Nitrate reductase activity in Cajanus cajan and Desmodium barbatum assay optimization

Abstract

Cajanus cajan L. and Desmodium barbatum (L.) Berth have been largely used as soil cover species for rubber tree plantations in tropical regions. In order to recommend this leguminous as cover plants to rubber tree plantations located out of the traditionally cultivated arcas, the in vitro nitrate reductase assay (NR) was optimized. The nitrate assimilation rates will be used as a physioecological parameter for C. cajan and D. barbatum growing in theses new production areas. Leaf sections of C. cajan and D. barbatum, grown respectively in greenhouse and in the field, were incubated in phosphate buffer plus KNO₃ and l-propanol, infiltrated under vacuum and transferred to a sharking water bath, in the dark. NR activity was estimated based on nitrite dosage through spectrophotometry at 540 nm. The following conditions were determined: optimal incubation time for nitrite release readings (30-60 minutes, for both species), optimal buffer pH (8,0 for C. cajan and 7,5 for D. barbatum) and nitrate concentration (200 mM for C. cajan and 100 mM for D. barbatum) and incubation temperature (32^oC for both leguminous) for maximum enzyme activity.