Pharmacokinetic properties of remimazolam in subjects with hepatic or renal impairment.

BACKGROUND: Remimazolam is a new benzodiazepine for procedural sedation and general anaesthesia. The aim of this study was to characterise its pharmacokinetic properties and safety in renally and hepatically impaired subjects. METHODS: Two separate trials were conducted in patients with hepatic (n=11) or renal impairment (n=11) compared with matched healthy subjects (n=9 and n=12, respectively). The hepatic impairment trial was an open-label adaptive 'Reduced Design' trial, using a single bolus of remimazolam 0.1 mg kg-1 i.v., whereas the renal impairment trial was an open-label trial of a single bolus dose of remimazolam 1.5 mg i.v. Remimazolam plasma concentrations over time were analysed by population pharmacokinetic modelling. RESULTS: Remimazolam pharmacokinetic properties were adequately described by a three-compartment, recirculatory model. Exposure in subjects with severe hepatic impairment was 38.1% higher (i.e. clearance was 38.1% lower) compared with healthy volunteers. This increase caused a slightly delayed recovery (8.0 min for healthy, 12.1 min for moderate, and 16.7 min for severe hepatic impairment). With renal impairment, plasma clearance was comparable with that measured in healthy subjects. Simulations of Cmax after a bolus dose of 10 mg showed no relevant impact of hepatic or renal impairment. The overall incidence of adverse events was low, and all adverse events were mild. CONCLUSIONS: As Cmax after a remimazolam bolus i.v. was not affected by hepatic or renal impairment, no dose adjustments are required. No unexpected adverse events related to remimazolam were seen in subjects with renal or hepatic impairment. CLINICAL TRIAL REGISTRATION: Hepatic impairment trial: ClinicalTrials.gov, NCT01790607 (https://clinicaltrials.gov/ct2/show/NCT01790607). Renal impairment trial: EudraCT Number: 2014-004575-23.

Metabolic diversity as a reason for unsuccessful detoxification from benzodiazepines: the rationale for serum BZD concentration monitoring.

PURPOSE: Many harms secondary to benzodiazepine (BZD) dependence force users towards detoxification treatment. However, even strongly motivated patients tolerate the process badly or experience early relapse. The detoxification procedure has not yet been standardized. The objective of this paper is to examine the hypothesis that faulty detoxification routines may have caused some failures. METHODS: The detoxification approaches found in the literature were compared stage by stage. The review was used to identify possible common, across-the-board systematic errors. RESULTS: The presented literature review confirms that the widespread divergence in the BZD metabolism rate is effectively neglected during detoxification routines. Without laboratory measurements, these differences, additionally interfered with by auxiliary drugs, undermine not only the scheduled but even the symptom-driven procedures. An initial substitution with a long-acting BZD, although recommended, may lead to over-accumulation. This excess, varying between patients and incompatible with the current tapering stage, may lead to repeated overestimation of the patient's adjustments to reduced doses. Consequently, the patient's good clinical presentation at withdrawal, resulting in a conclusion of detoxification, may actually reflect a persistently high serum BZD concentration. The low-concentration stage, if shifted past the end of treatment, exposes patients to unexpected, unassisted withdrawal crises. With laboratory feedback, these crises, unlike the symptoms related to deficient re-adaptation mechanisms, could be prevented. Moreover, by minimizing the high-concentration phase, time can be saved for properly assisted low-concentration challenges. CONCLUSION: A customized detoxification procedure driven not only by the intensity of withdrawal symptoms but also by serum BZD monitoring may prevent some failures. As the standard regimen, it would make detoxification from BZDs more reliable and effective.

Effects of Vitamin D Receptor, Cytochrome P450 3A, and Cytochrome P450 Oxidoreductase Genetic Polymorphisms on the Pharmacokinetics of Remimazolam in Healthy Chinese Volunteers.

