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Long-term stability data of atypical antipsychotics in different matrices are not widely available. The aim of this work was to assess the stability of amisulpride, aripiprazole and dehydroaripiprazole, clozapine and norclozapine, olanzapine, quetiapine, risperidone and 9-hydroxyrisperidone, and sulpiride in human EDTA plasma, heparinised haemolysed human whole blood, oral fluid, human serum, and newborn calf serum stored in tightly capped plastic containers under a range of conditions. Measurements were performed by LC-MS/MS. Analyte instability was defined as a deviation of 15% or greater from the expected concentration. All analytes were stable following 3 freeze-thaw cycles in human plasma, and were stable in this matrix for at least 5 days at ambient temperature (olanzapine, 3 days); 4 weeks at 2-8°C (olanzapine, 2 weeks), and 2 years at -20°C (except for dehydroaripiprazole, olanzapine, and quetiapine, 1 year). In human serum, aripiprazole, dehydroaripiprazole, norclozapine, olanzapine, quetiapine, risperidone, 9-hydroxyrisperidone, and sulpiride were unstable after 5 days at ambient temperature, 3 weeks at 2-8°C, and 9 months at -20°C. Olanzapine was unstable in whole blood and oral fluid under most conditions studied, although prior addition of ascorbic acid had a moderate stabilising effect. All other analytes were stable in whole blood and oral fluid for at least 2 days at ambient temperature, 1 week at 2-8°C, and 2 months at -20°C (clozapine and norclozapine, 1 month whole blood). These results confirm that plasma (EDTA anticoagulant) is the sample of choice for TDM of atypical antipsychotics. Delayed (more than 1 week) analysis of patient samples should be undertaken with caution, especially with serum and with haemolysed whole blood. With olanzapine, only plasma collected and stored appropriately is likely to give reliable quantitative results., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Therapeutic drug monitoring (TDM) of atypical antipsychotics is common, but published methods often specify relatively complex sample preparation and analysis procedures. The aim of this work was to develop and validate a simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the analysis of amisulpride, aripiprazole and dehydroaripiprazole, clozapine and norclozapine, olanzapine, quetiapine, risperidone and 9-hydroxyrisperidone, and sulpiride in small (200 μL) volumes of plasma or serum for TDM purposes. The applicability of the method as developed to haemolysed whole blood and to oral fluid was also investigated. Analytes and internal standards were extracted into butyl acetate:butanol (9+1, v/v) and a portion of the extract analysed by LC-MS/MS (100 mm × 2.1 mm i.d. Waters Spherisorb S5SCX; eluent: 50 mmol/L methanolic ammonium acetate, pH* 6.0; flow-rate 0.5 mL/min; positive ion APCI-SRM, two transitions per analyte). Assay calibration (human plasma, oral fluid, and haemolysed whole blood calibration solutions) was performed by plotting the ratio of the peak area of the analyte to that of the appropriate internal standard. Assay validation was as per FDA guidelines. Assay calibration was linear across the concentration ranges studied. Inter- and intra-assay precision and accuracy were within 10% for all analytes in human plasma. Similar results were obtained for oral fluid and haemolysed whole blood, except that aripiprazole and dehydroaripiprazole were within 15% accuracy at low concentration (15 μg/L) in oral fluid, and olanzapine inter-assay precision could not be assessed in these matrices due to day-by-day degradation of this analyte. Recoveries varied between 16% (sulpiride) and 107% (clozapine), and were reproducible as well as comparable between human plasma, human serum, calf serum and haemolysed whole blood. For oral fluid, recoveries were reproducible, but differed slightly from those in plasma suggesting the need for calibration solutions to be prepared in this medium if oral fluid is to be analysed. LLOQs were 1-5 μg/L depending on the analyte. Neither ion suppression/enhancement, nor interference from some known metabolites of the antipsychotics studied has been encountered. The method has also been applied to the analysis of blood samples collected post-mortem after dilution (1+1, 1+3; v/v) in analyte-free calf serum., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

The report presents a fatal poisoning of a neonate occurring in the final stage of gestational life and evoked by his mother, who, while 9 months pregnant, took a toxic dose of clozapine aiming at committing suicide. She was also severely poisoned, but ultimately was saved. The woman had been taking the medication due to schizophrenia and depression prior to conception, and the discontinuation of the drug in the course of pregnancy increased the risk of the woman attempting suicide. In the course of comprehensive toxicological analysis based on the developed analytical procedure with the use of LC-APCI-MS, clozapine and its two metabolites, norclozapine and clozapine-N-oxide, were determined in postmortem blood, liver and kidney in concentrations explaining the death of the neonate. The interpretation of the above-described case is complex and--apart from toxicological aspects--also involves issues associated with psychiatry, pharmacotherapy in pregnancy and medicolegal problems.

The long-term administration of clozapine could be verified by fine segmentation and analysis of single hairs of one person to examine the history of a multiple poisoning case. Segments of 1-2.5mm length were extracted by ultrasonification in 30 microl of the mobile phase (mixture of methanol+water, 50+50). By application of isocratic liquid chromatography and using narrow bore columns (Synergy Polar-RP, Phenomenex), an acceleration and miniaturization of the HPLC-MS-MS assay could be achieved. Total amounts of clozapine down to 30 fg (on column) and its desmethyl metabolite could be analysed in multiple reaction monitoring mode. According to typical sample amounts of approximately 16 microg, relevant hair concentrations higher than 1 pg/mg were detected. Significant and reproducible concentration profiles along the hair fibres revealed characteristic administration cycles. The administration time course - in particular the time of its termination - could be verified with a precision of a few days. The accuracy and reproducibility of the concentration profile was proven based on multiple investigations of single hairs. An individual hair growth rate of 0.55 mm/day was determined with a relative standard deviation of 8% by comparison of concentration profiles in hairs collected after a time span of 165 days.

Munchausen syndrome by proxy (MSBP) is a special form of child abuse in which an adult repeatedly produces symptoms of illness in a person under his/her care. In most cases the perpetrators are mothers who repeatedly and in different ways produce or feign symptoms of illness in their children in order to obtain medical treatment for them. MSBP is thus a special form of child abuse that is also of importance in the field of forensic medicine and a particular challenge to the medicolegal expert. We report two cases of poisoning with different substances (clozapine and clonidine) detected by toxicological investigations at our Department of Legal Medicine. The relevance of the problem for the medicolegal expert and the importance of an interdisciplinary co-operation are pointed out.

Until the first determination of opiates in human hair by GC/MS in 1985, radioimmunological results of hair examinations met with opposition. Since then, further GC/MS methods have been developed that have led not only to considerably increased sensitivity, but can also be used in hair analysis for screening purposes and can detect substances for which RIA kits are not available. In the present study different extraction methods were used along with two GC/MS processes which back-up each other. These methods include the enzymatic dissolution of the hair, incubation with a buffer solution followed by a solid-phase extraction and incubation with methanol without a further extraction. The GC/MS examination for heroin, cocaine, hashish and selected pharmaceuticals was carried out after derivatization with pentafluoropropionic acid anhydride or pentafluoro-1-propanol. Because of the higher sensitivity and versatility of GC/MS, two GC/MS processes were used to confirm each other instead of backing-up radioimmunological measurements with GC/MS.