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While they are theoretically conceptualized to restrict biodegradation of organic contaminants, bioavailability limitations are challenging to observe directly. Here we explore the onset of mass transfer limitations during slow biodegradation of the polycyclic aromatic hydrocarbon 2-methylnaphthalene (2-MN) by the anaerobic, sulfate-reducing strain NaphS2. Carbon and hydrogen compound specific isotope fractionation was pronounced at high aqueous 2-MN concentrations (60 μM) (ε carbon = -2.1 ± 0.1‰/ε hydrogen = -40 ± 7‰) in the absence of an oil phase but became significantly smaller (ε carbon = -0.9 ± 0.3‰/ε hydrogen = -6 ± 3‰) or nondetectable when low aqueous concentrations (4 μM versus 0.5 μM) were in equilibrium with 80 or 10 mM 2-MN in hexadecane, respectively. This masking of isotope fractionation directly evidenced mass transfer limitations at (sub)micromolar substrate concentrations. Remarkably, oil-water mass transfer coefficients were 60-90 times greater in biotic experiments than in the absence of bacteria ( k  org - aq 2-MN = 0.01 ± 0.003 cm h -1 ). The ability of isotope fractionation to identify mass transfer limitations may help study how microorganisms adapt and navigate at the brink of bioavailability at low concentrations. For field surveys our results imply that, at trace concentrations, the absence of isotope fractionation does not necessarily indicate the absence of biodegradation.

Mass Transfer Limitation during Slow Anaerobic Biodegradation of 2-Methylnaphthalene