Remimazolam is a new ultra-short-acting benzodiazepine used to induce and maintain anesthesia and procedural sedation. Its compound structure is similar to midazolam's. Midazolam metabolism might be affected by vitamin D receptor (VDR), cytochrome P450 3A, and cytochrome P450 oxidoreductase genetic polymorphisms. This study investigated the effects of VDR, cytochrome P450 3A, and cytochrome P450 oxidoreductase genetic polymorphisms on the pharmacokinetics of remimazolam in healthy Chinese volunteers after a single intravenous injection of remimazolam besylate. Blood samples were collected from subjects (n = 62) at scheduled time intervals before and after injection. High-performance liquid chromatography-tandem mass spectrometry was used to quantify plasma remimazolam and RF7054 (its inactive carboxylic acid metabolite) concentrations. The relationship between plasma remimazolam concentration, pharmacokinetic parameters, and polymorphic alleles was assessed for each subject. The rs4516035 allele affected the elimination half-life of RF7054 (P = .043), while the rs1544410 allele affected the dose-normalized maximum observed plasma concentration (Cmax /D) of remimazolam (P = .025) in 46 volunteers. Results showed that VDR genetic polymorphisms might affect the pharmacokinetics of remimazolam in the Chinese population.

Supramolecular solvent (SUPRASs) extraction method for detecting benzodiazepines and zolpidem in human urine and blood using gas chromatography tandem mass spectrometry.

OBJECTIVE: A high-throughput and sensitive method using supramolecular solvent (SUPRASs) for detecting 9 benzodiazepines and zolpidem in human urine and blood by gas chromatography-tandem mass spectrometry (GC-MS/MS) was newly established and applied to authentic human urine and blood samples in this study. METHODS: Urine and blood samples were subjected to liquid-liquid extractions with supramolecular solvent mixture which consists of tetrahydrofuran and 1-hexanol. The solvent layer was evaporated to dryness by stream of nitrogen. The residue was reconstituted with methanol, and subjected to analysis by GC-MS/MS in multiple reaction monitoring (MRM) mode; internal standard method was employed for quantifying of each targeted compound. RESULTS: The regression equation has a good linear relationship with correlation coefficients for all tested compounds were not lower than 0.9991. The lower limits of the quantification ranged from 0.20 to 5 ng/mL for tested compounds in urine; Meanwhile, the lower limits of the quantification in this method ranged from 1 to 50 ng/mL for tested compounds in blood. These results showed that excellent reproducibility and satisfactory extraction recovery rates could be obtained for the established analytical method for 10 drugs in both blood and urine samples. CONCLUSION: The established method in this study was high-throughput, simple and sufficiently sensitive for determining of benzodiazepinesand zolpidem in human urine and blood. Therefore, this newly established method could be of use for qualitative and quantitative determination of such drugs in urine and blood samples either for clinical poisoning monitoring or for forensic identification.

Identification and Quantification of Antipsychotics in Blood Samples by LC-MS-MS: Case Reports and Data from Three Years of Routine Analysis.

Antipsychotic drugs (AP) are widely prescribed for the treatment of schizophrenia and psychosis. The pharmacological treatment of schizophrenia is often performed with the simultaneous use of two or more antipsychotic agents to achieve the desired control of psychotic symptoms Available AP include both conventional (typical) and new (atypical) antipsychotic medications. Atypical AP, such as quetiapine, now account for the vast majority of AP prescriptions. In forensic toxicology, AP are of considerable interest because of their potential abuse and their involvement in intoxications and suicides. The authors retrospectively examined AP positive cases detected in samples collected during autopsies performed in the Forensic Clinical and Pathology Service of National Institute of Legal Medicine and Forensic Sciences Centre Branch or in other autopsies carried out in the central region of Portugal, between January 2016 and December 2018. A quantitative liquid chromatography-tandem mass spectrometry assay was developed for the simultaneous determination of 16 AP (amisulpride, aripiprazole, chlorpromazine, clozapine, cyamemazine, fluphenazine, haloperidol, levomepromazine, melperone, olanzapine, paliperidone, promethazine, quetiapine, risperidone, sulpiride and ziprasidone) in blood samples of postmortem cases. The Laboratory of Forensic Chemistry and Toxicology received 3,588 requests for toxicological analysis: 1,413 cases were positive for drugs from which 351 (24.8%) cases were positive for AP, 60.1% from male individuals and 39.9% from female. Quetiapine was the most prevalent AP (36.5%) followed by olanzapine (20.8%). During this period, there were 25 postmortem cases with AP blood concentrations above therapeutic range, in which 36% of those are in agreement with the information received (psychological history or acute intoxication suspicion) and the manner of death was suicide. Our results point that antipsychotics are an increasingly prevalent class of drugs. AP must be measured not only in toxic concentrations but also in therapeutic levels in postmortem cases; therefore, it is important to come up with a sensitive method to cover the low therapeutic range in which AP are usually present.

