Mercury stable isotope fractionation during microbial reduction of Hg(II) to Hg(0)

1. Quantify fractionation during transformations by • pure cultures of microbes and abiotic processes. • Determine isotope ratios for representative sources. Fig. 1 The Mercury Biogeochemical Cycle • Effect of change of environmental conditions • (T, pH, redox) on individual transformations • Fractionation by natural microbial community • Kinetic vs. Equilibrium change • Measure isotope ratios of an element in a natural ecosystem • Identify dominant sources & pathways • Date evolution of element’s microbial transformation • Determine environmental conditions at the time of deposition Fig. 4 Schematic of experimental set up Fig. 5 Measuring fractionation during Hg reduction by naturally occurring microbes • δ202Hg = [(202Hg/198Hg)Sample - 1] * 1000 ‰ (per mil) • (202Hg/198Hg)NIST 3133 Standard • Sample introduction:Cold vapor generation using Sn(II) reduction. • Mass Bias correction: Thallium (NIST 997) added to Hg(0) using desolvating nebulizer. • Precision: Typical internal precision < ±0.01‰ (2 SE) • Fractionation factor: • 202/198 = [(202Hg/198Hg)reactor/(202Hg/198Hg)trap] • Rayleigh Equation: • R(Reactor at t = i)/R(Reactor at t = 0) = f (1/ -1) (1) • RTrap/R(Reactor at t = 0) = (1/) f (1/ -1) (2) • Mass dependence:Multiple Hg isotope ratio (200Hg/198Hg, 204Hg/202Hg) measurement W-76 Kritee1, M. Johnson2, B. Bergquist2, J. D. Blum2, and T. Barkay1 1Rutgers University, 76 Lipman Drive, New Jersey 08901, 2University of Michigan, 1100 N. University Avenue, Michigan 48109 Email: kritee@eden.rutgers.edu Fig. 7 Fractionation of Hg isotopes by Hg(II) resistant microbes from a natural source Introduction Fig 2. Development of a stable isotope ratio based tool. • Biogeochemical concern • There are multiple sources (natural vs. anthropogenic, local vs. global) and transformations (microbial vs. abiotic) that can lead to buildup of methylmercury (Fig. 1). • In order to design/implement effective remediation strategies, we need tools to track the actual causes of Hg accumulation in a given ecosystem. Mercury stable isotope fractionation during microbial reduction of Hg(II) to Hg(0) Stage 1 • Enrichment of Hg(II) resistant microbes in a natural water sample (Increase in % of Hg(II) resistant colony forming units per ml (CFU/ml) with time7). B. Isotopic composition of the Hg(II) remaining in the reactor and Hg(0) produced by enriched microbes. Stage 2 Table 1 Stage 3 Methods Conclusions • Mercury resistant bacteria prefer lighter isotopes when reducing Hg(II) to Hg(0) (Fig. 6 and 7). • The similar values of  observed for the experiments done with two bacterial genera (B. cereus; preliminary results not shown) and a natural community (Table 1) suggest that the isotopic signature produced during biological Hg(II) reduction is unique. • Hg isotopes have the potential for distinguishing between different pathways leading to Hg(0) production based on the extent of fractionation and could help Hg remediation efforts. • Hg is the heaviest element4 for which mass dependent biological fractionation has been documented to date (Table 2). • The extent of fractionation observed per atomic mass unit (amu) is very high and is comparable to fractionation by much lighter elements (Table 2). • Stable isotopes fractionation • Ratio of abundance of a heavier to a lighter stable isotope as compared to a standard - reported as delta () per mil (see methods). • Measurement4 of isotope ratios of many elements (1H to 96Mo) has helped us determine: • Sources of pollutants or nutrients • Dominant pathways transforming the element • Paleo-environmental conditions (T, pH etc.) • Living organisms preferentially uptake lighter isotopes leaving heavier isotopes in the environment. • Why does Life like lighter isotopes?It takes less energy to uptake/process lighter molecules. • Hg has seven stable isotopes (Fig. 2). Significant Hg isotope ratio variations in natural samples from ores, sediment cores, and fish tissues have been reported1-3, but the processes leading to the fractionation have not yet been explored. Table 2 Multiple collector inductively coupled plasma mass spectrometry (MC-ICPMS)1,6 Research Objectives • Broader research question • Can Hg isotope ratios serve as a tool to differentiate between different types of sources and transformation pathways? • To answer this question the scheme depicted in Fig. 2 is followed. • Objectives of this study • Does reduction of Hg(II) to Hg(0) by pure cultures of Hg resistant bacteria (Fig. 3) cause fractionation? (Stage 1 in Fig. 2) Future research • To use Hg stable isotopes ratios as a successful bio-geochemical tool we need to: • Address additional transformations (methylation, de-methylation and long range transport) in the Hg cycle including abiotic processes (Stages 1-2; Fig. 2). • Improve instrumental sensitivity to get precise isotopic composition of natural samples with sub-ppb Hg concentrations. References • Smith C. et al. (2005), Geology 33(10), 825-828 • 2. Jackson T. A. (2004), Env. Sci. & Tech. 38(10) 2813-2821 • 3. Hintelman and Lu (2003), Analyst 128, 635-638 • 4. Johnson C. M. et al. (Ed.) (2004), Geochemistry of non-traditional isotopes. Reviews in Mineralogy & Geochemsitry 55 • 5. Barkay T. et al. (2003), FEMS Microbiol. Rev. 27, 355-384 • 6. Lauretta et al. (2001), Geochim. Cosmochim. Acta 65, 2807-18 • 7. Barkay T. (1987), Appl. & Env. Microbiol. 53(12), 2725-32 Results Fig. 6 The isotope data plotted as δ202/198Hg vs. f E. coli JM109/pPB117 at 370C [A] and 300C [B] Acknowledgements 2. What is the effect of changing incubation temperature on fractionation? (Stage 2) 3. Do naturally occurring microbes fractionate Hg when reducing Hg(II)? (Stage 2) Funding was provided by the NSF and NJWRRI. We thank Drs. Bjoërn Klaue, John Reinfelder, Paul Falkowski & Ariel Anbar for their helpful inputs at different stages of this project and Matt Meredith for help with performing experiments at Rutgers.