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[Study of the inhibitory effect of acetylene on the biological methane formation in a paddy soil (author's transl)].

A 0.05 atm partial pressure of acetylene stopped all the CH4 evolution during anaerobic paddy soil incubations when different carbon substances were added. It was shown proof that acetylene did not disturbe the volatil fatty acids yielding but prohibited the methane forming bacteria from doing future use of those. Acetylene does inhibit methane forming bacteria. Besides, the growth of a Methanosarcina strain was stopped by a 0.01 atm partial pressure of acetylene, and resting cells lost 98 per cent of their activity when the acetylene concentration was 0.5 x 10-3 M in the liquid. This acetylene inhibition can be used for studying the metabolism of methane forming bacteria.

Acetates↗

[Combined construction of rainwater catchment techniques with methane pool and greenhouse].

A combined structure of rainwater catchment, methane pool and reformed greenhouse was constructed to improve water use efficiency. The results showed that the combination of rainwater catchment, reserving techniques and water saving measures was efficient to resolve the problem of water deficiency in the semi-arid region with precipitation of 250-550 mm. The combined structure of water catchment engineering, greenhouse and methane pool redistributed light, temperature, water and heat to develop higher benefits economic crops, and improved water use efficiency, which made it possible to develop high yield, and best quality agriculture in this region. Through optimizing the ecological factor in greenhouse, the water use was reduced by 50%-70%, and the soil temperature and night temperature were raised. This combined construction also improved productivity in greenhouse, and efficiently controlled plant diseases and insect pests. Methane pool provided CO2 and part energy source. The shady area back of the greenhouse was exploited by growing shady crops and epiphyte, which contributed to exchange energy and CO2 between light and shady house. This model could be used as water catchment agricultural development model in the region, and some other new approaches to water catchment agriculture in semi-arid region were discussed in this paper.

Agriculture↗

[Ethanol formation by methane-utilizing bacteria at ethane co-metabolism].

It was established, that EDTA (1.0 mM) and formamide (100 mM) are inhibitors of methanol dehydrogenase in Methylobacter luteus 12b, Methylomonas rubra 15sh and Methylococcus thermophilus 111p. The investigated strains co-metabolised ethane with the use of formate as the co-substrate. The application of formamide (or EDTA) as inhibitors of methanol dehydrogenase prevented from further transformation of ethanol and resulted in accumulation of extracellular ethanol. It was shown, that M. rubra 15sh accumulated extracellular ethanol under cultivation in a chemostate. The carried out researches have shown a regulation path of co-metabolism process of hydrocarbons by methane utilizing bacteria. Using the specific inhibitors of methanol dehydrogenase and a source of reducing agent (energy) for methane monooxygenase with the help of the cells of methane-oxidizing bacteria it is possible to obtain from ethane or other hydrocarbons the products of their monooxygenation--alcohols.

Ethane↗

[Influence of products of thermophilous methane fermentation on the mixed lactic acid bacterial culture].

The effect of products resulting from thermophilous methane fermentation on the accumulation and ratio of different lactic acid bacteria (Str. lactis, Str. cremoris, Str. diacetilactis, Str. diacetil. var. acetoinicus, Leuc. citrovorum8 L. plantarum) was investigated upon their combined cultivation in milk and serum. In combined cultivation of the streptococcal culture the rate of growth of a single strain differed from that in separate cultivation. Str. cremoris was accumulated with the highest rate during its separate cultivation and with the lowest rate during its combined cultivation. An addition of products of thermophilous methane fermentation to the combined culture of streptococci of different strains increased the growth rate and accumulation of each strain. At the same time the products influenced differently the quantitative ratios of various strains in the resulting biomass. For instance, the relative content of Str. cremoris and Leuc. citrovorum decreased insignificantly and that of Str. lactis and Str. diacetil. var. acetoin.increased. The combined cultivation of the five streptococcal strains with L. plantarum in the medium containing no additions of thermophilous methane fermentation products increased the relative content of L. plantarum from the beginning of fermentation. In the medium containing the additions of this parameter varied sinusoidally with a minimum of 6 to 9 hours. Possible mechanisms of these changes are discussed.

Culture Media↗

[Utilization of methane and carbon dioxide by symbiotrophic bacteria in gills of Mytilidae (Bathymodiolus) from the Rainbow and Logachev hydrothermal fields on the Mid-Atlantic Ridge].

