap than did S-formate alone. Base deficit rose to about 20 mmol/L and S-formate rose to 12 mmol/L prior to a significant rise in S-lactate - most likely caused by formate inhibition of mitochondrial respiration (type B lactacidosis). The further rise in S-lactate was not linear to S-formate most likely due to the self-enhancing pathophysiology, but may also be associated with hypotension in critically ill patients and variable ethanol drinking habits. Our study suggests that the primary metabolic acidosis leads to a secondary lactic acidosis mainly due to the toxic effects of formate. The following decline in pH will further increase this toxicity. As such, a vicious and self-enhancing acidotic circle may explain the pathophysiology in methanol poisoning, namely the 'Circulus hypoxicus'.

An analytical method for the simultaneous determination of 13 mycotoxins in feed by magnetic dispersive solid-phase extraction combined with ultra high performance liquid chromatography and tandem mass spectrometry was developed. The samples were extracted with acetonitrile/water (80:20, v/v, containing 3% acetic acid), and separated by centrifugation after salting-out, and then treated with magnetic adsorbents to remove interferences. The separation of target mycotoxins was performed on an ACQUITY UPLC HSS T3 column using a mobile phase consisting of 1 mmol/L ammonium acetate with 0.1% formic acid and methanol by gradient elution. Good linearities for the 13 mycotoxins were achieved with correlation coefficients over 0.99, and the recoveries of mycotoxins were in the range of 89.3–112.6% at spiking at levels of 5, 20 and 100 μg/kg, with relative standard deviations of 0.9–10.4%. Based on the functional magnetic materials (MDN@Fe3O4, PSA@Fe3O4, ZrO2@Fe3O4) applied in dispersive solid-phase extraction, the pretreatment process is more convenient and it is beneficial to reduce the experimental cost by reusing the recycled magnetic materials. It is a simple, rapid and environmentally friendly analytical method for the determination of mycotoxins in feed. This article is protected by copyright. All rights reserved

The selective electroreduction of CO2 to formate (or formic acid) is currently of great interest in the field of renewable energy utilization. In this work, we designed a sulfur-doped Cu2O-derived Cu catalyst, and showed that the presence of sulfur could significantly tune the selectivity of Cu from producing a myriad of CO2 reduction products to almost exclusively formate. Sulfur is doped into the Cu catalysts by dipping the Cu substrates into ammonium polysulfide solutions. Catalyst films with the highest sulfur content of 2.7 atom% showed the largest formate current density (jHCOO-) of -13.9 mA/cm2 at -0.9 V vs. RHE, which is ~46 times larger than that previously reported on Cu(110) surfaces. At -0.8 V vs. RHE, the Faradaic efficiency of formate was maintained at ~75% for 12 h of continuous electrolysis. By analyzing how the jHCOO- and jH2 of the catalysts evolved with different sulfur content, we show that sulfur doping efficaciously increases formate production, while suppressing the hydrogen evolution reaction. Ag-S and Cu-Se catalysts did not exhibit any significant enhancement towards the reduction of CO2 to formate. This demonstrates clearly that sulfur and copper acted synergistically to promote the selective formation of formate. A hypothesis of the role of sulfur is proposed and discussed.

There has been much effort exerted to reduce one carbon (C1) gas emission to address climate change. As one promising way to more conveniently utilize C1 gas, several technologies have been developed to convert C1 gas into useful chemicals such as formic acid (FA). In this study, systems metabolic engineering was utilized to engineer Mannheimia succiniciproducens to efficiently utilize FA. 13 C isotope analysis of M. succiniciproducens showed that FA could be utilized through formate dehydrogenase (FDH) reaction and/or the reverse reaction of pyruvate formate lyase (PFL). However, the naturally favored forward reaction of PFL was found to lower the SA yield from FA. In addition, FA assimilation via FDH was found to be more efficient than the reverse reaction of PFL. Thus, the M. succiniciproducens LPK7 strain, which lacks in pfl, ldh, pta, and ack genes, was selected as a base strain. In silico metabolic analysis confirmed that utilization of FA would be beneficial for the enhanced production of SA and suggested FDH as an amplification target. To find a suitable FDH, four different FDHs from M. succiniciproducens, Methylobacterium extorquens, and Candida boidinii were amplified in LPK7 strain to enhance FA assimilation. High-inoculum density cultivation using 13 C labeled sodium formate was performed to evaluate FA assimilation efficiency. Fed-batch fermentations of the LPK7 (pMS3-fdh2 meq) strain was carried out using glucose, sucrose, or glycerol as a primary carbon source and FA as a secondary carbon source. As a result, this strain produced 76.11 g/L SA with the yield and productivity of 1.28 mol/mol and 4.08 g/L/h, respectively, using sucrose and FA as dual carbon sources. The strategy employed here will be similarly applicable in developing microorganisms to utilize FA and to produce valuable chemicals and materials from FA.

