¹³C-enriched PCE Study to Evaluate the CL-Out  Aerobic Cometabolic Pathway

CL Solutions partnered with Pelorus EnBiotech Corporation of Evergreen, Colorado and Microbial Insights Laboratory of Rockford, Tennessee to evaluate the cometabolic pathway that CL-Out utilizes to completely degrade PCE.  Field applications have demonstrated the effectiveness of CL-Out®, but laboratory investigations have lagged behind our field progress.  Other researchers have demonstrated an aerobic pathway for PCE and TCE utilized by pseudomonas sutuzeri.  This investigation showed that CL-Out organisms were able to completely degrade PCE using dextrose as a substrate.

Four microcosms were prepared with a phosphate-buffered medium, which also contained NaNO₃ and dextrose.  The media and reactor assemblies were autoclaved to ensure a sterile environment prior to being spiked with ¹³C-enriched PCE and CL-Out®.  Two reactors were attached to respirometers.  Samples were taken at initiation (t=0), 12, 24, 36, 48, 72 and 100 hours.  The samples were analyzed for PCE in liquid and vapor phase and the distribution of ¹³C-enriched PCE in dissolved inorganic carbon and PLFA.  In addition, DNA analysis was performed to determine whether the Pseudomonas in CL-Out was sutuzeri OX1, which has been shown to metabolize PCE.

During the study the PCE concentration decreased in both the dissolved and vapor phase in all of the microcosms, with the largest decreases in the microcosms attached to the respirometers.  The aqueous-phase PCE concentration decreased by 69 to 92% in the microcosms attached to the respirometers and 32 to 77% in the microcosms not attached to respirometers.  Interestingly, the vapor-phase PCE concentration also decreased, supporting the conclusion that the liquid-phase was destroyed and not volatilized.  The vapor phase PCE concentration decreased by 26 to 91% in the microcosms.

The isotope-labeled PCE was undertaken to determine where the carbon was distributed after the PCE was destroyed.  The ¹³C was found in both cell material and mineralized in CO₂.  A small proportion of the ¹³C was detected in two fatty acids of the PLFA biomass.  The dissolved inorganic carbon was enriched by ¹³C by a small amount.  While the enrichment by ¹³C in the fatty acid and dissolved inorganic carbon were low, the enrichment was in proportion to the amount of ¹³C available to total carbon added as dextrose.

The PLFA tests confirmed the organisms in the study were Proteobacteria, which includes Pseudomonas sp. The DNA analysis confirmed the presence of naphthalene dioxygenase, but did not confirm the presence of pseudomonas sutuzeri OX1.  ¹³C was also detected in the dissolved inorganic carbon.  These results confirm CL-Out follows a novel degradation pathway for the aerobic degradation of PCE.