OPTIMIZING METHODS FOR EXTRACTING LIPIDS FROM ORGANIC-RICH SUBSURFACE SHALE TO ESTIMATE MICROBIAL BIOMASS AND DIVERSITY

Deep, organic-rich shale located in formations across North America and abroad is currently being targeted for oil and gas development. The opportunity to characterize microbial life in this unique ecosystem is fundamentally important to our understanding of existing microbial metabolisms as well as in situ biogeochemical processes influencing energy production after shale development. Lipid-based investigations can provide insight about the state of a microbial community such as the amount of viable biomass, community structure, and possible environmental stressors. However, analysis is currently hindered by access to pristine samples, low microbial biomass, small sample size, and sample chemistry. Therefore, extraction methods should incorporate steps that maximize the efficiency of recovering representative biomarkers from shale. This research examines how various extraction methods influence phospholipid (PL) yield, biomarker diversity, and PL profiles in ground, non-pristine Marcellus Shale samples homogenized from cores that varied in carbon content and diagenesis history. We tested eight extraction protocols derived from the Modified Bligh and Dyer (mBD), Folch (FOL), and Microwave-Assisted Extraction methods (MAE), with treatments varying buffers (Phosphate [Phos] versus Citrate [Cit]) and adding spikes shown to aid in lipid recovery (e.g. Magnesium Chloride, E. coli biomass, and an intact phospholipid spike, [POPC]). Total extracted elemental P differed between the protocols and was used to guide the selection of triplicates from three treatments (mBD-Cit; mBD-Phos + POPC; and MAE) for detailed fatty acid methyl ester (FAME) profile analysis. Although all three treatments provided similar phospholipid yields, the mBD-Phos + POPC treatment produced less experimental variability and more consistent FAME profiles than the mBD-Cit and MAE treatments. Furthermore, significant differences in FAME profiles were observed, especially for low abundance FAMEs that may be important for differentiating communities from shale that differ in diagenesis history. The data garnered in this study will be used to select an appropriate extraction method for characterizing lipid profiles in pristine core from scientific Marcellus shale wells.