2.1 Sample preparation
Carbon sources; glucose (GA), Beta vulgaris (BA), and Beta vulgaris with cyanide (BCN) were used for the cultivation ofFusarium oxysporum in gold mine wastewater as shown in [26]. Briefly, Fusarium oxysporum was grown in a 1 litre continuously stirred tank reactor (CSTR) containing metal-laden synthetic gold mine wastewater at 25±2°C. The wastewater contains (per litre): 47 mg CuSO45H2O, 42 mg Fe2(SO4)3 H2O, 278 mg (NH4)2SO4, 27mg KH2PO4, 3mg ZnSO47H2O, 0.9 mg PbBr2, and 40 mg Na2HAsO4 7H2O. The reactor was inoculated with 10% (v/v) F. oxysporum and 0.3 g glucose as refined carbon source. Subsequently, experiments on 0.3 g pulverised B. vulgaris followed by 0.3 g pulverised B. vulgaris with 100 ppm CN-/L in form of KCN. Biomass was harvested once the carbon source was used up and/or when the microbial growth reached stationary phase. The harvested biomass was centrifuge at 4°C for 10 min at a speed of 10,000rpm in an Avanti® J-E centrifuge (Beckman Coulter, Inc. USA). Recovered biomass were washed thrice in sterile distilled water, dried in a Duran® vacuum desiccator (DURAN Group GmbH, Germany), and stored at -20 °C for further analyses.
Sample dried biomass was dissolved in sterile distilled water in a 1:1, weight: volume ratio and incubated at 298.15 K for 16 h to ferment any residual carbohydrates. The Durham tube method was used to test for any residual carbohydrates in the suspension [24]. The procedures were repeated until an appropriate quantity of dry biomass was obtained. The elemental analysis of the samples was determined by a Thermo Flash EA 1112 series analyser in a Helium carrier gas (Thermo Fisher Scientific Inc. Waltham, USA) subsequent to estimation of molar mass of dried biomass containing a unit carbon as 23.03, 33.14 and 27.06 g/C-mol for GA, BA and BCN samples, respectively. The detailed elemental analysis of the samples are presented in [26].