Suspected North Carolina counterfeit pill-involved deaths, 2020-2022.

The NC Office of the Chief Medical Examiner regularly assumes jurisdiction over deaths that are suspicious, unusual or unattended by a medical professional. In recent years, the presence of counterfeit pills is occasionally suggested by investigatory notes and/or scene findings that document reported consumption of prescription drugs, or prescription drugs on scene, which are not reflected in the final autopsy findings after toxicological analysis of the decedent's blood samples. Counterfeit pill consumption is a major public health hazard worthy of attention from the forensic toxicology community. Seventy-five cases from January 2020 to December 2022 serve as a convenience sample of cases where prescription pills including formulations of alprazolam, oxycodone and hydrocodone were specifically referenced during the death scene investigation as recently consumed, yet an unexpected substance was found during toxicological analysis rather than the expected pharmaceutical drug. Of note, novel benzodiazepines detected included flualprazolam, etizolam, clonazolam metabolite (8-aminoclonazolam), bromazolam, flubromazolam and desalkylflurazepam. Decedents' ages ranged from 16 to 69, across 33 different NC counties. Case notes indicated that eight of the decedents obtained pills through direct personal relationships, six decedents obtained them from "the street" and one decedent likely purchased pills online. Pills were largely consumed orally or through insufflation. Seven case reports contained indication that decedents knew or suspected the counterfeit nature of their pills. This study describes the context and characteristics of 2020-2022 suspected counterfeit pill-involved deaths in NC to further the understanding of the forensic science community, law enforcement partners, public health stakeholders and those potentially at risk through the consumption of counterfeit pills.

Interference of the novel designer benzodiazepine 4'-chloro deschloroalprazolam with alprazolam analysis in toxicology and seized drug DUID casework.

In recent years, the emergence of the novel designer benzodiazepine 4'-chloro deschloroalprazolam has presented a new challenge for forensic laboratories by interfering with the identification and quantitation of alprazolam. As an isomer of alprazolam, 4'-chloro deschloroalprazolam has similar physicochemical properties and can be misidentified in casework samples as alprazolam without a specific method to differentiate the two analytes. Starting in late 2021, the Houston Forensic Science Center (HFSC) received toxicological and seized drug evidence indicating the presence of 4'-chloro deschloroalprazolam. An interference study was performed to supplement the laboratory's validated benzodiazepines method for toxicological samples to differentiate alprazolam from 4'-chloro deschloroalprazolam. This study showed that while the isomers could not be chromatographically resolved using the current method, they could be differentiated based on their retention times relative to the internal standard, alprazolam-d5. Based on these findings, the HFSC toxicology laboratory reports test results as "unsuitable for analysis due to an interference" if a suspected alprazolam peak elutes before the alprazolam-d5 peak, even if all identification and quantification criteria (e.g., retention time) were acceptable. Additionally, the seized drug and toxicology laboratories re-evaluated previously analyzed alprazolam-positive casework to determine if suspected 4'-chloro deschloroalprazolam had been misidentified as alprazolam. This report presents three cases: one case with toxicological evidence indicating the presence of both 4'-chloro deschloroalprazolam and alprazolam, and two cases with both seized drug material and toxicology evidence indicating the presence of 4'chloro deschloroalprazolam with no detected alprazolam.

Validation of an HPLC-HR-MS Method for the Determination and Quantification of Six Drugs (Morphine, Codeine, Methadone, Alprazolam, Clonazepam and Quetiapine) in Nails.

Keratinized matrices, including nails, are among the most resistant matrices that can be analyzed in cases where remains are deeply decomposed and relatively non-invasive for living people. In order to exploit these new matrices in the search for exogenous substances, it is necessary to develop analytical technologies capable of achieving high levels of sensitivity. In this technical note, an easy method is presented for the simultaneous extraction and quantification of three narcotic substances (morphine, codeine and methadone), two benzodiazepines (BDZs) (clonazepam and alprazolam) and an antipsychotic (quetiapine) from nail matrix by analysis in ultra-high-performance liquid chromatography at high-resolution mass spectrometry. The method has been validated following the Standard Practices for Method Validation in Forensic Toxicology of the Scientific Working Group for Forensic Toxicology. Nail specimens from eight authentic postmortem (PM) cases and 13 living donor samples were extracted and analyzed. Of the eight PM samples, five resulted positive for at least one of the three substances searched. Ten of the 13 living donor specimens were positive for at least one of the targeted BDZs or quetiapine.

