EPRW 2002 - groundwater metabolite

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This work was presented at "EPRW 2002". It illustrates the identification of 3-sec-butyl-6-methyluracil, a metabolite isolated in groundwater, probably derived from herbicide "Bromacil" …

This work was presented at "EPRW 2002". It illustrates the identification of 3-sec-butyl-6-methyluracil, a metabolite isolated in groundwater, probably derived from herbicide "Bromacil" (3-sec-butyl-5-bromo-6-methyluracil).

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  • 1. IDENTIFICATION OF A “METABOLITE” IN GROUNDWATER, WITH STRUCTURE RELATED TO BROMACIL: 3-SEC-BUTYL-6-METHYLURACIL Tiziano VENDRAME, Alfredo MUSSATO ARPAV - Agenzia Regionale per la Prevenzione e Protezione Ambientale del Veneto - Servizio Laboratori del Dipartimento Provinciale ARPAV di Treviso - Via D’Annunzio 12, 31100 Treviso - Italy e-mail: tvendrame@arpa.veneto.it - amussato@arpa.veneto.it During groundwater controls it is not unusual detect compounds that are different from analysis’ object. Most of analytical “artifacts” (e.g. reagent contaminants, release from contact materials, stationary phase degradation in GC) are well known interferences. Typical examples are phthalates and silicon derivatives. A lot of other compounds really present in sample are usually overlooked, if they are not analysis’ targets, especially in routine analyses that are conditioned from time and cost. However, for an effective aquifer control, the simple search of compounds from “rigid” lists is inadequate. The appearance of new “pollutants” in known aquifers deserves a deeper study. In our case, attention was attracted from a new chromatographic peak (Fig. 1a - 1b), noted in routine analyses of triazinic herbicides (SPE technique, C8 phase), for two main reasons: • The presence of an unusual herbicide/pesticide nitrogen containing was suspected from positive response to NPD detector; • The distribution in a delimited area, near a small town, in repeated samples (this excluded an analytical artifact). Last peculiarity, since the town is aqueduct lacking, spurred the research on this unknown compound. Initial GC-MS screening was a poor aid: our library (Wiley 275.l) is lacking of wanted spectra (Fig. 1c). Other libraries also gave no useful data. The feeling on a possible new pesticide press to extend sampling, and then circumscribe “polluted” area, until exhausted gravel quarry. The lacking of upstream wells stopped the research for long time. Later the starting point of pollution was identified in a closed landfill, six kilometers away. The “unknown” compound was detected only in downstream groundwater wells and in landfill leachate. In spite of a well-known leak from this landfill, the finding of a pollutant, although in trace only, so remote from origin, was an unexpected finding (Fig. 2). Probably the landfill leak started two year ago, at least, with peaks of 100 mg/l of ammonia in the nearest wells. However, the “common” pollutants (e.g. ammonia, chloride etc.) are not detectable in aquifer some hundreds meters downstream, notwithstanding the gravel ground. After source’s discovery, it was urgent try compound identification, characterized from: • mass spectra with feeble molecular ion, mass = 182 (1-2%), near other peaks, sources of possible confusion; 1
