Substances taken for nonmedical reasons, usually for their mind-altering effects, are called drugs of abuse. The use of psychoactive plants as drugs of abuse has had a long tradition. Most commonly abused drugs extracted from or based on natural products are illicit substances, such as cannabis products, morphine, or cocaine but other herbal products used to produce a 'high' are becoming increasingly popular drugs of abuse. Unfortunately, these 'new herbal drugs' are falsely labeled as safe and legal. This article describes the classical herbal drugs of abuse and the emerging drugs of abuse of herbal origin in monographs. The monographs contain descriptions of the plants, their pharmacologically active compounds, and a summary of the current knowledge of their pharmacological properties.

Bei einer Rauchgasexposition ist an die Moglichkeit einer Mischintoxikation mit CN zu denken und bei Verdacht auf eine Zyanidintoxikation fruhzeitig eine Therapie zu beginnen. Andererseits sollten jedoch auch immer noch weitere toxische Substanzen in Betracht gezogen werden.

Background. Intravenous lipid emulsion (ILE) is recommended as a "rescue" treatment for local anaesthetic (LA) toxicity. A purported mechanism of action suggests that lipophilic LAs are sequestered into an intravascular "lipid-sink," thus reducing free drug concentration. There is limited data available correlating the effects of ILE on LAs. Aims. To compare the in vitro effect of ILE on LA concentrations in human blood/plasma and to correlate this reduction to LA lipophilicity. Method. One of four LAs (bupivacaine-most lipophilic-4mg/L, ropivacaine-6mg/L, lignocaine-14mg/L, and prilocaine-least lipophilic-7mg/L) was spiked into plasma or whole blood. ILE or control-buffer was added. Plasma was centrifuged to separate ILE and total-LA concentration assayed from the lipid-free fraction. Whole blood underwent equilibrium dialysis and free-LA concentration was measured. Percent reduction in LA concentration from control was compared between the LAs and correlated with lipophilicity. Results. ILE caused a significant reduction in total and free bupivacaine concentration compared with the other LAs. Ropivacaine had the least reduction in concentration, despite a lipophilicity similar to bupivacaine. The reduction in LA concentration correlated to increasing lipophilicity when ropivacaine was excluded from analysis. Conclusion. In this first in vitro model assessing both free-and total-LA concentrations exposed to ILE in human blood/plasma, ILE effect was linearly correlated with increasing lipophilicity for all but ropivacaine.

For many decades traditional alcohol congener analysis has provided the concentrations of fermentation by-product congeners found in blood, to ascertain if the claims of an individual regarding the alcoholic beverage(s) they have consumed were feasible, assisting in cases where after-drinking is involved. However, this technique does not provide information on the exact alcoholic beverage(s) consumed. More recently, ingredient biomarker congeners specific to certain alcoholic beverages have been detected in blood, making it possible to identify the particular alcoholic beverage consumed and therefore the source of alcohol (albeit only for a limited number of beverages). This novel approach may reduce current limitations that exist with traditional methods of detecting fermentation by-product congeners, which restrict the use of alcohol congener analysis internationally and for other medico-legal scenarios. This review examines the forensic application of alcohol congener analysis in determining the source of alcohol and other techniques.

Nonoxidative metabolites of ethanol (EtOH) have been the subject of much attention in recent years due to their diagnostic value. Due to the frequency of alcohol abuse throughout the world, forensic toxicologists are frequently confronted with the need to determine EtOH consumption. This article describes EtOH metabolites, specifically ethyl glucuronide, ethyl sulfate, ethyl phosphate, fatty acid ethyl esters, and phosphatidylethanol in humans. Methods of detection of the metabolites, interpretations of the measured EtOH metabolites in specimens such as plasma/blood, urine, hair, and vitreous humor, as well as information on the relative advantages of various specimens are described.

