Compound specific isotope analysis to evaluate in situ transformation of a complex mixture of substituted chlorobenzenes in a pilot constructed wetland system

Constructed wetlands can be a suitable remediation technique for the treatment of industrial contaminants via transfer (i.e., non-destructive) and transformation (i.e., destructive) processes. Providing direct evidence of in situ transformation using concentrations and biogeochemical parameters alone is challenging. Compound specific isotope analysis (CSIA) is a widely used tool to assess in situ transformation of contaminants based on changes in their stable isotope signatures. In this work, we evaluated the potential of CSIA to identify and possibly quantify the in situ transformation of six NO2- and NH2-chlorobenzenes in complex aqueous samples from a pilot constructed wetland system. No significant changes in δ13C, i.e., ≤2‰ were observed for any of the target compounds despite the contaminant concentration decreased by more than 99% between the inlet and outlet of the system. Using multi-element CSIA of carbon, hydrogen, and nitrogen and laboratory-derived isotope enrichment factors, we successfully identified and quantified the extent of in situ transformation of 2,3-dichloroaniline (2,3-DCA) in the pilot constructed wetlands. The isotopic trends provide evidence for aerobic biotransformation as a dominant pathway in the surface flow planted wetlands; whereas sorption was identified as the likely process in planted and unplanted upflow gravel bed wetlands during the initial wetland operation periods. Another major contaminant from the NO2-chlorobenzene group, i.e., 2-chloronitrobenzene (2-CNB), showed negligible δ13C, and small δ2H (±20‰) and δ15N (±2‰) isotope fractionation. No laboratory-controlled CSIA studies are yet available for 2-CNB biotransformation to characterize transfer and transformation processes. This study highlights the applicability of CSIA as a quantitative tool for 2,3-DCA in dynamic environmental conditions of wetlands and the need for pathway-specific isotope enrichment factors for the successful CSIA application of other target compounds.