This thesis describes the design of a convenient format of solid phase microextraction (SPME) for bioanalysis in pharmaceutical industry and the validation of the approach to the application. An automated in-tip SPME technique coupled with liquid chromatography (LC) and tandem mass spectrometry (MS/MS) for high throughout drug analysis has been developed and applied to the quantitative determination of various drug compounds in different biological fluids from drug discovery to clinical development.
The initial research in this thesis focused on a proof-of-concept study using manual multi-fiber approach to determine a drug compound in human plasma from a clinical trial. The proof-of-concept was achieved based on the validation data and a head-to-head comparison with conventional liquid-liquid extraction (LLE) method. An in-tip SPME technique was then proposed to explore the feasibility of SPME automation and two approaches of preparing in-tip SPME fibers were developed including fiber-packed and sorbent-packed fiber preparation. A simple and high throughput in-tip SPME fiber fabricating procedure based on polymer monoliths using photo-polymerization was introduced to prepare 96 fibers simultaneously. The biggest advantage of the in-tip SPME technique is that it is simple and easy to use for automation without introducing any additional devices and in the meantime, the simplicity of SPME is maintained.
Automated in-tip SPME was applied to routine drug analysis in drug discovery and development environment. One case study involved the determination of vitamin D3 in human serum with derivatization and the in-tip SPME approach was compared with traditional LLE method using either tubes or 96-well plate extraction. Another study was to use hydrophilic interaction chromatography (HILIC) –MS/MS to determine three polar compounds, imipenem (IMP), cliastatin (CIL) and -lactamase inhibitor (BLI) simultaneously in different biological fluids including rat plasma and mouse blood. The results from both studies clearly demonstrated that in-tip SPME could be used as an alternative sample preparation method in bioanalytical analysis. Matrix effects in bioanalysis using automated in-tip SPME and LC-MS/MS were then thoroughly evaluated for the first time. Our study indicated that the assumption that SPME should provide sample clean up as effective as or better than solid phase extraction (SPE) with no or minimal matrix effects might not be always true, and matrix effects should be investigated in any SPME assays in bioanalysis.
Comparisons between in-tip SPME and other automated SPME approaches such as blade/thin film geometries were performed, and the advantages and limitations of using SPME versus conventional sample preparation methods including protein precipitation (PPT), LLE and SPE were summarized. Strategies for in-tip SPME method development and validation and the potential applications and future directions of in-tip SPME in bioanalysis were discussed.
Finally, kinetic models were established to describe SPME extraction and desorption processes in a complex matrix with both liquid and solid fiber coatings. The models were successfully applied to different scenarios to estimate the boundary layer (BL) thickness, extraction equilibrium time and total amount of analytes extracted at a given time. The excellent agreements between the model prediction results and experimental data indicated that the SPME modeling approach had great potentials to speed up SPME method development and fiber selection.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OWTU.10012/6338 |
| Date | 12 September 2011 |
| Creators | Xie, Wei |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Thesis or Dissertation |
Page generated in 0.0019 seconds