Identification of Suitable Condition for Mannanase Production by Bacillus sp. GA2(1).

ABSTRACT

Aims: The effects of 1% (w/v) supplementation of additional 5 agricultural wastes, corn cob, bagasses, coffee residues, soybean meal, and copra meal for mannanase production by Bacillus sp.GA2(1) were studied. Hence, partial characterization of mannanase was determined. Methodology: The 1%(v/v) overnight cultured of Bacillus sp. GA2(1) was transferred into the basal medium and shaken at 150 rpm for 18 h at 37ºC. The additional of 5 AWs, corn cob, bagasses, coffee residues, soybean meal, and copra meal for the mannanase production were investigated. The cell suspension was centrifuged, and the crude mannanases were collected and stored at – 20ºC for enzyme assay. The mannanase activities were measured by the dinitrosalicylic acid method. The optimal pH of mannanase were studied by measuring enzyme activity at pH 3-10 using 50 mM of following buffers; citrate (pH 3.0-6.0), phosphate (pH 6.0-8.0), and glycine-NaOH (pH 8.0-10.0). The optimal temperature was measured at 30-80ºC. Under standard assay conditions, locust bean gum was used as substrate to determine the optimal pH and temperature of the reaction. Thermostability was determined by preincubating the enzyme at different temperatures (30-80ºC) for 1 h. The residual mannanase activities were measured under standard condition. Results: Among bagasses, coffee residues, soybean meal, corn cob and copra meal, the coffee residues was the most effective carbon source, the maximum yield of mannanase activity was 0.26 U/ml. The optimal temperature and pH for mannanase activity was pH 6.0 and 50ºC of 0.44 and 0.35 U/ml, respectively. The stability of enzyme was determined at 30-80ºC for 60 min. The results revealed that mannanase retained more than 96% of remaining activity after incubation of 60 min at 50ºC. Conclusion: The maximum mannanase production was found when the medium was supplemented with coffee residues. Crude mannanase showed the highest activities of 0.44 U/ml at pH 6.0 and of 0.35 U/ml at 50ºC. The mannanase from Bacillus sp. GA2(1) retained more than 90% of theirs activities at 30-60ºC after preincubated for 60 min and then rapidly decreased.