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Designer benzodiazepine groups, their reference classic benzodiazepine and their possible transformations: Asterisk indicates a possible removal of the chemical group and the arrow indicates a possible hydroxylation
Source publication
Designer benzodiazepines (DBZDs) have become of particular importance in the past few years. The metabolite monitoring of DBZD in biological fluids could be of great interest in clinical and forensic toxicology. However, DBZD metabolites are not known or not commercially available. The identification of some DBZD metabolites has been mostly explore...
Citations
... Refs. [74][75][76][77][78][79][80][81][82][83][84][85][86][87][88][89][90] ...
Benzodiazepines (BZDs), potent sedative and hypnotic drugs widely prescribed in psychiatry, pose a high risk of dependence and are globally abused. This study used wastewater-based epidemiology to investigate the consumption patterns of BZDs across four wastewater treatment plants (WWTPs) in Wuhu, China. A total of 16 BZDs and three metabolites were detected in influents and effluents, with concentrations reaching up to 90 ng/L (quetiapine fumarate) and 18.4 ng/L (diazepam). Most BZDs had a poor removal efficiency except quetiapine fumarate (>98% removal). The consumptions of BZDs in WWTPs ranged from <0.02 (lormetazepam) to 2700 mg/day/1000 people (quetiapine fumarate). Seasonal variation was found in BZD usage, where the consumptions in winter and spring were significantly higher than those in summer and autumn. It was worth noting that nimetazepam may be abused during the sampling campaign. Urban areas with higher housing prices match higher BZD consumption, correlating with greater stress and insomnia rates. This study reveals the relationship between socioeconomic factors and BZD consumption patterns, provide a new path to addressing community public health.
... Minor biotransformed reactions such as dehalogenation including debromination or dechlorination, desalkylation, and combinations of these minor reactions and the previously described reactions were also identified. el Balkhi et al. (2017) reported desnitronifoxipam and a metabolite of etizolam resulting from aldehyde formation in an in vitro investigation with pHlM. Ameline et al. (2019), also detected desnitroflunitrazolam in human urine. ...
Synthesis and illicit use of designer benzodiazepines are growing concerns, with these new psychoactive substances (NPS) posing serious health consequences and new hurdles for toxicologists. Consumption marker identification and characterization is paramount in confirming their use. The benzodiazepine core structure is a fusion of benzene and a seven-membered heterocycle with two nitrogen atoms forming the diazepine ring. Minor variations on the core structure produce different classes of benzodiazepines with marked differences in physiological effects. The present review provides a comprehensive designer benzodiazepines metabolism overview and suggests suitable human consumption biomarkers for toxicology casework. A systematic literature search of PubMed®, ScopusTM, Web of ScienceTM, and Cochrane databases was conducted independently by two coauthors adhering to PRISMA guidelines. Data from 30 in vitro and in vivo models of designer benzodiazepines metabolism from January 2007 to February 2023 were included. 1,4-benzodiazepines (n = 10), 2,3-benzodiazepines (n = 1), triazolo-benzodiazepines (n = 9), and thieno-triazolo-benzodiazepines (n = 3) study design, sample pretreatment, analytical techniques, and major metabolites detected in various matrices are addressed. Metabolites following hydroxylation and phase II glucuronide conjugation were the most prevalent analytes. N-Glucuronidation of parent azole-fused benzodiazepines, and nitro-reduced and N-acetylated metabolites of nitro-containing designer benzodiazepines were also common. From these data, we propose a generic metabolic pathway for designer benzodiazepines. The sporadic illicit market presents challenges in toxicological casework and necessitates comprehensive biomarker investigations, especially in cases with legal implications. There are few metabolism data for many designer benzodiazepines, emphasizing the need for research focusing on closing these gaps.
... In a case reported by Balkhi et al. (30), the corpse of a 31-yearold male drug abuser was found with injection materials and small plastic bags containing various DBZs, including deschloroetizolam. Autopsy revealed multi-organ congestion, likely of toxic origin, with no evident natural diseases or traumatic lesions related to death. ...
