All ETDs from UAB

Advisory Committee Chair

David E Graves

Advisory Committee Members

Stephen Barnes

Janusz Kabarowski

Donald Muccio

Sadanandan Velu

Document Type

Dissertation

Date of Award

2015

Degree Name by School

Doctor of Philosophy (PhD) College of Arts and Sciences

Abstract

The studies presented herein were born through the observations of the challenges that are faced by forensic toxicology laboratories. While these challenges were first observed in the forensic toxicology field, they are common across clinical, workplace drug testing, and pain management laboratories alike. The challenge is how to increase efficiency without sacrificing quality. Current drug screening protocols such as immunoassays (IA), gas chromatography mass spectrometry (GC/MS), and liquid-liquid chromatography tandem mass spectrometry (LC/MS/MS) are robust but inefficient and limited, with some analyses providing only class information. These analyses when coupled require up to four days to complete for a confirmed, reportable finding. Long analysis times contribute to the backlogs faced by many clinical and toxicological laboratories. In the study presented here, desorption ionization mass spectrometry (DI-MS) or, more specifically, Direct Analysis in Real Time mass spectrometry, DART™ MS, as a potential analysis platform for increasing analysis efficiency. DART™ enabled mass spectrometers are capable of providing specific analyte information in minutes. Therefore, this dissertation was designed to provide proof-of-concept that DART™ MS can be developed as a high throughput, cost effective, and highly selective platform for the analysis of select opioids in ante- and post-mortem biological specimens (i.e. blood, urine, and vitreous humor). This study focused on seven opioids analytes commonly encountered in drug testing laboratories in the three commonly submitted matrices. The selected analytes included: hydrocodone, methadone, morphine, 6-monoacetylmorphine (6MAM), morphine-6-β-glucuronide, oxymorphone and tapentadol. The selected matrices were whole blood, urine, and vitreous humor. Three key areas were evaluated as part of this dissertation: instrument selection/method development, comparison of sample processing techniques, and protocol verification. Two DART™ enabled mass spectrometers were selected: a DART™ enabled time of flight (TOF) mass spectrometer, was used for screening and a DART™ enabled triple quadrupole linear ion trap (QTRAP) mass spectrometer, used for confirmation. Both instruments rely on the mass spectrometer for separation rather than chromatography allowing for data acquisition with 2.0 and 2.5 minute methods, respectively. Acquisition methods were designed through optimization of each analyte, used to analyze fortified samples prepared for each matrix, and processed through a series of protocols. Results of this study indicated that the “best” protocol for each matrix to be dependent upon the instrument sensitivity, the analyte chemistries, and the matrix composition. For whole blood, a liquid-liquid extraction protocol utilizing a dichloromethane/chloroform extraction solvent provided the best coverage of selected analytes with the highest overall responses. This protocol was utilized to validate the designed DART™ QTRAP™ MS methods. Successful validation resulted in highly selective and specific methods for hydrocodone, methadone, tapentadol, and oxymorphone at therapeutic and sub-therapeutic concentrations. Therefore, we present DI-MS instruments such as the DART™ MS as a complementary platform for analysis of select opioids in biological specimens.

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