"Baiting" with Stable Isotopic Compounds

While the principles of Bio-Trap samplers provide a foundation for sound sample collection, the ability to amend or “bait” the Bio-Trap® samplers with specific compounds provides an excellent tool for answering questions about site design strategies, such as:

• Will HRC®, Molasses Acetate, etc effectively stimulate known dechlorinating bacteria?
• What is the relative rate of MTBE degradation?
• Will sulfate increase degradation rate of Benzene?

Absorbing known concentrations of contaminants and/or surrogates onto Bio-Trap® samplers allow for the ability to observe and quantify in-situ biodegradation.  The ability to use surrogates, particularly stable isotopes, has significantly increased our understanding of contaminant fate.  In nature, two stable forms of carbon exist of which carbon-12 or 12C  is significantly more abundant than carbon-13 (12C; 99% vs. 13C; 1%).  Chemical supply companies have the ability to produce 13C enriched contaminants (99% 13C), e.g. specialized forms of contaminants which have the same chemical structure but are 'labeled' by having an unusual isotope in their chemical composition (13c as opposed to 12c).  Labeled surrogates such as 13C enriched Benzene, are used to differentiate the stable isotope tracer compound from the contaminants already present in the environment. The ability to use stable isotope compounds within the “baited” Bio-Trap® samplers allows us to confirm biodegradation by showing incorporation of 13C within the microbial biomass and also to provide a relative rate of degradation (% loss of 13C).

Laboratory studies have shown that any leaching which may occur from the baited contaminants or surrogates are absorbed onto the beads. A study involving benzene (detailed below) demonstrates that contaminants and/or their surrogates should not easily leach from the bio-sep beads. These results have been used to gain regulatory approval for use of Bio-Trap® samplers in a variety of states across the US including California.  

The table below details analytical results from a study of Bio-Trap® samplers baited with benzene. Samples from Bio-Sep beads and sterilized water and containing 10mM sodium azide (to prevent microbial growth) were analyzed in triplicate over a simulated thirty day incubation period. 

Results obtained:

Following incubation, the Bio-Trap® samplers are analyzed to determine the fate of the 13C enriched compound in two key ways.

• Determination of a relative rate of degradation and % loss of the stable isotopic compound.

• Assimilation of 13C into the microbial biomass, which proves biodegradation of the compound of interest (through PLFA analysis discussed below)

Phospholipid fatty acids (PLFA) are essentially the “skin” of the microbe and provide an effective tool for monitoring microbial responses to their environment. PLFA analysis provides a broad understanding of the entire microbial community with information obtained in the following key areas:  viable biomass, community structure and metabolic activity.  Biodegradation of stable isotope compounds can be confirmed by measuring the isotopic abundance within specific fatty acids as this enrichment can only occur if the microbes have consumed the contaminant, incorporating the 13C into their membrane structure. | Click for more details on PLFA analysis