Site-, Technique-, and Time-Related Aspects of the Postmortem Redistribution of Diazepam, Methadone, Morphine, and their Metabolites: Interest of Popliteal Vein Blood Sampling.

Sampling site, technique, and time influence postmortem drug concentrations. In 57 cases, we studied drug concentration differences as follows: subclavian vein-dissection/clamping versus blind stick, femoral vein-dissection/clamping versus blind stick, right cardiac chamber, and popliteal vein-dissection and clamping only. Cases were distributed in group #1 (all cases with both techniques), group #2 (dissection/clamping), and group #3 (blind stick). Sampled drugs were diazepam, methadone, morphine, and their metabolites. To assess PMR, mean concentrations and ratios were calculated for each group. Time-dependent variations of blood concentrations and ratios were also assessed. Results indicate that site, method, and time may influence postmortem distribution interpretation in different ways. Popliteal blood seems less subject to PMR. In conclusion, our study is the first to evaluate concurrently three main aspects of PMR and confirms that the popliteal vein may represent a site that is more resistant to the changes seen as a result of PMR.

Anti-anxiety drugs and fish behavior: Establishing the link between internal concentrations of oxazepam and behavioral effects.

Psychoactive drugs are frequently detected in the aquatic environment. The evolutionary conservation of the molecular targets of these drugs in fish suggests that they may elicit mode of action-mediated effects in fish as they do in humans, and the key open question is at what exposure concentrations these effects might occur. In the present study, the authors investigated the uptake and tissue distribution of the benzodiazepine oxazepam in the fathead minnow (Pimephales promelas) after 28 d of waterborne exposure to 0.8 µg L-1 , 4.7 µg L-1 , and 30.6 µg L-1 . Successively, they explored the relationship between the internal concentrations of oxazepam and the effects on fish exploratory behavior quantified by performing 2 types of behavioral tests, the novel tank diving test and the shelter-seeking test. The highest internal concentrations of oxazepam were found in brain, followed by plasma and liver, whereas muscle presented the lowest values. Average concentrations measured in the plasma of fish from the 3 exposure groups were, respectively, 8.7 ± 5.7 µg L-1 , 30.3 ± 16.1 µg L-1 , and 98.8 ± 72.9 µg L-1 . Significant correlations between plasma and tissue concentrations of oxazepam were found in all 3 groups. Exposure of fish to 30.6 µg L-1 in water produced plasma concentrations within or just below the human therapeutic plasma concentration (HT PC) range in many individuals. Statistically significant behavioral effects in the novel tank diving test were observed in fish exposed to 4.7 µg L-1 . In this group, plasma concentrations of oxazepam were approximately one-third of the lowest HT PC value. No significant effects were observed in fish exposed to the lowest and highest concentrations. The significance of these results is discussed in the context of the species-specific behavior of fathead minnow and existing knowledge of oxazepam pharmacology. Environ Toxicol Chem 2016;35:2782-2790. © 2016 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals, Inc. on behalf of SETAC.

Population Pharmacokinetics of High-Dose Oxazepam in Alcohol-Dependent Patients: Is There a Risk of Accumulation?

BACKGROUND: According to the guidelines, benzodiazepines with a short half-life are the reference medication to treat alcohol withdrawal syndrome. The doses of oxazepam used in this population may reach up to 300 mg per day, significantly higher than usual doses. Its use in these patients deserves further information to confirm that the half-life remains constant and that no accumulation appears. The objective of this study was to investigate the pharmacokinetics of high doses of oxazepam in alcohol-dependent patients treated for alcohol withdrawal syndrome. METHODS: Overall, 63 outpatients [weight, 71.1 kg (45.0-118.0); age, 47.6 years (31-67)] followed in the addictology unit, were studied. Total mean dose of 96.0 mg per day (range, 20-300 mg/d) was administered by oral route. Therapeutic drug monitoring allowed the measurement of 96 plasma concentrations. The following covariates were evaluated: demographic data (age, body weight, height, gender) and biological data (creatinine, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase). Pharmacokinetic analysis was performed using a nonlinear mixed-effect population model. RESULTS: Data were modeled with a 1-compartment pharmacokinetic model. The population typical mean 90% confidence interval values for clearance, apparent volume of distribution (V), and duration of absorption (D1) were 6.8 L/h (range, 3.9-8.0 L/h), 159 L (range, 98.0-282 L), and 2 hours (fixed), respectively. The interindividual variability of clearance and V, and residual variability (90% confidence interval) were 74% (44%-96%), 69% (40%-89%), and 32% (20%-41%), respectively. The elimination half-life was 16 hours (range, 3-42 hours). CONCLUSIONS: Oxazepam exhibited a linear pharmacokinetics with a proportional relationship from 20 to 300 mg per day, the dose range currently used in alcohol-dependent patients treated for alcohol withdrawal syndrome. We did not find any evidence of drug accumulation with these doses.

