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Position-specific isotope analysis of the methyl group carbon in methylcobalamin for the investigation of biomethylation processes

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Abstract

In the environment, the methylation of metal(loid)s is a widespread phenomenon, which enhances both biomobility as well as mostly the toxicity of the precursory metal(loid)s. Different reaction mechanisms have been proposed for arsenic, but not really proven yet. Here, carbon isotope analysis can foster our understanding of these processes, as the extent of the isotopic fractionation allows to differentiate between different types of reaction, such as concerted (SN2) or stepwise nucleophilic substitution (SN1) as well as to determine the origin of the methyl group. However, for the determination of the kinetic isotope effect the initial isotopic value of the transferred methyl group has to be determined. To that end, we used hydroiodic acid for abstraction of the methyl group from methylcobalamin (CH3Cob) or S-adenosyl methionine (SAM) and subsequent analysis of the formed methyl iodide by gas chromatography (GC) isotope ratio mass spectrometry (IRMS). In addition, three further independent methods have been investigated to determine the position-specific δ 13C value of CH3Cob involving photolytic cleavage with different additives or thermolytic cleavage of the methyl-cobalt bonding and subsequent measurement of the formed methane by GC-IRMS. The thermolytic cleavage gave comparable results as the abstraction using HI. In contrast, photolysis led to an isotopic fractionation of about 7 to 9 ‰. Furthermore, we extended a recently developed method for the determination of carbon isotope ratios of organometal(loid)s in complex matrices using hydride generation for volatilization and matrix separation before heart-cut GC and IRMS to the analysis of the low boiling partly methylated arsenicals, which are formed in the course of arsenic methylation. Finally, we demonstrated the applicability of this methodology by investigation of carbon fractionation due to the methyl transfer from CH3Cob to arsenic induced by glutathione.

Position-specific isotope analysis of the methyl group in CH3Cob by abstraction using HI and subsequent analysis of formed CH3I by GC-IRMS

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References

  1. Thayer JS (2002) Biological methylation of less-studied elements. Appl Organomet Chem 16(12):677–691

    Article  CAS  Google Scholar 

  2. Thayer JS (1989) Methylation: its role in the environmental mobility of heavy elements. Appl Organomet Chem 3(2):123–128. doi:10.1002/aoc.590030202

    Article  CAS  Google Scholar 

  3. Qin J, Rosen BP, Zhang Y, Wang GJ, Franke S, Rensing C (2006) Arsenic detoxification and evolution of trimethylarsine gas by a microbial arsenite S-adenosylmethionine methyltransferase. Proc Natl Acad Sci U S A 103(7):2075–2080. doi:10.1073/pnas.0506836103

    Article  CAS  Google Scholar 

  4. Lin S, Shi Q, Nix FB, Styblo M, Beck MA, Herbin-Davis KM, Hall LL, Simeonsson JB, Thomas DJ (2002) A novel S-adenosyl-l-methionine: arsenic(III) methyltransferase from rat liver cytosol. J Biol Chem 277(13):10795–10803. doi:10.1074/jbc.M110246200

    Article  CAS  Google Scholar 

  5. Hayakawa T, Kobayashi Y, Cui X, Hirano S (2005) A new metabolic pathway of arsenite: arsenic-glutathione complexes are substrates for human arsenic methyltransferase Cyt19. Arch Toxicol 79(4):183–191. doi:10.1007/s00204-004-0620-x

    Article  CAS  Google Scholar 

  6. Bentley R, Chasteen TG (2002) Microbial methylation of metalloids: arsenic, antimony, and bismuth. Microbiol Mol Biol Rev 66(2):250. doi:10.1128/mmbr.66.2.250-271.2002

    Article  CAS  Google Scholar 

  7. Chasteen TG, Bentley R (2003) Biomethylation of selenium and tellurium: microorganisms and plants. Chem Rev 103(1):1–25. doi:10.1021/cr010210+