Bivalve mollusks Bathymodiolus asoricus and Bathymodiolus puteoserpentis collected from the Rainbow and Logachev hydrothermal fields during dives of Mir 1 and Mir 2 deep-sea manned submersibles were studied. Rates of methane oxidation and carbon dioxide assimilation in mussel gill tissue were determined by radiolabel analysis. During oxidation of 14Ch4, radiocarbon was detected in significant quantities not only in carbon dioxide but also in dissolved organic matter, most notably 14C-formate and 14C-acetate, occurring in a 2:1 ratio. Activities of hexulose-phosphate synthase, phosphoribulokinase, and ribulose 1,5-bisphosphate carboxylase were shown in the soluble fraction of gill tissue cells. At the same time, no activity of hydroxypyruvate reductase--the key enzyme of the serine pathway of C1-assimilation--was detected. The results of PCR amplification using genetic probes for membrane-bound methane monooxygenase (pmoA) and methanol dehydrogenase (mxaF) attest to the presence of the genes of these enzymes in the total DNA extracted from gill samples. However, no appropriate PCR responses were obtained with the mmoX primer system, which is a marker for soluble methane monooxygenase. All samples studied showed amplification with primers for the genera Methylobacter and Methylosphaera. At the same time, no genes specific to the genera Methylomonas, Methylococcus, Methylomicrobium, or Methylosinus and Methylocystis were detected. Electron microscopic examinations revealed the presence of two groups of endosymbiotic bacteria in the mussel gill tissue. The first group was represented by large cells possessing a complex system of cytoplasmic membranes, typical of methanotrophs of morphotype I. The other type of endosymbionts, having much smaller cells and lacking intracellular membrane structures, is likely to be constituted by sulfur bacteria.

Alcohol Oxidoreductases↗

[Radioisotope method of determining the intensity of bacterial methane formation].

The rate of methane formation by bacteria was determined by the isotope tracer technique using labeled carbon in the form of bicarbonate. The rate of methane production in the ooze deposits of the Kuznechikha lake was calculated by this technique to constitute 7.46 to 7.70 cm3 CH4 per 1 litre per day. The rate of the process was shown to depend on the number of methane-producing bacteria.

Bacteria↗

Liquefaction and methanization of solid and liquid coffee wastes by two phase anaerobic digestion process.

This study attempted to investigate the feasibility of volatile fatty acid (VFA) production from coffee pulp hydrolyse, and further to determine the potential of methanization of both the pre-acidified effluent and the coffee wastewater. The experiments were carried out in 2 completely mixed reactors, each one with a working volume of 4 litres. Coffee pulp was used as substrate in the acidogenic reactor and different mixtures of pulper and wash-water and pre-acidified effluent in the methanogenic one. The acidogenic and methanogenic reactors were operated at an organic loading rate of 5 COD g x l(-1) x d(-1) and 0.5 COD g x l(-1) x d(-1). The total, soluble and VFA's effluent COD concentrations of the acidogenic reactor present average values of 57.75, 17.00 and 13.92 g x l(-1) respectively. Under these experimental conditions, 23% (COD based) of coffee pulp was hydrolysed with a rate of 1.32 gCOD x l(-1) x d(-1) and the soluble fraction was transformed to VFA's with an acidification efficiency of 82%. Total VFA's concentration reached a value of 13.9 gCOD x l(-1), and acetate, propionate, butyrate and valerate represented 52%, 28%, 9% and 11% respectively of the liquid phase COD. In the methanogenic reactor, COD removal and methanization of fresh coffee wastewater, pre-acidified effluent and both combined occur with an efficiency of 85% to 95% respectively, with a characteristic biogas composition of 80% CH4 and 20% CO2. These results show that a humid coffee "Beneficio" processing daily 23 tons of cherry coffee (fresh fruit), equipped with a two stage anaerobic digestion process could generate at least 1,886 CH4 m3 x d(-1). This represents an increase in methane production by a factor 3 to 5 compared to a "Beneficio" using anaerobic digestion only for the treatment of its wastewater.

Bacteria, Anaerobic↗

[Gas chromatographic method of determining the intensity of microbiological oxidation of methane in reservoirs].

A method of gas chromatography is described for determining the rate of microbial methane oxidation. In the Rybinskoye Reservoir, the rate of methane oxidation in the microaerobic zone of water layers near the bottom over the silted former channel of the Mologa River was found to be 51.1 to 86.2 mcl of CH4 per litre per day at 2--3 degrees C, constituting 8--16% of the original methane content.

Chromatography, Gas↗

[The biogeochemical cycle of methane in the coastal zone and littoral of the Kandalaksha Bay of the White Sea].

