Field-scale 13C-labeling of phospholipid fatty acids (PLFA) and dissolved inorganic carbon: tracing acetate assimilation and mineralization in a petroleum hydrocarbon-contaminated aquifer

Abstract

This study was conducted to determine the feasibility of labeling phospholipid-derived fatty acids (PLFA) of an active microbial population with a (13)C-labeled organic substrate in the denitrifying zone of a petroleum hydrocarbon-contaminated aquifer during a single-well push-pull test. Anoxic test solution was prepared from 500 l of groundwater with addition of 0.5 mM Br(-) as a conservative tracer, 0.5 mM NO(3) (-), and 0.25 mM [2-(13)C]acetate. At 4, 23 and 46 h after injection, 1000 l of test solution/groundwater mixture were sequentially extracted. During injection and extraction phases we measured Br(-), NO(3) (-) and acetate concentrations, characterized the microbial community structure by PLFA and fluorescent in situ hybridization (FISH) analyses, and determined (13)C/(12)C ratios in dissolved inorganic carbon (DIC) and PLFA. Computed first-order rate coefficients were 0.63+/-0.08 day(-1) for NO(3) (-) and 0.70+/-0.05 day(-1) for acetate consumption. Significant (13)C incorporation in DIC and PLFA was detected as early as 4 h after injection. At 46 h we measured delta(13)C values of up to 5614 per thousand in certain PLFA (especially monounsaturated fatty acids), and up to 59.8 per thousand in extracted DIC. Profiles of enriched PLFA and FISH analysis suggested the presence of active denitrifiers. Our results demonstrate the applicability of (13)C labeling of PLFA and DIC in combination with FISH to link microbial structure and activities at the field scale during a push-pull test.