Sulfur isotopes as indicators of amended bacterial sulfate reduction processes influencing field scale uranium bioremediation.

Sulfur Isotopes as Indicators of Amended Bacterial Sulfate Reduction Processes Influencing Field Scale Uranium Bioremediation JENNIFER L. DRUHAN, 1 MARK E. CONRAD, 2 KENNETH H.WILLIAMS, 2 LUCIE N’GUESSAN, 3 PHILIP E. LONG, 4 SUSAN S. HUBBARD 2 Department of Earth and Planetary Science, University of California, Berkeley, California; 2 Lawrence Berkeley National Laboratory, Earth Sciences Division, Berkeley, California; University of Massachusetts, Amherst, Massachusetts; and Pacific Northwest National Laboratory, Richland, Washington Aqueous uranium (U(VI)) concentrations in a contaminated aquifer in Rifle CO, have been successfully lowered through electron donor amended bioreduction. Samples collected during the acetate amendment experiment were analyzed for aqueous concentrations of Fe(II), sulfate, sulfide, acetate, U(VI), and δ 34 S of sulfate and sulfide to explore the utility of sulfur isotopes as indicators of in situ acetate amended sulfate and uranium bioreduction processes. Enrichment of up to 7‰ in δ 34 S of sulfate in down-gradient monitoring wells indicates a transition to elevated bacterial sulfate reduction. A depletion in Fe(II), sulfate, and sulfide concentrations at the height of sulfate reduction, along with an increase in the δ 34 S of sulfide to levels approaching the δ 34 S values of sulfate, indicates sulfate-limited conditions concurrent with a rebound in U(VI) concentrations. Upon cessation of acetate amendment, sulfate and sulfide concentrations increased, while δ 34 S values of sulfide returned to less than-20‰ and sulfate δ 34 S decreased to near-background values, indicating lower levels of sulfate reduction accompanied by a corresponding drop in U(VI). Results indicate a transition between electron donor and sulfate- limited conditions at the height of sulfate reduction and suggest stability of biogenic FeS precipitates following the end of acetate amendment. Introduction Long-term uranium sequestration through amended microbial reduction has received much attention in recent years as a potentially fast and efficient means of groundwater remediation (1). In its oxidized form, U(VI), uranium exists as a highly soluble uranylion, UO 22+ . In its reduced form, U(IV), uranium is insoluble and precipitates as stable minerals such as uraninite, UO 2(s) . Since enzymatic reduction of U(VI) to U(IV) was first demonstrated by Lovley et al. (2), over 32 strains of bacterium, mostly iron and sulfate reducers, have been identified as capable of biological uranium reduction (1). Subsurface microbial activity is often electron-donor limited, and thus addition of organic compounds such as lactate, acetate, ethanol, and glucose offer a straightforward means of uranium sequestration (3–8). In situ stimulated bioremediation is now a key component in the research portfolio of DOE’s Environmental Remediation Sciences Program (ERSP)(http://www.lbl.gov/ERSP). Enhanced in situ uranium bioreduction has been successfully demonstrated at several DOE remediation sites, including the Oak Ridge National Laboratory, TN and the Old Rifle Site in western Colorado (3–8). At the Old Rifle Site, acetate has been utilized as the electron donor, and the efficiency of the associated bioremediation processes has been shown to depend directly on the redox conditions generated in the aquifer. During previous acetate amendments, the highest rates of uranium removal were observed at redox levels optimal for microbial iron reduction (3). U(VI) bioreduction continued at lower redox conditions favoring microbial sulfate reduction but at decreased rates. Following cessation of acetate amendment and a return to higher redox conditions, uranium precipitates may be susceptible to oxidation and remobilization (9–14). However, accumulation of biogenic Fe(II) may be capable of sustaining abiotic reduction of U(VI) as oxidizing conditions return to the aquifer (15). Furthermore, preferential oxidation of other bioreduction products such as FeS species may aid in long-term sequestration of uranium as biogenic uraninite (12,15,16). Clearly, understanding the chemical and mineralogical changes occurring within these systems is critical to achieving maximum, sustained bioreduction of uranium. Stable isotope studies have illustrated the benefit of using isotopic measurements to track the biological removal of contaminant species while separating out the effects of transport in through-flowing systems (17–20). Recent studies have also demonstrated enrichment of 238 U relative to 235 U in residual U(VI) during microbial uranium reduction (21). However, the use of sulfur isotopes as indicators of bioremediation progress has been limited to laboratory experiments and nonamended field-scale studies (22). The importance of sulfide species as products of amended bioreduction experiments suggests that stable isotopes of sulfur may prove to be an important tool in investigating in situ uranium bioremediation. Bacterial sulfate reduction (BSR) in the subsurface can be identified through characteristic fractionations in the stable isotope compositions of sulfate and sulfide. BSR preferentially utilizes sulfate containing 32 S, the lighter isotope of sulfur. This preference results in enrichment of S, the heavier isotope of sulfur, in the residual sulfate (23–25). The degree of enrichment of the residual sulfate pool may be influenced by a variety of parameters such as temperature and available organic substrates (25–27). U(VI) reduction by sulfate reducers has also been shown to depend on the type and variety of organic substrate, but, in general, sulfur isotope fractionation during BSR is principally dependent on the magnitude of the preference for reducing sulfate with the lighter sulfur isotope (the fractionation factor)and the extent of sulfate reduction that has occurred (25,28,29). Under most conditions, the difference between the δ 34 S of sulfate minus the δ 34 S of the sulfide produced is ≥ 10‰; however, under sulfate-limited conditions the fractionation factor reduces to almost zero (25). Precipitation of sulfide minerals