Profiling of performance-enhancing and illicit drugs by using comprehensive two-dimensional gas chromatography
Creators: name: Mitrevski, Blagoj
Publisher: name: RMIT University
Year: 2024

description: The emerging need for better separation has arisen from the limitation of conventional one dimensional (one column) GC separation, which was unable to solve the perpetual problem where coelution causes imprecise or erroneous analytical results. This becomes especially true in cases where detection and unambiguous identification of ever smaller amounts of analytes in the presence of a complex matrix becomes mandatory due to lower required detection limits. This thesis represents an original work in which the applicability of comprehensive two-dimensional gas chromatography (GC×GC) – which delivers considerable improvements in separation power – has been investigated for performance enhancing and illicit drug analysis. <br><br>Trimethylsilyl derivatized extracts of hydrolysed (anabolic agents) and non-hydrolysed (endogenous sterols) urine samples were separated on polar/non-polar and non-polar/polar column sets and detected using Time-of-Flight mass spectrometry. Additionally, headspace extraction was applied for sampling of volatiles in ecstasy, heroin and cocaine samples. A custom-created TOFMS library for the former, and commercial MS libraries in the latter study, were used for identification purpose.<br><br>The synergic effect of the highly selective and fast TOFMS detector, and the improved separation of GC×GC, was particularly advantageous over classical GC in terms of net information content and component identification. Many more analytes with higher similarity have been detected in GC×GC format. A new metric we called the minimum acceptable match (MAM) was proposed as an additional criterion when defining the limit of detection. Detection and identification of non-target components, whilst retaining full mass spectra of all components of the matrix are other benefits of the method. This benefits doping control and law enforcement since it enables retrospective search for new “designer” drugs or steroids through stored data, without need for sample re-analysis. <br><br>The proposed method was tested on the key WADA anabolic agents (clenbuterol, 19-norandrosterone, epimethendiol, M1/M2 metabolites and 3’hydroxystanozolol), the most difficult to analyse by using GC, and further evaluated against strict WADA criteria for identification and minimum required performance limits. Although GC×GC methods incorporate additional features when compared to classical GC-MS (i.e. second dimension retention time; full mass spectra) they still largely comply with WADA criteria. The bias of TOFMS towards higher masses when compared to the quadrupole mass spectra has been addressed as a major drawback of the method, and a custom-created TOFMS library is strongly recommended. <br><br>The general advantages of GC×GC method identified in doping control are fully applicable also in illicit drug profiling. By virtue of the improved separation and enhanced sensitivity of GC×GC a high number of chemically-resolved components have been detected in the profiles. This in turn requires further research to be undertaken on establishing links for their origin in drugs. Once a strong link is established, they can be used as new markers for particular geographic origin of plant materials used as precursors for drug production, for specific synthesis route of chemical production of synthetic drugs, or for establishing routes of trafficking.