Comments on the Role of H2S in the Chemistry of Earth's Early Atmosphere and in Prebiotic Synthesis

Free energy calculations and experimental measurements have been used to show that H₂S/CO₂ mixtures outgassing from a prebiotic Earth's crust would have produced a reducing gas mixture containing CO, H₂, H₂O, and S _x as principal components. Due to rapid recombination of H₂, CO, and S _x to H₂S and CO₂ on cooling from a high temperature to ambient conditions, reducing components would have been retained only if efficient quenching of the reduced gas mixture had been possible. Consequently, subsea vents or vents with efficient infusion of water would have been ideal sites for retention of reduced species and for prebiotic organic synthesis. It is suggested that C/H/O/S ratios are important factors in controlling the degree of prebiotic organic synthesis and, hence, the emergence of life, since if oxygen is abundant, CO₂ and SO₂ would have been dominant species. Received: 5 March 1997 / Accepted: 15 December 1997     

Evaluation of H2O2 and pH in exhaled breath condensate samples: methodical and physiological aspects

Abstract

This veterinary study is aimed at further standardization of H₂O₂ and pH measurements in exhaled breath condensate (EBC). Data obtained in the study provide valuable information for many mammalian species including humans, and may help to avoid general pitfalls in interpretation of EBC data. EBC was sampled via the ‘ECoScreen’ in healthy calves (body weight 63–98 kg). Serum samples and condensates of ambient (indoor) air were collected in parallel. In the study on H₂O₂, concentrations of H₂O₂ in EBC, blood and ambient air were determined with the biosensor system ‘ECoCheck’. In EBC, the concentration of H₂O₂ was found to be dependent on food intake and increased significantly in the course of the day. Physiologically, lowest H₂O₂ concentrations at 06:00 varied within the range 138–624 nmol l^?1 EBC or 0.10–0.94 nmol per 100 l exhaled breath and individual concentrations were significantly different indicating a remarkable intersubject variability. Highly reproducible results were seen within each subject (three different days within 4 weeks). No correlation existed between H₂O₂ concentrations in EBC and blood, and EBC–H₂O₂ was not influenced by variables of spontaneous breathing. Further results confirmed that standardization of H₂O₂ measurements in EBC requires (1) the re-calculation of the concentration exhaled per 100 l exhaled breath (because the analyzed concentration in the liquid condensate underlies multiple methodological sources of variability given by the collection process), and (2) subtracting the concentration of inspired indoor H₂O₂. In the study on pH use of the ISFET electrode (Sentron, the Netherlands) and a blood gas analyzer ABL 550 (Radiometer, Denmark) led to comparable results for EBC–pH (r=0.89, R²=79.3%, p≤0.001). Physiological pH data in non-degassed EBC samples varied between 5.3 and 6.5, and were not significantly different between subjects, but were significantly higher in the evening compared with the morning. EBC–pH was not dependent on variables of spontaneous breathing pattern or ambient conditions, and no significant correlation was found between serum and EBC for pH.     

Characterization of aMethanosarcina strain isolated from goat feces,and that grows on H2-CO2 only after adaptation

AMethanosarcina species, designated strain ChGul, was isolated from goat feces; this is the first fully described pure culture ofMethanosarcina obtained from feces. Antigenic fingerprinting suggests that isolate ChGul is a new immunotype. The mol% G + C content of DNA was 42.2%. Strain ChGul grew on methanol, methylamines, and acetate in a minimal salts medium. It grew on H₂-CO₂ only after adaptation. Growth occurred as a milky-white suspension and contained cells mostly in doublets and quadruplets of irregular cocci; many cells contained phase bright spots typical of gas vacuoles. The isolate did not grow on formate, or CO₂ plus isopropanol, ethanol, or acetone as substrates and did not produce methane from formate. The optimum growth temperature was 35–37°C, and optimum pH was 6.2–6.8. ChGul is unusually sensitive to sulfide and has low tolerance for NaCl. Optimal levels of total sulfide and NaCl for growth were 0.5 mM and 20–40 mM, respectively. Since ChGul requires adaptation for growth on H₂-CO₂ and cannot use formate, it may be restricted to methylotropic or acetoclastic methanogenesis in the rumen, a function not observed in previously isolated rumen methanogens that use H₂-CO₂ and formate. Our work suggests that improper NaCl and sulfide concentrations, and cell lysis, may have made isolation of rumenMethanosarcina difficult in the past. It also underscores the need to evaluate feed compositions and media components for most probable number studies, with respect to NaCl and sulfide levels, to understand the role ofMethanosarcina in the rumen.     