Ethyl glucuronide (EtG) quantification in hair was assessed using quality controls prepared by three methods: (a) spiking hair samples with known concentrations of EtG, (b) fortifying hair by incubation of blank hair with EtG for several days or (c) use of authentic hair samples positive for EtG. A liquid chromatography tandem mass spectrometry (LC–MS/MS) method was developed on a Shimadzu model 8030 instrument and validated for the quantification of EtG. For two concentration levels, approximately 50 and 500pg/mg QCs, EtG concentrations were measured in duplicate (N=2) on 8 days (N=16) and intra-assay precision (repeatability) and inter-assay precision determined using one-way analysis of variance. EtG concentrations measured in authentic hair exhibited poor intra-assay precision, with coefficients of variation of 25.1 and 20.9%, compared with 17.7 and 18.5% for fortified hair and 17.4 and 11.3% for spiked hair, for the lower and higher concentrations respectively. The inter-assay precision for authentic hair was also poorer, 35.7 and 22.5%, compared with fortified (28.2 and 19.8%) and spiked (18.4 and 13.2%) hair for the lower and higher concentrations. Although spiked QCs resulted in a better repeatability and inter-assay precision, the values obtained for QCs prepared from fortified and authentic hair are likely to be more representative of case specimens. These results have implications on the interpretation of EtG concentrations when spiked QCs are used to validate methods.

The post mortem redistribution of ten commonly prescribed antipsychotic drugs (APs) was investigated. Femoral blood was collected from 273 cases at admission to mortuary (AD) and at post-mortem (PM). The PM samples were collected at various times up to nine days after admission and the sample pairs analysed using LC-MS/MS. The drugs included in this study were 9OH-risperidone (paliperidone), amisulpride, chlorpromazine, clozapine, haloperidol, olanzapine, promethazine, quetiapine, risperidone, and zuclopenthixol. Haloperidol, quetiapine and risperidone showed minimal changes between AD and PM specimens, whereas the majority of drugs showed significant changes between the sample pairs collected at different time points post mortem (p<0.01) in addition to an average concentration change greater than the uncertainty of measurement of the applied method. Average increases in blood concentrations after admission to the mortuary ranged up to 112% (chlorpromazine and olanzapine) but also decreases up to -43% (9OH-risperidone) were seen. There were large standard deviations between sample pairs and substantial day-to-day unpredictable changes that highlight the difficulty in the interpretation of drug concentrations post-mortem. Based on the presented data, we recommend that specimens for toxicological analysis should to be taken as soon as possible after admission of a deceased person to the mortuary in order to minimise the effects of the PM interval on the drug concentration in blood.

A commercial enzyme immunoassay for the qualitative and semi-quantitative measurement of ethyl glucuronide (EtG) in urine was evaluated. Post-mortem (n=800), and clinical urine (n=200) samples were assayed using a Hitachi 902 analyzer. The determined concentrations were compared with those obtained using a previously published liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of EtG and ethyl sulfate. Using a cut-off of 0.5 µg/ml and LC-MS/MS limit of reporting of 0.1 µg/ml, there was a sensitivity of 60.8% and a specificity of 100% for clinical samples. For post-mortem samples, sensitivity and specificity were 82.4% and 97.1%, respectively. When reducing the cut-off to 0.1 µg/ml, the sensitivity and specificity were 83.3% and 100% for clinical samples whereas for post-mortem samples the sensitivity and specificity were 90.3 % and 88.3 %, respectively. The best trade-offs between sensitivity and specificity for LC-MS/MS limits of reporting of 0.5 and 0.1 µg/ml were achieved when using immunoassay cut-offs of 0.3 and 0.092 µg/ml, respectively. There was good correlation between quantitative results obtained by both methods but analysis of samples by LC-MS/MS gave higher concentrations than by enzyme immunoassay (EIA), with a statistically significant proportional bias (P<0.0001, Deming regression) for both sample types. The immunoassay is reliable for the qualitative and semi-quantitative presumptive detection of ethyl glucuronide in urine.

