Drugs in oral fluid Part I. Validation of an analytical procedure for licit and illicit drugs in oral fluid.
ABSTRACT A qualitative and quantitative analytical method was developed and validated for the determination of 49 licit and illicit drugs in oral fluid. Small oral fluid samples, volume 1mL, were collected from volunteers using a modified Omni-Sal device and the analytes were extracted from an oral fluid/buffer mixture using a single Bond Elut Certify solid phase extraction cartridge. Liquid chromatography-tandem mass spectrometry (LC-MS-MS) and gas chromatography-repetitive full scan mass spectrometry (GC-MS) were used in parallel to analyze the extracts for the targeted drugs. Extracts were analyzed by GC-MS in their underivatized form and as their pentafluoropropionyl derivatives. Deuterated internal standards were used for quantification of drugs of abuse by LC-MS-MS to minimize matrix effects. Methadone-d(9) and tumoxetine were used as the internal standards for quantification of non-derivatized and derivatized analytes respectively by GC-MS. Linearity was demonstrated over the range 5-200 ng/mL and limits of detection were less than 4 ng/mL for each drug analyzed. The method demonstrated acceptable recoveries for most of the analytes and good intra- and inter-day precision. Acquisition of data by repetitive full scan GC-MS allows the addition of further analytes to the target menu.
Article: Validation of analysis of amphetamines, opiates, phencyclidine, cocaine, and benzoylecgonine in oral fluids by liquid chromatography-tandem mass spectrometry.[show abstract] [hide abstract]
ABSTRACT: The aim of the present study was to develop and validate a method for the detection and quantitation of drugs of abuse in oral fluids. Fortified oral fluid samples (made in-house) and samples from donors collected with Quantasil oral fluid collection kits from Immunalysis were screened on an Olympus 5400 using reagents purchased from Immunalysis. Amphetamines (AMPs), opiates, phencyclidine (PCP), and cocaine and its metabolite benzoylecgonine (BE) in oral fluids were quantitated by an Applied Biosystems 3200 QTRAP liquid chromatograph-tandem mass spectrometer (LC-MS-MS). AMPs, opiates, PCP, cocaine, and BE were extracted from samples using liquid-liquid or solid-phase extractions and the extracts were separated on a Shimadzu high-performance liquid chromatograph prior to the MS-MS analysis. For each drug, two multiple reaction mode transitions were monitored using positive electrospray ionization coupled to an MS-MS detector. Corresponding d3, d5, d6, and d11 internal standards were used to quantitate the results. The limit of detection/quantitation for AMPs, opiates, PCP, cocaine, and its metabolite BE were 10, 10, 2, 2, and 2 ng/mL of oral fluid, respectively, on a signal-to-noise ratio > 4. This corresponded to 25, 25, 5, 5, and 5 pg on column. The method was verified by participating in the North America Oral Fluid Proficiency Testing administered by Research Triangle Institute and by analyzing real samples from donors. In conclusion, LC-MS-MS provided a simple way to analyze and quantitate drugs of abuse in oral fluids.Journal of analytical toxicology 11/2008; 32(8):605-11. · 2.02 Impact Factor
Article: Simultaneous detection and quantification of amphetamines, diazepam and its metabolites, cocaine and its metabolites, and opiates in hair by LC-ESI-MS-MS using a single extraction method.[show abstract] [hide abstract]
ABSTRACT: A liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous identification and quantification of amphetamines, diazepam and its metabolites, cocaine and its metabolites, and opiates from hair using a single extraction method. As part of the method development, Gemini C18, Synergi Hydro RP, and Zorbax Stablebond-Phenyl LC columns were tested with three different mobile phases. Analyte recovery and limit of detection were evaluated for two different solid-phase extraction methods that used Bond Elut Certify and Clean Screen cartridges. Phosphate buffer (pH 5.0) was chosen as the optimum hair incubation medium because of the high stability of cocaine and 6-monoacetylmorphine using this method and faster sample preparation. The optimized method was fully validated. Linearity was established over the concentration range 0.2-10 ng/mg hair, and the correlation coefficients were all greater than 0.99. Total extraction recoveries were greater than 76%, detection limits were between 0.02 and 0.09 ng/mg, and the intra- and interday imprecisions were generally less than 20% in spiked hair. The intra- and interbatch imprecision of the method for a pooled authentic hair sample ranged from 1.4 to 23.4% relative standard deviation (RSD) and 8.3 to 25.4% RSD, respectively, for representative analytes from the different drug groups. The percent matrix effect ranged from 63.5 to 135.6%, with most analytes demonstrating ion suppression. Sixteen postmortem samples collected from suspected drug-related deaths were analyzed for the 17 drugs of abuse and metabolites included in the method. The method was sufficiently sensitive and specific for the analysis of drugs and metabolites in postmortem hair samples. There is scope for the inclusion of other target drugs and metabolites in the method.Journal of analytical toxicology 10/2008; 32(7):457-69. · 2.02 Impact Factor
Article: Method for quantification of opioids and their metabolites in autopsy blood by liquid chromatography-tandem mass spectrometry.[show abstract] [hide abstract]
ABSTRACT: A method using liquid chromatography-electrospray ionization-tandem mass spectrometry was developed and validated for the determination of morphine, codeine, hydromorphone, dihydrocodeine, oxycodone, buprenorphine, and naloxone with their metabolites morphine-3-glucuronide, morphine-6-glucuronide, normorphine, 6-acetylmorphine, 6-acetylcodeine, codeine-6-glucuronide, norcodeine, hydromorphine-3-glucuronide, dihydrocodeine-6-glucuronide, dihydromorphine, dihydromorphine-3-glucuronide, dihydromorphine-6-glucuronide, oxymorphone, norbuprenorphine, buprenorphine-3-glucuronide, norbuprenorphine-3-glucuronide, and naloxone-3-glucuronide in human whole blood. Polar metabolites (glucuronides) and other analytes were extracted by SPE using Bond Elut C18. Chromatographic separation was performed on a Phenomenex Synergi reversed-phase column with gradient elution based on a mobile phase consisting of 10mM ammonium formate adjusted to pH 3 and acetonitrile. Intraday and interday precision for all analytes were between 0.6% and 13.8%, and recoveries were between 80.3% and 101.4%. Calibration curves were linear for all analytes over the concentration range 5-400 ng/mL, and correlation coefficients (R(2)) were better than 0.999. Limits of detection and quantitation were 0.16-1.2 ng/mL and 0.5-4.09 ng/mL, respectively. The method described consolidates previous work on opioids and their metabolites published in the literature and is the first to include the detection of naloxone-3-glucuronide. The method has been applied in routine postmortem cases after opiate overdose with the threefold purpose of providing interpretive information on the cause and type of death (rapid, sub-acute, or delayed death) and to distinguish heroin, morphine, and codeine users.Journal of analytical toxicology 10/2007; 31(7):394-408. · 2.02 Impact Factor