Oleochemicals industry effluence mainly contains a high chemical oxygen demand (COD) in a range of 6000-20,000 ppm. An effective biological wastewater treatment process must be carried out before wastewater is discharged into the environment. In this study, a submerged bed biofilm reactor (SBBR) was adapted to the biological oleochemical wastewater treatment plant observed in the present study. The effect of wastewater flow rate (100-300 mL/min), Cosmoball® percentage in the SBBR system (25-75%), and percentage of activated sludge (0-50%) were investigated in terms of COD reduction. The Box-Behnken design was used for response surface methodology (RSM) and to create a set of 18 experimental runs, which was needed for optimising the biological oleochemical wastewater treatment. A quadratic polynomial model with estimated coefficients was developed to describe COD reduction patterns. The analysis of variance (ANOVA) shows that the wastewater flow rate was the most effective factor in reducing COD, followed by activated sludge percentage and Cosmoball® carrier percentage. Under the optimum conditions (i.e., a wastewater flow rate of 103.25 mL/min a Cosmoball® carrier percentage of 71.94%, and an activated sludge percentage of 40.50%) a COD reduction of 98% was achieved. Thus, under optimum conditions, as suggested by the BBD, SBBR systems can be used as a viable means of biological wastewater treatment in the oleochemicals industry.
Endo-β-1,3-glucanase from alkalophilic bacterium, Bacillus lehensis G1 (Blg32) composed of 284 amino acids with a predicted molecular mass of 31.6 kDa is expressed in Escherichia coli and purified to homogeneity. Herein, Blg32 characteristics, substrates and product specificity as well as structural traits that might be involved in the production of sugar molecules are analysed. This enzyme functions optimally at the temperature of 70 °C, pH value of 8.0 with its catalytic activity strongly enhanced by Mn2+. Remarkably, the purified enzyme is highly stable in high temperature and alkaline conditions. It exhibits the highest activity on laminarin (376.73 U/mg) followed by curdlan and yeast β-glucan. Blg32 activity increased by 62% towards soluble substrate (laminarin) compared to insoluble substrate (curdlan). Hydrolytic products of laminarin were oligosaccharides with degree of polymerisation (DP) of 1 to 5 with the main product being laminaritriose (DP3). This suggests that the active site of Blg32 could recognise up to five glucose units. High concentration of Blg32 mainly produces glucose whilst low concentration of Blg32 yields oligosaccharides with different DP (predominantly DP3). A theoretical structural model of Blg32 was constructed and structural analysis revealed that Trp156 is involved in multiple hydrophobic stacking interactions. The amino acid was predicted to participate in substrate recognition and binding. It was also exhibited that catalytic groove of Blg32 has a narrow angle, thus limiting the substrate binding reaction. All these properties and knowledge of the subsites are suggested to be related to the possible mode of action of how Blg32 produces glucooligosaccharides.