The Acetylene-Ethylene Assay for N₂ Fixation: Laboratory and Field Evaluation

Abstract

A bstract. The methodology, characteristics and application of the sensitive C,H.,-C2H4 assay for N2 fixation by nitrogenase preparations and bacterial cultures in the laboratory and by legumes and free-living bacteria in situ is presented in this comprehensive report. This assay is based on the N9ase-catalyzed reduction of C2Ho to C2H4, gas chromatographic isolation of C2H2 and C9H4, and quantitative measurement with a H,-fiame analyzer. As little as 1 ,u,lmole C..H4 can be detected, providing a sensitivity 103-fold greater than is possible with '-5N analysis.A simple, rapid and effective procedure utilizing syringe-type assay chambers is described for the analysis of C,H2-reducing activity in the field. To meet the imminent crisis in the world food supply (38) it is imperative that the resources of this planet be mobilized as rapidly and effectively as possible. Basic to such mobilization is a knowledge of the magnitude of the dynamic processes in the biosphere which affect the availability of nitrogen, the one element most often limiting in the production of foodstuffs (37

Marcha de absorção do nitrogênio do solo, do fertilizante e da fixação simbiótica em feijão-caupi (Vigna unguiculata (L.) walp.) e feijão-comum (Phaseolus vulgaris L.) determinada com uso de 15N

“…A atividade da enzima nitrogenase foi medida indiretamente por meio do método da atividade de redução do acetileno (ARA) (Hardy et al, 1968). Após a coleta, as raízes nodulantes foram colocadas em frascos de vidro de 1.000 mL, os quais foram fechados hermeticamente.…”

Effects of Plant Growth Promoting Rhizobacteria (PGPRs) on the Biological Nitrogen Fixation, Nodulation, and Growth of Lupinus albus l. cv. Multolupa

“…The total amount of nitrogen was determined colorimetrically, following the Smith method [22], after a Kjeldhahl digestion by a microwave digester Maxidigest MX350 (Prolabo, Spain). Before the plants were pressed, acetylene reduction activity (ARA) was determined in the nodule [23]. The fixed nitrogen was calculated following the method of Zhang et al [2], total plant nitrogen content minus the amount of nitrogen in the original seed (32 mg per seed, n = 25 seeds).…”

Alkyne substrate interaction within the nitrogenase MoFe protein

“…This product profile is consistent with side-on binding of 2-butyne, with proton addition occurring at one face. Earlier work on wild-type nitrogenase revealed that proton addition to acetylene during its reduction to ethylene occurs with stereospecificity [27][28][29]. The product of acetylene (HC≡CH) reduction in the presence of D 2 O is 96 % cis-ethylene (HDC=CDH), with only 4 % of the trans-isomer [30].…”