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[Quantitative study of biological denitrification in soils with the aid of acetylene. II.--Evolution of inhibitory effect of acetylene on N2O-reductase; influence of acetylene on denitrification rate and on nitrate immobilisation (author's transl)].

Study of kinetics of nitrate or nitrite disappearance in soils incubating with atmosphere with or without acetylene, shows that the presence of this gas increases the rate of nitrate or nitrite reduction, and therefore very probably the denitrification process. The decrease of inhibitory effect on N2O-reductase after incubation during 15 days appears to be due to a great extent at a biological transformation of this gas with a consecutive increase of CO2 production. On the other hand, acetylene does not seem to affect perceptibly nitrate immobilisation.

Acetylene↗

The effect of incomplete acetylene washout on cardiac output measurement using open circuit acetylene uptake.

The open circuit acetylene uptake method is a useful non-invasive means of measuring cardiac output. However, because of accumulation of inhaled acetylene in tissues, the cardiac output uptake is underestimated, if residual acetylene is not allowed to wash out completely in between measurements. We determined the effect of applying a correction factor that estimates mixed venous acetylene concentration from endtidal values to the calculation of cardiac output. This accounts for mixed venous acetylene present during measurements made before complete washout. Six healthy subjects performed steady-state exercise at approximately 30% and 60% of V(O2 max). Cardiac output measurements were made at each exercise intensity using the open circuit acetylene uptake method (inspired [acetylene] approximately 1%), with the first and last measurements having no detectible levels of acetylene in expired gas (reference measurement). Data were also obtained with immediate pre-measurement endtidal concentrations ranging from 3% to 15% of the inspired [acetylene], in random order in between. Oxygen consumption, carbon dioxide production and heart rate did not change significantly during testing at each exercise intensity. Reference cardiac output also did not change significantly and averaged 11.1+/-0.8 L/min at 30% of V(O2 max) and 16.5+/-2.0 L/min at 60% of V(O2 max). Uncorrected cardiac output measurements progressively underestimated cardiac output by 15% at the 3% of inspired endtidal [acetylene] and by over 50% at 15% [acetylene] (p<0.0001). However, when corrected for residual endtidal [acetylene], cardiac outputs were not significantly different from the reference measurements. The results of this study suggest that by accounting for residual endtidal acetylene in mixed venous blood, cardiac output can be accurately measured even when washout of acetylene is incomplete, allowing measurements as often as every 10-15 s.

Acetylene↗

Repeatability of the acetylene rebreathing method in measuring cardiac output: influence of acetylene concentration.

The acetylene rebreathing method is a reliable noninvasive method for estimation of cardiac output. However, the method is not commonly used for clinical purposes. This is due mainly to the fact that acetylene is explosive at concentrations above 2% and that acetylene at concentrations above 0.5% has an unpleasant taste. In the present study we have investigated whether it is possible to reduce the concentration of acetylene to 0.3% in the rebreathing bag without degrading the repeatability of the estimated cardiac output. The problem was elucidated theoretically and experimentally. A theoretical investigation was carried out by producing an acetylene rebreathing curve from a single alveolus lung model. The theoretical study showed that the random noise arising from the gas analyzer gives rise to less than 10% of the total variation of the estimated cardiac output. The results of the experimental study showed that it is possible to reduce the acetylene concentration to 0.3% without decreasing the reproducibility of the acetylene rebreathing method.

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Acetylene degradation by new isolates of aerobic bacteria and comparison of acetylene hydratase enzymes.

Aerobic acetylene-degrading bacteria were isolated from soil samples. Two isolates were assigned to the species Rhodococcus opacus, two others to Rhodococcus ruber and Gordona sp. They were compared with known strains of aerobic acetylene-, cyanide-, or nitrile-utilizing bacteria. The acetylene hydratases of R opacus could be measured in cell-free extracts only in the presence of a strong reductant like titanium(III) citrate. Expression of these enzymes was molybdenum-dependent. Acetylene hydratases in cell-free extracts of R ruber and Gordona spp. did not require addition of reductants. No cross-reactivity could be found between cell-free extracts of any of these aerobic isolates and antibodies raised against the acetylene hydratase of the strictly anaerobic fermenting bacterium Pelobacter acetylenicus. These results show that acetylene hydratases are a biochemically heterogeneous group of enzymes.

Acetylene↗

Acetylene, Not Ethylene, Inactivates the Uptake Hydrogenase of Actinorhizal Nodules during Acetylene Reduction Assays.

Acetylene reduction assays were shown to inactivate uptake hydrogenase activity to different extents in one Casuarina and two Alnus symbioses. Inactivation was found to be caused by C(2)H(2) and not by C(2)H(4). Acetylene completely inactivated the hydrogenase activity of intact root systems of Alnus incana inoculated with Frankia strain Avcl1 in 90 minutes, as shown by a drop in the relative efficiency of nitrogenase from 1.0 to 0.73. The hydrogenase of Frankia preparations (containing vesicles) and of cell-free extracts (not containing vesicles) from the same symbiosis was much more susceptible to acetylene inactivation. Cell-free extracts lost all hydrogenase activity after 5 minutes of exposure to acetylene. The hydrogenase activity of intact root systems of Casuarina obesa was less sensitive to acetylene than that of root systems of A. incana, since the relative efficiency of nitrogenase changed only from 1.0 to 0.95 over 90 minutes. Frankia preparations and cell-free extracts of C. obesa still retained hydrogenase activity after a 10 minute-exposure to acetylene.

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Acetylene inhibition of Azotobacter vinelandii hydrogenase: acetylene binds tightly to the large subunit.