Low doses of ethanol markedly potentiate the anti-seizure effect of diazepam in a mouse model of difficult-to-treat focal seizures.

Ethanol is commonly used as a solvent in injectable formulations of poorly water-soluble drugs. The concentrations of ethanol in such formulations are generally considered reasonably safe. It is long known that ethanol can potentiate central effects of sedatives and tranquillizers, particularly the benzodiazepines, most likely as a result of a synergistic interaction at the GABAA receptor. However, whether this occurs at the low systemic doses of ethanol resulting from its use as solvent in parenteral formulations of benzodiazepines is not known. In the present study we evaluated whether a commercial ethanol-containing aqueous solution of diazepam exerts more potent anti-seizure effects than an aqueous solution of diazepam hydrochloride or an aqueous emulsion of this drug in the intrahippocampal kainate model of temporal lobe epilepsy in mice. Spontaneous epileptic seizures in this model are known to be resistant to major antiepileptic drugs. Administration of the ethanol-containing formulation of diazepam caused an almost complete suppression of seizures. This was not seen when the same dose (5 mg/kg) of diazepam was administered as aqueous solution or emulsion, although all three diazepam formulations resulted in similar drug and metabolite concentrations in plasma. Our data demonstrate that ethanol-containing solutions of diazepam are superior to block difficult-to-treat seizures to other formulations of diazepam. To our knowledge, this has not been demonstrated before and, if this finding can be translated to humans, may have important consequences for emergency treatment of acute seizures, series of seizures, and initial treatment of status epilepticus in patients.

Direct determination of diazepam and its glucuronide metabolites in human whole blood by µElution solid-phase extraction and liquid chromatography-tandem mass spectrometry.

A µElution solid-phase extraction (SPE) liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of diazepam, nordiazepam, oxazepam, oxazepam glucuronide, temazepam and temazepam glucuronide in human whole blood is presented. 200 µL of whole blood samples were loaded onto a Waters Oasis HLB 96-well µElution SPE plate using 75 µL of methanol as the elution solvent, and the eluents were injected into an Eclipse XDB C18 column. No hydrolysis, solvent transfer, evaporation or reconstitution was involved in the sample preparation procedures. Tandem mass spectrometric detection with Turbo Ion Spray was conducted via multiple reaction monitoring (MRM) under positive ionization mode. The method was validated and proved to be accurate (accuracy within 93-108%), precise (intra-day RSD<9.9% and inter-day RSD<7.2%) and sensitive with limits of detection (LOD) in the range of 0.05-0.25 ng/mL for all the compounds. Extraction recoveries were in the range of 31-80% for all the analytes. This method demonstrated to be reproducible and reliable. The applicability of the method was demonstrated by analysis of several forensic cases involving diazepam and its metabolites.

Pharmacokinetics of the cytochrome P-450 substrates phenytoin, theophylline, and diazepam in healthy Greyhound dogs.

The purpose of this study was to determine the pharmacokinetics of phenytoin, theophylline, and diazepam in six healthy Greyhound dogs. Additionally, the pharmacokinetics of the diazepam metabolites, oxazepam and nordiazepam, after diazepam administration was determined. Phenytoin sodium (12 mg/kg), aminophylline (10 mg/kg), and diazepam (0.5 mg/kg) were administered IV on separate occasions, and blood was collected at predetermined time points for the quantification of plasma drug concentrations by fluorescence polarization immunoassay (phenytoin, theophylline) or mass spectrometry (diazepam, oxazepam, and nordiazepam). The terminal half-life was 4.9, 9.2, and 1.0 h, respectively, for phenytoin, theophylline, and diazepam, and 6.2 and 2.4 h for oxazepam and nordiazepam after IV diazepam. The clearance was of 2.37, 0.935, and 27.9 mL · min/kg, respectively, for phenytoin, theophylline, and diazepam. The C(MAX) was 44.7 and 305.2 ng/mL for oxazepam and nordiazepam, respectively, after diazepam administration. Temazepam was not detected above 5 ng/mL in any sample after IV diazepam.