    Article  CAS  Google Scholar 

  8. Ridley W, Dizikes L, Wood J (1977) Biomethylation of toxic elements in the environment. Science 197(4301):329–332. doi:10.1126/science.877556

    Article  CAS  Google Scholar 

  9. Challenger F (1945) Biological methylation. Chem Rev 36(3):315–361

    Article  CAS  Google Scholar 

  10. Thomas F, Diaz-Bone RA, Wuerfel O, Huber B, Weidenbach K, Schmitz RA, Hensel R (2011) Connection between multimetal(loid) methylation in methanoarchaea and central intermediates of methanogenesis. Appl Environ Microbiol 77(24):8669–8675. doi:10.1128/aem.06406-11

    Article  CAS  Google Scholar 

  11. Schrauzer GN, Seck JA, Holland RJ, Beckham TM, Rubin EM, Sibert JW (1973) Reductive dealkylation of alkylcobaloximes, alkylcobalamins, and related compounds: simulation of corrin dependent reductase and methyl group transfer reactions. Bioinorg Chem 2(2):93–124

    Article  CAS  Google Scholar 

  12. Zakharyan RA, Aposhian HV (1999) Arsenite methylation by methylvitamin B-12 and glutathione does not require an enzyme. Toxicol Appl Pharmacol 154(3):287–291

    Article  CAS  Google Scholar 

  13. Nakamura K, Hisaeda Y, Pan L, Yamauchi H (2009) Methyl transfer from a hydrophobic vitamin B12 derivative to arsenic trioxide. J Organomet Chem 694(6):916–921

    Article  CAS  Google Scholar 

  14. Elsner M (2010) Stable isotope fractionation to investigate natural transformation mechanisms of organic contaminants: principles, prospects and limitations. J Environ Monit 12(11):2005–2031. doi:10.1039/c0em00277a

    Article  CAS  Google Scholar 

  15. Matthews DE, Hayes JM (1978) Isotope-ratio-monitoring gas chromatography–mass spectrometry. Anal Chem 50(11):1465–1473

    Article  CAS  Google Scholar 

  16. Sessions AL (2006) Isotope-ratio detection for gas chromatography. J Sep Sci 29(12):1946–1961

    Article  CAS  Google Scholar 

  17. Caytan E, Botosoa EP, Silvestre V, Robins RJ, Akoka S, Remaud GS (2007) Accurate quantitative C-13 NMR spectroscopy: repeatability over time of site-specific C-13 isotope ratio determination. Anal Chem 79(21):8266–8269. doi:10.1021/ac070826k

    Article  CAS  Google Scholar 

  18. Schrauzer GN, Sibert JW, Windgassen RJ (1968) Photochemical and thermal cobalt-carbon bond cleavage in alkylcobalamins and related organometallic compounds. Comparative study. J Am Chem Soc 90(24):6681–6688. doi:10.1021/ja01026a021

    Article  CAS  Google Scholar 

  19. Fanchiang YT, Pignatello JJ, Wood JM (1983) Demethylation of methylcobalamin by platinum(IV)/platinum(II) couples. Formation of methylplatinum(IV) products. Organometallics 2(12):1748–1751. doi:10.1021/om50006a008

    Article  CAS  Google Scholar 

  20. Manley SL (1994) The possible involvement of methylcobalamin in the production of methyl iodide in the marine environment. Mar Chem 46(4):361–369

    Article  CAS  Google Scholar 

  21. Dolphin D, Johnson AW, Rodrigo R (1964) 606. Reactions of the alkylcobalamins. Journal of the Chemical Society (Resumed)

  22. Yamada RH, Shimizu S, Fukui S (1966) Factors affecting the anaerobic photolysis of the cobalt–carbon bond of cobalt-methylcobalamin. Biochim Biophys Acta 124(1):195

    Article  CAS  Google Scholar 

  23. Zeisel S (1885) Über ein Verfahren zum quantitativen Nachweise von Methoxyl. Monatshefte für Chemie 6(1):989–997. doi:10.1007/bf01554683