This review article takes a closer look at a new class of psychoactive substances called designer benzodiazepines (DBZs) and the challenges of their detection. These are adinazolam, clonazolam, deschloroetizolam, diclazepam, etizolam, flualprazolam, flubromazepam, flubromazolam, phenazepam, and pyrazolam. They are central nervous system depressants and sedatives that can cause psychomotor impairment and increase the overdose risk when combined with other sedatives. DBZs undergo phase I and II metabolism similar to traditional benzodiazepines, but their specific metabolic pathways and the influence of genetic polymorphisms are yet to be clarified. Advances in liquid chromatography-tandem mass spectrometry (LC-MS/MS) have enhanced the method's sensitivity for DBZs and their metabolites in biological samples and coupled with improved blood sampling methods require less blood for drug monitoring. Further research should focus on elucidating their pharmacokinetic properties and metabolism in humans, especially in view of genetic polymorphisms and drug interactions that could inform clinical treatment choices. Even though we have witnessed important advances in DBZ detection and measurement, further refinements are needed to expand the scope of detectable DBZs and their metabolites. All this should help toxicological research to better identify and characterise the risks of chronic and polydrug abuse and facilitate clinical, forensic, and regulatory responses to this growing issue.
... Currently, there are few reports on the detection of etizolam and its metabolites, the instruments used for etizolam detection are mainly limited to LC-MS/MS, GC-MS/MS and HPLC-MS [8][9][10]. Compared with these three traditional analytical instruments, UPLC-Q-Exactive-MS has higher column efficiency and better separation effect, but there is no relevant literature report on the detection of etizolam and its metabolites. ...
... mRNA levels, intracellular protein levels, secreted/released proteins (e.g., cytokine), and the type of cell death (e.g., necrosis or apoptosis) can be determined with PCR [88,89], Western blot [90,91], slot blot [66], ELISA [92][93][94], MALDI/ESI-MS [90], microscope [95,96], or flow cytometry [95,[97][98][99]. Specific investigation methods such as the measurement of membrane potential, DNA biosynthesis assessment, cell viability assay, and enzyme assay in microsomes can also be applied because researchers can obtain each reagent and instrument required for examination [100][101][102][103]. Extracellular AGEs may be able to promote ulcers in three kinds of epithelial cells via the AGEs-RAGE/TLR4 axis, although this may also be caused by other factors (such as anti-cancer drug side effects [7]). ...
Kampo medicines are Japanese traditional medicines developed from Chinese traditional medicines. The action mechanisms of the numerous known compounds have been studied for approximately 100 years; however, many remain unclear. While components are normally affected through digestion, absorption, and metabolism, in vitro oral, esophageal, and gastric epithelial cell models avoid these influences and, thus, represent superior assay systems for Kampo medicines. We focused on two areas of the strong performance of this assay system: intracellular and extracellular advanced glycation end-products (AGEs). AGEs are generated from glucose, fructose, and their metabolites, and promote lifestyle-related diseases such as diabetes and cancer. While current technology cannot analyze whole intracellular AGEs in cells in some organs, some AGEs can be generated for 1–2 days, and the turnover time of oral and gastric epithelial cells is 7–14 days. Therefore, we hypothesized that we could detect these rapidly generated intracellular AGEs in such cells. Extracellular AEGs (e.g., dietary or in the saliva) bind to the receptor for AGEs (RAGE) and the toll-like receptor 4 (TLR4) on the surface of the epithelial cells and can induce cytotoxicity such as inflammation. The analysis of Kampo medicine effects against intra/extracellular AGEs in vitro is a novel model.
... There have also been reports on the metabolic patterns of Etizolam in vivo and in vitro. Balkhi [7] et al. simulated the in vitro metabolism of Etizolam using human liver microsomes and identified five metabolites. Erin [8] et al. studied the metabolites of Etizolam in horses using two metabolic models, namely, horses and horse liver microsomes, and identified 23 metabolites with α-OH-Etizolam as a potential metabolic marker. ...
As one of the most widely abused designer benzodiazepines worldwide, Etizolam is characterized by its high addiction potential, low production cost, and difficulty in detection. Due to the rapid metabolism of Etizolam in the human body, the probability of detecting the Etizolam parent drug in actual case samples by forensic personnel is low. Therefore, without detecting the parent drug, analysis of Etizolam metabolites can help forensic personnel provide references and suggestions on whether the suspect has taken Etizolam. This study simulates the objective metabolic process of the human body. It establishes a zebrafish in vivo metabolism model and a human liver microsome in vitro metabolism model to analyze the metabolic characteristics of Etizolam. A total of 28 metabolites were detected in the experiment, including 13 produced in zebrafish, 28 produced in zebrafish urine and feces, and 17 produced in human liver microsomes. The UPLC-Q-Exactive-MS technology was used to analyze the structures and related metabolic pathways of Etizolam metabolites in zebrafish and human liver microsomes, and a total of 9 metabolic pathways were identified, including monohydroxylation, dihydroxylation, hydration, desaturation, methylation, oxidative deamination to alcohol, oxidation, reduction acetylation, and glucuronidation. Among them, metabolites involving hydroxylation reactions (including monohydroxylation and dihydroxylation) accounted for 57.1% of the total number of potential metabolites, indicating that hydroxylation may be the major metabolic pathway of Etizolam. Based on the response values of each metabolite, monohydroxylation (M1), desaturation (M19), and hydration (M16) were recommended as potential biomarkers for Etizolam metabolism. The experimental results provide reference and guidance for forensic personnel in identifying Etizolam use in suspects.