Quantification of total and unbound concentrations of lorazepam, oxazepam and temazepam in human plasma by ultrafiltration and LC-MS/MS.

BACKGROUND: A fast and sensitive assay for quantifying total and unbound concentrations of lorazepam (Lzp), oxazepam (Ozp) and temazepam (Tzp) in human plasma was needed for a plasma protein binding study. RESULTS: Plasma samples were precipitated with acetonitrile for determination of total concentrations or subjected to ultrafiltration for determination of unbound concentrations. An LC-MS/MS assay was developed with an Allure® PFP propyl column and a mobile phase of 35% acetonitrile/0.1% formic acid over 4.5 min and ESI+-MS/MS detection. Matrix effects were negligible in plasma and approximately 70% in ultrafiltrate but were accounted for by the internal standards Lzp-d4, Ozp-d5 and Tzp-d5. The assay was validated for total concentrations of 10-100 ng/ml Lzp, 200-2000 ng/ml Ozp and 100-1000 ng/ml Tzp, and for unbound concentrations of 1-10 ng/ml Lzp, 20-200 ng/ml Ozp and 10-100 ng/ml Tzp. Precision was <14% CV and accuracy was 96-110% throughout the calibration range. The mean precision of triplicate analysis of 60 study samples was <4% CV for total and <8% CV for unbound concentrations. CONCLUSION: A fast and sensitive assay was developed and validated. It has been applied successfully to a protein binding study.

A case of extreme agitation and death after the use of mephedrone in The Netherlands.

A 36-year old man, having injured himself severely by smashing windows in a rage of fury, was arrested by the police. He died despite resuscitation attempts. The forensic autopsy showed many superficial skin lacerations, bruises and minor brain swelling, but there was no definitive cause of death. Toxicological analysis showed a high concentration of mephedrone in femoral blood (5.1mg/L) and traces of cocaine, MDMA and oxazepam. The remaining dose of mephedrone in the stomach contents was estimated at 113 mg. Tablets that were found in the house of the deceased also contained mephedrone. We attribute this man's death to a fatal oral intake of mephedrone, which probably led to a state of excited delirium. This was aggravated by blood loss from multiple wounds.

Diazepam pharmacokinetics after nasal drop and atomized nasal administration in dogs.

The standard of care for emergency therapy of seizures in veterinary patients is intravenous (i.v.) administration of benzodiazepines, although rectal administration of diazepam is often recommended for out-of-hospital situations, or when i.v. access has not been established. However, both of these routes have potential limitations. This study investigated the pharmacokinetics of diazepam following i.v., intranasal (i.n.) drop and atomized nasal administration in dogs. Six dogs were administered diazepam (0.5 mg/kg) via all three routes following a randomized block design. Plasma samples were collected and concentrations of diazepam and its active metabolites, oxazepam and desmethyldiazepam were quantified with high-performance liquid chromatography (HPLC). Mean diazepam concentrations >300 ng/mL were reached within 5 min in both i.n. groups. Diazepam was converted into its metabolites within 5 and 10 min, respectively, after i.v. and i.n. administration. The half lives of the metabolites were longer than that of the parent drug after both routes of administration. The bioavailability of diazepam after i.n. drop and atomized nasal administration was 42% and 41%, respectively. These values exceed previously published bioavailability data for rectal administration of diazepam in dogs. This study confirms that i.n. administration of diazepam yields rapid anticonvulsant concentrations of diazepam in the dog before a hepatic first-pass effect.

[Determination of 10 sedative-hypnotics in human plasma using pulse splitless injection technique and gas chromatography-mass spectrometry].

A simple, precise and sensitive gas chromatography-mass spectrometry (GC-MS) method coupled with pulse splitless injection technique was developed for the determination of 10 sedative-hypnotics (barbital, amobarbital, phenobarbital, oxazepam, diazepam, nitrazepam, clonazepam, estazolam, alprazolam, triazolam) in human plasma. The drugs spiked in plasma were extracted with ethyl acetate after alkalization with 0.1 mol/L NaOH solution. The organic solvent was evaporated under nitrogen stream, and the residues were redissolved by ethyl acetate. The separation was performed on an HP-5MS column (30 m x 250 microm x 0.25 microm). The analytes were determined and identified using selected ion monitoring (SIM) mode and scan mode, respectively. The internal standard method was used for the determination. The target analytes were well separated from each other on their SIM chromatograms and also on the total ion current (TIC) chromatograms. The blank extract from human plasma gave no peaks that interfered with all the analytes on the chromatogram. The calibration curves for 10 sedative-hypnotics showed excellent linearity. The correlation coefficients of all the drugs were higher than 0.9954. The recoveries of the drugs spiked in human plasma ranged from 92.28% to 111.7%, and the relative standard deviations (RSDs) of intra-day and inter-day determinations were from 4.09% to 14.26%. The detection limits ranged from 2 to 20 microg/L. The method is simple, reliable, rapid and sensitive for the determination and the quantification of 10 sedative-hypnotics in human plasma and seems to be useful in the practice of clinical toxicological cases.