Antipsychotic drugs (APs) are prescribed for a wide range of psychotic illnesses. With more than 35 APs currently available worldwide, this drug class has rapidly gained importance in both clinical and forensic settings. On account of their chemical properties, many APs are present in human specimens at very low concentrations, which complicate their detection using standard gas chromatography–mass spectrometry (GC‐MS) procedures that often cannot provide the required sensitivity. Recent advances in liquid chromatography‐(tandem) mass spectrometry LC‐MS(/MS) technology have enabled accurate detection and quantification of these compounds in various human specimens, indicated by the increasing number of published methods. Method validation has been a particular focus of analytical chemistry in recent times. Recommendations set by several guidance documents are now widely accepted by the toxicology community, as reflected by the guidelines drafted by leading toxicological societies. This review provides a critical review of single‐stage and tandem LC‐MS procedures for the detection and quantification of APs, with a particular emphasis on appropriate method validation. The quality of published methods is inconsistent throughout the literature. While the majority of authors incorporate some validation experiments in their respective method development, a large number of published methods lack essential components of method validation, which are considered mandatory according to the guidelines. If adapting a method for the detection of APs for use in a laboratory, analysts should ensure successful validation experiments for appropriateness and completeness have been conducted, and perform additional experiments when indicated. Copyright © 2012 John Wiley & Sons, Ltd.

In recent years, derivatives of cathinone, a naturally occurring beta-keto phenylethylamine, have entered the illicit drug market. These compounds have been marketed over the internet or in so-called head shops as "legal highs" and have gained popularity among drug users. Numerous fatalities due to the abuse of these drugs in recent years have increased the need for their detection in human blood samples. For detection and determination of 25 designer cathinones and their related ephedrines in blood samples, a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed using only 100 µL of blood. The blood was extracted using liquid-liquid extraction with 1 mL of 1-chlorobutane containing 10% of isopropanol. The final extract was analyzed using a Shimadzu 8030 LC-MS-MS system operated in electrospray positive ionization multiple reaction monitoring mode. The method has been validated according to international guidelines and was found to be selective for all tested compounds. Calibration for all 25 studied analytes was satisfactory from 10-1,000 ng/mL. Accuracy data were within the acceptance interval of ±15% [±20% at the lower limit of quantification (LLOQ)] of the nominal values for all drugs. Within-day (repeatability) and intermediate precision data were within the required limits of 15% relative standard deviation (RSD) (20% RSD at LLOQ).

Olanzapine (OLZ) is amongst the most commonly prescribed antipsychotic drugs and is associated with substantial instability. The aim of this study was to investigate the instability of OLZ and to identify the degradants formed from its breakdown. Three experiments were conducted to monitor the degradation of OLZ and the formation of degradants in blood (1), water (2), and post-extraction at 4 °C (3). All three sample sets were analysed in duplicate and repeated in the absence (A) and presence (B) of 0.25% ascorbic acid. One degradant was identified in sample sets 2A and 3A with m/z 329 and confirmed as 2-hydroxymethyl-OLZ (2-OH-OLZ) using LC-MS techniques. The addition of 0.25% ascorbic acid slowed the degradation of OLZ down in all three experiments and inhibited the formation of 2-OH-OLZ in sample sets 2A and 3A. To investigate the influence of oxygen on the degradation of OLZ and the formation of 2-OH-OLZ in water, an additional experiment (4) was conducted. Sample sets were prepared containing different vortexing or sonication steps in order to alter the oxygen content in the samples. Statistical analysis confirmed that degradation increased significantly following vortexing for 1 min while sonication did not affect the rate of degradation of OLZ further suggesting the involvement of oxygen in the degradative processes. 2-OH-OLZ was only identified as a degradant of OLZ in aqueous solutions. It also degrades over time but its product is currently unknown and is under investigation.

Postmortem redistribution (PMR) is an accepted toxicological phenomenon that may affect the interpretation of postmortem blood concentrations. The extent of PMR is not well understood for some drugs. This report describes the PMR of selected substances resulting from the analysis of 149 cases comparing blood specimens taken at admission of the deceased to the mortuary and then at autopsy. Blood was collected in preserved tubes containing 1 % sodium fluoride/potassium oxalate. All cases were subject to a full autopsy and blood extracts were analyzed using a targeted screen by LC-MS/MS. 30 drug or drug metabolites that were detected with an incidence of 6 or more were included in this study. The pre-autopsy interval ranged from 0.5 to 164 h (6.4 days) with an average of 64 h for the cases analyzed. The increase in drug concentration from mortuary admission to autopsy ranged from 30 % for drugs such as citalopram, mirtazapine, and sertraline to 300 % for doxylamine. Only 7 drugs of the 30 studied showed increases of greater than 20 % when comparing autopsy to mortuary admission blood irrespective of the length of the postmortem interval. Drugs including methadone, EDDP, fluoxetine, mirtazapine, and sertraline all showed statistically significant increases during the pre-autopsy interval (p < 0.05) while 6-acetylmorphine, 9-hydroxy-risperidone, and caffeine showed significant decreases (p < 0.05) from mortuary admission to autopsy. While femoral blood is thought to reduce PMR, this data shows that for some drugs significant redistribution can occur even when taking peripheral specimens irrespective of the delay in the postmortem interval.

