Identification of the nickel uranyl phosphate deposits on Citrobacter sp. cells by electron microscopy with electron probe X-ray microanalysis and by proton induced X-ray emission analysis

Immobilized cells of a Citrobacter sp. can remove heavy metals from wastewaters by deposition of metals with enzymatically liberated phosphate. Nickel is not removed effectively by this technique, but Ni2+ can be intercalated into cell-bound, crystalline HUO2PO4 previously deposited enzymatically. This technique for efficient removal of Ni from solution has been generically termed microbially enhanced chemisorption of heavy metals(MECHM). The nickel uranyl phosphate deposits bound to Citrobacter sp. cells immobilized in polyacrylamide gel (FAG) were analyzed using scanning transmission electron microscopy with electron probe X-ray microanalysis (EPXMA) and proton-induced X-ray emission analysis(PIXE). Both methods gave the molar ratios of nickel, uranium, and phosphorus in the deposits as close to 1:2:2 in all analyzed parts of the sample. EPXMA proved that the deposits were localized on the surface of cells inside FAG particles as well as those immobilized on the edge. Small deposits of nickel uranyl phosphate were also found in FAG between the cells, indicating the possible involvement of extracellular polymeric substances (EPS) in the creation of intercellular deposits. These findings confirm the mechanism of MECHM and show that this mechanism operates throughout the immobilized cell matrix. The use of two independent methods of solid-state analysis in a common sample provides validation of both techniques for the spatial and quantitative analysis of biomass-bound elements.