Here, we found that formate, an essential one-carbon metabolite, activates superoxide (O2·-)/hydrogen peroxide (H2 O2 ) release from mitochondria. Sodium formate (30 μm) induces a significant increase in O2·-/H2 O2 production in liver mitochondria metabolizing pyruvate (50 μm). At concentrations deemed to be toxic, formate does not increase O2·-/H2 O2 production further. It was observed that the formate-mediated increase in O2·-/H2 O2 production is not associated with cytochrome c oxidase (COX) inhibition or changes in membrane potential and NAD(P)H levels. Sodium formate supplementation increases phosphorylating respiration without altering proton leaks. Finally, it was observed that the 2-oxoglutarate dehydrogenase (OGDH) inhibitors 3-methyl-2-oxovaleric acid (KMV) and CPI-613 inhibit the formate-induced increase in pyruvate-driven ROS production. The importance of these findings in one-carbon metabolism and physiology are discussed herein.

The enzymatic electrosynthesis of formic acid from the reduction of carbon dioxide (CO2) by using formate dehydrogenase (FDH) as a catalyst at the cathode in both its free and immobilized forms was studied in detail in a bioelectrochemical system (BES). The essential role of solubilizing CO2 for its conversion was also studied by adding carbonic anhydrase (CA) to the FDH enzyme in both its free and immobilized forms. FDH alone in the free form showed large variation in the reduction current [(−6.2±3.9) A m−2], whereas the immobilized form showed less variation [(−3.8±0.5) A m−2] due to increased enzyme stability. The addition of CA with FDH increased the consumption of the current in both forms due to the fact that it allowed rapid dissolution of CO2, which made it available for the catalytic reaction with FDH. Remarkably, stable consumption of the current was observed throughout the operation if both CA and FDH were immobilized onto the electrode [(−3.9±0.2) A m−2]. Product formation by the immobilized enzyme was also continued for three repetitive cycles, which revealed the longevity of the enzyme after immobilization. The recyclability of NADH (NAD=nicotinamide adenine dinucleotide) was also clearly evidenced on the derivative voltammetric signature. Extension of this study for continuous and long-term operation may reveal more possibilities for the rapid capture and conversion of CO2.

Fabric-based microfluidic fuel cells (MFCs) serve as a novel, cost-efficient alternative to traditional FCs and batteries, since fluids naturally travel across fabric via capillary action, eliminating the need for an external pump and lowering production and operation costs. Building on previous research with Y-shaped paper-based MFCs, fabric-based MFCs mitigate fragility and durability issues caused by long periods of fuel immersion. In this study, we describe a microfluidic fabric-based direct formate fuel cell, with 5 M potassium formate and 30% hydrogen peroxide as the anode fuel and cathode oxidant, respectively. Using a two-strip, stacked design, the optimized parameters include the type of encasement, the barrier, and the fabric type. Surface contact of the fabric and laminate sheet expedited flow and respective chemical reactions. The maximum current (22.83 mA/cm2 ) and power (4.40 mW/cm2 ) densities achieved with a 65% cotton/35% polyester blend material are a respective 8.7% and 32% higher than previous studies with Y-shaped paper-based MFCs. In series configuration, the MFCs generate sufficient energy to power a handheld calculator, a thermometer, and a spectrum of light-emitting diodes.

A series of new imidazoline-based iridium complexes has been developed for hydrogenation of CO2 and dehydrogenation of formic acid. One of the proton-responsive complexes bearing two −OH groups at ortho and para positions on a coordinating pyridine ring (3 b) can catalyze efficiently the chemical fixation of CO2 and release H2 under mild conditions in aqueous media without using organic additives/solvents. Notably, hydrogenation of CO2 can be efficiently carried out under CO2 and H2 at atmospheric pressure in basic water by 3 b, achieving a turnover frequency of 106 h−1 and a turnover number of 7280 at 25 °C, which are higher than ever reported. Moreover, highly efficient CO-free hydrogen production from formic acid in aqueous solution employing the same catalyst under mild conditions has been achieved, thus providing a promising potential H2-storage system in water.

Direct synthesis of dimethyl isosorbide (DMI) from d-sorbitol via dimethyl carbonate (DMC) chemistry is herein first reported. High yield of DMI was achieved using the nitrogen superbase 1,5,7-triazabicyclo[4.4.0]dec-5-ene (TBD) as catalyst and performing the reaction in a stainless steel autoclave by increasing the temperature from 90 to 200 °C. In this procedure, DMC features its full capacity acting in the different steps of the process as carboxymethylating, leaving-group (cyclization), and methylating agent; DMC is also employed as the reaction media.