Anaerobic biodegradation of the isoprenoid biomarkers pristane and phytane Katherine S. Dawson ⇑ , Irene Schaperdoth, Katherine H. Freeman, Jennifer L. Macalady Pennsylvania State University, Department of Geosciences, University Park, PA 16802, USA article info Article history: Received 26 June 2013 Received in revised form 27 September 2013 Accepted 16 October 2013 Available online 22 October 2013 abstract 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 hydrocar- bon 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 micro- bial degraders are lacking. Here we describe pristane (Pr) and phytane (Ph) degradation associated with NO  3 reduction. We confirmed isoprenoid conversion to CO 2 using 13 C-labeled Ph. After 120 days, dis- solved inorganic carbon (DIC) produced in incubations grown with 13 C-labeled Ph had a d 13 C 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 NO  3 reduc- tion 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. Ó 2013 Elsevier Ltd. All rights reserved. 1. Introduction Several reviews have highlighted progress in understanding the microbiology and biochemistry associated with anaerobic hydro- carbon degradation (Spormann and Widdel, 2000; Lovley, 2001; Head et al., 2003; Widdel et al., 2006; Heider, 2007; Foght, 2008; Grossi et al., 2008; Widdel et al., 2010; Mbadinga et al., 2011). Additional studies have investigated biodegradation of hydrocar- bon mixtures in subsurface petroleum reservoirs and contami- nated aquifers (Head et al., 2003; Larter et al., 2003; Townsend et al., 2003; Jones et al., 2008; Gieg et al., 2010). Most studies have focused on the anaerobic biodegradation of pollutants such as ben- zene and toluene and gaseous hydrocarbons such as methane and propane. In contrast, anaerobic biodegradation of linear and cyclic isoprenoid hydrocarbons has received less attention (Hylemon and Harder, 1998) despite the relevance of these compounds as essen- tial biomarkers for taxonomic groups (Brocks and Pearson, 2005; Peters et al., 2005; Dutkiewicz et al., 2006) and paleoenvironmen- tal conditions such as temperature and salinity (Wuchter et al., 2006; Turich and Freeman, 2011), as well as petroleum maturity (Petrov and Abryutina, 1989). Isoprenoid hydrocarbons can repre- sent up to 1% of a crude oil, with a major contribution from pris- tane (Pr) and phytane (Ph; Tissot and Welte, 1978). Hydrocarbons in general are recalcitrant to anaerobic biodegra- dation and require activation by conversion to more oxidized inter- mediates prior to catabolism via b-oxidation (Heider et al., 1998; Heider, 2007). Variation in susceptibility to biodegradation de- pends on features such as chain length, branching and cyclization (Head et al., 2003; Peters et al., 2005). Isoprenoids derive from lip- ids made by organisms from all three domains of life and are syn- thesized from branched, C 5 isoprene units. Branching can impart additional recalcitrance by hindering b-oxidation (Cantwell et al., 1978; Schaeffer et al., 1979). In studies of aerobic branched alkane degradation, this hindrance is overcome through additional deca- rboxymethylation steps (Seubert and Fass, 1964; Pirnik et al., 1974; Pirnik and McKenna, 1977; Cantwell et al., 1978). The need for repeated decarboxymethylation or alternative demethylation steps before b-oxidation can proceed effectively reduces the meta- bolic energy yield from isoprenoid substrates and may explain why degradation in natural environments appears slower than for equivalent non-isoprenoid hydrocarbons (Head et al., 2003). In addition, iso-methyl branching at the chain ends (e.g. Pr, archaeol) likely prevents some activation mechanisms, such as fumarate addition (Heider, 2007), due to steric hindrance, while low solubil- ity likely reduces bioavailability (Fichan et al., 1998; Hylemon and Harder, 1998). 0146-6380/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.orggeochem.2013.10.010 ⇑ Corresponding author. Present address: California Institute of Technology, Division of Geological and Planetary Sciences, Pasadena, CA 91125, USA. Tel.: +1 626 395 6894; fax: +1 626 568 0935. E-mail address: kdawson@caltech.edu (K.S. Dawson). Organic Geochemistry 65 (2013) 118–126 Contents lists available at ScienceDirect Organic Geochemistry journal homepage: www.elsevier.com/locate/orggeochem