A system to quantitatively recover bacterioplankton respiratory CO2 for isotopic analysis to trace sources and ages of organic matter consumed in freshwaters

S. Leigh McCallister, François Guillemette,Paul A. del Giorgio

Limnol. Oceanogr. Methods 4:406-415 (2006) | DOI: 10.4319/lom.2006.4.406

ABSTRACT: We present a new system and method to measure the natural abundance isotopic (δ13C, Δ14C) values of respiratory CO2 produced by bacterioplankton, to directly assess the sources and ages of organic carbon (OC) respired by bacteria in aquatic ecosystems. The Respiratory Carbon Recovery System (ReCReS) and operating procedure was designed to reduce background dissolved inorganic carbon values by > 98% and then quantitatively recover the CO2 derived from bacterial respiration after incubation of freshwater samples. The two-component ReCReS consists of an airtight incubation system (20L) for short term regrowth incubations of filtered water samples inoculated with ambient bacteria, and a harvest system to recover the respiratory CO2 produced during these incubations. The multi-step operating procedure involves the following: 1) filling of incubation system with 0.2 µm-filtered sample water; 2) addition of 1 N HCl (pH to ~2.8), 3) sparge with ultra high purity (UHP) He to remove dissolved inorganic carbon, 4) sparge with UHP, volatile organic carbon-, -CO2-free air to replenish oxygen, 5) neutralization (1 N carbonate free NaOH), re-inoculation with the ambient bacterial assemblage and incubation (80-132 hours), 6) acidification to pH 2.8 and 7) UHP He sparge for > 12 hours. The evolved CO2 is sent through 2 water traps (dry ice slurries) prior to cryogenic trapping in liquid N2. Control incubations were processed concurrently to evaluate extraction efficiencies, potential methodological contamination, and fractionation artefacts. Collectively our results suggest that respiratory CO2 is quantitatively recovered and the isotopic fidelity (δ13C, Δ14C) between the OC respired and the CO2 harvested is retained. Moreover, the system allows the recovery of sufficient C to measure both δ13C and Δ14C values, even in the most oligotrophic systems.