Biotransformation of diazepam in a clinically relevant flat membrane bioreactor model using primary porcine hepatocytes.

In vitro biotransformation of drug using commercial culture medium with serum may not be the ideal culture medium for clinical application in extracorporeal bioartificial liver support (BAL) systems. In these systems, patient's blood or plasma is plumbed to primary hepatocytes within a seeded bioreactor, creating interaction between plasma and seeded hepatocytes. To address this situation, we investigated the biotransformation potential of diazepam in primary porcine hepatocytes with a flat membrane bioreactor (FMB); we used human plasma exposure and serum-free media in organotypical double gel culture model for long-term culture. We investigated diazepam clearance and all major metabolites of diazepam, such as oxazepam, temazepam, and desmethyldiazepam, in conventional single gel and organotypical sandwich models and compared them to the FMB model. Diazepam elimination was higher in double gel cultures with exposure to both SF 3 medium conditions and plasma, when compared to the single gel model in a Petri dish. It was observed that in the FMB, diazepam elimination was stable at about 3 pg/h/cell in plasma and SF 3 exposure. Oxazepam synthesis in the bioreactor was approximately one quarter less than in the Petri dish, but there were no differences between N-desmethyldiazepam and temazepam synthesis in double gel culture. In the flat membrane bioreactor, there was no decrease in the biotransformation of diazepam in plasma exposure compared with the control group. Our results suggest that this plasma exposure bioreactor may offer a useful approach in clinical use of extracorporeal BAL, as well as for drug metabolite investigation into toxicological research.

Quantitation of benzodiazepines in blood and urine using gas chromatography-mass spectrometry (GC-MS).

The benzodiazepine assay utilizes gas chromatography-mass spectrometry (GC-MS) for the analysis of diazepam, nordiazepam, oxazepam, temazepam, lorazepam, alpha-hydroxyalprazolam, and alpha-hydroxytriazolam in blood and urine. A separate assay is employed for the analysis of alprazolam. Prior to solid phase extraction, urine specimens are subjected to enzyme hydrolysis. The specimens are fortified with deuterated internal standard and a five-point calibration curve is constructed. Specimens are extracted by mixed-mode solid phase extraction. The benzodiazepine extracts are derivatized with N-methyl-N-(tert-butyldimethylsilyl)trifluoroacetamide (MTBSFTA) producing tert-butyldimethyl silyl derivatives; the alprazolam extracts are reconstituted in methanol without derivatization. The final extracts are then analyzed using selected ion monitoring GC-MS.

LC-MS/MS analysis of 13 benzodiazepines and metabolites in urine, serum, plasma, and meconium.

We describe a single method for the detection and quantitation of 13 commonly prescribed benzodiazepines and metabolites: alpha-hydroxyalprazolam, alpha-hydroxyethylflurazepam, alpha-hydroxytriazolam, alprazolam, desalkylflurazepam, diazepam, lorazepam, midazolam, nordiazepam, oxazepam, temazepam, clonazepam and 7-aminoclonazepam in urine, serum, plasma, and meconium. The urine and meconium specimens undergo enzyme hydrolysis to convert the compounds of interest to their free form. All specimens are prepared for analysis using solid-phase extraction (SPE), analyzed using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), and quantified using a three-point calibration curve. Deuterated analogs of all 13 analytes are included as internal standards. The instrument is operated in multiple reaction-monitoring (MRM) mode with an electrospray ionization (ESI) source in positive ionization mode. Urine and meconium specimens have matrix-matched calibrators and controls. The serum and plasma specimens are quantified using the urine calibrators but employing plasma-based controls. Oxazepam glucuronide is used as a hydrolysis control.

Simultaneous determination and quantification of 12 benzodiazepines in serum or whole blood using UPLC/MS/MS.