Flubromazolam: Detection in five post-mortem cases.

Flubromazolam is a potent triazole benzodiazepine with moderately long-lasting central nervous system-depressant effects relative to other benzodiazepines such as commonly prescribed diazepam. Flubromazolam has been studied in the living. However, there are no published reports including measured drug concentrations in post-mortem cases. We report five cases in which flubromazolam was detected in a systematic screen using high-resolution mass spectrometry and then quantified in femoral blood. In none of the five cases was the cause of death directly attributed to flubromazolam toxicity, as there was a variety of both sedative and stimulant drugs also present. However, it is important that the drug concentrations that were measured are made available for future post-mortem forensic interpretation.

Development and Validation of an LC-MS-MS Method for the Detection of 40 Benzodiazepines and Three Z-Drugs in Blood and Urine by Solid-Phase Extraction.

An analytical method for the detection of 40 benzodiazepines, (±)-zopiclone, zaleplon and zolpidem in blood and urine by solid-phase extraction liquid chromatography-tandem mass spectrometry was developed and validated. Twenty-nine of 43 analytes were quantified in 0.5 mL whole blood for investigating postmortem, drug-facilitated sexual assault (DFSA) and driving under the influence of drugs cases (DUID). The four different dynamic ranges of the seven-point, linear, 1/x weighted calibration curves with lower limits of quantification of 2, 5, 10 and 20 µg/L across the analytes encompassed the majority of our casework encountered in postmortem, DFSA and DUID samples. Reference materials were available for all analytes except α-hydroxyflualprazolam, a hydroxylated metabolite of flualprazolam. The fragmentation of α-hydroxyflualprazolam was predicted from the fragmentation pattern of α-hydroxyalprazolam, and the appropriate transitions were added to the method to enable monitoring for this analyte. Urine samples were hydrolyzed at 55°C for 30 min with a genetically modified ß-glucuronidase enzyme, which resulted in >95% efficiency measured by oxazepam glucuronide. Extensive sample preparation included combining osmotic lysing and protein precipitation with methanol/acetonitrile mixture followed by freezing and centrifugation resulted in exceptionally high signal-to-noise ratios. Bias and between-and within-day imprecision for quality controls (QCs) were all within ±15%, except for clonazolam and etizolam that were within ±20%. All 29 of the 43 analytes tested for QC performance met quantitative reporting criteria within the dynamic ranges of the calibration curves, and 14 analytes, present only in the calibrator solution, were qualitatively reported. Twenty-five analytes met all quantitative reporting criteria including dilution integrity. The ability to analyze quantitative blood and qualitative urine samples in the same batch is one of the most useful elements of this procedure. This sensitive, specific and robust analytical method was routinely employed in the analysis of >300 samples in our laboratory over the last 6 months.

Pharmacokinetic Variability During Long-Term Therapeutic Drug Monitoring of Valproate, Clobazam, and Levetiracetam in Patients With Dravet Syndrome.