Occurrence of Z-drugs, benzodiazepines, and ketamine in wastewater in the United States and Mexico during the Covid-19 pandemic.

Z-drugs, benzodiazepines and ketamine are classes of psychotropic drugs prescribed for treating anxiety, sleep disorders and depression with known side effects including an elevated risk of addiction and substance misuse. These drugs have a strong potential for misuse, which has escalated over the years and was hypothesized here to have been exacerbated during the COVID-19 pandemic. Wastewater-based epidemiology (WBE) constitutes a fast, easy, and relatively inexpensive approach to epidemiological surveys for understanding the incidence and frequency of uses of these drugs. In this study, we analyzed wastewater (n = 376) from 50 cities across the United States and Mexico from July to October 2020 to estimate drug use rates during a pandemic event. Both time and flow proportional composite and grab samples of untreated municipal wastewater were analyzed using solid-phase extraction followed by liquid chromatography-tandem mass spectrometry to determine loadings of alprazolam, clonazepam, diazepam, ketamine, lorazepam, nordiazepam, temazepam, zolpidem, and zaleplon in raw wastewater. Simultaneously, prescription data of the aforementioned drugs were extracted from the Medicaid database from 2019 to 2021. Results showed high detection frequencies of ketamine (90 %), lorazepam (87 %), clonazepam (76 %) and temazepam (73 %) across both Mexico and United States and comparatively lower detection frequencies for zaleplon (22 %), zolpidem (9 %), nordiazepam (<1 %), diazepam (<1 %), and alprazolam (<1 %) during the pandemic. Average mass consumption rates, estimated using WBE and reported in units of mg/day/1000 persons, ranged between 62 (temazepam) and 1100 (clonazepam) in the United States. Results obtained from the Medicaid database also showed a significant change (p < 0.05) in the prescription volume between the first quarter of 2019 (before the pandemic) and the first quarter of 2021 (pandemic event) for alprazolam, clonazepam and lorazepam. Study results include the first detections of zaleplon and zolpidem in wastewater from North America.

A method for improved detection of 8-isoprostaglandin F2α/ß and benzodiazepines in wastewater.

Wastewater-based epidemiology is a tool incorporating biomarker analysis that can be used to monitor the health status of a population. Indicators of health include endogenous oxidative stress biomarkers and hormones, or exogenous such as alcohol and nicotine. 8-Iso-prostaglandin F2α/ß is a biomarker of endogenous metabolism that can be used to measure oxidative stress in a community. Benzodiazepines are a harmful subclass of anxiolytics either prescribed or sourced illegally. The analysis of oxidative stress markers and uptake of benzodiazepines in wastewater may provide information about distress in the community. A method has been applied to detect 8-isoPGF2α/ß and the illicit benzodiazepines clonazolam, flubromazolam and flualprazolam in addition to other prescribed benzodiazepines in wastewater. These substances have been sold as counterfeit pharmaceutical products, such as Xanax, which was formulated to include alprazolam. Deconjugation was initially performed on wastewater samples, followed by liquid-liquid extraction for isoprostanes and solid phase extraction for benzodiazepines to determine the total levels of these analytes. Limits of quantification were in the range of 0.5-2 ng/L for all the analytes except 8-isoPGF2α/ß which was 50 ng/L. Stability, recovery and matrix effect studies were also conducted. Finally, this method was applied to influent wastewater from South Australia which showed the prevalence of 8-isoPGF2α/ß and benzodiazepines.

Adsorptive Cathodic Stripping Voltammetry for Quantification of Alprazolam.