Synthetic cannabinoids sprayed on herbal mixtures have been abused as a new designer drug all over the world since 2004. In 2008, the first compounds, CP 47,497 and JWH-018, were identified as active ingredients in these mixtures. Most of the compounds have been synthesized for research purposes and are potent CB1 and/or CB2 receptor agonists. To investigate the presence of synthetic cannabinoids in blood samples, a liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed using only 100 µL of blood. After the addition of 0.2 mL of trizma buffer, the blood was extracted using liquid-liquid extraction with 1 mL of 1-chlorobutane containing 10% of isopropanol for 5 min on a shaker at 1,500 rpm. After centrifugation at 12,000 rpm for 1 min, the separated solvent layer was transferred to an autosampler vial and evaporated to dryness under N₂. The residue was reconstituted in methanol and injected into a Shimadzu 8030 LC-MS-MS system to separate and detect 25 synthetic cannabinoids. The method has been validated according to international guidelines and was found to be selective for all tested compounds. Calibration was satisfactory from 0.5-100 ng/mL, and from 5.0-500 ng/mL. for HU-210, CP 47,497 and the CP 47,497 C-8 homolog, respectively. The extraction efficiencies ranged from 30-101% and the matrix effects from 67-112%. Accuracy data were within the acceptance interval of ±15% (±20% at the lower limit of quantification) of the nominal values for all drugs.

Detection of the alcohol metabolites ethylglucuronide (EtG) and ethylsulfate (EtS) has become routine in many forensic laboratories over the last few years. Most previously published methods using liquid chromatography coupled with electrospray tandem mass spectrometry require a post-chromatographic addition of solvent and/or extensive sample preparation prior to analysis. The aim of the study was to develop a simplified method. To 20 μL urine, internal standard containing EtG-d5 and EtS-d 5 was added and the mixture was treated with elution buffer internal standard. EtG and EtS were separated using a Shimadzu Prominence high performance liquid chromatography (HPLC) system with a C18 separation column (Restek Ultra Aqueous C18, 4.6 × 150 mm, 5 μm), using isocratic elution with a mobile phase consisting of 10 mM ammonium acetate buffer pH 7 (total run time, 6 min). The compounds were detected using an Applied Biosystems API 5000 liquid chromatography tandem mass spectrometry system (atmospheric pressure chemical ionization, multiple-reaction monitoring mode). The method was fully validated according to international guidelines. The assay was found to be selective for the compounds of interest. It was linear from 0.1 to 10 mg/L for all analytes (R 2 > 0.99). Matrix effects studies showed the presence of a slight but consistent ion enhancement (n = 10 different urine samples) at low concentrations and no effects at higher concentrations. Accuracy data were between 0.75% and 8.1% bias for EtG and between −5.0% and −11.3% bias for EtS. Precision data were between 4.3% and 6.9% relative standard deviations (RSD) for EtG and between 6.0% and 7.5% RSD for EtS. No instability was observed after repeated freezing and thawing. This fast, reliable, and accurate method enables the detection and quantification of alcohol metabolites in urine. The method is easier to use and more sensitive than previously published methods.

