In recent years there has been a great deal of scientific interest in processes that affect the fate of organic chemicals in the environment. one main reason for this increased interest is due to greater environmental concern over accidental or purposeful release of these chemicals into the environment by man. A major environmental concern is the increased use of pesticides over the last few years. In the thirty years prior to 1978 the use of pesticides has increased by a factor of forty (Ridgeway et al., 1978). Recently the use of herbicides has been increasing, but that of insecticides has stabilized (Willis, 1983). Detectable amounts of organic pesticides can be found in many areas of the biosphere. For toxic organic chemicals to be used safely, researchers must have a clear understanding of the fate and persistence of these chemicals when they are released into the environment. This understanding will also allow the development of new products that, when properly used, will not produce adverse effects to man or the environment (Weber, 1972). According to the Toxic Substance Control Act (TSCA) any new or expanded-use chemical that might be released into the environment must be tested for environmental hazard.

Thesis--University of California, Davis, 1950. / Typescript (carbon copy). Also available via the World Wide Web. (Restricted to UC campuses).

Thesis (Ph. D.)--Illinois State University, 1972. / Title from title page screen, viewed Sept. 23, 2004. Dissertation Committee: Mathew J. Nadakavukaren (chair), Robert M. Chasson, Dorothy E. Fensholt, Anthony E. Liberta, E. Ione Rhymer. Includes bibliographical references (leaves 157-170) and abstract. Also available in print.

This document only includes an excerpt of the corresponding thesis or dissertation. To request a digital scan of the full text, please contact the Ruth Lilly Medical Library's Interlibrary Loan Department (rlmlill@iu.edu).

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Graduation date: 1997

Thesis (Ph. D.)--West Virginia University, 2003. / Title from document title page. Document formatted into pages; contains xii, 243 p. : ill. Vita. Includes abstract. Includes bibliographical references (p. 213-240).

The cytotoxic effects of exposure to low concentrations of the herbicide, 2,4-dichlorophenoxyacetic acid (2,4-D) that are typically found in groundwater were investigated, in vitro. Most 2,4-D toxicology studies use high concentrations of the herbicide that are above those typically found in groundwater and measure overt biological endpoints. In contrast, this thesis examines the effects of low concentrations of 2,4-D and measures more subtle and sensitive endpoints such as gene expression and the generation of reactive oxygen species. This work derives from recent cDNA microarray analysis conducted in our laboratory that revealed significant alterations in the expression of 238 genes in cells exposed to nanomolar (nM) concentrations of a commercial formulation of 2,4-D. These findings are extended in this thesis to include the in vitro cytotoxic effects of low concentrations of both technical and commercial 2,4-D on two cell lines. Cells derived from liver (HepG2) and kidney (HEK293) respectively, were chosen, since liver and kidney are known to metabolize 2,4-D in vivo. Cell viability was measured using the Resazurin assay, reactive oxygen species (ROS) were measured with 2,7-dichlorofluorescin diacetate (2,7-DCFH-DA), and real timepolymerase chain reaction (RT-PCR) was used to assess changes in mRNA expression while protein expression was examined by Western blot.<p>Cell viability studies revealed that low environmental concentrations (0.1 to 100 nM) of 2,4-D induced small, but statistically significant decreases in cell viability. No concentration or time-dependent decreases in cell viability were observed in cells exposed to either forms of low environmental 2,4-D concentrations. HEK293 cells were more susceptible than HepG2 cells to the toxic effects of both forms of 2,4-D, having statistically significant lower viability at all exposure concentrations and durations. Both forms of 2,4-D reduced cell viability in both cell lines, suggesting that cytotoxicity was induced directly by 2,4-D, and not by the inert ingredients in the commercial formulation.<p>The ROS assays illustrated that 2,4-D induced statistically significant ROS production in HepG2 and HEK293 cell cultures at concentrations greater than 10 µM and 100 nM respectively. This was both a concentration and time-dependent effect in both cell lines. Although HEK293 cells were more susceptible to 2,4-D, they had 50 to 70% less ROS production than HepG2 cells, at all exposure concentrations and times.<p>The RT-PCR and Western blot analyses showed that exposure of HepG2 and HEK293 cells to low 2,4-D concentrations induced (< 2 fold) alterations in mRNA and protein levels of FTL, FTH1 and PCNA however these changes did not consistently vary with concentration.<p>Taken together, cell viability, ROS and gene expression studies show that low environmental 2,4-D concentrations induced subtle in vitro cytotoxic effects. However we have no evidence that these subtle changes pose a serious health threat to exposed humans.

