Graduate Thesis Or Dissertation
 

Transformation of Phenanthrene by Mycobacterium sp. ELW1 and the Formation of Toxic Metabolites

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  • The ability of Mycobacterium sp. ELW1, a novel microbe capable of alkene oxidation, to co-metabolize phenanthrene (PHE) was studied. ELW1 was able to completely co-metabolize PHE, at different concentrations below its water solubility limit, in an aqueous environment. The alkene monooxygenases in ELW1, used to initiate oxidation of PHE, were effectively inhibited by 1-octyne despite some PHE transformation observed. PHE metabolites consisted of only hydroxyphenanthrenes (OHPHEs) with trans-9,10-dihydroxy-9,10-dihydrophenanthrene (trans-9,10-PHE), the primary product, comprising more than 90% of the total metabolites formed in both PHE-exposed cells and 1-octyne controls. Mass balance was estimated by summing the zero-order formation rates of OHPHE metabolites and comparing these to the zero-order transformation rates PHE in PHE-exposed cells. The transformation rates of PHE and were in good agreement with the formation rates of the metabolites. PHE transformation followed first-order rates that, when normalized by biomass, were in the range of those estimated by the ratio of the Michaelis-Menten kinetic variables of maximum transformation rate (k[subscript max]) to the half-saturation constant (K[subscript s]). Estimated values for k[subscript max] to K[subscript s] obtained through both non-linear and linearization methods resulted in k[subscript max]/K[subscript s] estimates that were a factor of ~3 lower compared to experimental values. Both experimental and estimated values of k[subscript max], K[subscript s], and k[subscript max]/K[subscript s] were 2-3 magnitudes lower than literature values determined for microbes other than Mycobacterium sp. using different models that incorporated additional parameters. OHPHE standards, including 1-hydroxyphenanthrene (1-PHE), 3-hydroxyphenanthrene (3-PHE), 4-hydroxyphenanthrene (4-OHE), 9-hydroxyphenanthrene (9-PHE), and 1,9-dihydroxyphenanthrene (1,9-PHE), were developmentally toxic to embryonic zebrafish. However, PHE and trans-9,10- PHE. were not toxic. OHPHE metabolite mixtures formed by ELW1 were also tested for toxicity using embryonic zebrafish. The embryonic zebrafish were exposed to OHPHE metabolite mixtures that were at least 1.5 times less than the concentration need to elicit a toxic response. However, toxicity was observed in the two latest time points, 76 and 122 hr, in PHE-exposed cells. The toxicity may have been caused by an unidentified toxic metabolite.
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