... Fonazepam and nifoxipam, the desmethyl-and 3-hydroxy-desmethyl-derivatives of flunitrazepam (a 7-nitrobenzodiazepine), respectively, are 2 of their active metabolites. 17,47 Finally, generating 1,4-benzodiazepine analogs using triazolo or thienotriazolo-benzodiazepine backbones is an endless source of new DBZDs. ...
Purpose:
Although designer benzodiazepines (DBZDs) constitute a minor part of new psychoactive substances, they deserve the greatest attention because of their popularity among drug users and increasing number and availability. This review covers the effects of different DBZDs, available pharmacological evaluation tools, and their reported toxicity and potential pharmacodynamic and pharmacokinetic interactions with other drugs commonly co-abused with DBZDs.
Methods:
For this narrative review, a nonsystematic search was performed on PubMed, EMBASE, Google Scholar, and PubMed Central databases between June and July 2021.
Results:
The current consensus hypothesis suggests that DBZDs mediate their effects through interactions with the GABAA receptor, producing similar effects to benzodiazepines used in therapy, including sedation, hypnosis, anxiolysis, muscle relaxation, euphoria, amnesia, and addiction. Owing to the complexity of their action mechanism and the numerous GABAA subtype receptors, the pharmacodynamic metrics of DBZDs are very difficult to establish. The pharmacological effects of DBZD are related to their structure, influencing their binding to GABAA receptor subunits. Quantitative structure-activity relationship studies successfully predicted the biological activity and relative potency of DBZD but could not predict the main pharmacological effect of a given DBZD. Exploring the effects by netnographic studies is one of the available alternatives, despite its limitations. DBZDs are usually identified in the context of polysubstance use. Pharmacodynamic interactions between DBZDS and other CNS depressants, such as opioids, have been extensively reported. However, pharmacokinetic interactions between DBZDs and opioids are considered less important, and contradictory conclusions about their clinical significance have been reported.
Conclusions:
Understanding the mechanism of action and other pharmacological metrics is highly important in the clinical management of DBZDs.
... The metabolism of eight designer BZD metabolites-including etizolam-was identified in vitro by incubation with human liver microsomes and analysis using a triple quadrupole mass spectrometer (QQQ-MS) to confirm the phase I metabolites. The study confirmed the presence of other etizolam-related metabolites; 2 ′ -OH-deschloroetizolam was identified in urine and whole blood, while the two other metabolites (mono-hydroxy and di-hydroxyetizolam) were also present but with lower concentrations (49). A study on the inhibition of CYP3A4 using the antifungal itraconzole shows a significant increase in the plasma concentration and elimination half-life of etizolam, which proves that CYP3A4 is at least partly involved in etizolam metabolism (see the "Etizolam interactions" section) (50). ...
Etizolam is a benzodiazepine (BZD). Etizolam is structurally different from BZDs as a thiophene replaces the benzene ring, and a triazole ring is fused to the diazepine ring, but etizolam's pharmacological profile is similar. Etizolam has been used to treat anxiety and panic disorders, to reduce depressive and somatization symptoms, and to induce muscle relaxation. Etizolam is used recreationally due to etizolam's reinforcing and sedative effects. Etizolam is available in tablet or powder form, or administered on blotter paper that can be placed on the tongue for oral absorption. Etizolam metabolises into two major metabolites α-hydroxyetizolam and 8-hydroxyetizolam and all three compounds can be detected in different biological specimens using various common analytical techniques such as immunoassay, chromatography and mass spectrometry. Etizolam is a controlled drug in many countries around the globe but is approved for medical use in some countries, such as Japan, South Korea and Italy. This work is a collation and review of available literature on etizolam to help improve the fundamental understanding of its toxicology, outline best analytical practice, and aid interpretation of toxicology results.
