Date of Award

Spring 2006

Project Type

Dissertation

Program or Major

Chemistry

Degree Name

Doctor of Philosophy

First Advisor

Sterling A Tomellini

Abstract

The initial phase of this project investigates the use of copper(II) complexes of tryptophan and its analogues for indirect fluorescence detection in high-performance liquid chromatography. First, indirect photometric and fluorometric detection in high performance liquid chromatography (HPLC) is reviewed, giving the functional definition of indirect detection employed for this project, distinguishing different approaches to indirect photometric and fluorometric detection in HPLC, and explaining the basis of indirect fluorescence detection using copper(II) complexes of tryptophan and its analogues as postcolumn interaction components. Subsequent studies were conducted to develop and apply indirect HPLC detection for detecting specific mono-amino sugars, glucosamine, galactosamine, and mannosamine. Two tryptophan analogues, 5-hydroxy-L-tryptophan and DL-5-methoxytryptophan were evaluated as potential alternatives to L-tryptophan for the detection of these mono-amino sugars. This indirect fluorescence detection method was also evaluated for the analysis of glucosamine in commercial dietary supplements following chromatographic separation.

The second part of this project focuses on separating copper(II)-azamacrocyclic complexes with the use of high-performance liquid chromatography. Interest in these complexes arises from the increasing use of copper radioisotopes in imaging and therapy leading to the synthesis of ligands which form stable copper complexes. The effects of the concentration of the buffer, the pH of the buffered mobile phase, and the concentration of the organic modifier, methanol, on the separations produced were investigated. Separation of these copper complexes by ion-pair HPLC with the use of a mass spectrometry-compatible ion-pair reagent is also presented. The reversed phase chromatographic conditions utilized also allow the pa's and the lipophilicity parameter of the complexes studied to be estimated. The separation and retention of several copper(II)-azamacrocyclic complexes on a graphitic carbon stationary phase, a very hydrophobic reversed-phase stationary phase with the capability of retaining highly polar compounds, was also investigated.

Finally the potential application of the indirect fluorescence detection method described here for detecting some of the ligands of the copper(II)-azamacrocyclic complexes, which do not absorb in the UV-vis range or fluoresce, following separation by liquid chromatography is discussed. Future studies are proposed for estimating the lipophilicity of metal complexes by reversed-phase high-performance liquid chromatography and capillary electrophoresis.

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