Microbial dehalogenation of trichlorinated dibenzo-p-dioxins by a Dehalococcoides-containing mixed culture is coupled to carbon isotope fractionation.

An anaerobic enrichment culture reductively dehalogenated 1,2,4- and 1,2,3-trichlorodibenzo-p-dioxin (TrCDD) almost exclusively at peripheral positions forming the main products 1,3-dichloro-(DiCDD) and 2-monochlorodibenzo-p-dioxin (MCDD) from 1,2,4-TrCDD and 2,3-DiCDD from 1,2,3-TrCDD. Dehalococcoides was monitored in the mixed culture by quantitative real-time PCR. A yield of 2.5 x 10(8) to 2.75 x 10(8) copies of 16S rRNA genes per micromole of chloride released suggested growth by dehalorespiration with dibenzo-p-dioxins. For the analysis of carbon isotope fractionation, the dioxin congeners were isolated by solid-phase microextraction (SPME) from the headspace of the cultures. The delta13C composition of 1,2,4-TrCDD did not change remarkably during the course of reductive dehalogenation; however, the intermediate 1,3-DiCDD became enriched, and the final product 2-MCDD significantly depleted in 13C with a discrimination of 2.5-3.6 per thousand between 1,3-DiCDD and 2-MCDD. 1,2,3-TrCDD and its main product 2,3-DiCDD became slightly enriched in 13C, whereas the formed low concentrations of 2-MCDD were depleted in 13C by 5.5-4.8 per thousand. This study demonstrates carbon isotope fractionation during sequential reductive dehalogenation of chlorinated dibenzo-p-dioxins, whereby isotope fractionation upon dehalogenation of the intermediate was substantial. This can provide a basis for the development of a new method to monitor the fate of dioxins in the environment using compound specific stable isotope analyses.