Reduction of Uranium(VI) under Micro-aerobic Conditions using an

Indigenous Mine ConsortiumUniversity of

Pretoria

Energy Postgraduate Conference 2013

Aim & ObjectivesTo utilise indigenous cultures of bacteria from the localenvironment to biologically reduce U(VI) to U(IV)

– Isolation and purification of microorganisms for use in further experiments

– Characterization of microorganisms in order to identify and classify the microorganisms involved in the reduction of uranium (VI)

– Investigation of the reduction potential of microorganisms that reduce uranium (VI) to uranium (IV) using the consortium in a batch system to establish kinetic parameters for use in reactor scale-up

Introduction– Among all elements currently in use in the energy

industry worldwide, uranium is the most abundant

– Uranium-containing wastes are produced at various steps of the nuclear fuel cycle, and vary considerably from low level radioactive effluents produced during uranium mining to intensely radioactive levels in nuclear power plant, spent fuel, and liquid wastes

– Discharge of radio-nuclides such as uranium from contaminated sites and their subsequent mobility in the environment is a subject of paramount concern

– The primary radiation health effect of concern is an increased probability of the exposed individual developing cancer during their lifetime

Introduction

Physical/chemical processes

Treatment OptionsPump and treat processes of U(VI)-contaminated waterInvolves extraction of contaminated water, followed by aseparation process on the surface

Separation processes include:• Ion exchange • Chemical precipitation• Reverse osmosis

Limitation: • Expensive to apply

Introduction

Biological processes

Offer the potential for removing metal/radionuclidepollutants from dilute solutions, where physicalchemical methods may not be feasible.

4 mechanisms by which bacteria immobilize metals orRadio-nuclides namely;

• Bio-sorption, • Bioaccumulation,• Precipitation by reaction with inorganic ligands• Microbial reduction

Introduction

Bioreduction

• Involves the reduction of an element from a higher to a lower oxidation state or to an elemental form affects its solubility, resulting in its precipitation using bacteria

Advantages:

• It is not limited by saturation

• Many radio-nuclides are less soluble when reduced

Materials & MethodsElemental Analysis of Soil• Uranium contaminated soil was collected from a

closed uranium mine was analyzed by ICP-OES• Isolation of Indigenous Bacteria • Mixed culture was obtained by inoculating basal

mineral medium (BMM) amended with glucose with mine soil

• Bacterial cultures were purified and then incubated

Batch studies• Pure culture batch studies were conducted in basal

mineral medium (BMM) supplemented with glucose as a carbon source with different concentrations of U(VI) 75, 100, 200, 400, 600 and 800 mg/L

Materials & MethodsSampling• A 0.5 mL sample of the homogenous solution was

collected using a syringe and then centrifuged

• The sample was then diluted with 4.5 mL of BMM (1:10 dilution), mixed with 2 mL of complexing reagent and analyzed in duplicate for U6+

immediately on the UV spec

• Total uranium level in each sample (U(IV) and U(VI)) was determined by oxidizing an unfiltered sample with nitric acid prior to uranium measurement

Results • Elemental analysis of soil

• Soil was found to contain 168 mg/kg Uranium

• Preliminary culture characterization• Gram staining – Gram- & Gram+

• Culture characterization• Purified and sequenced rRNA genes from

bacteria 4 species were found:• Pantoea agglomerans• Enterobacter cloacae• Pseudomonas stutzeri

Removal of Uranium at Low Concentrations

Figure 2. Uranium(VI) reduction for the three pure cultures of bacteria Pseudomonas stutzeri, Pantoea agglomerans and

Enterobacter cloacae under an initial concentration of 75 mg/L

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Pseudomonas stutzeri

Pantoea agglomerans

Enterobacter cloacae

Removal of Uranium at High Concentrations

Figure 3. Uranium(VI) reduction for the three pure cultures of bacteria Pseudomonas stutzeri, Pantoea agglomerans and Enterobacter cloacae under varying

concentrations; A: 100 mg/L B: 200 mg/L C: 400 mg/L and D: 600 and 800 mg/L

(B)

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Pseudomonas stutzeri

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Conclusion

• Microorganisms play important roles in the environmental fate of toxic - array of physical-chemical and biological mechanisms effecting transformations between soluble and insoluble phases

• Further work - Identify and characterize of cytosolic and outer membrane proteins involved in U(VI) reduction at a cellular level

• Kinetic modelling of uranium reduction and cumulative removal studies should help us to better predict and model how uranium will behave in situ

Contribution to Scientific Community• Achieving U(VI) reduction reaction under near zero

oxidation reduction potential (ORP) under facultative conditions

• Developing a simplified and more reliable method for measuring U(VI) using the Arsenazo III method

oProf EMN Chirwa (University of Pretoria)oNational Research Foundation (NRF)oSouth African Nuclear Human Asset & Research Programme (SANHARP)

Acknowledgements