Interaction of trout hemoglobin with H2O2: a chemiluminescence study

Erythrocytes from trout Salmo irideus are characterized by four different hemoglobin components (HbI, HbII, HbIII and HbIV), HbI and HbIV being predominant. In this study we describe the interaction between trout hemoglobin (HbI and HbIV) and H₂O₂ using a chemiluminescence assay. Our data show that the reaction of hemoglobins with H₂O₂ produces a time-limited and significant increase of chemiluminescence signal. The half-life of the decay of this chemiluminescence signal was characteristic for each type of hemoglobin used. These results indicate the formation of excited molecules related to the interaction between trout hemoglobin and H₂O₂. © 1997 John Wiley & Sons, Ltd.     

No acetogen is equal: Strongly different H2 thresholds reflect diverse bioenergetics in acetogenic bacteria

Acetogens share the capacity to convert H₂ and CO₂ into acetate for energy conservation (ATP synthesis). This reaction is attractive for applications, such as gas fermentation and microbial electrosynthesis. Different H₂ partial pressures prevail in these distinctive applications (low concentrations during microbial electrosynthesis [<40 Pa] vs. high concentrations with gas fermentation [>9%]). Strain selection thus requires understanding of how different acetogens perform under different H₂ partial pressures. Here, we determined the H₂ threshold (H₂ partial pressure at which acetogenesis halts) for eight different acetogenic strains under comparable conditions. We found a three orders of magnitude difference between the lowest and highest H₂ threshold (6 ± 2 Pa for Sporomusa ovata vs. 1990 ± 67 Pa for Clostridium autoethanogenum), while Acetobacterium strains had intermediate H₂ thresholds. We used these H₂ thresholds to estimate ATP gains, which ranged from 0.16 to 1.01 mol ATP per mol acetate (S. ovata vs. C. autoethanogenum). The experimental H₂ thresholds thus suggest strong differences in the bioenergetics of acetogenic strains and possibly also in their growth yields and kinetics. We conclude that no acetogen is equal and that a good understanding of their differences is essential to select the most optimal strain for different biotechnological applications.     

Relative Importance of H2 and H2S as Energy Sources for Primary Production in Geothermal Springs

Geothermal waters contain numerous potential electron donors capable of supporting chemolithotrophy-based primary production. Thermodynamic predictions of energy yields for specific electron donor and acceptor pairs in such systems are available, although direct assessments of these predictions are rare. This study assessed the relative importance of dissolved H₂ and H₂S as energy sources for the support of chemolithotrophic metabolism in an acidic geothermal spring in Yellowstone National Park. H₂S and H₂ concentration gradients were observed in the outflow channel, and vertical H₂S and O₂ gradients were evident within the microbial mat. H₂S levels and microbial consumption rates were approximately three orders of magnitude greater than those of H₂. Hydrogenobaculum-like organisms dominated the bacterial component of the microbial community, and isolates representing three distinct 16S rRNA gene phylotypes (phylotype = 100% identity) were isolated and characterized. Within a phylotype, O₂ requirements varied, as did energy source utilization: some isolates could grow only with H₂S, some only with H₂, while others could utilize either as an energy source. These metabolic phenotypes were consistent with in situ geochemical conditions measured using aqueous chemical analysis and in-field measurements made by using gas chromatography and microelectrodes. Pure-culture experiments with an isolate that could utilize H₂S and H₂ and that represented the dominant phylotype (70% of the PCR clones) showed that H₂S and H₂ were used simultaneously, without evidence of induction or catabolite repression, and at relative rate differences comparable to those measured in ex situ field assays. Under in situ-relevant concentrations, growth of this isolate with H₂S was better than that with H₂. The major conclusions drawn from this study are that phylogeny may not necessarily be reliable for predicting physiology and that H₂S can dominate over H₂ as an energy source in terms of availability, apparent in situ consumption rates, and growth-supporting energy.     

