Simultaneous Determination of Methylphenidate, Amphetamine and their Metabolites in Urine using Direct Injection Liquid Chromatography-Tandem Mass Spectrometry (original) (raw)
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Journal of Analytical Toxicology, 2010
Human Services posted a final notice in the Federal Register authorizing the use of liquid chromatography-tandem mass spectrometry (LC-MS-MS) and other technologies in federally regulated workplace drug testing (WPDT) programs. These rules are expected to become effective in May 2010. To support this change, it is essential to explicitly demonstrate that LC-MS-MS as a technology can produce results at least as valid as gas chromatography-mass spectrometry (GC-MS), the longaccepted standard in confirmatory analytical technologies for drugs of abuse and currently the only confirmatory method allowed for use in support of federally regulated WPDT programs. A series of manufactured control urine samples (n = 10 for each analyte) containing benzoylecgonine, morphine, codeine, and 6-acetylmorphine at concentrations ranging from 10% to 2000% of federal cutoffs were analyzed with replication by five federally regulated laboratories using GC-MS (five replicate analyses per lab) and at RTI International using LC-MS-MS (10 replicate analyses). Interference samples as described in the National Laboratory Certification Program 2009 Manual were also analyzed by both GC-MS and LC-MS-MS. In addition, matrix effects were assessed for LC-MS-MS, and both analytical technologies were used to analyze previously confirmed urine specimens of WPDT origin. Results indicated that LC-MS-MS analysis produced results at least as precise, accurate, and specific as GC-MS for the analytes investigated in this study. Matrix effects, while evident, could be controlled by the use of matrix-matched controls and calibrators with deuterated internal standards. LC-MS-MS data parameters, such as retention time and product ion ratios, were highly reproducible.
Journal of Analytical Toxicology, 2005
The use of fast gas chromatography-mass spectrometry (FGC-MS) was investigated to improve the efficiency of analysis of urine specimens that previously screened presumptively positive for amphetamine (AMP), methamphetamine (MAMP), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), and/or 3,4 methylenedioxyethylamphetamine (MDEA) by immunoassay testing. Specimens were pretreated with basic sodium periodate, extracted using a positive-pressure manifold/cation-exchange solidphase cartridge methodology, and derivatized using 4-carbethoxyhexafluorobutyryl chloride (4-CB). The analytical method was compared to traditional GC-MS analysis and evaluated with respect to assay chromatography, linearity, sensitivity, precision, accuracy, and reproducibility. The limits of detection were 62.5 ng/mL for MDA and 31.25 ng/mL for AMP, MAMP, MDMA, and MDEA. All of the target analytes were linear to 12,000 ng/mL with the exception of MAMP which was linear to 10,000 ng/mL. The intra-assay precision of a 500 ng/m/multiconstituent control (n-15) ranged from 522.6 to 575.9 ng/mL with a coefficient of variation of less than 3.8%. Authentic human urine specimens (n-187) previously determined to contain the target analytes were re-extracted and analyzed by both FGC-MS and the currently utilized GC-MS method. No significant differences in specimen concentration were observed between these analytical methods. No interferences were seen when the performance of the FGC-MS method was challenged with ephedrine, pseudoephedrine, phenylpropanolamine, and
2005
The use of fast gas chromatography-mass spectrometry (FGC-MS) was investigated to improve the efficiency of analysis of urine specimens that previously screened presumptively positive for amphetamine (AMP), methamphetamine (MAMP), 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxymethamphetamine (MDMA), and/or 3,4 methylenedioxyethylamphetamine (MDEA) by immunoassay testing. Specimens were pretreated with basic sodium periodate, extracted using a positive-pressure manifold/cation-exchange solidphase cartridge methodology, and derivatized using 4-carbethoxyhexafluorobutyryl chloride (4-CB). The analytical method was compared to traditional GC-MS analysis and evaluated with respect to assay chromatography, linearity, sensitivity, precision, accuracy, and reproducibility. The limits of detection were 62.5 ng/mL for MDA and 31.25 ng/mL for AMP, MAMP, MDMA, and MDEA. All of the target analytes were linear to 12,000 ng/mL with the exception of MAMP which was linear to 10,000 ng/mL. The intra-assay precision of a 500 ng/m/multiconstituent control (n-15) ranged from 522.6 to 575.9 ng/mL with a coefficient of variation of less than 3.8%. Authentic human urine specimens (n-187) previously determined to contain the target analytes were re-extracted and analyzed by both FGC-MS and the currently utilized GC-MS method. No significant differences in specimen concentration were observed between these analytical methods. No interferences were seen when the performance of the FGC-MS method was challenged with ephedrine, pseudoephedrine, phenylpropanolamine, and
Journal of Analytical Toxicology, 2000
