A new pH indicator dye-based method for rapid and efficient screening of l -asparaginase producing microorganisms

Abstract

l-asparaginase is a pharmaceutically and industrially important enzyme as it has potential to treat different cancers and inhibit acrylamide formation in fried and baked food products. In the present study, an attempt was made to screen for new and novel l-asparaginase producers using a widely applied phenol red and bromothymol blue (BTB)1 dye-based plate assay. Screening of four different soil samples for l-asparaginase producers resulted in the isolation of three new potential l-asparaginase producing bacteria. These three strains identified (by 16S rRNA sequencing) as a Pseudomonas resinovorans strain IGS-131, a Bacillus safensis strain IGS-81, and a Glutamicibacter arilaitensis strain ICS-13 with enzyme activities of 10.91 IU/ml, 6.65 IU/ml, and 1.47 IU/ml, respectively. These three strains of bacteria have not been reported as l-asparaginase producers previously. Also, we developed a new pH indicator dye-based plate assay for the screening of l-asparaginase producers after testing eight different pH indicator dyes. This cresol red dye-based method gave a better differentiable zone of hydrolysis and consistent results as compared to previously reported phenol red and BTB-based plate assay. It was also found to be efficient in comparison to all other dyes studied. It produced a bright yellow color at acidic pH (5.5) and turned into a dark red or maroon color when pH was increased (above 7.5). This finding is expected to make screening of all kinds of l-asparaginases more comfortable, rapid, and efficient.