•Pristane and phytane were degraded by a NO3- reducing microbial enrichment.•The likely degrading bacterium was Pseudomonas stutzeri.•Degradation was confirmed by addition of 13C-labeled phytane.Isoprenoids, a diverse class of compounds synthesized by all three domains of life, comprise many of the biomarker compounds used in paleoenvironmental and paleoecological reconstruction of Earth history. These biomarkers include hopanoids, sterols and archaeal membrane lipids. While changes in hydrocarbon profiles in anoxic sediments and oilfields indicate that anaerobic microbial metabolism is involved in the disappearance or alteration of isoprenoids, direct links between specific compounds and their microbial degraders are lacking. Here we describe pristane (Pr) and phytane (Ph) degradation associated with NO3- reduction. We confirmed isoprenoid conversion to CO2 using 13C-labeled Ph. After 120 days, dissolved inorganic carbon (DIC) produced in incubations grown with 13C-labeled Ph had a δ13C value of +76.7 ± 11.9‰, significantly higher than values for incubations with unlabeled Ph (−35.7 ± 2.0‰) and those without an added carbon substrate (−30.0 ± 2.1‰). Additional incubations, displayed NO3- reduction after amendment with archaeal diphytanyl glycerol diether (DGD) core lipids, but not in those amended with glycerol diphytanyl glycerol tetraether (GDGT) core lipids. Both 16S rRNA clone libraries and whole cell rRNA-targeted fluorescent in situ hybridization (FISH) indicated that the likely Pr and Ph degrading Bacteria were Gamma proteobacteria, with > 99% similarity to Pseudomonas stutzeri.