Microbiological and biogeochemical investigations of the processes of methane production (MP) and methane oxidation (MO) in the coastal waters and littoral of the Kandalaksha Bay of the White Sea were carried out. The studies were conducted in the coastal zones and in the water areas of the Kandalaksha Preserve, Moscow University White Sea Biological Station, and Zoological Institute (RAS) Biological Station in August, 1999, 2000, and 2001 and in March, 2001. The rate of CO2 assimilation in the shallow and littoral sediments was 35-27800 microg C/(dm3 day) in summer and 32.8-88.9 microg C/(dm3 day) in winter. The maximal rates of MP were observed in the littoral sediments in the zone of macrophyte decomposition, in local depressions, and in the estuary of a freshwater creak (up to 113 microl/(dm3 day)). The maximal level of MO was observed in the shallow estuarine sediments (up to 2450 microl/(dm3 day)). During the winter season, at the temperature of -0.5 to 0.5 degrees C, the MP rate in the littoral sediments was 0.02-0.3 microl/(dm3 day), while MO rate was 0.06-0.7 microl/(dm3 day). The isotopic data obtained indicate that the C(org) of the mats and of the upper sediment layers is enriched with the heavy 13C isotope by 1-4 per thousand as compared to the C(org) of the suspension, comprised on 33.5-34.3% of phytoplankton. A striking difference was found between the levels of methane emission by the typical littoral microlanscapes. In fine sediments, the average emission was 675 microl CH4/(m2 day), in the stormy discharge stretch sediments it was 1670 microl CH4/(m2 day), and under the stones and in silted pits, 1370 microl CH4/(m2 day). The calculation performed with consideration of the microlandscape areas with a high production allowed the CH4 production of 1 km2 of the littoral to be estimated as 192-300 1 CH4/(km2 day).

Bacteria↗

Production of surface active compounds from methane.

Methane utilizing bacterial strain was isolated and found to produce a surface active compound while growing on methane. The maximum production of the surface active compound depended on pressure of methane and air in the reactor. The crude surface active compound was extracted from the cell free broth and its surface active and emulsification properties were studied. The purified surface active compound had a critical micelle concentration of 150 mg/l at pH 7.0. On the basis of surface tension reduction, emulsification property and IR-spectrophotometric results this surface active compound was characterized as a glycolipid.

Methane↗

Production of hydrogen and methane from wastewater sludge using anaerobic fermentation.

The hydrogen and methane were produced from wastewater sludge using a Clostridium strain. The original sludge and the pre-treated (acidified, sterilized, freeze/thawed, and sonicated) sludges were tested. Some pre-treatment could enhance hydrogen yield, and the other tests could enhance methane yield. Hydrogen yield followed freeze/thawed>acidified>sterilized>original sludge>sonicated; while methane yield followed sonicated>freeze/thawed>sterilized>acidified>original sludge. The production and consumption of acetate correlated closely with the trends in both yields.

Acetic Acid↗

Methane production in a 100-L upflow bioreactor by anaerobic digestion of farm waste.

Manure waste from dairy farms has been used for methane production for decades, however, problems such as digester failure are routine. The problem has been investigated in small scale (1-2 L) digesters in the laboratory; however, very little scale-up to intermediate scales are available. We report production of methane in a 100-L digester and the results of an investigation into the effect of partial mixing induced by gas upflow/recirculation in the digester. The digester was operated for a period of about 70 d (with 16-d hydraulic retention time) with and without the mixing induced by gas recirculation through an internal draft tube. The results show a clear effect of mixing on digester operation. Without any mixing, the digester performance deteriorated within 30-50 d, whereas with mixing continuous production of methane was observed. This study demonstrates the importance of mixing and its critical role in design of large scale anaerobic digesters.

Agriculture↗

[Localization of energy generators in methane oxidizing bacteria].

Cytochromes a, b and c, and their quantitative distribution in the cells, were studied by means of differential spectra in obligate methane oxidizing bacteria, Methylosinus trichosporium with the serine pathway of methane carbon assimilation and Methylomonas agile with the ribulose phosphate pathway of methane carbon assimilation, and different types of topography of intracytoplasmic membranes. The membranes are involved in processes of coupled respiration which was confirmed by cytochemical reactions employed for studying the terminal step of the electron transport chain and Mg2+-stimulated ATPase.

Cytochromes↗

Accumulation of methane, acetone, and nitrogen in the inspired gas during closed-circuit anesthesia.

