Δ9-Tetrahydrocannabinol (THC) is the most psychoactive substance out of the over 80 active cannabinoids. Due to its psychoactive and impairing properties, detection and quantitation is important to determine impairment levels of individuals. With an increased use of recreational marijuana, the risk of Driving Under the Influence of Drugs (DUID) is steadily increasing. Current legislation outlaws driving under the influence of Marijuana however there exists limitations with current methods of detection of drug analyte. Δ9-Tetrahydrocannabinol, 11-Hydroxy- THC, and 11-nor-carboxy-THC, were used in detection because these analytes are produced in the metabolism of THC. Since THC is very lipid soluble, it is present in lipid rich environments in the body. Due to the lipid rich nature of meibomian fluid, a component of tears, and the presence of Fatty Acid Binding Protein (5) FAPB5, a protein known to bind to cannabinoids, tear fluid could be used as a less-invasive biological matrix to test for the presence of THC and its metabolites.
This project optimized a collection of tear fluid, along with a simple buffer extraction, to create a method suitable for direct injection using LC-MS/MS. Collection was completed by BVI Weck-Cel® Sterile Cellulose strips, measuring approximately 2 x 20 mm, and placed in Thompson eXtreme PVDF 0.2 𝜇m, pre-slit, red cap, filter vials containing Quantisal buffer solution for extraction. All analysis and calibrations were completed with fortified matrix standards with concentrations ranging from 0.25 - 250 ng/mL. Validation was consistent with American Academy of Forensic Sciences Academy Standards Board (ASB) Standards of Forensic Toxicology Standard 036, First Edition 2018.
Tear samples were collected from volunteer patients according to Institutional Review Board (IRB) standards before and after administration of Marijuana. Samples were collected approximately 30 minutes post administration in order to capture tears when the analyte is most potent in the body. Samples and calibration standards were analyzed using Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS) with the QSight® 220 CR LC-MS/MS and the Halo® C18 3.0x50 mm (2.7 µm) column. Limit of Detection (LOD) and Limit of Quantitation (LOQ) for THC was calculated at 0.25 ng/mL. Limit of Detection of THCOOH was detected at 0.25 ng/mL and Limit of Quantitation was calculated at 1 ng/mL.
Upon analysis of Patient Samples, it was determined that THC and metabolites could be detected and quantitated in tear fluid. However, it is noted that insufficient sample volume in collection of this type of sample is an issue that leads to poor quantitation and should be optimized in future research.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/43464 |
| Date | 24 November 2021 |
| Creators | Mello Jr., Allen J. |
| Contributors | Botch-Jones, Sabra |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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