Isolation, optimization, characterization and application of bioflocculant BA-CGB produced by novel Bacillus atrophaeus isolated from Richards Bay Harbour, South Africa

A novel bacterium with bioflocculant-producing potential was amongst bacterial isolates isolated from soil sample at Tuzi Gazi, Richards Bay Harbour, Kwazulu-Natal, Republic of South Africa and screened for bioflocculant producing potential after cultivation using a production medium at 30 degrees C for 72 hours (160 rpm against 4g/L Kaolin clay suspension. The bacterium with an initial flocculating activity of 74% was further identified using 16S rRNA molecular sequencing method as Bacillus atrophaeus SRCM 101359 (with 98.64% similarity). Optimisation of Bacillus atrophaeus culture conditions were carried out resulting in a yield of 3.165 g pure BA-CGB, recovered after 96 hours of fermentation from 1 litre of production broth in the presence of 4% inoculum size, C/N ratio (40:1) glucose (carbon source), ammonium chloride (nitrogen source), at pH 9 and temperature of 35 degrees C as well as shaking speed of 110 rpm. Flocculating activity was stimulated with 1% Ba2+ solution. Characterisation of the pure bioflocculant using Fourier transform infrared spectroscopy (FT-IR) of purified BA-CGB revealed the presence of functional groups such as hydroxyl, carboxyl, amine and amide. Elements such as carbon and oxygen among others, were present in the molecular chain of bioflocculant BA-CGB. This could serve as a confirmation that the bioflocculant is with a polysaccharide backbone; accounting for its thermal stability character as it managed to attain more than 60% flocculating activity at 121 degrees C for 15 min, with 0.4 mg/mL dosage size and Ca (2+) as a stimulating agent. Images of BA-CGB from scanning electron microscopy (SEM) showed that it is an amorphous and irregularly shaped biopolymer. Further analysis of the bioflocculant BA-CGB showed a chemical composition of carbohydrates (65%), uronic acid (25%) and trace proteins (10%). The flocculating activity of purified BA-CGB was greatly influenced by a wide pH range (3-12). This bioflocculant holds a promising future to replace in-use chemical flocculants in wastewater treatment.