| Fluorescence spectroscopy is an extremely powerful analytical tool for the spectral characterization of human serum. It offers a highly sensitive method to study subnanomolar concentrations of serum components in their native, unperturbed microenvironments. The spectral characterization of human serum in the near-UV/visible was previously limited due to the highly overlapping spectra of serum components. However, the incorporation of the independent dimension of fluorescence lifetime greatly increases the information in fluorescence measurements. This dissertation describes the first application of phase-resolved fluorescence spectroscopy (PRFS) to the spectral characterization of human serum.;PRFS, a multidimensional fluorescence lifetime technique, offers several advantages for the characterization of human serum. Lifetime selectivity, the removal of scattered light contributions, and the multidimensional information afforded through both excitation and emission wavelengths, make PRFS more powerful than conventional steady-state techniques. In this work, PRFS was applied to the characterization of the intrinsic fluorescence of pooled human serum samples that were either stripped of or enriched with certain components. Chemometric techniques were employed to analyze the multidimensional data arrays.;In addition, extrinsic probe techniques were used to gain additional insight into the differences between serum samples and the relationship of these differences to lipid and albumin components in the serum, through steady-state vibronic band intensity ratios (VBIRs) and fluorescence lifetime distributions for pyrene and benzo(ghi)perylene. This dissertation also features the introduction of lifetime-resolved vibronic band intensity ratios (LRVBIRs), a new technique that allows resolution of the polarities of different probe microenvironments in complex samples. The microenvironment polarities can be related to specific lifetime components in the sample.;Finally, intrinsic and extrinsic probe techniques were used to study the effects of two antihyperlipidemic pharmaceuticals, Lovastatin and Simvastatin, on different serum samples and solutions of lipoproteins and albumin. Results show that Lovastatin and Simvastatin have definite effects on the lipids and the lipoproteins in serum, which have not been previously considered as a factor in the therapeutic activity of these drugs. |
