Isolation, Characterization and Biodegradability Tests of Cellulase from Bacterium Obtained from the Soil of Agro Dumpsite

Abstract

The study investigated the waste biodegradation potential of cellulase produced by bacteria isolated from soil at an agro dumpsite, along  with their physicochemical properties. The bacterium was identified as Bacillus subtilis and subjected to extracellular cellulase production via shaking flask fermentation. The production conditions such as pH, temperature, and incubation time were optimized  using random surface methodology (RSM). The response was predicted by second-order polynomial model. In addition, the enzyme produced was partially purified, characterized, and explored in the biodegradation of agricultural waste products using standard  methods. Cellulase yield in the experiment varied from 3.00 to 10.30 U/mL and under the optimum conditions of pH 7.0, 40 oC and 18 hr incubation time, the value for cellulase yield was 10.70 U/mL. The enzyme purification resulted in a specific activity of 3.3 Units/ mg of  protein and 20 % yield. The Km and Vmax of the partially purified enzyme were 3.33 mg/mL and 52.0 U/mL respectively. The optimum pH and temperature of B. subtilis cellulase were at pH 5.0 and 50^oC respectively. The inhibitory study showed that Na+ did not inhibit the  activity of the enzyme while the enzyme activity was moderately inhibited by Mg²⁺, Sn²⁺ and Hg²⁺. Potassium ions (K+) strongly inhibited enzyme activity. The substrate specificity test showed that the enzyme was able to utilize other substrates. CMC, orange peel, pectin,  pineapple peel and maize cob all showed 100% activity while rice husk and maize starch had an activity of 89 and 14 % respectively. The study concluded that response surface methodology (RSM) could be used to optimize bacteria cellulase production conditions and that  the enzyme is suitable for various biotechnological processes such as waste management.