July 18, 2006 Presented by Jeff Fitts Environmental Research & Technology Division

1. Synchrotron-based investigations of biogeochemical transformations of priority contaminants in soils at DOE sties July 18, 2006 Presented by Jeff Fitts Environmental Research & Technology Division

2. DOE Environmental Management Sites http://web.em.doe.gov/em94/sitesum.html • 3,000 inactive waste sites; • DOE EM (Environmental Management) FY06 appropriation $6.6 billion • The Big Three: Hanford, WA; Oak Ridge, TN; Savannah River, SC

3. Research funding from ERSP within DOE Environmental Remediation Sciences Program mission is to advance our understanding of the fundamental biological, chemical, and physical processes that control contaminant behavior in the environment in ways that help solve DOE’s intractable problems in environmental remediation and stewardship. ERSP Field Research Center at ORNL • S-3 disposal ponds • U processing waste • In-use 1951-1983 • Neutralized 1984 • Paved 1988 http://www.lbl.gov/NABIR/

4. Two remediation approaches • Extract uranium from the ground • Stabilize uranium in the ground Groundwater Flow Uranium Plume Manipulate Uranium oxidation state to control U solubility • Oxidized uranium, U(IV), is soluble and mobile • Reduced Uranium, U(IV), is insoluble and immobile • Soil Bacteria often control the redox state of soil

5. FRC conditions present challenges to in-situ bioremediation strategies Problematic conditions Result • Too high redox for stimulating sulfate and Fe reducers • Affects metal bioavailability, and thus, toxicity • Inhibit nitrate reducers • High nitrate (~1000 ppm in Area 2) • Acidity • Heavy metals (Ni, Al…) Will introduction of nickel resistance into indigenous microbial community have a positive effect on nitrate reducers and stimulate iron and sulfate reducers?

6. ERSP project overview Goal:Immobilize uranium in contaminated sediments via microbial reduction and precipitation Problem: Active uranium reducers are inhibited by co-contaminants in complex waste streams (e.g., heavy metals) S3 ponds at ORNL Major project objectives Demonstrate application of natural gene transfer to improve community function under increased levels of toxic metal stress (Dr. Daniel van der Lelie, BNL Biology Dept.) Demonstrate ability to enhance uranium immobilization in ORNL sediments by indigenous microorganisms that have adopted the toxic metal resistance marker(Fitts)

7. Project design schematic Nickel stress 1. Community structure 2. Improved nitrate red. FRC soils Total community Horizontal gene transfer (in vivo) – soil columns Model organisms Ni resistance mechanisms Isolated from FRC fluidized bed reactor Strain construction (in vitro) 1. S and Fe reduction 2. Uranium reduction and precipitation

8. What is really happening to the uranium? • How stable is the uranium associated with the soil? • Could it become a problem in the future?

9. 3 2 1 Contaminant speciation in complex real-world systems • Heterogeneous materials such as soils • Low contaminant concentrations Speciation on model soil particles and in pure bacterial cultures • Focus on predominant processes • Molecular details of transient species Improve predictions of contaminant behavior Single-crystal oxide/water interfaces • Derive atomic-level picture • Test predictive models

10. Role of soil bacteria and their mechanisms of toxic metal resistance Pure culture 1. Growth media 2. Common soil bacteria, Pseudomonas w/ and w/o gene for Ni resistance 3. As a function of Ni concentration 2-5 ml sample 1. Monitor bacterial growth (UV-Vis) 2. Measure Ni uptake by bacteria X-ray transparent window

11. 0.8mm Optical Density at 290eV Ni resistance mechanisms 2.5mm Scanning Transmission X-ray Microscope imaging at the carbon K-edge Washed cells exposed to Ni for 2 hrs Rinsed cells are dried on microscope window carbonate C=O Pseudomonas DMY2::ncc-nre exposed to 2 mM NiCl2 Cluster image Carbonates not observed O K-edge will be sensitive to NiO formation C=C Carbonate in organic matrix NSLS Beamline X1A

12. C U(VI) reduction by Fe(II)-containing minerals Green rust: Fe(II)-containing mineral Green rust after Rxn w/ U(VI)-containing aqueous solution

13. Remobilization potential of sequestered Uranium After initial rxn with Green rust Reduced U, sequestered as U(IV) After exposure to oxygen Mixture of U(VI) & U(IV) Aqueous U(VI) Bioavailable Uranium How stable is U(IV) that is associated with Fe-oxide? What is the role of aqueous Fe(II)?

14. Pseudomonas DMY2 tested in column studies 2,6 - FRC community 3,7 - FRC community + Pseudomonas wild type 4,8 - FRC community + Pseudomonas pMol222 5,9 - FRC community + Pseudomonas::ncc-nre 1 2 7 8 3 4 5 6 9 Kill Media + Formaldehyde Media + 1 mM NiCl2 Media no Nickel added

15. Geochemical interrogation: S, Fe & U at time zero U-Fe correlation U distribution S speciation and redox state port B U oxidation state at M5 edge Area 2 FRC soil Area 2 FRC soil U4+ standard Typical soil Organic matter U6+ standard inorganic sulfate sulfate reduced organic S species NSLS beamlines X27A & X15B

16. Geochemistry of columns after 65 days Column effluent indicators Ni distribution in Column 2 • Initial mobilization of Uranium • Nickel breakthrough observed but significant adsorption occurs Soil indicators by x-ray absorption spectroscopy • Small increase in sulfide relative to kill • (oxidation during transfer may be problem) • Fe(III) oxides still dominate • No reduction of Uranium observed

17. Summary: Applications of synchrotron-based studies in nuclear waste management • Determine chemical state of untreated wastes • Assess and optimize treatment technologies and remediation strategies • Assess end-states to improve long-term performance predictions • Conduct basic science research to seed future innovation in cleanup technologies

18. Acknowledgements BNL Environmental Sciences Dept. Garry Crosson BNL Biology Department Niels van der Lelie David Moreels, Safiyh Taghavi, Craig Garafola NSLS Measurements Paul Northrup (BNL Environmental Sci. Dep.) – XAS & Microprobe Bjorg Larson, Sue Wirick (Stony Brook University) – STXM James Ablett (BNL NSLS) – Microprobe beamline X27A Oak Ridge FRC Dave Watson (ORNL)