Biomass fast pyrolysis in a fluidized bed reactor under N2, CO2, CO, CH4 and H2 atmospheres

Biomass fast pyrolysis is one of the most promising technologies for biomass utilization. In order to increase its economic potential, pyrolysis gas is usually recycled to serve as carrier gas. In this study, biomass fast pyrolysis was carried out in a fluidized bed reactor using various main pyrolysis gas components, namely N₂, CO₂, CO, CH₄ and H₂, as carrier gases. The atmosphere effects on product yields and oil fraction compositions were investigated. Results show that CO atmosphere gave the lowest liquid yield (49.6%) compared to highest 58.7% obtained with CH₄. CO and H₂ atmospheres converted more oxygen into CO₂ and H₂O, respectively. GC/MS analysis of the liquid products shows that CO and CO₂ atmospheres produced less methoxy-containing compounds and more monofunctional phenols. The higher heating value of the obtained bio-oil under N₂ atmosphere is only 17.8 MJ/kg, while that under CO and H₂ atmospheres increased to 23.7 and 24.4 MJ/kg, respectively.     

Chlorobium limicola forma thiosulfatophilum: Biocatalyst in the Production of Sulfur and Organic Carbon from a Gas Stream Containing H2S and CO2

Chlorobium limicola forma thiosulfatophilum (ATCC 17092) was grown in a 1-liter continuously stirred tank reactor (800-ml liquid volume) at pH 6.8, 30°C, saturated light intensity, and a gas flow rate of 23.6 ml/min from a gas cylinder blend consisting of 3.9 mol% H₂S, 9.2 mol% CO₂, 86.4 mol% N₂, and 0.5 mol% H₂. This is the first demonstration of photoautotrophic growth of a Chlorobium sp. on a continuous inorganic gas feed. A significant potential exists for applying this photoautotrophic process to desulfurization and CO₂ fixation of gases containing acidic components (H₂S and CO₂).     

Soil H2-uptake in relation to soil properties and rhizobial H2-production

Summary The biological nature of soil H₂-consumption has been investigated. Soil microorganisms were capable to remove H₂ present in the gas phase at concentrations in the range of 200 ppm at rates varying between 0.2 and 1.0 l.min^–1. 100 g^–1. Free soil enzymes did not contribute significantly at the H₂ concentrations tested. Oxygen seemed to be the predominant electron acceptor. The influence of microbiological and physical soil properties on the H₂-uptake activity was examined for 38 soils.A highly significant correlation between biomass-C and H₂-uptake rate of the soil was noted, suggesting that the latter parameter might be useful as an indirect estimation of soil microbial biomass. The correlation was however not applicable for soils recently grown with legumes. Indeed, soya plants nodulated with aRhizobium strain with a weak hydrogen uptake capability, strongly increased the hydrogen oxidizing capability of the surrounding soil.     

