Kim Haynes (Presenter)
Waters Corporation
Authorship: Kim Haynes, Xin Zhang, Jonathan Danaceau, and Erin Chambers
Waters Corporation
| Short Abstract A novel reversed-phase SPE sorbent has been used to rapidly analyze urinary THC and its major metabolites. Extraction of pretreated samples was rapid and UPLC/MS/MS analysis was complete in less than 3 minutes. Extraction recoveries were efficient and consistent, and matrix effects were minimal. All quantitative parameters such as linearity, accuracy, and precision were well within established limits for bioanalytical and forensic methods. This method represents a simplified and highly efficient approach for the analysis of these important compounds. |
Long Abstract
Background: Cannabis continues to be a highly abused recreational drug. In addition, the increasing number of states legalizing it for medical use combined with the trend towards legalization for recreational purposes, means than analytical methods for the quantification of of Ä-9-tetrahydrocannabinol (THC) and its metabolites and related cannabinoids continue to be necessary. Among drugs of abuse and pain management compounds, natural cannabinoids present some unique analytical challenges. Excreted THC and related metabolites are highly glucuronidated, requiring efficient deconjugation before analysis. In addition, the highly hydrophobic nature of natural cannabinoids makes them exceptionally susceptible to loss via non-specific binding, meaning that more care than usual must be taken with sample handling and processing of prepared extracts. This method details the extraction and analysis of THC and its major metabolites, 11-hydroxy Ä-9-THC (THC-OH) and 11-nor-9-Carboxy-Ä-9-THC (THC-COOH) from urine using a novel reversed-phase sorbent in a µElution format, followed by UPLC/MS/MS. Analysis is rapid and highly consistent, with all analytes eluting in less than 3 minutes. Recoveries were excellent and matrix effects were minimal for all compounds. Quantitative results were highly reproducible. Quality control results were within 12% of expected concentrations and %RSDs were less than 2%.
Methods: Urine samples were treated with β-glucuronidase followed by alkaline hydrolysis to fully deconjuate glucuronide metabolites. After pretreatment, samples were directly loaded onto the wells of SPE plates. All wells were then washed with 25% MeOH and eluted with 60:40 ACN:IPA. The sample eluates were then diluted 1:1 with water and analyzed by UPLC/MS/MS.
Results: The chromatographic method was rapid, with all compounds eluting in less than 3 minutes. Recoveries were efficient and consistent for all compounds in both panels, with %RSDs of less than 10%. Matrix effects were under 15% for all anlaytes. The method was very sensitive. Limits of quantification (LOQs) were 0.2 ng/mL for THC and THC-OH and 0.1 ng/mL for THC-COOH. The curves were linear up to 100 ng/mL for all compounds. Accuracies of QC samples were also excellent, with biases under 12% and %RSDs under 5% for all analytes.
Conclusion: This work highlights a rapid, simple and efficient analysis method for THC and urinary metabolites. Pretreated samples were directly loaded onto µElution extraction plates, without the need for traditional conditioning and equilibration. The samples are also eluted in a minimal volume, allowing direct analysis and eliminating the risk of loss from evaporation and reconstitution. Finally, extraction recoveries, matrix effects, linearity, accuracy, and precision all were well within accepted method validation parameters for bioanalytical and forensic analyses.
For Research Use Only, Not for Use in Diagnostic Procedures.
References & Acknowledgements:
| Description | Y/N | Source |
| Grants | no | |
| Salary | yes | Waters Corporation |
| Board Member | no | |
| Stock | no | |
| Expenses | no |
IP Royalty: no
| Planning to mention or discuss specific products or technology of the company(ies) listed above: | yes |