vi, 208 leaves : / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Thesis (M.D.)--University of Adelaide, Dept. of Medicine, 1971

The work described in this Thesis was conducted to investigate aspects of copper metabolism in the plaice Pleuronectes platessa (L). Three major aspects were studied. The first was to investigate and partially characterize the distribution of copper and copper-dependent enzymes in the various tissues of the plaice. The second was to study the effect of cadmium on copper-dependent enzymes as a base line for understanding the competition between an essential metal ion and a non-essential toxic metal ion. This has established a foundation for the interpretation of the toxicity of pollutants to a marine organism. The third and the major part of the investigation was to purify and characterize the copper-dependent enzyme, ceruloplasmin, and to compare it with the isolated enzyme from the mammalian system. The results from the first part of the investigation showed that (i) Copper is relatively evenly distributed in the plaice. Apart from brain which has the highest concentration of copper, the concentrations of copper in the other tissues generally reflect the copper-dependent enzyme activities. (ii) The presence of the copper-dependent enzymes in the plaice are confirmed. The assay system used for the mammalian enzyme systems apply reasonably well to the plaice enzymes. (iii) The limited study on the relatively crude preparations of copper-dependent enzymes show that they are similarly distributed in mammals and that they have similar properties. Cytochrome oxidase, however, has a higher pH optimum than that of the mammalian enzymes. Exposure of the plaice to sublethal concentrations of cadmium as cadmium chloride results in initial increase in ceruloplasmin, cytochrome oxidase, monomaine oxidase and tryptophan oxygenase activities followed by a decrease after a prolonged exposure. Changes in the copper-dependent enzyme activities are accompanied by parallel changes in the concentrations of copper in the serum and liver. Thus cadmium alters copper metabolism in the plaice.

Plicatic acid and 5-isopropyltropolone, which are found in the heartwood of western red cedar (Thuja plicata, Donn), are known chelators of copper. A study has been made of their effects on some aspects of copper metabolism in rats and mice. As a basis of comparison similar experiments were carried out using penicillamine (PEN), a chelating agent with wide clinical use. In the experiments the sodium salt of 5-isopropyltropolone (T-Na) and the potassium salt of plicatic acid (P-K) were used. When administered daily, P-K and PEN caused an increase in urinary and fecal excretion of copper in rats on a normal diet and an increase in the level of copper in the liver and kidney and a decreased level of copper in the heart and brain. The increased urinary excretion is much more marked with PEN than with P-K. T-Na administered daily had no effect on the excretion of copper in rats on a normal diet and causes a rise in copper content of liver and kidney and a lowering of copper content of heart and brain. It appears that the copper level elevation in kidney and liver caused by P-K and PEN is due to an increased utilization of the routes of excretion while the increased level due to T-Na is probably due to deposition in the cells. When these compounds are administered daily to copper-fed rats similar observations were made with the difference that in organs where copper levels are increased the increases are greater and in organs where copper levels are decreased the decreases are smaller. With T-Na there is one marked difference in that the copper level in the brain is increased. It is felt that this increase is due to a greater penetrating ability of the T-Na-copper chelate because of its lipid solubility. P-K and PEN have very little effect on the toxicity of a single large dose of copper. Earlier experiments with T-Na showed that when T-Na is given shortly after a single large dose of copper the toxicity is greatly increased. The explanation of this is very likely that T-Na increases the penetration of the copper into the CNS and it is here that the toxic effect is exerted. It is felt that this is due to a solubility factor, T-Na and its chelate being lipld-soluble and P-K, PEN and their chelates being water-soluble. When T-Na, P-K and PEN are given to rats in small daily doses over a long period, the atria from such rats exhibited a reduced chronotropic response to tyramine. If at the same time as the rats are being given the chelating agent, they are given excess copper in drinking water the tyramine response remains close to normal. In this regard T-Na possesses the most activity. These observations support the theory that chelating agents inhibit dopamine-β-hydroxylase by rendering copper inaccessible to the enzyme and that subsequent to this inhibition the biosynthesis of the catecholamines is inhibited at the dopamine stage preventing the formation of norepinephrine and epinephrine. As the levels of endogenous norepinephrine and epinephrine fall, the result is a reduced tyramine response. If T-Na, P-K, PEN and D-Na are given in a large single dose the inhibition of dopamine-β-hydroxylase is not evident while it appears that COMT may be being inhibited. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

Call number: LD2668 .T4 1964 A55 / Master of Science

Pregnant and weanling rats were fed liquid diets with or without 30 percent of total kcal from ethanol and varying levels of copper in order to determine if ethanol ingestion would exaggerate a marginal copper status to an obvious copper deficiency. Pregnant albino rats were fed either 0.75 or 3.75 mg Cu/L throughout gestation and the first 15 days of lactation while female weanling rats received 0.5 or 2.5 mg Cu/L for 5 weeks. Ethanol consumption exaggerated a marginal copper status during reproduction as evidenced by significant reductions in maternal liver copper concentration and enzymatic activity of the copper metalloprotein Copper-Zinc superoxide dismutase in offspring liver. Ethanol had little or no effect upon copper status in weanling rats. In addition, serum copper failed to reflect a developing depletion of liver copper when ethanol was being consumed. Since it is known that the average American diet is just adequate in copper content, and that copper balance is difficult to achieve during times of increased metabolic demand, pregnant subjects may be at a great risk to develop a copper deficiency when ethanol is being consumed. This ethanol and copper interaction, however, will likely go undetected if only serum copper is used as an indicator of copper status. / Graduation date: 1989

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Copper is one of the most important trace elements in ruminant nutrition. Its deficiency causes certain pathologies that can be cured by supplementation, by either five ("inorganic") or complexed ("chelated") forms. With the objective of being able to quantitatively compare the distribution of copper between the two forms of supplements, the speciation of copper in ruminant fluids was studied here. For this study, copper lysine supplement was used. It was necessary to first study the acid-base and complexation chemistry of lysine with copper (II). In addition, the complexation chemistry of glycine and histidine, were investigated. Mass balance equations were used to describe the distribution of copper (II) amongst different ligands. The results of the distribution of copper (II) ions in McDougall's solution (a simulated form of bovine saliva), indicate no significant differences in the distribution of copper using the different form of supplements. 1H NMR was used to validate the results from the computer model. Using a combination of the results from the saliva simulation model and the chemical shifts from the NMR studies, the chemical shift changes that would accompany the addition of copper (II) to McDougall's solution were predicted. Results from the models do not show any appreciable differences from experimental values. Rumen samples were collected. Important peaks in the 1H NMR spectrum were assigned. The spectrum indicated that acetic acid, resulting from the fermentation in the rumen, was a good probe for monitoring the speciation pattern. Speciation calculation indicated that the bulk of the copper would be bound to ammonia in the rumen. Changes in chemical shifts that result from the introduction of copper (II) to the rumen contents were predicted. Results were compared with experimental values. Agreement between the two sets of results was found to be satisfactory. The study shows that any advantages that result from the use of copper lysine supplement are not as a result of its remaining intact. Though metal bioavailabilities are hard to predict this approach could help better our understanding of this process. The methods developed here could be extended to other metal complexation problems in biological fluids. (Abstract shortened by UMI.)