Isolation and identification of fluoranthene biodegradation products

Abstract

Analytical procedures for isolating and identifying the biodegradation products of fluoranthene, a model four ring polycyclic aromatic hydrocarbon, were improved for ex-situ experiments using the pure bacterial strain Pasteurella sp. IFA. The stable degradation products were determined over a 10 day incubation period. The most efficient method for isolating the metabolites was a two step liquid–liquid extraction using ethyl acetate. Because of the polar nature of fluoranthene metabolites, separation by GC proved inefficient. Their chromatographic properties, however, were improved by derivatisation: silylation with N-methyl-N-(trimethylsilyl) trifluoroacetamide for hydroxy and carboxy groups, oxime formation with O-(2,3,4,5,6-pentafluorobenzyl) hydroxylamine hydrochloride for carbonyl groups, or a combination of both. The metabolites were identified using gas chromatography with mass selective detection (GC-MS/SCAN, SIM). Using this method we were able to identify and quantify six stable fluoranthene metabolites (9-fluorenone-1-carboxylic acid, 9-fluorenone, 9-hydroxy-1-fluorene-carboxylic acid, 2-carboxybenzaldehyde, benzoic acid and phenylacetic acid).

References

1. M. A. Hietkamp, J. P. Freeman, D. W. Miller and C. E. Cerniglia, Appl. Environ. Microbiol., 1988, 54, 2556.
2. W. D. Weissenfels, M. Beyer and J. Klein, Appl. Microbiol.Biotechnol., 1990, 32, 479.
3. I. Kelley, J. P. Freeman, F. E. Evans and C. E. Cerniglia, Appl.Environ. Microbiol., 1991, 57, 636.
4. B. Boldrin, A. Tiehm and C. Fritzsche, Appl. Environ. Microbiol., 1993, 59, 1927.
5. X.-F. Li, W. R. Cullen, K. J. Reimer and X.-C. Le, Sci. Total Environ., 1996, 177, 17.
6. I. Kelley and C. E. Cerniglia, J. Ind. Microbiol., 1991, 7, 19.
7. I. Kelley, J. P. Freeman, F. E. Evans and C. E. Cerniglia, Appl.Environ. Microbiol., 1993, 59, 800.
8. J. G. Muller, P. J. Chapman and P. H. Pritchard, Appl. Environ.Microbiol., 1989, 55, 3085.
9. C. L. Holder, W. A. Korfmacher, W. Slikker, D. A. Miller and A. W. Gosnell, Biomed. Mass Spectrom., 1985, 12, 151.
10. E. Šepič, M. Bricelj and H. Leskovšek, J. Appl. Microbiol., 1997, 83, 561.
11. E. Šepič, M. Bricelj and H. Leskovšek, J. Appl. Microbiol., 1998, 85, 554.
12. E. Heath, PhD Thesis, University of Ljubljana, Ljubljana, Slovenia, 1998.
13. C. F. Poole, in Handbook of Derivatives for Chromatography, ed. K. Blau and G. S. King, Heyden & Son, London, 1977, ch. 4.
14. S. Sloan, D. J. Harvey and P. Vouros, Org. Mass Spectrom., 1971, 5, 789.
15. M. Ende and H. Luftmann, Tetrahedron, 1984, 40, 5167.
16. P. Kováč and D. Anderle, in Handbook of Derivatives for Chromatography, ed. K. Blau and G. S. King, Heyden & Son, London, 1977, ch. 5.
17. M. A. Hietkamp, J. P. Freeman and C. E. Cerniglia, Appl. Environ.Microbiol., 1987, 53, 129.
18. C. E. Cerniglia, in Petroleum Microbiology, ed. R. M. Atlas, Macmillan, New York, 1984, ch. 3.
19. D. T. Gibson and V. Subramanian, Microbial Degradation of Organic Compounds, ed. D. T. Gibson, Marcel Dekker, New York, 1984, ch. 7.
20. C. E. Cerniglia, Biodegradation, 1992, 3, 351.