Acetylene reduction technique

Burris, R.H. The acetylene reduction technique. In Nitrogen Fixation by Free-living Micro-organisms, VoL 6, W.D.P. Stewart,... 

Burris, R.H. (1975). The acetylene-reduction technique. Nitrogen fixation by free-living microorganisms. The International Biological Programme 6, 249-257. 

Nitrogen fixation has been demonstrated only in those lichens with blue-green phycobionts. Of the eight genera of blue-green algae listed by Ahmad-jian (1967) and Duncan (1970) as phycobionts, only lichens with Nostoc xid Calothrix have so far been shown to fix nitrogen. The lichens in which fixation has been shown either by means of or the acetylene reduction technique... 

However, critical examination of the acetylene-reduction technique, often... 

This new assay technique was developed independently by American and Australian workers who first established the connection between nitrogenase and hydrogenase, and showed the former could reduce a number of substrates other than nitrogen, for example nitrous oxide, azide, cyanide and, most usefully, acetylene. The latter is reduced to ethylene and this forms the basis of the acetylene reduction test. The material to be examined is briefly gassed with acetylene and the ethylene formed measured by gas chromatography. While this technique was being refined,... 

Indirect methods used to estimate DNF include the acetylene block method (S0rensen, 1978), metabolite stoichiometry (Dollar et al., 1991), or stable isotope tracers (Nielsen, 1992). Acetylene (C2H2) blocks the terminal step of DNF, the conversion of N2O to N2, and the DNF rate is estimated by quantifying the production of N2O on a gas chromatograph with an electron capture detector. Problems with the acetylene block technique include blockage of NTR (which means that rates of coupled NTR—DNF cannot be obtained), inefficacy at low N03 concentration, and interference by H2S. Sulfide appears to alleviate the acetylene block of nitrous oxide reductase and permit full reduction of N2O to N2. 

In the acetylene inhibition technique, acetylene is added to a water sample, which inhibits the reduction of N2O to N2 (Sorensen, 1978). The accumulation of N2O is then measured using gas chromatography and an electron capture detector and the denitrification rate is taken to be equal to the total N2O flux. One potential problem is incomplete inhibition of N2O reduction to N2, particularly in the presence of hydrogen sulfide, a compound commonly found under anaerobic conditions. Another potential problem with the technique is that acetylene also inhibits nitrification, a process that often supplies the NOs and N02 substrates for denitrification. To inhibit nitrification is to inhibit denitrification if it is at aU substrate limited (Hynes and Knowles, 1978). 

Welsh, D. T., CastadeUi, G., Bartoli, M., Poli, D., Careri, M., de Wit, R., and Viaroh, P. (2001). Denitrification in an intertidal seagrass meadow, a comparison of N-isotope and acetylene-block techniques DissimUatory nitrate reduction to ammonia as a source of N2O Mar. Biol. 139, 1029-1036. 

A wide range of techniques are used to determine catabolic nitrate reduction rates. These include mass balance methods using input-outputs, acetylene inhibition techniques, dinitrogen production rates, nitrate consumption rates, nitrate pore water profiles, and stable isotope tracer techniques. The limitations and advantages of these methods are discussed by Seitzinger (1988) and Herbert (1999). 

Denitrification is the reduction of N03 N02 NO N2O —> N2 gas that is mediated by bacteria under anaerobic conditions, most generally in microbial mats and sediments. There are a number of methods to measure denitrification acetylene inhibition, isotope pairing, changes in N2 fluxes, and changes in the N2 to argon (Ar) ratio. Each of the techniques has their pros and cons and none is clearly superior under all conditions (see reviews by Cornwell et al, 1999 Chapter 6 by Devol, this volume). 

The radiolysis of propane has been studied extensively in experiments that have included a wide range of techniques. The gas-phase radiolysis in the absence of inhibitors yields the products hydrogen, ethane, propene, 2,3-dimethylbutane, methane, ethylene, isobutane, acetylene, isopentane and n-butane as well as small quantities of butene-1, -pentane, 2-methylpentane and -hexane ° ° . At high conversions the yield of ethylene, propene, 2,3-dimethylbutane and isobutane are all reduced. The reduction in ethylene arises from hydrogen atom addition, while the reduction in the other products may arise from the reaction of propyl ions with propene to remove both C3H6 and the source of isopropyl radicals. 

At last, the inhibition technique takes advantage of the property of acetylene to block the reduction of N O to N after it is injected into the sediment. The total amount of N O produced is then the measure for the denitrification rate as it is easy to determine by gas chromatography (Andersen et al. 1984) or by microsensors (Christensen et al. 1989). The advantage of this method is that analyses can be carried out rapidly and sensitively. Problems are (a) N O reduction is sometimes incomplete, (b) a homogenous distribution of in the pore water is difficult to maintain, (c) inhibits nitrification in the sediment meaning that the coupled system (nitrification / denitrification) might be seriously affected due to the applied method, and... 

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