BACKGROUND: The use of therapeutic drug monitoring (TDM) for antiseizure medications (ASMs) may contribute to treatment optimization in individual patients. This study included patients with Dravet syndrome as they often require close monitoring because of polypharmacy with various ASMs. The aim was to use long-term TDM to investigate pharmacokinetic variability of ASMs in these patients. METHODS: Retrospective data from patients with Dravet syndrome were collected from the TDM database at the Section for Clinical Pharmacology, National Center for Epilepsy in Norway (2008-2018). Concentration/(dose/kg)ratios (C/D ratios) were calculated for the ASMs and the concentration (C/C ratio) for N-desmethylclobazam. In patients with at least 3 measurements, the CV for C/D ratios for intrapatient and interpatient variability was calculated. RESULTS: Fifty-three patients (30 male patients/23 female patients) between 2 and 50 years of age (mean, 16 years) were included. Pharmacokinetic variability of the total number of measurements of valproate (n = 417), clobazam and N-desmethylclobazam (n = 328), and levetiracetam (n = 238) was determined. Interpatient variability was more pronounced than intrapatient variability (coefficient of variations: valproate, 65% vs. 24%; levetiracetam, 71% vs. 27%; and clobazam/N-desmethylclobazam, 47%/77% vs. 35%/55%) (P < 0.01). Comedication with stiripentol (n = 16) increased the C/D ratio of valproate by 63% and of clobazam by 133% and the C/C ratio of N-desmethylclobazam/clobazam by 104% (P < 0.05). Younger age also contributed to pharmacokinetic variability. CONCLUSIONS: Long-term TDM revealed extensive variability in serum concentrations over time; the variability was lowest for levetiracetam, moderate for valproate, and highest for clobazam. Pharmacokinetic variability and interactions can thus be identified and adjusted to facilitate decision making to achieve the optimal treatment outcome.

Blood Concentrations of Designer Benzodiazepines: Relation to Impairment and Findings in Forensic Cases.

The use of designer benzodiazepines appears to be increasing in many countries, but data concerning blood concentrations are scarce, making interpretation of concentrations difficult. The aim of this study was to report blood concentrations of clonazolam, diclazepam, etizolam, flualprazolam, flubromazepam, flubromazolam and phenazepam and to investigate the relationship between blood concentrations and impairment. The concentration data are from blood samples collected from living cases (apprehended drivers and other drug offences) and medico-legal autopsies. The blood samples were analysed for the seven designer benzodiazepines mentioned above by ultra high performance liquid chromatography-tandem mass spectrometry. Positive cases from between 1 June 2016 and 30 September 2019 were included. Blood concentrations and the conclusion from a clinical test of impairment (when available) are reported. The presented seven benzodiazepines were detected in a total of 575 cases, where 554 of these cases concerned apprehended drivers or other criminal offenders. The number of findings and the median (range) concentrations were as follows: clonazolam, n = 22, 0.0041 mg/L (0.0017-0.053 mg/L); diclazepam, n = 334, 0.0096 mg/L (0.0016-0.25 mg/L); etizolam, n = 40, 0.054 mg/L (0.015-0.30 mg/L); flualprazolam, n = 10, 0.0080 mg/L (0.0033-0.056 mg/L); flubromazepam, n = 5, 0.037 mg/L (0.0070-0.70 mg/L); flubromazolam, n = 20, 0.0056 mg/L (0.0004-0.036 mg/L); and phenazepam, n = 138, 0.022 mg/L (0.0018-0.85 mg/L). A designer benzodiazepine was the only drug detected with relevance for impairment in 25 of the 554 living cases. The physician concluded with impairment in 19 of the 25 cases. Most of the concentrations in these cases were relatively similar to or higher than the median reported concentrations. The most frequent other drugs detected were amphetamine, tetrahydrocannabinol, clonazepam and methamphetamine. The presented blood concentrations can be helpful with the interpretation of cases involving one or more of these seven benzodiazepines. The results indicate that concentrations commonly observed in forensic cases are associated with impairment.

Multifaceted Bioanalytical Methods for the Comprehensive Pharmacokinetic and Catabolic Assessment of MEDI3726, an Anti-Prostate-Specific Membrane Antigen Pyrrolobenzodiazepine Antibody-Drug Conjugate.