In mid 2009 Victoria introduced compulsory drug testing of blood taken from all injured drivers taken to hospital. Δ(9)-Tetrahydrocannabinol (THC), methylamphetamine (MA) and 3,4-methylenedioxy-methylamphetamine (MDMA) are prohibited and if drivers are positive to any amount an automatic penalty is enforced. Laboratory screens were conducted on preserved blood using ELISA testing for cannabis metabolite and methylamphetamines and a fully validated LC-MS/MS method for 105 drugs including THC, amphetamines, opioids, benzodiazepines, antidepressants and antipsychotics and a number of other psychoactive substances using a minimum of two transitions per drug. Conventional GC-testing for ethanol was used to screen and quantify the presence of alcohol. 1714 drivers were tested and showed alcohol in 29% (≥ 0.01 g/100mL) and drugs in 35%. The positive rate for the three drugs prohibited by legislation was 12.5%. The prevalence of THC, MA and MDMA was 9.8%, 3.1%, and 0.8%, respectively. The range of THC concentrations in blood was 2-42 ng/mL (median 7) of which 70% had a concentration of 10 ng/mL or higher. The range of concentrations for MA and MDMA was 0.02-0.4 and 0.03-0.3mg/L (median for both drugs was 0.05 mg/L). Drugs of any type were detected in 35% of cases. The other drugs were largely prescribed drugs such as the antidepressants (9.3%) and benzodiazepines (8.9%). Neither 6-acetylmorphine nor cocaine (or benzoylecgonine) was detected in these cases.

The stability of 30 common antipsychotics (APs) in spiked whole blood was investigated over ten weeks in a preliminary experiment (designated “P experiment”). Pools of blank blood spiked with drugs at two different therapeutic levels were stored at four different temperatures: 20°C, 4°C, −20°C, and −60°C and extracted once weekly in duplicate, using a previously published method. A loss of >15% of the initial drug concentration was considered to indicate possible instability and the respective drugs were selected for further investigation in a final experiment (designated “F experiment”).Eight APs (chlorpromazine, chlorprothixene, fluspirilene, droperidol, olanzapine, thioridazine, triflupromazine, and ziprasidone) were incorporated into the F experiment. The same conditions were used in both experiments, however only a high therapeutic drug concentration was chosen for the F experiment and the storage time was extended to 20 weeks.All drugs of interest in the F experiment showed significant losses after 20 weeks of storage under at least one storage condition. The most notable results involved olanzapine, where losses of almost 100% in all storage temperatures were observed. Drug degradation in fluspirilene samples was significant after 20 weeks under all storage conditions. Overall, extensive degradation was seen with approximately 80% drug loss when stored at 20°C and 4°C with samples also seriously affected by degradation of up to 50% when stored at −20°C and −60°C, respectively. Ziprasidone remained stable when stored at 4°C, −20°C, and −60°C over 9 weeks, however significant degradation was observed when stored at 20°C, with a loss of almost 100% after 20 weeks of storage.The time period and temperature of storage of biological samples can have a significant influence on the stability of several APs. It is therefore important to be aware of potential changes in drug concentrations during storage when interpreting analytical results.

Oral fluid (OF) has become a popular specimen to test for presence of drugs, particularly in regards to road safety. In Victoria, OF specimens from drivers have been used to test for the presence of methylamphetamine (MA) and Δ(9)-tetrahydrocannabinol (THC) since 2003 and 3,4-methylenedioxy-N-methylamphetamine (MDMA) since 2006. LC-MS/MS has been used to test the most recent 853 submitted OF specimens from Victoria Police for 31 drugs of abuse including those listed in the Australian Standard AS4760-2006. At least one proscribed drug was detected in 96% of drivers, of which MA was the most common (77%), followed by THC (42%), MDMA (17%) and the combination of all three (3.9%). Opioids were detected in 14% of drivers of which 4.8% were positive for 6-acetylmorphine and 3.3% for methadone. The incidence of the opioids tramadol (1.2%) and oxycodone (1.1%) were relatively low. Cocaine (8.0%) was as commonly detected as benzodiazepines (8.0%), and was almost always found in combination with MA (7.9%). Samples positive to benzodiazepines were largely due to diazepam (3.5%) and alprazolam (3.4%), with only 0.2% of drivers combining the two. Ketamine was also detected in 1.5% of cases. While the incidences of the proscribed drugs itself are concerning, it is clear that many drivers are also using other drugs capable of causing impairment.

