This thesis describes a developmental process required for the construction and optimisation of two-dimensional liquid chromatographic systems. The text encompasses much of the difficulties and provides a frame work for achieving positive results. The hardware requirements and the concepts for successful completion of heart-cutting and comprehensive modes of separation are addressed. Low molecular weight polystyrenes play an essential role in providing a model framework or theme for the development of two-dimensional liquid chromatographic systems. Low-molecular weight oligostyrenes were initially used to provide a theoretical assessment of combinations of liquid chromatographic systems. Interpretation of theoretical results then allowed for the development of a heart-cutting two-dimensional liquid chromatographic system for the separation of oligostyrene complex samples. This allowed for practice to be compared with the theoretical results, which are presented and provided direction as to what attributes of a two-dimensional liquid chromatographic system are beneficial. Petroleum oils are one of the most complex mixtures found in the natural environment and are included to provide a real-world application that demonstrates the potential power of comprehensive two-dimensional liquid chromatographic systems. Orthogonality is a central theme throughout the whole thesis. Separation correlation is shown to be advantageous in certain instances in the evaluation of hybrid stationary phases, as shown in the analysis of a low molecular weight polystyrene sample. The thesis finishes by the incorporation of monolithic separation media as an alternative second dimension stationary phase support. A method is discussed that allows for a substantial increase in the volume and concentration loading of two-dimensional liquid chromatographic systems that employ monolithic chromatography columns in the second dimension. This is important since volume overloading and concentration overloading in the second dimension can reduce separation performance and hence the final chapter provides a new direction for research. / Doctor of Philosophy (PhD)

Thesis (Ph.D.)--University of Delaware, 2006. / Principal faculty advisor: Abraham M. Lenhoff, Dept. of Chemical Engineering. Includes bibliographical references.

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The principle of the combined reversed-phase chromatographic electrochemical detection assay of the major serum thyroid hormones is demonstrated. The separation of 3,3',5-triiodothyronine (T(,3)), 3,3',5'-triiodothyronine (rT(,3)) and 3,5,3',5'-tetraiodothyronine (T(,4)) has been studied as a function of solvent strength, pH, temperature and ionic strength. Detection was carried out using a thin-layer electrochemical detection cell set at 1.500 V containing a new material, low temperature isotropic carbon, as a working electrode. This material was evaluated against carbon paste and using it, subnanogram detection limits were noted for the above analytes under defined conditions. Investigation of the feasibility of this analysis applied to 1.0 mL serum samples was carried out using a combined solvent extraction-cation exchange resin clean-up approach. The data indicate that under the defined conditions, elevated total T(,3) levels can be discerned and total T(,4) levels quantitated demonstrating the potential utility of this approach in screening thyrometabolic status.

Ion exchange chromatography methods which have been developed in recent years appear to offer a sensitive technique that can be utilized very advantageously in studies on various proteins and their properties. The application of such a procedure for the successful fractionation of bovine sarcoplasmic proteins should stimulate interest and research in characterizing the changes occurring in beef muscle during the post-mortem aging period. The research described herein pertains to the development and application of a DEAE-cellulose ion exchange chromatography procedure for the fractionation of bovine sarcoplasmic proteins. Results of preliminary experiments indicated that columns packed under pressure possessed superior fractionational qualities than did columns packed without pressure. Also in the preliminary experiments, a Tris buffer system (a starting buffer of 0.04 M Tris phosphate, pH 9.0, and a limiting buffer of 0.5 M Tris H₂PO₄, pH 3.6) and a concave gradient elution procedure were developed which were found to separate the sarcoplasmic proteins satisfactorily. At least 16 components were recognized to be fractionated in the chromatography of beef sarcoplasmic proteins extracted two hours post-mortem. Duplication of the chromatographic results was found to be quite good. Some changes that had occurred in the sarcoplasmic proteins during a post-mortem aging period of 10 days were detected by the chromatographic technique. The changes observed were the appearance of new components, and disappearance of some fractions while others diminished. Data obtained from experiments on the pre-chromatographic treatment of the samples showed that any deviation in procedure always resulted in chromatographic differences. Hence, strict uniformity must be maintained throughout the chromatographic procedure in order to obtain reproducible results. Although the experimental evidence obtained in this study indicates that further research must be completed on the chromatographic procedure, the method does offer a sensitive technique for gaining new information on the properties of the sarcoplasmic proteins. / Graduation date: 1965

Thesis (M.S.)--Rochester Institute of Technology, 1990. / Spine title: Chromatographic retention and donor and acceptor numbers. Typescript. Includes bibliographical references.