Developments and applications of solid phase microextraction (SPME) coupled to direct analysis in real time (DART) and approaches towards small volume analysis

Abstract

Solid phase microextraction (SPME) has been rapidly developing since its invention in 1990. Due to a flexible design and the ability to implement various extractive coatings, SPME has found use in numerous applications. One such application is the coupling of SPME to Direct Analysis in Real Time (DART), an ambient ionization mass spectrometry (AIMS) technique. AIMS techniques completely eliminate the use of chromatography, reducing lengthy workflows and improving time-efficiency. SPME coupled to DART-MS has been quite successful in achieving low detection limits and reproducible results in bioanalytical applications. These successes are mostly due to the excellent preconcentrating and interference resisting abilities of SPME coatings and detection capabilities of DART-MS. Certainly, as a novel technique, many aspects of SPME-DART-MS are yet to be explored. Hence, this thesis investigated fundamental aspects of SPME-DART-MS, in addition to exploring novel applications, such as detection of prohibited substances in small bio-volumes (i.e., ≤ 25 µL of oral fluid (OF) and human blood) and implementation of SPME-DART-MS as high-throughput technology for quantitation of opioids in urine and human plasma in less than 1.5 h for a set of 96 samples. Coupling of SPME to portable DART-MS instrumentation was investigated for on-site, in vivo drug detection in samples of OF as a potential tool of law enforcement, reporting detection limits that are below those proposed by regulatory agencies. Additionally, the use of biocompatible plastic materials such as polyetheretherketone (PEEK) was investigated as a potential alternative material for the manufacturing of SPME devices for DART-MS, which are usually made of stainless steel (SS). Performance of the PEEK devices was similar to those of SS, allowing for their use in quantification of drugs-of-abuse (DoAs) in OF and urine at low parts-per-billion levels (ppb). In addition, OF samples following coffee consumption from a female volunteer were used to monitor caffeine half-life. The results were in compliance with established half-life of caffeine reported in literature, and highlighted the potential of the PEEK devices for in vivo buccal swabbing testing. Another novel application of SPME described in this thesis was the development of miniaturized SPME devices “minitips” for analysis of small volumes. The analysis of small sample volumes (biofluids, tissues, and cells) has become more common within the scientific community in the last few years. For this type of analysis, very sensitive instrumental platforms and sample preparation methods are required to obtain qualitative and quantitative information. Hence, the SPME minitip format was introduced, and the tip apex was coated with 1 millimeter (mm) of polyacrylonitrile (PAN) and in-house-synthesized N-vinylpyrrolidone-co-divinylbenzene, also known as hydrophilic lipophilic balance (HLB) particles. Robustness, extraction efficiency, repeatability, reusability, and matrix effect (ME) assessment yielded good results. Benzodiazepines were extracted from 15 µL of phosphate buffer saline (PBS) and urine, and certain DoAs were extracted from 1 µL of OF using LC-MS/MS. Very good figures of merit were obtained, with limits of detection (LODs) and quantitation (LOQs) in the lower ppb range. DoAs were also extracted from 1 µL of blood by directly coupling the SPME minitips to nanoelectrospray ionization (nESI) devices. The SPME HLB minitips were also used for untargeted metabolomic profiling of caviar eggs via liquid chromatography high resolution mass spectrometry (LC/HRMS), thereby showing that despite introduced dilution, satisfactory results can be obtained. Indeed, good statistical separation of caviar eggs was observed, in addition to extracting well over a hundred significant features. Lastly, this thesis also details application of SPME for untargeted metabolomics profiling of patient affected with a genetic condition known as malignant hyperthermia (MH). Small human muscles obtained from Toronto General Hospital were sampled using SPME fibers and analysed via LC/HRMS using an untargeted metabolomics approach. Potential disease related compounds (i.e., biomarkers) were found, aiding in the elucidation of the biochemical pathways affected in patients with MH. The accepted diagnostic tool of MH is a highly invasive procedure which involves excision of a portion of the Gracilis muscle. Hence the prospect of using SPME as minimally invasive diagnostic tool was discussed. As this thesis shows, the opportunities of SPME are many; and its continuous development will most likely require efforts and collaboration from experts in numerous fields.