Previous studies have shown that rainbow trout exposed to dieldrin via diet for 9 to 12
weeks increased biliary excretion of a subsequent dose of [¹⁴C]dieldrin by 500%. This
was not explained by induction of the cytochrome P-450 (CYP) system involved in
oxidative metabolism of these compounds. We hypothesized that epoxide hydrolase
activity increased in dieldrin fed-fish. Epoxide hydrolase is an enzyme that catalyzes the
hydrolysis of epoxide compounds to their corresponding diols. For instance, dieldrin is
metabolized to 6,7 trans-aldrindihydrodiol. This study investigated the activity of
epoxide hydrolase in microsomes and cytosol of rainbow trout fed a diet that contained 0
or 15 ppm dieldrin. Fish were fed control or dieldrin diet (0.324 ug/g body weight/day)
for 3, 6, or 9 weeks. There was a small increase in mortality and decrease in body
weight among dieldrin-fed fish after 9 weeks. After week 9, dieldrin-fed fish were fed a
control diet for an additional 3 weeks because of these signs of toxicity. At week 12, the
difference of body weight between control and treated was not significant. Microsomal
and cytosolic epoxide hydrolase activities were measured with a radiometric assay which
determined differential partitioning of the parent compound (epoxide) in dodecane and
the metabolite (diol) in the aqueous phase. Assays were run at optimal pH and
temperature using [³H]trans-stilbene oxide (pH 7) as substrate for cytosol and [³H]cis-stilbene
oxide (pH 8) as substrate for microsomes. In order to prevent competition for
reaction with stilbene oxide, depletion of glutathione was efficiently achieved by dialysis
at 4°C for 2 hours at room temperature in buffer [pH 7.5, potassium phosphate 10 mM,
KCL 0.15 M, EDTA 1 mM, BHT 0.1 mM, 0.1 mM PMSF]. Protein quantification was
determined by using BCA assay and concentrations were always between 5 and 25
ug/ml in the final assay volume. Epoxide hydrolase activities were not significantly
different in cytosol or microsomes from control and dieldrin-fed fish. Dieldrin residues
in liver were analyzed by gas chromatography with electron capture detection
(GC/ECD). The concentration in the liver increased with time of exposure and declined
markedly in fish fed dieldrin for 9 weeks and then fed control diet. No dieldrin was
detected in livers from control fish. / Graduation date: 2003
| Identifer | oai:union.ndltd.org:ORGSU/oai:ir.library.oregonstate.edu:1957/30809 |
| Date | 30 April 2002 |
| Creators | Rosemond, Marie Victoire M. |
| Contributors | Curtis, Lawrence R. |
| Source Sets | Oregon State University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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