The Development of Enhanced Raman Scattering for the Analysis of Biomedical and Pharmaceutical Compounds in Biofluids

Abstract

Surface-enhanced Raman scattering (SERS) is a surface-sensitive extension of conventional Raman spectroscopy. In SERS molecules of interest, when excited in the vicinity of rough nanoscale substrates, exhibited an enhancement in Raman scattering through the interaction of surface charges (plasmon) with the incident electromagnetic radiation (laser). The majority of the work developed and described in this thesis focusses on the detection and quantification of pharmaceuticals as well as studies to improve the reproducibility of SERS. Here within, a nanoparticle-based SERS approach was used to detect the β-blocker propranolol in multiple human biofluids. Initial investigations involved careful control for the optimisation of SERS conditions taking into consideration the complex biological matrix, as this optimisation plays a crucial role in maximising both the enhancement effect and reproducibility. The detection and quantification of propranolol spiked into human serum, plasma and urine at physiologically relevant concentrations in combination with chemometrics was then successfully achieved. Additionally, the results from SERS and LC-MS for propranolol quantification in plasma were in excellent agreement. Multiple factors may be affected during SERS quantification and these may include number of nanoparticles in the collection volume and incident laser fluence; therefore SERS was combined with an isotopic labelling strategy in order to improve the quantification of tryptophan and caffeine. Results confirmed that the inclusion of an isotopolog internal standard enabled SERS detection to be further developed by improving its reproducibility, accuracy and precision based on the ratio of analyte/internal standard, and this was the case for both 2H for tryptophan and 13C for caffeine. Moreover, this strategy was further applied to a complex biological matrix where human plasma was spiked with codeine and codeine-d6 was used as an internal standard. These findings clearly demonstrate the potential application of SERS in complex biological matrices when combined with internal isotope standards. Finally, a novel SERS detector combined with high-performance liquid chromatography (HPLC) (LC-SERS) was built. The LC-SERS approach is based on the introduction of nanoparticles into the LC eluent followed by the introduction of the aggregation agent and allows for more intense and reproducible SERS spectra to be generated. The ability to quantify methotrexate and it metabolites was demonstrated, from mixtures in water and in patient urine samples, even when the analytes were co-eluting. This quantitative analysis was conducted on-line and in real-time, making it applicable to high-throughput applications. It is hoped that the advances through the research projects presented in this thesis have made a significant contribution to the field of SERS and advanced its sensitivity and reproducibility as a quantitative analytical technique for the analysis of biomedical compounds in biofluids.

Date of Award	1 Aug 2017
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Royston Goodacre (Supervisor)