    Article  Google Scholar 

  24. Zeisel S (1886) Zum quantitativen Nachweise von Methoxyl. Monatshefte für Chemie / Chemical Monthly 7(1):406–409. doi:10.1007/bf01516585

    Article  Google Scholar 

  25. Keppler F, Kalin RM, Harper DB, McRoberts WC, Hamilton JTG (2004) Carbon isotope anomaly in the major plant C-1 pool and its global biogeochemical implications. Biogeosciences 1(2):123–131

    Article  CAS  Google Scholar 

  26. Greule M, Mosandl A, Hamilton JTG, Keppler F (2009) A simple rapid method to precisely determine C-13/C-12 ratios of plant methoxyl groups. Rapid Commun Mass Spectrom 23(11):1710–1714. doi:10.1002/rcm.4057

    Article  CAS  Google Scholar 

  27. Wuerfel O, Diaz-Bone RA, Stephan M, Jochmann MA (2009) Determination of C-13/C-12 isotopic ratios of biogenic organometal(loid) compounds in complex matrixes. Anal Chem 81(11):4312–4319. doi:10.1021/ac8027307

    Article  CAS  Google Scholar 

  28. Diaz-Bone RA, Hitzke M (2008) Multi-element organometal(loid) speciation by hydride generation-GC-ICP-MS: overcoming the problem of species-specific optima by using a pH-gradient during derivatisation. J Anal At Spectrom 23:861–810

    Article  CAS  Google Scholar 

  29. Merijani A, Zingaro RA (1966) Arsine oxides. Inorg Chem 5(2):187

    Article  Google Scholar 

  30. Werner RA, Brand WA (2001) Referencing strategies and techniques in stable isotope ratio analysis. Rapid Commun Mass Spectrom 15(7):501–519

    Article  CAS  Google Scholar 

  31. Jochmann MA, Blessing M, Haderlein SB, Schmidt TC (2006) A new approach to determine method detection limits for compound-specific isotope analysis of volatile organic compounds. Rapid Commun Mass Spectrom 20(24):3639–3648. doi:10.1002/rcm.2784

    Article  CAS  Google Scholar 

Download references

Acknowledgments

We thank David Widory of the BGRM (Orleans, France) for measurement of our reference CO2 gas. In addition, we thank Roland Diaz-Bone and Marcel Schulte. Furthermore, we acknowledge financial support by the German Research Foundation (DFG) and the Centre for Water and Environmental Research (ZWU). Marcus Greule and Frank Keppler were supported by the DFG (KE 884/6-1).

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Authors and Affiliations

  1. Instrumental Analytical Chemistry, University of Duisburg-Essen, Universitätsstrasse 5, 45141, Essen, Germany

    Oliver Wuerfel, Maik A. Jochmann & Torsten C. Schmidt

  2. Max-Planck-Institute for Chemistry, Hahn-Meitner-Weg 1, 55128, Mainz, Germany

    Markus Greule & Frank Keppler

  3. Centre for Water and Environmental Research (ZWU), University of Duisburg-Essen, Universitätsstraße 2, 45141, Essen, Germany

    Torsten C. Schmidt

Authors
  1. Oliver Wuerfel
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  2. Markus Greule
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  3. Frank Keppler
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  4. Maik A. Jochmann
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  5. Torsten C. Schmidt
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Corresponding author

Correspondence to Maik A. Jochmann.

Additional information

Published in the topical collection Isotope Ratio Measurements: New Developments and Applications with guest editors Klaus G. Heumann and Torsten C. Schmidt.

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Wuerfel, O., Greule, M., Keppler, F. et al. Position-specific isotope analysis of the methyl group carbon in methylcobalamin for the investigation of biomethylation processes. Anal Bioanal Chem 405, 2833–2841 (2013). https://doi.org/10.1007/s00216-012-6635-x

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  • DOI: https://doi.org/10.1007/s00216-012-6635-x

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