Complex biotherapeutic modalities, such as antibody-drug conjugates (ADC), present significant challenges for the comprehensive bioanalytical characterization of their pharmacokinetics (PK) and catabolism in both preclinical and clinical settings. Thus, the bioanalytical strategy for ADCs must be designed to address the specific structural elements of the protein scaffold, linker, and warhead. A typical bioanalytical strategy for ADCs involves quantification of the Total ADC, Total IgG, and Free Warhead concentrations. Herein, we present bioanalytical characterization of the PK and catabolism of a novel ADC. MEDI3726 targets prostate-specific membrane antigen (PMSA) and is comprised of a humanized IgG1 antibody site-specifically conjugated to tesirine (SG3249). The MEDI3726 protein scaffold lacks interchain disulfide bonds and has an average drug to antibody ratio (DAR) of 2. Based on the structural characteristics of MEDI3726, an array of 4 bioanalytical assays detecting 6 different surrogate analyte classes representing at least 14 unique species was developed, validated, and employed in support of a first-in-human clinical trial (NCT02991911). MEDI3726 requires the combination of heavy-light chain structure and conjugated warhead to selectively deliver the warhead to the target cells. Therefore, both heavy-light chain dissociation and the deconjugation of the warhead will affect the activity of MEDI3726. The concentration-time profiles of subjects dosed with MEDI3726 revealed catabolism of the protein scaffold manifested by the more rapid clearance of the Active ADC, while exhibiting minimal deconjugation of the pyrrolobenzodiazepine (PBD) warhead (SG3199).

Fatal α-PVP and amphetamine poisoning during a sauna and autoerotic practices.

We describe the sudden death of a middle-aged man while having a sauna under the influence of α-pyrrolidinovalerophenone (α-PVP) (PM blood concentration: 0.8 mg/L), amphetamine (0.34 mg/L), and other drugs (buprenorphine, benzodiazepines), and engaging in solitary sexual activities. The drugs' effects on the cardio-circulatory system and on body thermoregulation combined with the high temperatures are likely to have been central mechanisms leading to death. The high levels of adrenaline triggered by sexual arousal and the respiratory depression caused by buprenorphine, in association with benzodiazepines, may have also contributed to his death. This previously unreported type of accidental autoerotic death illustrates the risk of using amphetamine-like sympathomimetic drugs (e.g. cathinone derivates) in hot environments such as a sauna, and during sexual activities therein.

The implementation of per-se limits for driving under the influence of benzodiazepines and related drugs: No increased risk for arrest during therapeutic use in Norway.

Objective: To investigate whether the use of recommended therapeutic doses of medicinal drugs has led to suspicion of driving under the influence of drugs (DUID) after implementation of legislative limits for illicit and medicinal drugs in 2012.Methods: Data from suspected drug-impaired drivers apprehended by the police from 2013 to 2015 were selected from the Norwegian Forensic Toxicology Database. The blood samples had been analyzed for benzodiazepines (BZDs), z-hypnotics, opioids, stimulants, certain hallucinogens, and alcohol. Drivers who tested positive for one BZD or a z-hypnotic only, were included in the study. Drug concentrations measured in their blood samples were compared to the maximal obtainable steady state concentrations if the drug had been used in accordance with the recommendations set by the Norwegian Directorate of Health.Results: BZDs or z-hypnotics were found in 10 248 samples, representing 59.6% of the total number of drivers arrested for suspected DUID (n = 17 201). Only one BZD or z-hypnotic with a blood drug concentration above the legislative limit was detected in 390 (2.3%) of the total number of samples. Clonazepam was the most frequently detected BZD (n = 4656), while as a single drug above the legislative limit, it was detected in only 3.6% (n = 168) of the clonazepam-positive blood samples. For drivers testing positive for only one z-hypnotic, drug concentrations above the legislative limit were found in 27% (n = 55) of the blood samples that tested positive for zolpidem and 12.4% (n = 53) of the samples that tested positive for zopiclone. In total, 155 subjects out of 10 248 testing positive for BZDs or z-hypnotics displayed concentrations above the legislative limit but within the concentration ranges that are expected when taking recommended therapeutic drug doses, and 77 below the legislativel limit.Conclusions: The results show that the implementation of legislative limits for BZDs and z-hypnotics may have contributed to DUID suspicion for a small group of patients using therapeutic drug doses; only 1.3% of the suspected DUID offenders had concentrations of only one of those drugs in-line with recommended therapeutic dosing.