A rapid, sensitive, and solvent-free procedure for the simultaneous determination of amphetamine, methamphetamine, 3,4-methyleoedioxyamphetamlne (MDA), and 3,4-methylenedioxymethamphetamine (MDMA) in urine was developed using solid-phase microextractlon (SPME) and gas chromatography-mass spectrometry (GC-MS) in the selected ion monitoring mode. A headspace vial containing the urine sample, NaOH, NaCI, and amphetamine-d] as the internal standard was heated at 100~ for 20 rain. A polydimethylsiloxane fiber was maintained in the vial headspace for 10 rain in order to adsorb the amphetaminic compounds, which were subsequently derivatized by exposing the fiber to trifluoroacetic anhydride for 20 rain in the headspace of another vial maintained at 60~ for 20 rain. The trifluoroacetyl derivatives were desorbed in the GC injection port for 5 rain. Several parameters were considered during the method optimization process. These included a comparison of SPME with or without headspace, the required derivatization procedure, and the influence of temperature on the headspace extraction and derivatization methods. The optimized method was validated for the four compounds tested. Calibration curves showed linearity in the range 50-1000 ng/mL (r = 0.9946-0.9999). Recovery data were 71.89-103.24%. The quantitation limits were 10 ng/mL for amphetamine and methamphetamine and 20 ng/mL for MDA and MDMA. All of these data recommend the applicability of the method for use in the analytical routine of a forensic laboratory.
Journal of Chromatography B-analytical Technologies in The Biomedical and Life Sciences, 2002
The need for analytical screening tests more reliable and valid to detect amphetamine and related ''designer drugs'' in biological samples is becoming critical, due to the increasing diffusion of these drugs on the European illegal market. The most common screening procedures based on immunoassays suffer a number of limitations, including low sensitivity, lack of specificity and limited number of detectable substances. This paper describes a screening method based on gaschromatography-mass-spectrometry (GC / MS) using positive chemical ionisation (PCI) detection. Methanol was used as reactant gas in the ionisation chamber. Molecular ions of different compounds were monitored, allowing a sensitivity of 5-10 ng / ml with high selectivity. The sensitivity of the method gives positive results in samples taken 48-72 h after intake of one dose of 50-100 mg. The method is simple and rapid. Sample preparation was limited to one liquid-liquid extraction, without any hydrolysis and derivatisation. Hydrolysis is critical to identify metabolites excreted as conjugates. Blank urine samples spiked with known amounts of amphetamine (AM), methylamphetamine (MA), methylenedioxyamphetamine (MDA), methylenedioxymethylamphetamine (MDMA), methylenedioxyethylamphetamine (MDEA) and methylenedioxyphenyl-N-methyl-2-butanamine (MBDB) were analysed. The method was successfully tested on real samples of urine from people, whose use of amphetamine was suspected, and results were compared with results obtained with immunoassays.
Journal of Analytical Toxicology
Heroin, methamphetamine and ketamine have been the most commonly abused drugs in Taiwan. The presence of these drugs and their metabolites in postmortem specimens has been routinely monitored in our laboratory mostly by gas chromatographic-mass spectrometric methods. This study aimed to evaluate a more effective approach to simultaneously quantify these analytes (i.e., amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine (MDMA), morphine, codeine, 6-acetylmorphine, 6-acetylcodeine, ketamine and norketamine) in postmortem urine and blood specimens by liquid chromatography-tandem mass spectrometry (LC-MS-MS). Samples (1 mL) were extracted via solid-phase extraction, evaporated and reconstituted in the mobile phase for injection into the LC-MS-MS system. Respective deuterated analogs of these analytes were used as internal standards. Chromatographic separation was achieved by an Agilent Zorbax SB-Aq analytical column at 50°C. Mass spectrometric analysis was performed by electrospray ionization in positive-ion dynamic multiple reaction monitoring mode with optimized collision energy for respective precursor ion selected for each analyte, and the monitoring of two transition ions. Performance characteristics were assessed using drug-free samples that were fortified with 50-1,000 ng/mL of the 10 analytes. Analytical parameters evaluated and resulting data are as follows: (i) average extraction recoveries (n = 3) were better than 80%, except for MDMA (71%) and morphine (74%); (ii) inter-day and intra-day precision ranges (%CV) were 1.59-8.80% and 0.57-3.89%, respectively; (iii) calibration linearity (r 2), detection limit and quantitation limit for all analytes were >0.999, 1 and 5 ng/mL, respectively; (iv) matrix effects (ion suppression) were observed for three analytes, but were satisfactorily compensated for by the deuterated internal standards adopted in the analytical protocol. This method was successfully applied to the analysis of specimens collected from unknown death cases from various district prosecutors' offices in Taiwan, and was also found helpful to understanding whether the detected opiates were derived from heroin or legal morphine/codeine-containing medications.