  • 2. • base ion mass = 127 (100%) • characteristic masses: 167 (5-6%), 153 (10%), 110 (23%), 84 (30%), 70 (8-9%); • halogen lacking from spectra and ECD detector behaviour. Structure assignation started from a molecule with spectra similar to unknown compound, the 3-propyl-6-methyluracil, selected (visually) among those singled from research program of our library (Fig. 3). This have MW (molecular weight) 168, whereas unknown have MW 182, but shows similar fragmentation feature, at lower mass. Then it was hypotized an additional CH3 or CH2 in “unknown”, and it was outlined some possible isomers and keto-enolic tautomers (12 with MW 182 ad 4 with MW 168 for comparison). From “Beilstein Handbuch der Organische Chemie” (“on-line” version), resulted 5 known compounds, but the only one with right spectrum is 3-sec-butyl-6-methyluracil (ref. 1 for spectrum description - Fig. 5 for structure) This compound is not commercially available, but starting from few articles, which cited this substance in last years, it was contacted at last Prof. J. Suflita (Univ. Oklahoma). He supplied kindly a sample of this compound, which shows chromatographic behaviour and spectra identical with “unknown” (Fig. 4). Later, the agreement of fragmentation pattern with specialized literature (2-3) was checked. With this standard was possible to measure concentrations in groundwater which increase from about 0,1 µg/l near the town (six kilometers away from landfill), to 10 µg/l in wells near the landfill, and 5000 µg/l in landfill leachate (see ref. 5 for comparison standards). The origin of this compound in landfill is still obscure. From patent literature results a possible use as intermediate in “Bromacil” synthesis (3-sec-butyl-5-bromo-6-methyluracil), via bromuration. Bromacil is a well-known herbicide, but with limited use in North Italy. No other industrial use was quoted in available literature. A possible source is also microbiological debromination of Bromacil, in anoxic conditions (see ref. 4 - A. Neal, J. Suflita), similar to landfill interior. Then, the compound can be buried directly in landfill, or it can derive from Bromacil residues degraded in landfill. The Bromacil research gives negative results; however could not be ruled out the second hypothesis, because of possible fast degradation of Bromacil in anoxic environment. The topical problem is to decide if polluted water (at low level) is allowable as drinking water. Bibliography and notes: 1. A. Acher, C. Hapeman J. Agric. Food Chem. 1994, 42, pp. 2040-2047: Comparison of formation and biodegradation of bromacil oxidation products in aqueous solutions; 2. J. M. Rice J. of the American Chemical Society, (1965) pp. 4569-4576: Mass spectra of nucleic acid derivatives. Pyrimidines; 3. E. Falch Acta Chemica Scandinavica, 24 (1970) pp.137-144: Mass spectra of Pyrimidines (Part I. N-Alkiluracils); 4. A. Neal, J. Suflita Appl. Environ. Microbiol. 56 (1), pp. 292-294: Reductive dehalogenation of a nitrogen heterocyclic herbicide in anoxic aquifer slurries. 5. A second sample of standard was synthesized at laboratory of "Istituto Tecnico Industriale Statale E. Fermi" in Treviso, from R. Scandiuzzi (Laboratory Teacher) and his students. 2
  • 3. Chromatographic behaviour and spectrum was identical with sample from Prof. J. Suflita (Fig. 7). Illustration Index: Fig. 1 a: chromatogram of typical water well extract (SPE technique, C8 phase). Fig. 1 b: as 1 a, enlarged; the "unknown" peak is at 8,9 min (red). Fig. 1 c: spectrum of 3-sec-butyl-6-methyluracil from the same extract of 1 a. Fig. 2: spatial distribution of 3-sec-butyl-6-methyluracil (red: positive; blue: negative). Fig. 3: spectrum of 3-propyl-6-methyluracil (Wiley 275.l). Fig. 4: spectrum of 3-sec-butyl-6-methyluracil (sample from Prof. J. Suflita). Fig. 5: structures of 3-sec-butyl-6-methyluracil and "Bromacil" (5-bromo3-sec-butyl-6- methyluracil). Fig. 6: possible relations between 3-sec-butyl-6-methyluracil and "Bromacil" . Fig. 7: spectrum of 3-sec-butyl-6-methyluracil (sample from I.T.I.S. "E. Fermi" - Treviso). Fig 1a Index 3
  • 4. Fig 1b Index 4
  • 5. Fig1c Index 5
  • 6. 6
  • 7. Fig 2 Index 7
  • 8. Fig 3 Index 8
  • 9. Fig 4 Index Fig 5 Index 9
  • 10. Fig 6 Index 10
  • 11. Fig 7 Index 11
  • 12. 12
  • 13. 13