Year

2005

Degree Name

Master of Science - Research

Department

Department of Chemistry - Faculty of Science

Abstract

Electrospray ionization mass spectrometry (ESI-MS) was assessed as a tool for the analysis of different glycosylated and iron forms of transferrin (tf). Carbohydrate deficient transferrin (CDT) is clinically relevant in the diagnosis of congenital disorders of glycosylation (CDG) and in monitoring compliance in rehabilitation programs for alcoholism. The iron load of transferrin is an indicator of iron deficiency (or sufficiency). The concentration ranges (5 – 15 μM) over which the ESI-MS response for tf was λινεαρ were determined in 0.1 M NH4HCO3, pH 8.2, and in 1% formic acid. In order to develop a method for analysis by ESI-MS, optimal conditions were determined for obtaining CDT from commercial apo-tf by treatment with neuraminidase or PNGase. The resulting CDT forms were separated by HPLC and analysed by ESIMS. The abundant human serum albumin was removed from plasma by loading 25-75 μl of plasma onto a Micro Bio-Spin column containing Cibacron Blue-agarose. A high quality ESI mass spectrum of tf was obtained from the eluent. Using ESI-MS it was possible to distinguish between commercial holo-tf (Fe2-tf) and apo-tf (Fe0-tf). The rapid analysis of transferrin from plasma by ESI-MS lays the foundation for development of a clinical method for analysing CDT and possibly for determining the iron load of circulating tf.

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Unless otherwise indicated, the views expressed in this thesis are those of the author and do not necessarily represent the views of the University of Wollongong.