Amphetamine and methamphetamine are commonly abused synthetic drugs which act as central nervous stimulants and have sympathomimetic properties. Because of their abuse, their use is closely monitored in the United State and elsewhere. Both drugs are also administered legitimately for common ailments. Amphetamine can be taken for the treatment of hypotension, narcolepsy and obesity. The "d" isomer of methamphetamine can also be prescribed for obesity, and the "l" isomer is commonly taken as a nasal decongestant through the use of Vicks ® Vapor Inhalers ® inhalers. When abused, synthesized amphetamine and d-methamphetamine are typically ingested or taken intravenously. l-methamphetamine has also been abused by its removal from inhalers followed by oral or intravenous administration 1 .
Mass Selective Detection of Amphetamine, Methamphetamine, and Related Compounds in Urine
Journal of Chromatographic Science, 2004
A method is presented for the routine analysis of amphetamine, methamphetamine, and related compounds in urine with gas chromatography coupled with mass spectrometry operated in the selective ion monitoring mode. The analytes are isolated by liquid-liquid extraction and are derivatized with trifluoroacetic anhydride. 3,4-Methylenedioxy-methamphetamine-D 5 is employed as the internal standard. Standard solutions are prepared using spiked urine samples, which are subjected to all phases of sample preparation. Disposable deactivated glass containers are employed throughout the process.
Journal of Chromatography B, 2002
A method using liquid chromatography coupled to tandem mass spectrometry is described for the determination of drugs of abuse in oral fluid. The method is able to simultaneously quantify amphetamines (amphetamine, methamphetamine, MDA, MDMA and MDEA), opiates (morphine and codeine), cocaine and benzoylecgonine. Only 200 micro of oral fluid is spent for analysis. The sample preparation is easy and consists of mixed mode phase solid-phase extraction. Reversed-phase chromatography is carried out on a narrow bore phenyl type column at a flow-rate of 0.2 ml/min. A gradient is applied ranging from 6 to 67.6% methanol with ammonium formate (10 mM, pH 5.0) added to the mobile phase. The column effluent was directed into a quadrupole-time-of-flight instrument by electrospray ionization, without the use of a splitter. A validation study was carried out. Recovery ranged from 52.3 to 98.8%, within-day and between-day precision expressed by relative standard deviation were less than 11.9 and 16.8%, respectively, and inaccuracy did not exceed 11.6%. The limit of quantification was 2 ng/ml (0.66 x 10(-5)-1.48 x 10(-5) M) for all compounds. Internal standards were used to generate quadratic calibration curves (r(2)>0.999). The method was applied to real samples obtained from suspected drug users. An interference was observed from the device used to sample the oral fluid, consequently this was excluded from the method which was validated on oral fluid obtained by spitting in a test-tube.
Electrophoresis, 2000
Monitoring of amphetamines and designer drugs in human urine is a timely topic in clinical toxicology, surveillance of drug substitution, forensic science, drug testing at the workplace, and doping control. Confirmation testing of urinary amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine (MDMA, Ecstasy) and 3,4-methylenedioxyamphetamine (MDA) by capillary electrophoresis (CE) combined with atmospheric pressure electrospray ionization and ion trap mass spectrometry (MS) is described. Using an aqueous pH 4.6 buffer composed of ammonium acetate/acetic acid, CE-MS and CE-MS 2 provided data that permitted the unambiguous confirmation of these drugs in external quality control urines. Furthermore, other drugs of abuse present in alkaline urinary extracts, including methadone and morphine, could also be monitored. The data presented illustrate that the sensitivity achieved with the benchtop MS is comparable to that observed by CE with UV absorption detection. CE-MS 2 is further shown to be capable of identifying comigrating compounds, including the comigration of amphetamine with nicotine.