Anaerobic Biodegradation of n -Hexadecane by a Nitrate-Reducing Consortium

Abstract

Nitrate-reducing enrichments, amended with n -hexadecane, were established with petroleum-contaminated sediment from Onondaga Lake. Cultures were serially diluted to yield a sediment-free consortium. Clone libraries and denaturing gradient gel electrophoresis analysis of 16S rRNA gene community PCR products indicated the presence of uncultured alpha- and betaproteobacteria similar to those detected in contaminated, denitrifying environments. Cultures were incubated with H ₃₄ -hexadecane, fully deuterated hexadecane ( d ₃₄ -hexadecane), or H ₃₄ -hexadecane and NaH ¹³ CO ₃ . Gas chromatography-mass spectrometry analysis of silylated metabolites resulted in the identification of [H ₂₉ ]pentadecanoic acid, [H ₂₅ ]tridecanoic acid, [1- ¹³ C]pentadecanoic acid, [3- ¹³ C]heptadecanoic acid, [3- ¹³ C]10-methylheptadecanoic acid, and d ₂₇ -pentadecanoic, d ₂₅ -, and d _{2 4} -tridecanoic acids. The identification of these metabolites suggests a carbon addition at the C-3 position of hexadecane, with subsequent β-oxidation and transformation reactions (chain elongation and C-10 methylation) that predominantly produce fatty acids with odd numbers of carbons. Mineralization of [1- ¹⁴ C]hexadecane was demonstrated based on the recovery of ¹⁴ CO ₂ in active cultures.