Date of Award


Document Type


Degree Name

Master of Science (MS)

Legacy Department

Environmental Engineering and Science

Committee Chair/Advisor

Freedman, David L.


The Twin Lakes chlorinated ethene plume at the Department of Energy's
Savannah River Site (SRS) emerges in a wetland in which trichloroethene (TCE) is
completely reduced to ethene and ethane. Novel strains of Dehalococcoides have been
detected from the wetland area. The objectives of this study were 1) to develop an
enrichment culture capable of completely dechlorinating TCE and tetrachloroethene
(PCE) to ethene using samples from the wetland; 2) to evaluate the use of lactate,
emulsified vegetable oil and corn syrup as possible electron donors for biostimulation of
the P-area chlorinated ethene plume at SRS that is not undergoing natural attenuation;
and 3) to evaluate the use of the enrichment culture for bioaugmentation of the P-area
chlorinated ethene plume.
The enrichment culture was started with samples from microcosms that were used
to confirm reductive dechlorination activity in the SRS Twin Lakes area. Samples were
transferred to an anaerobic mineral medium, repeatedly fed with TCE and PCE, and
transferred a second time to mineral medium. TCE and PCE concentrations of 35-40
mg/L and 4-8 mg/L, respectively, were completely consumed by the enrichment culture
within three to five weeks. Quantitative polymerase chain reaction (PCR) analysis
indicated a linear increase in Dehalococcoides as increasing amounts of PCE and TCE
are reduced to ethene. The Dehalococcoides cell density in the enrichment culture has
stabilized at approximately 4.5 x 108 cells per mL. Lactate was used as the electron
donor and carbon source. An electron donor balance indicated that most of the lactate is
fermented to acetate and propionate, with less than 3% used for reductive dechlorination.
Methanogenesis in the enrichment culture is insignificant.
A laboratory study was conducted to compare biostimulation and
bioaugmentation for removal of PCE and TCE from the SRS P-area plume. Microcosms
were prepared with sediment and groundwater from the site, at PCE and TCE
concentrations close to the reported maxima of 5 mg/L and 35 mg/L, respectively. The
pH of the groundwater was adjusted from 5.7 to 7 with NaOH and resazurin was added as
a redox indicator. Lactate, corn syrup and emulsified vegetable oil were used for
biostimulation. Bioaugmentation was assessed using varying doses of the SRS
enrichment culture (1.0, 0.1, 0.01, and 0.001% v/v), along with one treatment using a
commercial bioaugmentation culture (1.0% v/v). After more than eight months of
incubation, there was no evidence of PCE or TCE dechlorination in any of the
biostimulated treatments, even though the amount of donor added was in considerable
excess and redox and pH conditions were favorable. The microcosms were
bioaugmented after three months of incubation (to establish favorable redox conditions)
and the response was immediate. In the treatment that received the 1.0% dose of SRS
culture, reduction of the PCE and TCE to ethene was completed in approximately three
weeks. The lower doses of SRS culture were also effective, although up to seven months
of incubation was needed before dechlorination was complete. In contrast,
dechlorination of PCE and TCE was much slower and incomplete in the treatment that
received the commercial bioaugmentation culture, over the same incubation period. The
microcosm results indicate that the SRS enrichment culture holds promise for use in
bioaugmentation of the P-area groundwater plume. Efforts are underway to scale up the
enrichment culture in preparation for a pilot-scale field test.



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