A simple and highly sensitive electrochemical sensor was developed for adsorptive cathodic stripping voltammetry of alprazolam. Based on an electrochemically pretreated glassy carbon electrode, the sensor demonstrated good adsorption and electrochemical reduction of alprazolam. The morphology of the glassy carbon electrode and the electrochemically pretreated glassy carbon electrode were characterized by scanning electron microscopy/energy dispersive X-ray spectroscopy, Fourier transform infrared spectroscopy, cyclic voltammetry, and electrochemical impedance spectroscopy. The electrochemical behaviors of alprazolam were determined by cyclic voltammetry, and the analytical measurements were studied by adsorptive cathodic stripping voltammetry. Optimized operational conditions included the concentration and deposition time of sulfuric acid in the electrochemical pretreatment, preconcentration potential, and preconcentration time. Under optimal conditions, the developed alprazolam sensor displayed a quantification limit of 0.1 mg L-1, a detection limit of 0.03 mg L-1, a sensitivity of 67 µA mg-1 L cm-2 and two linear ranges: 0.1 to 4 and 4 to 20 mg L-1. Sensor selectivity was excellent, and repeatability (%RSD < 4.24%) and recovery (82.0 ± 0.2 to 109.0 ± 0.3%) were good. The results of determining alprazolam in beverages with the developed system were in good agreement with results from the gas chromatography-mass spectrometric method.

Single-step extraction for simultaneous quantification of desvenlafaxine and alprazolam in human spiked plasma by RP-HPLC.

Desvenlafaxine (DES) and Alprazolam (ALP) are the drugs commonly prescribed together for the treatment of Major Depressive Disorders (MDD). A literature survey revealed, there is no method for the simultaneous determination of these two drugs. The purpose of this research was to develop and validate a simple, accurate, precise, robust, and isocratic RP-HPLC method for simultaneous determination of DES and ALP in human spiked plasma using UV-detector in short analysis time. The method utilized Hypersil BDS C18 (250 mm×4.6 mm, 5 µm) through an isocratic mode of elution using HPLC grade acetonitrile and 0.02M KH2PO4 buffer (65:35) and 0.1% Tri Fluoro Acetic acid (TFA) with pH 4.00 adjusted with 1M KOH. The flow rate was 1.00 mLmin-1 and elution of the drugs was monitored at 230nm. The elution time of DES and ALP was 4.011 and 5.182 minutes respectively. The method was linear for the concentration range 10-150 µgmL-1 for DES and 5.0-75.0 µgmL-1 for ALP. According to the validation results, the method is sensitive with Limit of Detection (LOD) 4.740 µgmL-1 and Limit of Quantification (LOQ) of 14.365 µgmL-1 for DES and LOD 1.891 µgmL-1 & LOQ 5.730 µgmL-1 for ALP. The reproducibility of results with minute deliberate variations in method parameters has proven that the method is robust. The data from stability studies show a non-significant change in drugs solutions for 2 months. The optimized method was validated as per International Conference for Harmonisation (ICH) Q2(R1) guidelines. This method can be used for the estimation of DES and ALP in plasma and can evaluate pharmacokinetic parameters of both drugs simultaneously.

An unusual detection of tert-butyl-4-anilinopiperidine-1-carboxylate in seizures of falsified 'Xanax' tablets and in items in a suspected heroin seizure submitted by Irish law enforcement.

The identification of tert-butyl-4-anilinopiperidine-1-carboxylate (4-anilinopiperdine-t-BOC or 4-AP-t-BOC) in many seized falsified 'Xanax' tablets has been reported after being encountered in forensic casework in late 2019 and early 2020 in Ireland. This substance was also detected in a pink powder submitted for analysis in March 2020. The pink powder was part of a larger seizure comprising brown powders which contained morphine or diamorphine (heroin) or a type of counterfeit heroin or heroin adulterant (known as 'bash'). Novel benzodiazepines and other substances are being detected as ingredients in falsified benzodiazepine tablets more frequently on the illicit market. The detection of 4-AP-t-BOC in benzodiazepine tablets is noteworthy and 4-AP-t-BOC is added to the list of adulterants found in benzodiazepine tablets emerging in Europe. The presence of 4-AP-t-BOC in both falsified 'Xanax' and powdered seizures is unusual, and analytical data are presented to assist with the identification of this compound in suspected illicit substances. The presence of 4-AP-t-BOC in the tablets was confirmed using gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry analyses, and spectral fragmentation pathways were suggested. To the authors' best knowledge, information about the biological activity of 4-AP-t-BOC is not available. The removal of the t-BOC protecting group yields 4-anilinopiperidine which has been reported to be involved in the synthesis of fentanyl.