Selection of Vaginal H2O2-Generating Lactobacillus Species for Probiotic Use

Lactobacilli are believed to contribute to the control of the vaginal microflora by different mechanisms such as production of antagonistic substances like lactic acid, bacteriocins, and H₂O₂. This paper describes the selection of H₂O₂-generating lactobacilli among 35 hydrophobic isolates from the human vagina. Lactobacillus crispatus F117, which generated the highest H₂O₂ level, was chosen to study: (a) the kinetics of H₂O₂ production considering different culture conditions, and (b) the effect of this metabolite on the growth of urogenital tract pathogens. The levels of H₂O₂ in L. crispatus supernatant increased during its growth and were maximum at the early stationary phase (3.29 mmol H₂O₂L⁻¹) under aerated conditions (agitated cultures). In nonagitated cultures there were no detectable levels of H₂O₂. L. crispatus F117 spent supernatant inhibited Staphylococcus aureus growth in plaque assay. Inhibition was due to H₂O₂ since catalase treatment of the supernatant suppressed inhibition. In mixed cultures performed with L. crispatus and S. aureus a significant decrease in pathogen growth was observed. The inhibitory effect depended on the initial inoculum of S. aureus. Further evaluation of the properties of L. crispatus F117 will be performed to consider its inclusion in a probiotic for local use in the vaginal tract. Received: 17 November 1998 / Accepted: 17 December 1998     

H2-CO2 Recirculation and pH Control for Growth of Methanogens in Mass Culture

We modified a fermentor (10-liter liquid volume) for the growth of anaerobic, H₂-CO₂-catabolizing bacteria. Gas in the fermentor (ca. 10% CO₂, 50% H₂, 40% CH₄) was recirculated by a diaphragm pump. During growth, the gas composition was maintained by the addition of a mixture of 80% H₂ and 20% CO₂, and this addition was controlled by a pH auxostat. During gas addition, gas was discharged from the recirculating gas stream and was collected by the displacement of an acidified salt solution.     

Microbiological hydrogen (H2) thresholds in anaerobic continuous-flow systems: Effects of system characteristics

Hydrogen (H₂) concentrations that were associated with microbiological respiratory processes (RPs) such as sulfate reduction and methanogenesis were quantified in continuous-flow systems (CFSs) (e.g., bioreactors, sediments). Gibbs free energy yield (ΔǴ ~ 0) of the relevant RP has been proposed to control the observed H₂ concentrations, but most of the reported values do not align with the proposed energetic trends. Alternatively, we postulate that system characteristics of each experimental design influence all system components including H₂ concentrations. To analyze this proposal, a Monod-based mathematical model was developed and used to design a gas–liquid bioreactor for hydrogenotrophic methanogenesis with Methanobacterium bryantii M.o.H. Gas-to-liquid H₂ mass transfer, microbiological H₂ consumption, biomass growth, methane formation, and Gibbs free energy yields were evaluated systematically. Combining model predictions and experimental results revealed that an initially large biomass concentration created transients during which biomass consumed [H₂]_L rapidly to the thermodynamic H₂-threshold (≤1 nM) that triggerred the microorganisms to stop H₂ oxidation. With no H₂ oxidation, continuous gas-to-liquid H₂ transfer increased [H₂]_L to a level that signaled the methanogens to resume H₂ oxidation. Thus, an oscillatory H₂-concentration profile developed between the thermodynamic H₂-threshold (≤1 nM) and a low [H₂]_L (~10 nM) that relied on the rate of gas-to-liquid H₂-transfer. The transient [H₂]_L values were too low to support biomass synthesis that could balance biomass losses through endogenous oxidation and advection; thus, biomass declined continuously and disappeared. A stable [H₂]_L (1807 nM) emerged as a result of abiotic H₂-balance between gas-to-liquid H₂ transfer and H₂ removal via advection of liquid-phase.     

H2O2 in plant peroxisomes: an in vivo analysis uncovers a Ca2+‐dependent scavenging system