Acetylene is a slow-binding inhibitor of the Ni- and Fe-containing dimeric hydrogenase isolated from Azotobacter vinelandii. Acetylene was released from hydrogenase during the recovery from inhibition. This indicates that no transformation of acetylene to another compound occurred as a result of the interaction with hydrogenase. However, the release of C2H2 proceeds more rapidly than the recovery of activity, which indicates that release of C2H2 is not sufficient for recovery of activity. Acetylene binds tightly to native hydrogenase; hydrogenase and radioactivity coelute from a gel permeation column following inhibition with 14C2H2. Acetylene, or a derivative, remains bound to the large 65,000 MW subunit (and not to the small 35,000 MW subunit) of hydrogenase following denaturation as evidenced by SDS-PAGE and fluorography of 14C2H2-inhibited hydrogenase. This result suggests that C2H2, and by analogy H2, binds to and is activated by the large subunit of this dimeric hydrogenase. Radioactivity is lost from 14C2H2-inhibited protein during recovery. The inhibition is remarkably specific for C2H2: propyne, butyne, and ethylene are not inhibitors.

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Nitrogenase. VI. Acetylene reduction assay: Dependence of nitrogen fixation estimates on component ratio and acetylene concentration.

Acetylene reduction, an assay for nitrogenase activity (nitrogen:(acceptor) oxidoreductase, EC 1.7.99.2), Is dependent on the ratio of the two protein components of nitrogenase as well as on C2H2 concentration. As the component I : component II ratio (based on activity) is increased, the C2H2 reduction : N2 fixation ratio decreases to a minimum of 3.4 and then increases. The minimum is found at a ratio near 1 : 1. At a component I : component II ratio of 20 : 1, the C2H2 reduction : N2 fixation ratio is 5.3. Acetylene exhibits substrate inhibition in assays for nitrogenase activity. Both the apparent Km and Ki for acetylene vary as a function of the relative concentrations of components I and II present in the assay. When the more labile component II is limiting in the assay and "saturating" levels of C2H2 (above 0.1 atm) are used, N2-fixation capacity may be greatly under-estimated.

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Analysis of acetylene reduction rates of soybean nodules at low acetylene concentrations.

It has been previously proposed that acetylene reduction data at subsaturating acetylene concentrations could be interpreted by use of the Michaelis-Menten equation, based on the acetylene concentration external to the nodules. One difficulty of this view is that the assumption that the system is not diffusion limited is violated when studying intact nodules. The presence of a gas diffusion barrier in the nodule cortex leads to an alternate expression for the gas exchange rates at subsaturating gas concentrations. A theoretical comparison of the ;apparent' Michaelis-Menten model and diffusion model illustrated the difficulties observed in the former model of overestimating the Michaelis-Menten coefficient and yielding a correlation between the Michaelis-Menten coefficient and the maximum rate. On the other hand, use of a diffusion model resulted in (a) estimates of the Michaelis-Menten coefficient consistent with enzyme studies, (b) stability of the estimates of the Michaelis-Menten coefficient independent of treatment, and (c) a sensitivity of the diffusion barrier conductance to plant drought stress. It was concluded that all studies of nodule gas exchange need to consider possible effects caused by the presence of a diffusion barrier.

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Effect of acetylene on root respiration and acetylene reducing activity in nodulated soya bean.

Acetylene decreased root and nodule respiration, as measured by CO(2) evolution of nodulated or non-nodulated Glycine max. An inhibition of 25 to 35% in 15 to 30 minutes occurred when 13% C(2)H(2) was introduced in the gas flux which aerated the root nutrient solution. When the light intensity was doubled to 800 microeinsteins per square meter per second, the inhibition increased to 50% and nodule acetylene reduction activity was inhibited 50%.

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Rotational isomerism involving an acetylenic carbon IV: synthesis and structure of bis(1,1';3',1"-terphenyl-2'-yl)ethynes: molecular design of sterically congested alkynes toward restricted rotation about acetylenic axis.

The title diphenylethyne derivative with 4-methylphenyl (tolyl) groups at all the ortho positions was synthesized by the Stille or Sonogashira coupling from the corresponding iodide. The X-ray structure revealed that the two terminal phenyl groups at the sp carbons are twisted by 63 degrees out of the coplanar conformation to avoid steric interactions between the tolyl groups. The relative stabilities of possible conformers were analyzed by the PM3 calculations. The axially chiral derivative with two methoxymethyl groups showed no evidence of restricted rotation about the acetylenic axis by VT NMR measurements, its barrier being less than 35 kJ mol(-1). The spectroscopic features and reactivities of this sterically congested alkyne are also described.

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Denitrification, acetylene reduction, and methane metabolism in lake sediment exposed to acetylene.

Samples of sediment from Lake St. George, Ontario, Canada, were incubated in the laboratory under an initially aerobic gas phase and under anaerobic conditions. In the absence of added nitrate (NO(3)) there was O(2)-dependent production of nitrous oxide (N(2)O), which was inhibited by acetylene (C(2)H(2)) and by nitrapyrin, suggesting that coupled nitrification-denitrification was responsible. Denitrification of added NO(3) was almost as rapid under an aerobic gas phase as under anaerobic conditions. The N(2)O that accumulated persisted in the presence of 0.4 atm of C(2)H(2), but was gradually reduced by some sediment samples at lower C(2)H(2) concentrations. Low rates of C(2)H(2) reduction were observed in the dark, were maximal at 0.2 atm of C(2)H(2), and were decreased in the presence of O(2), NO(3), or both. High rates of light-dependent C(2)H(2) reduction occurred under anaerobic conditions. Predictably, methane (CH(4)) production, which occurred only under anaerobiosis, was delayed by added NO(3) and inhibited by C(2)H(2). Consumption of added CH(4) occurred only under aerobic conditions and was inhibited by C(2)H(2).

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