Clonazolam a new designer benzodiazepine intoxication confirmed by blood concentration.

BACKGROUND: Recently the number of new psychoactive substances have significantly increased, becoming popular among experienced users of designer drugs. A significant group includes benzodiazepine derivatives, which have not been introduced as medications but are abused by people experimenting with new and classical psychoactive substances. CASE PRESENTATION: The aim of this paper was to present the case of a clonazolam ingestion by a person who was not habituated to benzodiazepines. The intake caused only prolonged coma, decreased muscle tone, and deep tendon reflexes without any other concomitant toxicity and cardio-respiratory failure. CONCLUSIONS: Clonazolam concentrations in patient's blood, measured three times were 0.077 mg/L, 0.015 mg/L, 0.009 mg/L after 4, 8 and 12 h, respectively. Clonazolam's human toxicity has not been well established, so any case of poisoning should be closely monitored.

Toxicokinetics and Analytical Toxicology of Flualprazolam: Metabolic Fate, Isozyme Mapping, Human Plasma Concentration and Main Urinary Excretion Products.

An increasing number of benzodiazepine-type compounds are appearing on the new psychoactive substances market. 8-Chloro-6-(2-fluorophenyl)-1-methyl-4H-[1,2,4]triazolo[4,3-a][1,4]benzodiazepine (well known as flualprazolam) represents a potent 'designer benzodiazepine' that has been associated with sedation, loss of consciousness, memory loss and disinhibition. The aims of the present study were to tentatively identify flualprazolam metabolites using in vitro incubations with pooled human liver S9 fraction or HepaRG cells by means of liquid-chromatography-high resolution tandem mass spectrometry. Isozymes involved in phase I and II biotransformation were identified in vitro. Results were then confirmed using human biosamples of an 18-year old male who was admitted to the emergency department after suspected flualprazolam ingestion. Furthermore, the plasma concentration was determined using the standard addition method. Seven flualprazolam metabolites were tentatively identified. Several cytochrome P450 and UDP-glucuronosyltransferase isozymes, amongst them CYP3A4 and UGT1A4, were shown to be involved in flualprazolam biotransformation reactions, and an influence of polymorphisms as well as drug-drug or drug-food interactions cannot be excluded. Alpha-hydroxy flualprazolam glucuronide, 4-hydroxy flualprazolam glucuronide and the parent glucuronide were identified as most abundant signals in urine, far more abundant than the parent compound flualprazolam. These metabolites are thus recommended as urine-screening targets. If conjugate cleavage was performed during sample preparation, the corresponding phase I metabolites should be added as targets. Both hydroxy metabolites can also be recommended for blood screening. The flualprazolam plasma concentration determined in the intoxication case was as low as 8 µg/L underlining the need of analytical methods with sufficient sensitivity for blood-screening purposes.

Pharmacokinetics and Pharmacodynamics of Remimazolam (CNS 7056) after Continuous Infusion in Healthy Male Volunteers: Part I. Pharmacokinetics and Clinical Pharmacodynamics.