Oxidative stress is a major challenge for all cells living in an oxygen‐based world. Among reactive oxygen species, H₂O₂, is a well known toxic molecule and, nowadays, considered a specific component of several signalling pathways. In order to gain insight into the roles played by H₂O₂ in plant cells, it is necessary to have a reliable, specific and non‐invasive methodology for its in vivo detection. Hence, the genetically encoded H₂O₂ sensor HyPer was expressed in plant cells in different subcellular compartments such as cytoplasm and peroxisomes. Moreover, with the use of the new green fluorescent protein (GFP)‐based Cameleon Ca²⁺ indicator, D3cpv–KVK–SKL, targeted to peroxisomes, we demonstrated that the induction of cytoplasmic Ca²⁺ increase is followed by Ca²⁺ rise in the peroxisomal lumen. The analyses of HyPer fluorescence ratios were performed in leaf peroxisomes of tobacco and pre‐ and post‐bolting Arabidopsis plants. These analyses allowed us to demonstrate that an intraperoxisomal Ca²⁺ rise in vivo stimulates catalase activity, increasing peroxisomal H₂O₂ scavenging efficiency.     

H2-Photoproduction by green algae: The significance of anaerobic pre-incubation periods and of high light intensities for H2-photoproductivity ofChlorella fusca

Using sodium-dithionite as an oxygen scavenger, the influences of different light intensities and periods of anaerobic pre-incubation in the dark on H₂-photoproductivity were studied with the green algaChlorella fusca. By measuring hydrogen production in the light using manometric and gas chromatographic methods the effectiveness of sodium dithionite in stabilizing photoproduction was established. For high rates of H₂-photoproduction high light intensities up to 30,000 lux (580 W m^-2) were necessary; these are comparable to those required for light saturation of oxygen photoproduction by this alga. AlthoughChlorella fusca produces H₂ immediately after transition to anaerobic conditions, the optimum rate of H₂ production was reached after a 5 h dark adaptation period only. The results obtained are discussed with respect to characteristics of H₂-photoproduction by green algae: the initial burst kinetics, the light saturation, and the obligate period of anaerobic adaptation. It is concluded that H₂-photoproduction byChlorella is an anaerobic photosynthetic process which occurs in the absence of CO₂ and can be experimentally stabilized by exogenous oxygen scavengers.Abbreviations DCMU (3-(3,4-Dichlorophenyl)-1,1-dimethylurea) - HEPES (2-[4-(2-Hydroxyethyl)-1-piperazinyl]ethanesulfonic acid)     

Effect on methane digestion of decreased H2 partial pressure by means of phototrophic or sulfate-reducing bacteria grown in an auxiliary reactor

Summary H₂ is a central metabolite in the process of methane digestion. In this study, the partial pressure of H₂ was decreased by sparging the gas phase of the digester through an auxiliary reactor in which a Rhodomicrobium vaniellii culture or a mixed culture of sulfate-reducing bacteria was allowed to develop at the expense of H₂ and CO₂ present in the biogas. The decrease of the H₂ concentration in the gas phase was significant. A 18–23 percent increase of the gas production rate and a concomitantly improved removal of volatile fatty acids from the mixed liquor was obtained. The sulfate-reducing bacteria appeared to be slightly more effective than the phototrophs. The results suggest that the increased biogas production rate is due to the decrease of propionic acid formation and the concomitant stimulation of propionate degradation.Abbreviations COD_t Total chemical oxygen demand - COD_s Soluble chemical oxygen demand - SS Suspended solids - DM Dry matter - VFA Volatile fatty acids     

Crystallographic and mutational analyses of cystathionine β‐synthase in the H2S‐synthetic gene cluster in Lactobacillus plantarum

