Production and Optimization of Pullulanase by Stenotrophomonas maltophilia under Submerged Fermentation Using Response Surface Methodology (RSM)

Abstract

Pullulanase is a starch debranching enzyme, that cleaves the α-1,6, glucosidic linkage. The research investigates the screening, production, and optimization of pullulanase from bacterial isolates. Five different soil samples were collected from Choba (River State) and Oshodi (Lagos State). The total culturable heterotrophic bacterial counts ranged from 2.21x10⁶ to 3.2 x 10⁶Cfu/ml. A total of eight bacterial isolates exhibited the potential to degrade pullulan which produced a clear halo zone at 70% around the colonies. The isolate with the highest pullulanase degradation was further investigated for production using wheat bran as substrate under submerged fermentation (SmF) applying RSM. Identification and characterization of the isolate were done using both conventional and molecular techniques. This isolate was identified molecularly as Stenotrophomonas maltophilia. The 16S rRNA sequence was deposited in GenBank under accession number MH225448.1. The optimum production of pullulanase obtained using One-factor-at-a-time (OFAT) was achieved with wheat bran (2.469U/ml) at 48h, pH 7 and temperature of 37⁰C. Optimization using RSM revealed the following optimum conditions; temperature 36ºC, pH 7.5, incubation period of 36h, and pullulan concentration of 1.5%. The result gathered from this study showed that the bacterial isolate can successfully utilize agro-waste materials for production of pullulanase. This poses great potential in ameliorating the environmental pollution arising from their poor disposal as well as encouraging the production of the enzyme at a low cost. The data from this study also makes the isolate a good candidate for the production of pullulanase for industrial purposes.