High Throughput Detection of 327 Drugs in Blood by LC-MS-MS with Automated Data Processing.

The described procedure provides a rapid technique for the detection and semi-quantitation of a large number of drugs in blood. This procedure uses a minimal sample volume and employs a one-step liquid extraction and automated data processing to yield rapid turnaround times. A total of 327 of the most commonly used medicinal and illicit drugs in Australia were selected including various amphetamines, anesthetics, antidepressants, antipsychotics, anticonvulsants, benzodiazepines, beta blockers, opioid and nonopioid analgesics, stimulants, THC and a large number of synthetic cannabinoids and other novel psychoactive substances. The extracts were subject to 5-minute chromatography using a Kinetex C18 50 × 4.6 mm 2.6 µm solid-core analytical column and analyzed using a Sciex 3200 Q-TRAP MS-MS (+ ESI, MRM mode, two transitions per analyte). The method was fully validated in accordance with international guidelines. Matrix effects and extraction efficiencies were acceptable with most analytes showing > 80% response and low variation (within 25%RSD). Cannabinoids were most affected by the matrix and yielded poorest recovery values but were still detectable. Precision, accuracy, repeatability and multipoint linearity were assessed for all analytes. The method has been used in routine practice in the forensic toxicology service at the Victorian Institute of Forensic Medicine in over 6000 coronial investigations using both postmortem and clinical blood specimens. This technique has greatly increased throughput, reduced turnaround times and allowed for rapid same-day analysis of results when needed. The method is routinely used in routine overnight testing with results reported to pathologists within 4 h of data acquisition. This rapid toxicological technique is used in conjunction with other investigative processes such as full-body CT imaging, review of case circumstances and medical histories to provide an efficient death investigation process.

Ultra-rapid targeted analysis of 40 drugs of abuse in oral fluid by LC-MS/MS using carbon-13 isotopes of methamphetamine and MDMA to reduce detector saturation.

The number of oral fluid samples collected by the road policing authority in Victoria, Australia, requiring confirmatory laboratory analysis for drugs proscribed under Victorian legislation (methamphetamine, MDMA and Δ9-tetrahydrocannabinol) has greatly increased in recent years, driving the need for improved analysis techniques to enable expedient results. The aim of this study was to develop an LC-MS/MS-based targeted oral fluid screening technique that covers a broad range of basic and neutral drugs of abuse that can satisfy increased caseload while monitoring other compounds of interest for epidemiological purposes. By combining small sample volume, simple extraction procedure, rapid LC-MS/MS analysis and automated data processing, 40 drugs of abuse including amphetamines, benzodiazepines, cocaine and major metabolites, opioids, cannabinoids and some designer stimulants were separated over 5 min (with an additional 0.5 min re-equilibration time). The analytes were detected using a Sciex® API 4500 Q-Trap LC-MS/MS system with positive ESI in MRM mode monitoring three transitions per analyte. The method was fully validated in accordance with international guidelines and also monitored carbon-13 isotopes of MDMA and MA to reduce detector saturation effects, allowing for confirmation of large concentrations of these compounds without the need for dilution or re-analysis. The described assay has been successfully used for analysis of oral fluid collected as part of law enforcement procedures at the roadside in Victoria, providing forensic results as well as epidemiological prevalence in the population tested. The fast and reliable detection of a broad range of drugs and subsequent automated data processing gives the opportunity for high throughput and fast turnaround times for forensic toxicology.

Fast targeted analysis of 132 acidic and neutral drugs and poisons in whole blood using LC-MS/MS.

The aim of this study was to develop an LC-MS/MS based screening technique that covers a broad range of acidic and neutral drugs and poisons by combining a small sample volume and efficient extraction technique with simple automated data processing. After protein precipitation of 100µL of whole blood, 132 common acidic and neutral drugs and poisons including non-steroidal anti-inflammatory drugs, barbiturates, anticonvulsants, antidiabetics, muscle relaxants, diuretics and superwarfarin rodenticides (47 quantitated, 85 reported as detected) were separated using a Shimadzu Prominence HPLC system with a C18 separation column (Kinetex XB-C18, 4.6mm×150mm, 5µm), using gradient elution with a mobile phase of 25mM ammonium acetate buffer (pH 7.5)/acetonitrile. The drugs were detected using an ABSciex(®) API 2000 LC-MS/MS system (ESI+ and -, MRM mode, two transitions per analyte). The method was fully validated in accordance with international guidelines. Quantification data obtained using one-point calibration compared favorably to that using multiple calibrants. The presented LC-MS/MS assay has proven to be applicable for determination of the analytes in blood. The fast and reliable extraction method combined with automated processing gives the opportunity for high throughput and fast turnaround times for forensic and clinical toxicology.

Comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry (GC x GC-TOFMS) for drug screening and confirmation.

Comprehensive two-dimensional gas chromatography (GC x GC) is applied to analysis of drug standard mixtures containing 78 drugs of interest in forensic samples. For this study, underivatised drugs were employed. While several of the drugs were not detected at the low concentrations employed in the samples, most could be satisfactorily assigned their first and second dimension retentions in the GC x GC retention plane. For this study, time-of-flight mass spectrometry (TOFMS) detection was used. The enhanced separation possible in GC x GC is demonstrated, and typical linearity and apparatus precision are shown for tramadol, diazepam, olanzapine and desipramine using selected qualifier ions. Mass spectral library search quality for the detection of drugs in a selection of authentic forensic cases, along with retention position in the 2D retention plane, is used to support positive identification of the presence of the drugs. The analysis of 'difficult' drugs paracetamol and phenytoin is shown to produce anomalous chromatographic peak shape in the 2D plane, whereas most drugs gave acceptable peak shapes. The GC x GC technique was applied to screening drugs in forensic samples, with either flame ionisation (FID) or TOFMS detection, and compared favourably with conventional single column GC-MS analysis when tested for diazepam in an authentic forensic study.