Stable carbon isotopic composition of lipids in Euglena-dominated biofilms from an acid mine drainage site: Implications of carbon limitation, microbial physiology, and biosynthetic pathways

Abstract

This study addresses carbon isotope ratios (δ13C) of lipid biomarkers isolated from Euglena-dominant biofilms in an acid mine drainage (AMD) site in western Indiana, USA. Overall, the hydrocarbons, including n-alkenes, phytadienes, and wax esters, showed heavier δ13C values than what would be expected for microeukaryotic biomarkers recorded in past studies. The primary cause for the 13C-enrichment is the existence of a carbon-limiting system in the AMD, which in turn is regulated by the pH of the system. Floating biofilms showed more depleted δ13C values for phytadienes and n-alkenes (average of −23.6‰) as compared to benthic biofilm (average of −20.8‰). This pattern was also observed in the δ13C values of wax esters, suggesting that microbial physiology plays an important role in carbon isotopic fractionation. But carbon-limitation and biofilm physiology may not be the only causes of 13C-enrichment. Biosynthetic pathways involved in the production of microeukaryotic biomarkers can lead to heavy δ13C values as well. 13C-enriched values of wax esters resulted from the metabolic pathway Euglena used to generate the esters—the reverse β-oxidation pathway. Kinetic isotope effect at a branch point at acetyl CoA in the reverse β-oxidation pathway is the plausible cause for the heavier δ13C values of wax esters. A combination of specific microeukaryotic biomarkers in AMD, including short chain wax esters (C25–32) and n-alkenes, that shows unique carbon isotope ratios, may provide useful biosignatures for reconstructing similar extreme environments that may have existed on early Earth during Late Archean–Early Proterozoic, when eukaryotes first arose and evolved.