The benzodiazepines are a large, commonly prescribed family of psychoactive drugs. We describe a method permitting the simultaneous detection and quantification of 12 benzodiazepines in serum using ultra-performance liquid chromatography (UPLC) coupled with tandem mass spectrometry (MS/MS). Analytes included alprazolam, temazepam, oxazepam, nordiazepam, clonazepam, lorazepam, diazepam, chlordiazepoxide, midazolam, flunitrazepam, 7-aminoclonazepam, and 7-aminoflunitrazepam. Sample pretreatment is simple consisting of protein precipitation using cold acetonitrile (ACN) mixed with the deuterated internal standards. Samples were capped and vortexed for 5 min to ensure maximum precipitation. Following a 5-min centrifugation period, 400 microL of the supernatant was transferred to a clean tube and evaporated down under nitrogen. Samples were reconstituted in 200 microL of a deionized water:ACN (80:20) mixture and transferred to appropriate vials for analysis. Chromatographic run time was 7.5 min, and the 12 analytes were quantified using multiple reaction monitoring (MRM) and 6-point calibration curves constructed for each analyte at concentrations covering a clinically significant range.

Simultaneous analysis of diazepam and its metabolites in rat plasma and brain tissue by HPLC-UV and SPE.

Diazepam is frequently used as an adjuvant during antidepressant therapy. Recently, some studies have suggested that the treatment with benzodiazepines could have different efficacy in depressed patients as opposed to non-depressed ones. To clarify the matter, a study is currently underway, regarding the drug metabolism in rats. In order to obtain a more complete and significant set of data, the main diazepam metabolites have also been considered, namely: nordiazepam, temazepam and oxazepam. A feasible and reliable HPLC method has been developed for the simultaneous determination of these compounds in plasma and brain tissue of rats. The method has been applied to "normal" rats and to genetic rat models of depression in order to estimate drug metabolism in different breeds. Analyte separation was achieved on a C8 reversed phase column using an acidic phosphate buffer/acetonitrile mixture as the mobile phase. The detection wavelength was 238 nm. An original sample pre-treatment, based on solid-phase extraction (SPE) was developed in order to eliminate endogenous interference, using only 250 microL of matrix (brain homogenate or plasma) for a complete analysis. The method has been validated with good results in terms of precision, extraction yield, sensitivity, selectivity and accuracy on both matrices and has been successfully applied to samples from some rats subjected to the preliminary study. The obtained data will hopefully contribute to the clarification of possible differences between depressed and non-depressed subjects with respect to benzodiazepine biotransformation.

[Simultaneous determination of 5 sedative hypnotics in human plasma by reversed phase high-performance liquid chromatography].

OBJECTIVE: To determine diazepam, nitrazepam, oxazepam, estazolam, and alprazolam simultaneously in human plasma by reversed phase high-performance liquid chromatography (RP-HPLC). METHODS: Ten microliter carbamazepine (50 mg/L)as the internal standard was added into 1 mL sample, which contained the 5 mixed sedative hypnotics as standard substance and human plasma as ground substance. They were extracted with acetoacetate from plasma samples, and then were dissolved by 100 microL mobile phase. The blood drug levels were analyzed by high performance liquid chromatograph with 20 microL sample injection on a chromatographic column C18 (4.6 mm x 250 mm) at 30 degree. The mobile phase consisted of methanol and water (65:35),and the flow rate was 1.0 mL/min.The ultraviolet detection wavelength was 230 nm. RESULTS: The linearity range of the 5 drugs was 5-1,200 microg/L (r> or =0.9966, P<0.05). The recovery rate was 95.5%-105.6%. The extraction recovery rate was more than 75%. The relative standard deviation (RSD) of intra-day and inter-day was less than 10% (n=5). CONCLUSION: RP-HPLC method is convenient, accurate and sensitive for simultaneous determination of the concentration of diazepam, nitrazepam, oxazepam, estazolam, and alprazolam in human plasma.

Cardiac and peripheral blood similarities in the comparison of nordiazepam and bromazepam blood concentrations.

Concomitant heart and peripheral blood determinations were performed on 40 fatal cases involving nordiazepam (20 cases) and bromazepam (20 cases). The heart blood concentration for the two drugs (588 ng/mL for nordiazepam and 802 ng/mL for bromazepam) does not differ from the corresponding peripheral blood concentration (587 ng/mL for nordiazepam and 883 ng/mL for bromazepam). The mean ratios for the heart and peripheral blood concentrations were 0.95 for nordiazepam and 0.86 for bromazepam. No postmortem redistribution was observed for these two benzodiazepines. The authors thus suggest that corresponding heart blood can be proposed in the quantitative analysis of these drugs when peripheral blood is unavailable. The present study also shows the stability of the two drugs after a year of storage.