Detailed quantum mechanical studies on bioactive benzodiazepine derivatives and their adsorption over graphene sheets.

Estazolam (Z1) and related derivatives, adinazolam (Z2), alprazolam (Z3), 4-hydroxyalprazolam (Z4) and triazolam (Z5) have been studied by using various computational tools to analyze their geometry and spectral characteristics. The compounds were found to interact with graphene monolayer results shows that there is enhancement in various physico-chemical descriptors and surface enhanced Raman spectra (SERS). The various reactive descriptors obtained from the FMO analysis predict the reactive nature of the compound. The various lone pair/sigma to pi conjugation was analyzed using NBO formalism, which provides valuable information about intra molecular electron transfer which is vital in predicting the inherent stability of the molecule. Simulated electronic spectra using TD-DFT and CAM-B3LYP functional are discussed in detail with respect to electronic transitions and light harvesting efficiency. Suitability of candidates as a photo sensitizer in dye sensitized solar cells was studied and 4-Hydroxyalprazolam is identified as a suitable candidate. Nucleophilic and electrophilic regions of the molecules are identified using MESP, which adds to the reactivity information. It can be seen that the highest interaction energy has been obtained in the case of the Z5-graphene system, while the lowest interaction energy has been obtained in the case of the Z1-graphene system. Docking indicates that the ligands adsorbed over graphene also form stable complexes with the receptors as indicated by the high binding affinity energy values.

Liquid chromatographic method for simultaneous determination of alprazolam with NSAIDs in bulk drug, pharmaceutical formulation and human serum.

High performance liquid chromatography with UV/vis detection was optimized and validated for simultaneous quantification of alprazolam with celecoxib and diclofenac sodium in pharmaceutical formulation and human serum. Chromatographic separation was achieved at detection wavelength of 230 nm on Shimadzu Shim-pack CLC-ODS (M) 25M column employing 80:20 (v/v) methanol: water (pH 3.5) as mobile phase with elution rate 1.0mL min-1. Analytes were quantified in the ranges 0.2-15, 0.3-20 and 0.6-40 µg mL-1 with detection limits 19.76, 17.29 and 11.83ng mL-1 respectively. Recoveries were in the range 98.15-101.15, 99.24-99.90 and 98.87-101.19% in pharmaceutical formulation and 98.05-101.01, 98.72-99.49 and 98.25-99.47% in human serum respectively and precision ranged from 0.19-1.84%. The analytes were successfully detected without any observable interference commonly present in pharmaceutical formulation and human serum demonstrating applicability of method.

Validation of a sensitive UHPLC-MS/MS method for cytochrome P450 probe substrates caffeine, tolbutamide, dextromethorphan, and alprazolam in human serum reveals drug contamination of serum used for research.

To evaluate the potential for interactions between botanical dietary supplements and drug metabolism, Phase I clinical pharmacokinetics studies are conducted using an oral cocktail of probe substrates of cytochrome P450 (CYP) enzymes. A sensitive, specific, and fast ultra-high performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for determination of caffeine (probe of CYP1A2), tolbutamide (probe of CYP2C9), dextromethorphan (probe of CYP2D6), and alprazolam (probe of CYP3A4/5) in human serum. Stable isotope-labelled analogs were used as internal standards, and sample preparation involved only rapid protein precipitation and centrifugation. The method of standard addition was used for the measurement of caffeine, because commercially available pooled human serum contains caffeine. Out of 18 lots of pooled human serum tested, caffeine was detection in all lots, alprazolam was detected in 13 lots, 8 lots contained dextromethorphan, and no tolbutamide was detected. Only serum prepared from the blood of select individuals was determined to be drug-free. The analytical method was validated with respect to linearity, accuracy and precision, recovery, stability, and matrix effects. The calibration curves were linear over the range of 25-12,000 ng/mL for caffeine, 75-36,000 ng/mL for tolbutamide, 0.05-30 ng/mL for dextromethorphan, and 0.1-60 ng/mL for alprazolam. The intra-assay and inter-assay coefficients of variation (%CV) and %Bias were <13 % (<17 % at the lower limit of quantitation). The recovery of each probe substrate ranged from 84.2%-98.5 %. All analytes were stable during sample storage and handling. Matrix effects were minimized by using stable isotope-labeled internal standards. The method was successfully applied to clinical studies investigating the pharmacokinetic alterations of probe substrates caused by chronic consumption of botanical dietary supplements.