... Although meclonazepam functions as a potent anti-parasitic agent, its sedative side effects have prevented its use. Two double-blind placebo-controlled studies reported arousal, psychomotor performance, and subjective mood following single oral doses of 1, 2, and 4 mg of meclonazepam [94,95]. The study showed a significant impairment in cognitive and psychomotor functions in all doses exceeding 1 mg [95]. ...
... Two double-blind placebo-controlled studies reported arousal, psychomotor performance, and subjective mood following single oral doses of 1, 2, and 4 mg of meclonazepam [94,95]. The study showed a significant impairment in cognitive and psychomotor functions in all doses exceeding 1 mg [95]. Sedative effects peaked at 3 h, but moderate sedation persisted until 6 h post-ingestion [95]. ...
... The study showed a significant impairment in cognitive and psychomotor functions in all doses exceeding 1 mg [95]. Sedative effects peaked at 3 h, but moderate sedation persisted until 6 h post-ingestion [95]. New research targets involve investigating new DBZs that maintain anti-parasitic effects while mitigating the significant alterations in mood and consciousness. ...
As tranquilizers, benzodiazepines have a wide range of clinical uses. Recently, there has been a significant rise in the number of novel psychoactive substances, including designer benzodiazepines. Flubromazolam(8-bromo-6-(2-fluorophenyl)-1-methyl-4H-[1,2,4]triazolo[4,3-a][1,4]benzodiazeZpine) is a triazolo-analogue of flubromazepam. The most common effects noted by recreational users include heavy hypnosis and sedation, long-lasting amnesia, and rapid development of tolerance. Other effects included anxiolysis, muscle-relaxing effects, euphoria, loss of control, and severe withdrawals. Clonazolam, or 6-(2-chlorophenyl)-1-methyl-8-nitro-4H-[1,2,4]triazolo[4,3-α]-[1,4]-benzodiazepine, is a triazolo-analog of clonazepam. It is reported to be over twice as potent as alprazolam. Deschloroetizolam (2-Ethyl-9-methyl-4-phenyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepine) is part of the thienodiazepine drug class, which, like benzodiazepines, stimulates GABA-A receptors. Meclonazepam ((3S)-5-(2-chlorophenyl)-3-methyl-7-nitro-1,3-dihydro-1,4-benzodiazepin-2-one) is a designer benzodiazepine with additional anti-parasitic effects. Although it has proven to be an efficacious therapy for schistosomiasis, its sedative side effects have prevented it from being marketed as a therapeutic agent. The use of DBZs has been a subject of multiple recent clinical studies, likely related to increasing presence and availability on the internet drug market and lack of regulation. Many studies have aimed to identify the prevalence of DBZs and their effects on those using them. This review discussed these designer benzodiazepines and the dangers and adverse effects that the clinician should know.
... 3 Benzodiazepines are depressants that are widely prescribed to treat anxiety, insomnia, muscle spasms, alcohol withdrawal, and epilepsy, and the number of benzodiazepine analogs has increased in recent years. 4,5 Benzodiazepine NPSs, also known as "designer benzodiazepines," are not used for legal purposes 6 ; at least 24 of these compounds have been identified as of November 2018. 7 Designer benzodiazepines are sold by online marketplaces at low prices without restrictions, thus posing a threat to public health worldwide. ...
The use of many benzodiazepines is controlled worldwide due to their high likelihood of abuse and potential adverse effects. Flubromazepam—a designer benzodiazepine—is a long-acting gamma-aminobutyric acid subtype A receptor agonist. There is currently a lack of scientific evidence regarding the potential for flubromazepam dependence or other adverse effects. This study aimed to evaluate the dependence potential, and cardiotoxicity via confirmation of the QT and RR intervals which are the factors on the electrical properties of the heart of flubromazepam in rodents. Using a conditioned place preference test, we discovered that mice treated intraperitoneally with flubromazepam (0.1 mg/kg) exhibited a significant preference for the flubromazepam-paired compartment, suggesting a potential for flubromazepam dependence. In addition, we observed several cardiotoxic effects of flubromazepam; 100-μM flubromazepam reduced cell viability, increased RR intervals but not QT intervals in the electrocardiography measurements, and considerably inhibited potassium channels in a human ether-à-go-go-related gene assay. Collectively, these findings suggest that flubromazepam may have adverse effects on psychological and cardiovascular health, laying the foundation for further efforts to list flubromazepam as a controlled substance at both national and international levels.