During closed-circuit anesthesia, the patient's inspired gas may become progressively contaminated by nonanesthetic gases. We studied the concentrations of methane, acetone, and nitrogen as nonanesthetic gas contaminants in the circuit gas of 16 cases during closed-circuit anesthesia. After a "short" period of denitrogenation (6-8 min), average nitrogen concentration in the closed circuit increased from 6.4 to 16.2%, methane from 4.3 to 22.4 ppm, and acetone from 0.3 to 2.2 ppm. After "long" denitrogenation (33 min), average nitrogen concentration in the closed circuit increased from 1.0 to 5.1%, methane from 3.7 to 17.9 ppm, and acetone from 1.3 to 5.9 ppm. It is concluded that gases stored in tissues or produced within the body can appear in the patient's expired gas during closed-circuit anesthesia. Intermittent flushing of the circuit with high flow gases is suggested to remove these contaminants.

Acetone↗

[Catalytic properties of immobilized cells of methane-oxidizing bacteria].

The article deals with conditions for immobilization of methane-oxidizing bacteria cells as well as with catalytic properties of the immobilized cells. The method of immobilization in polyacrylamide gel is shown to be not suitable for cells of methane-oxidizing bacteria. The greatest number of cells (85%) is immobilized on sylochrome modified by cyanuric chloride. However, the catalytic properties of the methane-oxidizing bacteria are better retained when the bacteria are immobilized on sylochrome modified by isocyanate. A stand installation is created for studying the catalytic properties of the immobilized cells oxidizing substrates in the gas phase.

Acrylamides↗

[Transformation of substrates not used for growth by the immobilized methane-oxidizing bacteria].

The article deals with the theoretical problems of obtaining certain substrates transformation products using immobilized cells of the methane-oxidizing bacteria. This process is based on the ability of these bacteria to cometabolize the substrates which are not used for growth. The methane-oxidizing bacteria are shown to utilize ethane during growth on methanol. Cometabolism of ethane and methanol occurs due to the presence of conjugated points of these substances oxidizing reactions as well as of the oxidation products assimilation reactions. The optimal concentrations of ethane and methanol are established for the methane-oxidizing bacteria growth and for formation of the ethane oxidation products by means of the immobilized cells.

Ethane↗

Carbon dioxide reduction factor and methanopterin, two coenzymes required for CO2 reduction to methane by extracts of Methanobacterium.

Carbon dioxide reduction (CDR) factor is contained in a low molecular weight fraction of cell extract that is required for methane production from CO2 by resolved cell extracts. This fraction has been separated into two components both of which have been highly purified. One component is methanopterin, and for the other component the name CDR factor is retained. No known coenzymes tested substitute in the methane-producing assay for CDR factor and methanopterin, both of which are stable to boiling and exposure to air. The ultraviolet-visible spectrum of CDR factor has a peak at 273 nm, a shoulder at 280 nm, and at pH 1, a peak at 219 nm. The ultraviolet-visible spectrum of methanopterin isolated from the CDR fraction is similar to the spectrum previously reported for this compound (Keltjens, J. T., and Vogels, G. D. (1981) in Microbial Growth on C1 Compounds (Dalton, H., ed) pp. 152-158, Heyden and Son, Ltd., London). The addition of CDR factor (0.8 micrograms) and methanopterin (50 micrograms) to the assay mixture increased by 12-fold the amount of methane formed from CO2.

Carbon Dioxide↗

[Comparative characteristics of the enzymatic systems of methane-utilizing bacteria that oxidize NH2OH and CH3OH].

Methane hydroxylase (MH) from the obligate methane assimilating culture of Methylococcus thermophilus catalyses oxygenation of both CH4+ and NH4+; therefore, we studied the specificity of enzyme systems catalysing the subsequent oxidation of compounds produced upon the oxygenation of these substrates (CH3OH and NH2OH). CH3OH and NH2OH were shown to be oxidized by different enzymes, viz. methanol dehydrogenase (MD) and hydroxylamine oxidase (HO), respectively. Similar to MH, MD is characterized by the absence of strict substrate specificity, and catalyses oxidation of primary alcohols other than methanol, rather than hydroxylamine. HO catalyses oxidation of hydroxylamine rather than methanol and possesses the activity of hydroxylamine:cytochrome c oxidoreductase. The constitutive character of HO from the methane assimilating bacteria and the substrate specificity of the enzyme suggest that a lithotrophic pathway for producing energy operates in these bacteria. The HO of Methylococcus thermophilus is similar in certain properties to the HO of the nitrifying bacterium Nitrosomonas europaea.

Alcohol Oxidoreductases↗