High-throughput screening for distinguishing nitrilases from nitrile hydratases in Aspergillus and application of a Box-Behnken design for the optimization of nitrilase.

Abstract

Nitrilases and nitrile hydratases/amidases hydrolyze nitriles into carboxylic acids and/or amides, which are used in industrial chemical processes. In the present study, 26 microorganisms, including yeasts and filamentous fungi, in a minimum solid mineral medium supplemented with glucose and phenylacetonitrile were screened to evaluate their biocatalytic potential. Of these microorganisms, five fungi of the genus Aspergillus were selected and subjected to colorimetry studies to evaluate the production and distinction of nitrilase and nitrile hydratase/amidase enzymes. Aspergillus parasiticus Speare 7967 and A. niger Tiegh. 8285 produced nitrilases and nitrile hydratase, respectively. Nitrilase optimization was performed using a Box-Behnken design (BBD) and fungus A. parasiticus Speare 7967 with phenylacetonitrile volume (μl), pH, and carbohydrate source (starch:glucose; g/g) as independent variables and nitrilase activity (Uml-1 ) as dependent variable. Maximum activity (2.97 × 10-3 Uml-1 ) was obtained at pH 5.5, 80μl of phenylacetonitrile, and 15g of glucose. A. parasiticus Speare 7967 showed promise in the biotransformation of nitriles to carboxylic acids.