Quantitation of benzodiazepines in urine, serum, plasma, and meconium by LC-MS-MS.

A single method for confirmation and quantitation of a panel of commonly prescribed benzodiazepines and metabolites, alpha-hydroxyalprazolam, alpha-hydroxyethylflurazepam, alpha-hydroxytriazolam, alprazolam, desalkylflurazepam, diazepam, lorazepam, midazolam, nordiazepam, oxazepam, temazepam, clonazepam, and 7-aminoclonazepam, was developed for three specimen types, urine, serum/plasma, and meconium. Quantitation was by liquid chromatography tandem-mass spectrometry (LC-MS-MS) using a Waters Alliance-Quattro Micro system. The instrument was operated in multiple reaction monitoring mode with an electrospray ionization source in positive ionization mode. The method was evaluated for recovery, imprecision, linearity, analytical measurement range, specificity, and carryover. Average recovery and imprecision (within-run, between-run, and total % CV) were within +/- 15% of the target concentrations for urine (10 to 5000 ng/mL) and serum/plasma (10 to 2500 ng/mL) and within +/- 20% for meconium (10 to 5000 ng/g). In all, 205 patient specimens were analyzed, and the results compared to a previous in-house gas chromatography-MS method or LC-MS-MS results from an outside laboratory. Oxazepam glucuronide was evaluated as a hydrolysis control for the urine and meconium specimens.

Fatal intoxication with milnacipran.

The antidepressant milnacipran is a double serotonin/noradrenalin reuptake inhibitor. The low reported incidence of intoxication indicates excellent tolerance in comparison with tricyclic and second generation antidepressants. We report a fatal intoxication associating milnacipran, at blood levels (femoral=21.5 mg/l, cardiac=20 mg/l) 40-fold higher than the usual treatment concentration, and six other molecules (fluoxetine, norfluoxetine, sertraline, cyamemazine, nordazepam and oxazepam) at therapeutic levels. To the best of our knowledge, this is the first reported fatal intoxication involving milnacipran.

Associations between use of benzodiazepines or related drugs and health, physical abilities and cognitive function: a non-randomised clinical study in the elderly.

OBJECTIVE: To describe associations between the use of benzodiazepines or related drugs (BZDs/RDs) and health, functional abilities and cognitive function in the elderly. METHODS: A non-randomised clinical study of patients aged > or =65 years admitted to acute hospital wards during 1 month. 164 patients (mean age +/- standard deviation [SD] 81.6 +/- 6.8 years) were admitted. Of these, nearly half (n = 78) had used BZDs/RDs before admission, and the remainder (n = 86) were non-users. Cognitive ability was assessed by the Mini-Mental State Examination (MMSE). Patients scoring > or =20 MMSE sum points were interviewed (n = 79) and questioned regarding symptoms and functional abilities during the week prior to admission. Data on use of BZDs/RDs before admission, current medications and discharge diagnoses were collected from medical records. Health, physical abilities and cognitive function were compared between BZD/RD users and non-users, and adjustments were made for confounding variables. The residual serum concentrations of oxazepam, temazepam and zopiclone were analysed. RESULTS: The mean +/- SD duration of BZD/RD use was 7 +/- 7 years (range 1-31). Two or three BZDs/RDs were concomitantly taken by 26% of users (n = 20). Long-term use of these drugs was associated with female sex and use of a higher number of drugs with effects on the CNS, which tended to be related to diagnosed dementia. After adjustment for these variables as confounders, use of BZDs/RDs was not associated with cognitive function as measured by the MMSE. However, use of BZDs/RDs was associated with dizziness, inability to sleep after awaking at night and tiredness in the mornings during the week prior to admission and with stronger depressive symptoms measured at the beginning of the hospital stay. Use of BZDs/RDs tended to be associated with a reduced ability to walk and shorter night-time sleep during the week prior to admission. A higher residual serum concentration of temazepam correlated with a lower MMSE sum score after adjustment for confounding variables. CONCLUSIONS: Long-term use and concomitant use of more than one BZD/RD were common in elderly patients hospitalised because of acute illnesses. Long-term use was associated with daytime and night-time symptoms indicative of poorer health and potentially caused by the adverse effects of these drugs.