BACKGROUND: Remimazolam (CNS 7056) is a new ultra-short-acting benzodiazepine for intravenous sedation and anesthesia. Its pharmacokinetics and pharmacodynamics have been reported for bolus administration. This study aimed to investigate the pharmacokinetics and pharmacodynamics of remimazolam after continuous infusion. METHODS: Twenty healthy male volunteers (20 to 38 yr, 64 to 99 kg) received remimazolam as continuous intravenous infusion of 5 mg/min for 5 min, 3 mg/min for the next 15 min, and 1 mg/min for further 15 min. Pharmacokinetics of remimazolam and its metabolite were determined from arterial plasma concentrations. Sedation was assessed using the Modified Observer's Assessment of Alertness and Sedation scale. Pharmacokinetic-pharmacodynamic modeling was performed by population analysis. Hemodynamics and the electrocardiogram were also investigated. RESULTS: Pharmacokinetics was best described by a three-compartment model for remimazolam and a two-compartment model with transit compartment for the metabolite. Remimazolam showed a high clearance (1.15 ± 0.12 l/min, mean ± SD), a small steady-state volume of distribution (35.4 ± 4.2 l) and a short terminal half-life (70 ± 10 min). The simulated context-sensitive halftime after an infusion of 4 h was 6.8 ± 2.4 min. Loss of consciousness was observed 5 ± 1 min after start, and full alertness was regained 19 ± 7 min after stop of infusion. Pharmacodynamics of Modified Observer's Assessment of Alertness and Sedation score was best described by a sigmoid probability model with effect site compartment. The half-maximum effect site concentration for a Modified Observer's Assessment of Alertness and Sedation score less than or equal to 1 was 695 ± 239 ng/ml. The equilibration half-time between central and effect compartment was 2.7 ± 0.6 min. Mean arterial blood pressure decreased by 24 ± 6%, and heart rate increased by 28 ± 15%. Spontaneous breathing was maintained throughout the study. There was no significant prolongation of the QT interval of the electrocardiogram observed. CONCLUSIONS: Remimazolam was characterized by a pharmacokinetic-pharmacodynamic profile with fast onset, fast recovery, and moderate hemodynamic side effects.

Pharmacokinetics and Pharmacodynamics of Remimazolam (CNS 7056) after Continuous Infusion in Healthy Male Volunteers: Part II. Pharmacodynamics of Electroencephalogram Effects.

BACKGROUND: Remimazolam (CNS 7056) is a new ultra-short acting benzodiazepine for IV sedation. This study aimed to investigate the electroencephalogram (EEG) pharmacodynamics of remimazolam infusion. METHODS: Twenty healthy male volunteers received remimazolam as continuous IV infusion of 5 mg/min for 5 min, 3 mg/min for the next 15 min, and 1 mg/min for further 15 min. Continuous EEG monitoring was performed by a neurophysiologic system with electrodes placed at F3, F4, C3, C4, O1, O2, Cz, and Fp1 (10/20 system) and using the Narcotrend Index. Sedation was assessed clinically by using the Modified Observer's Assessment of Alertness and Sedation scale. Pharmacodynamic models were developed for selected EEG variables and Narcotrend Index. RESULTS: EEG changes during remimazolam infusion were characterized by an initial increase in beta frequency band and a late increase in delta frequency band. The EEG beta ratio showed a prediction probability of Modified Observer's Assessment of Alertness and Sedation score of 0.79, and could be modeled successfully using a standard sigmoid Emax model. Narcotrend Index showed a prediction probability of Modified Observer's Assessment of Alertness and Sedation score of 0.74. The time course of Narcotrend Index was described by an extended sigmoid Emax model with two sigmoid terms and different plasma-effect equilibration times. CONCLUSIONS: Beta ratio was identified as a suitable EEG variable for monitoring remimazolam sedation. Narcotrend Index appeared less suitable than the beta ratio for monitoring the sedative effect if remimazolam is administered alone.

Protein precipitation method for determination of clobazam and N-desmethylclobazam in human plasma by LC-MS/MS.

A protein precipitation method for the determination of clobazam (CLB) and its major active metabolite N-desmethylclobazam (N-CLB) in human plasma by liquid chromatography tandem mass spectrometry (LC-MS/MS) was established. CLB and N-CLB were extracted from human plasma samples by protein precipitation with methanol. Analyte separation was done using a Phenomenex Kinetex™ Biphenyl (50 × 2.1 mm, 1.7 µm) column using isocratic elution with a mobile phase of 5 mm ammonium formate with 0.01% ammonium hydroxide (40%) and methanol (60%) at a flow rate of 0.4 mL/min and an injection volume of 10 µL. The detection was performed on a triple quadrupole mass spectrometer in multiple reaction monitoring mode to monitor the precursor-to-product ion transitions of m/z 301.1 → 259.0, 306.0 → 263.9 for CLB and CLB-D5 and 287.0 → 245.0, 292.0 → 250.0 for N-CLB and N-CLB-D5 in positive electrospray ionization mode, respectively. The method was validated over a concentration range of 2.0-750 ng/mL for CLB and 0.7-200 ng/mL for N-CLB on SCIEX Triple Quad 4500 MS System. Total run time was 5 min. This method has been designed for bioequivalence study for formulations containing 20 mg of CLB.

