Synthesis and Mass Spectrometry of Bisconjugate Phase II Steroids

Abstract

The work described in this thesis details the efforts towards the synthesis of steroid bisconjugate materials. Specifically, a library of ten steroid sulfate glucuronide and ten bisglucuronide reference materials were prepared. In addition, a study of their mass spectrometry (MS) behaviour was conducted, with the goal of using these reference materials to identify new urinary markers that can then be used in anti-doping or diagnostic applications. Two extra steroid bis(sulfates) were also synthesised as reference materials for prenatal diagnosis. Chapter 1 introduces steroids in general, their metabolism in mammalian systems, the strengths and limitations of the current steroid detection methods, and how they can be improved. Chapter 2 discusses the synthetic steps that were taken to produce the steroid sulfate glucuronides and bisglucuronides, including stable isotope labelled derivatives. This includes a discussion of the sulfation, reduction, and glucuronylation reactions. This chapter also explains the MS study of the reference materials that were synthesised, especially using tandem mass spectrometry (MS/MS). It also details an LC-MS method developed to identify new steroid markers in human urine and efforts to confirm their identities against the prepared materials. Chapter 3 discusses the synthetic steps that were taken to produce the steroid bis(sulfates) to be used as reference materials in prenatal diagnosis. Experimental procedures for the synthesis of these compounds are also presented. Chapter 4 concludes the work presented in this thesis followed by suggestions for future work in this field.