OVERCOMING THE CHALLENGE OF IDENTIFYING BIODEGRADATION IN SUBARCTIC REGIONS USING ANALYSIS OF BENZYLSUCCINATES AND STABLE ISOTOPIC RATIOS

Isotopic fractionation of contaminants and the identification of distinctive metabolites of toluene (benzylsuccinate) and xylenes (methylbenzylsuccinates) at field sites can be used as indicators of in situ biodegradation. At a contaminated groundwater site near Fairbanks, Alaska, isotopic analysis of benzene and toluene and measurement of benzylsuccinate and methylbenzylsuccinates was carried out to assess in situ biodegradation. The combined use of both techniques was particularly valuable in this subarctic environment where biodegradation rates can be slow and demonstrating biodegradation can be difficult using other methods.  

d¹³C and d²H analysis of benzene suggested that biodegradation of this contaminant was unlikely at this site. In contrast, an isotopic enrichment in d ¹³C of 2‰ and enrichment in d²H of 70‰ in toluene occurred, suggesting in situ toluene biodegradation. Xylene and toluene biodegradation was also supported by the presence of 17-50 µg/L of methylbenzylsuccinate and 10-49 µg/L of benzylsuccinate.  

A comparison of isotopic and benzylsuccinate analysis for demonstrating anaerobic toluene biodegradation at this site is useful. Toluene biodegradation was indicated by the presence of benzylsuccinate in the three wells with the highest concentrations of toluene. Benzylsuccinate concentrations are typically much lower than that of toluene in groundwater. As such, in the wells with lower concentrations of toluene, benyzylsuccinate concentrations were below detection limits. In contrast, isotopic analysis works best for samples in which a large fraction of the toluene has been degraded, since significant isotopic fractionation is often not resolvable until a large extent of biodegradation occurs. Hence, in the higher concentration wells, d¹³C and d²H values for toluene agreed within error and provided no evidence of biodegradation. In contrast, in samples with lower toluene concentrations, while benzylsuccinate was below detection limits, isotopic enrichment in ¹³C and ²H of toluene provided evidence of biodegradation. Analysis of metabolites may therefore provide the earliest evidence of biodegradation, while stable carbon and hydrogen isotopes may continue to provide support for biodegradation in less contaminated wells where metabolites may be below detection limits.