Cystathionine β‐synthase (CBS) catalyzes the formation of l ‐cystathionine from l ‐serine and l ‐homocysteine. The resulting l ‐cystathionine is decomposed into l ‐cysteine, ammonia, and α‐ketobutylic acid by cystathionine γ‐lyase (CGL). This reverse transsulfuration pathway, which is catalyzed by both enzymes, mainly occurs in eukaryotic cells. The eukaryotic CBS and CGL have recently been recognized as major physiological enzymes for the generation of hydrogen sulfide (H₂S). In some bacteria, including the plant‐derived lactic acid bacterium Lactobacillus plantarum, the CBS‐ and CGL‐encoding genes form a cluster in their genomes. Inactivation of these enzymes has been reported to suppress H₂S production in bacteria; interestingly, it has been shown that H₂S suppression increases their susceptibility to various antibiotics. In the present study, we characterized the enzymatic properties of the L. plantarum CBS, whose amino acid sequence displays a similarity with those of O‐acetyl‐l ‐serine sulfhydrylase (OASS) that catalyzes the generation of l ‐cysteine from O‐acetyl‐l ‐serine (l ‐OAS) and H₂S. The L. plantarum CBS shows l ‐OAS‐ and l ‐cysteine‐dependent CBS activities together with OASS activity. Especially, it catalyzes the formation of H₂S in the presence of l ‐cysteine and l ‐homocysteine, together with the formation of l ‐cystathionine. The high affinity toward l ‐cysteine as a first substrate and tendency to use l ‐homocysteine as a second substrate might be associated with its enzymatic ability to generate H₂S. Crystallographic and mutational analyses of CBS indicate that the Ala70 and Glu223 residues at the substrate binding pocket are important for the H₂S‐generating activity.     

H2S degradation is reflected by both the activity and composition of the microbial community in a compost biofilter

In this study, 16S rRNA- and rDNA-based denaturing gradient gel electrophoresis (DGGE) were used to study the temporal and spatial evolution of the microbial communities in a compost biofilter removing H₂S and in a control biofilter without H₂S loading. During the first 81 days of the experiment, the H₂S removal efficiencies always exceeded 93% at loading rates between 4.1 and 30 g m⁻³ h⁻¹. Afterwards, the H₂S removal efficiency decreased to values between 44 and 71%. RNA-based DGGE analysis showed that H₂S loading to the biofilter increased the stability of the active microbial community but decreased the activity-based diversity and evenness. The most intense band in both the RNA- and DNA-based DGGE patterns of the H₂S-degrading biofilter represented the sulfur oxidizing bacterium Thiobacillus thioparus. This suggested that T. thioparus constituted a major part of the bacterial community and was an important primary degrader in the H₂S-degrading biofilter. The decreasing H₂S removal efficiencies near the end of the experiment were not accompanied by a substantial change of the DGGE patterns. Therefore, the decreased H₂S removal was probably not caused by a failing microbiology but rather by a decrease of the mass transfer of substrates after agglutination of the compost particles.     

Determination of free salsolinol concentrations in human urine using gas chromatography—mass spectrometry

The urine concentrations of free salsolinol were determined in six healthy volunteers, using a gas chromatographic—mass spectrometric method with electron-capture negative-ion chemical ionization after derivatization with pentafluoropropionyl anhydride. The sensitivity of this method allows the quantification of salsolinol concentrations of 0.55 pmol/ml. The synthesis of [²H₄]salsolinol from dopamine and [²H₄]acetaldehyde via a Pictet—Spengler condensation is described; [²H₄]salsolinol was used as the internal standard for salsolinol quantification. The urine concentrations of free salsolinol ranged from ca. 1 to 6 pmol/ml.     

Sensitive determination of bromhexine hydrochloride based on its quenching effect on luminol/H2O2 electrochemiluminescence system

In this paper, the electrochemiluminescence (ECL) behavior of luminol/H₂O₂ system in the presence of bromhexine hydrochloride (BrH) was investigated. It was found that the ECL intensity of luminol/H₂O₂ system on a platinum electrode could be intensely quenched by BrH owing to the scavenging superoxide radical ability of BrH, and therefore the sensitive determination of BrH was possible. Under optimal conditions, the quenched ECL intensity was linear to the concentration of BrH in a wide range of 0.08 to 500 μM, with a detection limit of 0.02 μM (signal‐to‐noise ratio (S/N) = 3). This ECL method possessed the merits of rapid, simple and sensitive, and was successfully applied to the BrH quantification in pharmaceutical preparations with satisfactory recoveries of 91.0 ± 4.0 to 106.5 ± 3.4%. The possible route of the quenched ECL of luminol/H₂O₂ in the presence of BrH was also discussed.