Wood, et al., reported in 2001 that Thiobacillus ferrooxidans can be immobilised on sand (size 0.85 mm to 1.18 mm) for use in a repeated batch and continuously operated packed-bed bioreactor which has not been previously reported in the literature. Repeated batch operation resulted in the complete oxidation of ferrous to ferric iron. The bacteria were active immediately after 3–4 weeks in a non-aqueous medium; i.e. the sand was allowed to dry out, demonstrating the stability of the system. A lag phase of 28 days was recorded when the sand was stored dried in a sealed container for 16 weeks compared with a lag phase of 13 days for a sample frozen for 18 weeks. After a period of 10 days, continuous operation of the reactor at a dilution rate of 0.64 h–1 resulted in 95–99% oxidation of ferrous iron or 0.31–0.33 kg m–3 h–1. With the use of a scanning electron microscope, images were recorded of Thiobacillus ferrooxidans on sand.
The authors concluded that contrary to the results published by Halfmeier et al. (1993b), Thiobacillus ferrooxidans has been immobilized on quartz sand particles. The method simply requires introducing the sand to a culture and the cells soon adhere to the particles shortly afterwards. The immobilized culture oxidizes iron at a slightly faster rate than the liquid culture, but the variation is probably due to a difference in population rather than an effect of immobilization. The number of cells adsorbed per gram of sand is lower than on other carrier matrices previously investigated: 250–500-µm activated carbon, 6.8× 109 cells/g; <63-µm coal, 4.09×1010 cells/g; 120–180-µm pore glass, 2.18×1010 cells/g; calcium alginate gel, 2× 1010 cells/ml gel, κ-carrageenan gel, 3.3×109 cells/ml gel; gelrite gel, 6.1×109 cells/ml gel. However, the cell concentration could be increased by using smaller particles of sand.
It is not possible to accurately compare the packed sand column with other systems due to the low Fe2+ concentrations used to test the column, the low flow rate and lack of aeration. However, comparing the oxidation rate with other studies, i.e. chemostat free cells at 28 °C, 0.42 kg m–3 h–1 oxidized, inlet loading 0.49 kg m–3 h–1 (MacDonald and Clark 1970); an aerated packed bed with sintered glass at 30 °C, 3.6 kg m–3 h–1 oxidized, inlet loading 4 kg m–3 h–1; packed bed with calcium alginate at 25 °C, 0.53 kg m–3 h–1 oxidized, inlet loading 0.56 kg m–3 h–1, shows that sand is not inhibitory to the oxidation of Fe2+ by Thiobacillus ferrooxidans. Using an aerated column at higher temperatures may produce results comparable with other systems. The packed column used in this study was stable for up to 1 month when the sand was allowed to completely dry out and could be reactivated immediately simply by the introduction of 9K medium. Longer periods of shutdown would require a greater length of time to allow the
In continuous experiments, the reactor was operated at a dilution rate of 0.64 h–1 and an average inlet value of 0.33 kg Fe2+ m–3 h–1, resulting in the oxidation of 0.31–0.33 kg Fe2+ m–3 h–1. Oxidation rates were calculated using the retained volume, 0.188 l. Sand from bacteria to adapt to the changed environment. For longterm storage, the inoculated sand particles could be frozen and introduced to 9K medium when required and recovered within 2 weeks. Both methods, drying and freezing of Thiobacillus cells, would be advantageous to industry where it may be necessary to transport inoculated sand to various sites.
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