Abstract
Summary Iso ‐alkanes comprise a substantial proportion of petroleum and refined products that impact the environment, but their fate is cryptic under methanogenic conditions. We investigated methanogenic biodegradation of C 7 and C 8 iso ‐alkanes found in naphtha, specifically 2‐methylhexane, 3‐methylhexane, 2‐methylheptane, 4‐methylheptane and 3‐ethylhexane. These were incubated as a mixture or individually with enrichment cultures derived from oil sands tailings ponds that generate methane from naphtha components; substrate depletion and methane production were monitored for up to 663 days. 3‐Methylhexane and 4‐methylheptane were degraded both singly and in the mixture, whereas 2‐methylhexane and 2‐methylheptane resisted degradation as single substrates but were depleted in the iso ‐alkane mixture, suggesting co‐metabolism. 3‐Ethylhexane was degraded neither singly nor with co‐substrates. Putative metabolites consistent with succinylated C 7 and C 8 were detected, suggesting activation by addition of iso ‐alkanes to fumarate and corresponding to detection of alkylsuccinate synthase‐like genes. 454 pyrotag sequencing, cloning and terminal restriction fragment length polymorphism of 16 S rRNA genes revealed predominance of a novel member of the family P eptococcaceae (order C lostridiales) and Archaea affiliated with M ethanoregula and M ethanosaeta . We report here isomer‐specific metabolism of C 7 ‐ C 8 iso ‐alkanes under methanogenic conditions and propose their activation by a novel P eptococcaceae via addition to fumarate.