The cytotoxic effects of exposure to low concentrations of the herbicide, 2,4-dichlorophenoxyacetic acid (2,4-D) that are typically found in groundwater were investigated, in vitro. Most 2,4-D toxicology studies use high concentrations of the herbicide that are above those typically found in groundwater and measure overt biological endpoints. In contrast, this thesis examines the effects of low concentrations of 2,4-D and measures more subtle and sensitive endpoints such as gene expression and the generation of reactive oxygen species. This work derives from recent cDNA microarray analysis conducted in our laboratory that revealed significant alterations in the expression of 238 genes in cells exposed to nanomolar (nM) concentrations of a commercial formulation of 2,4-D. These findings are extended in this thesis to include the in vitro cytotoxic effects of low concentrations of both technical and commercial 2,4-D on two cell lines. Cells derived from liver (HepG2) and kidney (HEK293) respectively, were chosen, since liver and kidney are known to metabolize 2,4-D in vivo. Cell viability was measured using the Resazurin assay, reactive oxygen species (ROS) were measured with 2,7-dichlorofluorescin diacetate (2,7-DCFH-DA), and real timepolymerase chain reaction (RT-PCR) was used to assess changes in mRNA expression while protein expression was examined by Western blot.<p>Cell viability studies revealed that low environmental concentrations (0.1 to 100 nM) of 2,4-D induced small, but statistically significant decreases in cell viability. No concentration or time-dependent decreases in cell viability were observed in cells exposed to either forms of low environmental 2,4-D concentrations. HEK293 cells were more susceptible than HepG2 cells to the toxic effects of both forms of 2,4-D, having statistically significant lower viability at all exposure concentrations and durations. Both forms of 2,4-D reduced cell viability in both cell lines, suggesting that cytotoxicity was induced directly by 2,4-D, and not by the inert ingredients in the commercial formulation.<p>The ROS assays illustrated that 2,4-D induced statistically significant ROS production in HepG2 and HEK293 cell cultures at concentrations greater than 10 µM and 100 nM respectively. This was both a concentration and time-dependent effect in both cell lines. Although HEK293 cells were more susceptible to 2,4-D, they had 50 to 70% less ROS production than HepG2 cells, at all exposure concentrations and times.<p>The RT-PCR and Western blot analyses showed that exposure of HepG2 and HEK293 cells to low 2,4-D concentrations induced (< 2 fold) alterations in mRNA and protein levels of FTL, FTH1 and PCNA however these changes did not consistently vary with concentration.<p>Taken together, cell viability, ROS and gene expression studies show that low environmental 2,4-D concentrations induced subtle in vitro cytotoxic effects. However we have no evidence that these subtle changes pose a serious health threat to exposed humans.

Microcystis aeruginosa is known widely for the obnoxious nature of its bloom. Problems resulting from blooms of the alga include: death of fish and other aquatic life, clogging of filter systems in water treatment plants, taste and odor problems, and death of cattle and water fowl via extracellular polypeptides. Blooms of this nuisance alga have historically occurred in waters receiving pollution from sewage effluents or runoff from agricultural lands. Studies have shown that 2,4-0ichlorophenoxyacetic acid (2,4-0) and 2,4,5-Trichlorophenoxyacetic acid (2,4,S-T) mimic the metabolic activities of the plant hormone indoleacetic Acid (IAA). IAA has been shown to stimulate the growth of some green and blue-green algae. Recent evidence suggests that blooms of Microcystis aeruginosa were triggered when the herbicides 2,4-D and 2,4,5-T were introduced to lakes and rivers. The results of this study showed that batch cultures of non-axenic Microcystis aeruginosa containing 2,4-D and 2,4,5-T exhibited significantly more growth than controls over a wide range of herbicide concentrations. Growth in 2,4-D treated cells was statistically significant from the control at a = 0.05 at concentrations as low as 10 -7 M (0.020 mg/l) and 10 -5M (2.5 mg/l) for 2,4,5-T. Also, a plot of maximum cell yield values versus herbicide concentrations resulted in a linear relationship. / Master of Science