Enzymatically‐mediated uranium accumulation and uranium recovery using a Citrobacter sp. Immobilised as a biofilm within a plug‐flow reactor

Abstract

Biofilm‐immobilised Citrobacter sp. removed uranyl ion from flows supplemented with glycerol 2‐phosphate. The metal uptake mechanism was mediated by the activity of a cell‐surface bound phosphatase that precipitated liberated inorganic phosphate with uranyl ion as HUO2PO4·4H2O at the bacterial surface. A modified integrated form of the Michaelis–Menten equation is proposed to describe the removal of metal ion by a columnar bioreactor, where the efficiency of metal removal is semi‐quantitatively related to the input flow rate, the total enzyme loading (E0) and the bioreactor activity. With biofilm‐immobilised bacteria, E0 was further divisible (split) into subparameters of phosphatase titre per bacterium and total biomass surface area. Varying the split E0 and the reaction temperature modified the bioreactor performance. The immobilised bacteria retained high metal loads without loss in steady‐state activity. Accumulated metal was recovered as a concentrated solution.

Phosphate release and heavy metal accumulation by biofilm-immobilized and chemically-coupled cells of acitrobacter sp. pre-grown in continuous culture

“…The production of a cellbound, atypical, acid phosphatase enzyme by a Citrobacter sp. has been well documented (Jeong et al, 1997a;Macaskie and Dean, 1989;Macaskie et al, 1992Macaskie et al, , 1994Macaskie et al, , 1995aMacaskie et al, ,b, 1997 and previous studies have harnessed the release of inorganic phosphate (HPO 4 2− ) from an organic phosphate ''donor'' molecule to the stoichiometric precipitation of metal cations (M) as insoluble metal phosphates (MHPO 4 ) (Macaskie et al, 1992(Macaskie et al, , 1994. The enzyme system is robust, functioning between temperatures of 2-45°C, over a pH range from 4.5-8.5 and tolerating elevated metal concentrations.…”

“…strain N14 (Basnakova and Macaskie, 1997;Jeong et al, 1997a;Macaskie et al, 1995aMacaskie et al, ,b, 1997 was used under license from Isis Innovation, Oxford. Minimal medium comprised (g/L): Tris buffer, 12.0; MgSO 4 .7H 2 O, 0.63; FeSO 4 .7H 2 0, 0.00032 and KCl, 0.62.…”

“…Uranyl and phosphate are removed stoichiometrically [the precipitated compound was identified as HUO 2 PO 4 .4H 2 O (Basnakova and Macaskie, 1997;Macaskie et al, 1992;Yong and Macaskie, 1995)], but phosphate is required to excess for efficient precipitation . Uranyl uptake was determined by assay of UO 2 2+ in the column inflow and exit solutions using arsenazo III (Basnakova and Macaskie 1997;Macaskie et al, 1995aMacaskie et al, , 1997Tolley et al, 1995;Yong and Macaskie, 1995). The loaded uranyl was estimated as the difference between the input and exit UO 2 2+ (mg/L) multiplied by the total volume passed (L).…”

“…The phosphatase specific activity of the culture was monitored in fermentor-outflow samples by the release of pnitrophenol from the chromogenic substrate p-nitrophenyl phosphate as described previously (Basnakova and Macaskie, 1997;Jeong et al, 1997a;Macaskie et al, 1995aMacaskie et al, ,b, 1997. The specific activity (1 unit) is defined as that liberating 1 nmol of p-nitrophenol/min/mg bacterial protein.…”

“…The presence of inorganic solid supports during growth often results in the ''selfimmobilization'' of cells as a biofilm; such biofilms have been used in large scale metal wastewater treatment (e.g., Mudder and Whitlock, 1984; also other processes reviewed by Macaskie and Dean, 1989). Inexpensive materials of large surface area are generally chosen, e.g., anthracite, sand particles, polyvinyl chloride beads, porous glass, stainless steel mesh, wood chips, and reticulated foams Webb, 1986, Dempsey et al, 1986;Macaskie and Dean, 1989;Macaskie et al, 1995a). Alternatively, the cells can be grown in liquid culture, the biomass concentrated, and chemically coupled to suitable carriers using techniques adapted from immobilized enzyme technology.…”