Odds of culpability associated with use of impairing drugs in injured drivers in Victoria, Australia.

Culpability analysis was conducted on 5000 drivers injured as a result of a vehicular collision and in whom comprehensive toxicology testing in blood was conducted. The sample included 1000 drivers for each of 5 years from approximately 5000-6000 drivers injured and taken to hospital in the State of Victoria. Logistic regression was used to investigate differences in the odds of culpability associated with alcohol and drug use and other selected crash attributes using the drug-free driver as the reference group. Adjusted odds ratios were obtained from multivariable logistic regression models in which other potentially explanatory driver and crash attributes were included. Drivers with alcohol present showed large increases in the odds of culpability similar to that seen in other studies investigating associations between blood alcohol concentration and crash risk. Methylamphetamine also showed a large increase in the odds of culpability (OR 19) compared to the reference group at both below and above 0.1 mg/L, whereas those drivers with Δ9-tetrahydrocannabinol (THC) present showed only modest increase in odds when all concentrations were assessed (OR 1.9, 95 %CI 1.2-3.1). Benzodiazepines in drivers also gave an increase in odds (3.2, 95 %CI 1.6-6.1), but not other medicinal drugs such as antidepressants, antipsychotics and opioids. Drivers that had combinations of impairing drugs generally gave a large increase in odds, particularly combinations of alcohol with THC or benzodiazepines, and those drivers using both THC and methamphetamine.

Femoral blood concentrations of flualprazolam in 33 postmortem cases.

Flualprazolam is a novel designer benzodiazepine, structurally related to alprazolam, flubromazolam and triazolam. In the last couple of years, it has been frequently detected in seizures and in forensic cases in Sweden and Finland. However, there is a lack of published blood concentrations for the drug, which presents difficulties when assessing its relevance for the cause of death. A quantitative method for the determination of flualprazolam in post-mortem blood was developed and validated, and subsequently used to analyse samples from 33 deaths previously screened as testing positive for flualprazolam in Sweden and Finland. Most of the cases in the study were accidental deaths (61 %) or suicides (18 %). The median (range) flualprazolam concentration was 18.0 (3.0-68) ng/g. The majority of the deceased were male (82 %) and the median age was 30 years. The median age in the Swedish cases was significantly higher (35 years) than in the Finnish cases (23 years) (p< 0.05). Poly-drug use and particularly the concomitant use of flualprazolam and opioids were very common in the study population. Most of the cases that were positive for flualprazolam were fatal poisonings by a drug (N=23), and in 13 cases, flualprazolam was implicated in the cause of death. Combining the resources of two countries in which all post-mortem toxicology is centralised provided a more comprehensive insight into the toxicology of flualprazolam. Research on novel psychoactive substances, such as flualprazolam, is required in order to be able to provide scientific evidence on the risks of these new substances for drug administration and potential users.

The challenge that chlorine presented during the development of a benzodiazepine assay.

During the routine validation of a benzodiazepine method (performed on a Liquid Chromatography - Tandem Mass Spectrometer), it was noted that lorazepam, triazolam, and α-hydroxytriazolam showed a quadratic shift/bias in the calibration curve, particularly at high concentrations. The ultimate cause of this bias was determined to be due to the natural presence of chlorine (Cl) isotopes (35Cl and 37Cl) in these benzodiazepines. The presence of the heavy (37Cl) isoforms of Cl resulted in the analyte's mass being the same as the internal standard which, in turn, caused the internal standard to appear "falsely increased", thus skewing the calibration curve. One solution to this potential issue was to take advantage of this natural phenomenon and use the Cl heavy isoforms of the respectively labeled internal standards.