Absorption, Distribution, and Transformation of Radiolabeled Trinitrotoluene

1
Absorption, Distribution, and Transformation of
Radiolabeled Trinitrotoluene

• Amy Palmer
• Dr. A. Morrie Craig
• Department of Biomedical Sciences

2
Relevance

• Nitroaromatic compounds such as TNT contaminate
  soil and groundwater across the US.
• -- 700,000 cubic yards of
• soil
• -- 10 BILLION gallons of
• groundwater
• Most of the contamination occurs on more than
  16,000 Department of Defense facilities.

3
Background
4
Background

• The cost of decontamination as it stands now is
  35 billion.
• TNT toxicity has human health risks

5
Past Work

• Plants alone There have been many experiments on
  different types of plants, from the lower forms
  to higher plants, each showing that plants are
  able to handle only certain levels of TNT.
• Microbes There have been different types of
  microbe studies from bioslurries to rumen fluid
  microbes which have been performed by Dr. Craig
  previously.

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Strategy

• Plants have been shown to break down TNT
• into metabolites, but are only metabolized
• down to the monoamines are which are still
• toxic. Ruminal animals, such as sheep, are
• able to ingest the plants that take in the TNT
• and its metabolites and break down TNT into
• the non-toxic metabolites with the help of
• rumen microbes.

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Purpose

• Determine if cool season grasses take in TNT from
  the contaminated soil and which of the three
  grasses does it the best
• Determine if the plant had broken down the TNT to
  other metabolites and what those metabolites are.

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Hypothesis

• Cool season grasses such as tall fescue,
  perennial ryegrass, and orchard grass, will take
  in TNT contaminated soil and break it down into
  other metabolites.

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Three Candidates
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Methods

• Determine the absorption, distribution, and
  transformation of 14C-TNT from soil in three
  species of grasses
• How
• -- Verify uptake of 14C-TNT through
  autoradiography and HPLC
• -- Quantify uptake and bioremediation of TNT by
  the grasses with HPLC

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Methods

• Soil preparation
• -- Each replicate had the same soil nutrients
  that are measured carefully.
• -- The 14C- TNT mixture with cold TNT that is
  dissolved in acetone was then S added.
• Planting of the three types of seed (Tall Fescue,
  Perennial Ryegrass, Orchardgrass)
• -- 24 pots of soil
• -- Each pot yields between 20 to 50 individual
  seedlings

Mg
P
N
K
Zn
B
Cu
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Methods

• Harvesting of plant materials
• -- collect clippings every 60 days or when the
• seedlings are 6 inches tall

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Methods
Clippings
Autoradiography
Each clipping was divided into five subgroups
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Methods

• At each harvest and at the beginning of the
  experiment a plug of soil had been taken from two
  pots of each species designated for destructive
  soil sampling. The extraction had followed a
  modified EPA method 8330.
• -- Concentrations of TNT were monitored
• -- TNT had been determined by the radioactivity
  of the carbon

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Results to date

• During the method development for soil
  extraction, TNT was extractable in minute
  amounts.
• These amounts of TNT show an accurate measurement
  of ppb that would be found in most contaminated
  areas.
• The plants have now been planted and are starting
  to sprout seedlings.

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Future Work

• The plant experiment will continue through 4-5
  harvests and then the results will be compiled.
• The next step is to combine the plant portion of
  the experiment with the animal portion on actual
  sites.

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Acknowledgements

• Howard Hughes Medical Institute
• Dr. A. Morrie Craig
• Dr. Jennifer Duringer
• USDA
• Dr. Craigs Lab
• Dr. Kevin Ahern