Introduction Targeted screening methods that cover a broad range of common drugs and drugs of abuse are essential for forensic laboratories. The most common techniques used are immunoassay techniques, GC-MS, and more recently LC-MS(MS). The major advantages of immunoassay techniques are their easy use and fast turnaround time, however, mass spectrometric techniques are more selective and/or sensitive, and technically more demanding. The aim of this study was to develop a LC-MS/MS based screening technique that covers the most common drugs observed in forensic analysis, using fast extraction method combined with fully automated data processing. Methods After liquid-liquid extraction (LLE) of 100 μL of post-mortem blood, 105 of the most common drugs and drugs of abuse were separated using a Shimadzu Prominence HPLC system with an C18 separation column (Eclipse XBD C18, 4.6 × 150 mm, 5 μm), using gradient elution with a mobile phase of 50 mM ammonium formate buffer pH 3.5 / acetonitrile. The drugs were detected using an Applied Biosystems 3200 Q-TRAP LC-MS-MS system (ESI, MRM mode). The method was fully validated according to international guidelines. Quantification data obtained using calibration curves were compared to a one point calibration. Results The assay was found to be selective for the compounds of interest. The calibration range spanned from therapeutic to potentially toxic concentrations described for these drugs. With a few exceptions, recoveries were typically > 70%. Accuracy, repeatability and intermediate precision were within the required limits for most analytes. Using a one point calibration, most analytes obtained similar accuracy and precision compared to a full calibration. Data processing was automated based on custom built settings. All quantifier transition peaks matching the retention times within 2% maximum difference that resulted in a calculated concentration above the lowest calibrator were reported as tentatively positive for further checking. In the scenario where the qualifier ion matched the MRM ratio obtained from in batch quality control samples within the EU mass spectrometry guidelines, the drug was highlighted in green. In the scenario where the MRM ratio did not match, the drug was still shown as tentatively positive, but highlighted in yellow. Peak review including MRM ratio was assessed for all tentatively positive peaks. Conclusions The presented LC-MS-MS assay has proven to be applicable for determination of the studied analytes in blood. The fast and reliable extraction method combined with automated processing gives the opportunity for overnight turnaround times. The method has been applied to routine analysis at the Victorian Institute of Forensic Medicine since April 2009 and approximately 3500 blood samples have been analysed.

Over the last decade, the prescription rates of antipsychotic (AP) drugs have increased worldwide. Studies have shown that the risk of sudden cardiac death is threefold higher among patients treated with APs. To investigate the presence of APs in postmortem cases, a liquid chromatography (LC)-MS/MS method was developed using only 0.1 ml of blood sample with 10 microl of internal standard (IS) (haloperidol-d(4), 1 microg/ml). After the addition of 0.2 ml of Trizma buffer, the blood sample was extracted using liquid-liquid extraction (LLE) with 1 ml of 1-chlorobutane for 5 min on a shaker at 1500 rpm. After centrifugation at 12,000 rpm for 1 min, the separated solvent layer was transferred to an autosampler vial and evaporated to dryness under N(2). The residue was reconstituted in 0.05 ml acetonitrile containing 0.1% formic acid, vortexed for 30 s and an additional 0.45 ml of 50 mmol/l ammonium formate pH 3.5 was added and the sample vortexed; 0.1 ml of the final extract was injected into a Shimadzu Prominence HPLC system, with detection of drugs achieved using an Applied Biosystems 3200 Q-TRAP LC-MS/MS system equipped with a Turbo V ion source [electron spray ionization (ESI), multiple reaction monitoring (MRM) mode]. The method has been validated according to international guidelines and was found to be selective for all tested compounds. Calibration was satisfactory for all drugs, except olanzapine, from subtherapeutic to toxic concentrations. The lower limits of quantifications (LLOQs) corresponded to the lowest concentrations used for the calibration curves. With the exception of the lowest concentrations of bromperidol, buspirone and perphenazine, accuracy data were within the acceptance interval of +/- 15% (+/- 20% at LLOQ) of the nominal values for all drugs. The method has been proven to be useful for the routine analysis of APs in postmortem blood samples.