Quantification of eight benzodiazepines in human breastmilk and plasma by liquid-liquid extraction and liquid-chromatography tandem mass spectrometry: Application to evaluation of alprazolam transfer into breastmilk.

Breastfeeding is strongly encouraged for infant and maternal health. Benzodiazepines (BZDs) are widely prescribed drugs for symptoms, such as anxiety and insomnia, which many women could experience during the postpartum period. However, limited information is currently available to evaluate the transfer of different BZDs into breastmilk. In order to assess the proprieties of this medication during breastfeeding, robust and sensitive analytical methods to quantify BZDs are required. For this purpose, we developed a method for quantification of BZDs, including alprazolam, bromazepam, clonazepam, clotiazepam, etizolam, flunitrazepam, lorazepam, and CM7116 (a metabolite of ethyl loflazepate), in human breastmilk and plasma using liquid chromatography/tandem mass spectrometry (LC/MS/MS). Sample preparation was performed by a simple liquid-liquid extraction (LLE) with ethyl acetate. For sample preparation of CM7116, the pretreatment process to completely obtain the metabolite was added before the LLE step. The BZDs were separated by a C18 column using a gradient elution of acetonitrile in aqueous ammonium acetate solution, and were detected in the positive ion electrospray mode with multiple reaction monitoring (MRM). Lower limits of quantification (LLOQs) in breastmilk ranged from 0.25 to 0.5 ng/mL, and those in plasma ranged from 0.5 to 1.0 ng/mL. The intra-day and inter-day precision, and accuracy of data were assessed and found to be acceptable. The developed method was successfully applied to measure the concentration of alprazolam in breastmilk and plasma, which were donated by a lactating woman who had been regularly treated with alprazolam. Milk to plasma (M/P) ratios were calculated as 0.52 (before oral administration) and 0.49 (2 h after administration) 3 days after delivery. The M/P ratio 1 month after delivery was calculated as 0.41 (2 h after administration). We estimated that the relative infant dose (RID) values of alprazolam ranged from 3.11 to 4.61%.

Alprazolam and Zolpidem in Skeletal Tissue of Decomposed Body Confirms Exposure.

In several medico-legal cases, bone samples analysis may provide the only source of toxicological information. This case study reports the analysis of a human bone specimen, belonging to a 46-year-old man, found 3 months after his death due to cervical-thoracic injuries in a motorcycle accident. Bone specimen was the only available material for toxicological analysis, among few skull hair and rotten skin. Analysis was performed by a newly developed and validated ultra-high-pressure liquid chromatography-mass spectrometry/mass spectrometry (UHPLC-MS/MS) method, following simple and efficient sample pretreatment. The results were in accordance with the man's medical record: Alprazolam and zolpidem were found at 2.2 and 5.4 ng/g of bone, respectively. Both these drugs were prescribed to the deceased.

Toxicological findings in 1000 cases of suspected drug facilitated sexual assault in the United States.

The purpose of this study was to identify the extent and types of drugs found in alleged drug facilitated sexual assaults (DFSA) in 37 states and 1 territory of the United States. In total, 1000 cases were reviewed. Between the cases that gender was provided (613), most of the victims (91.68%) were woman, mean age of 26.8 years old. Blood and/or urine samples were tested. Twenty-one point six percent of the cases were negative for intoxicating substances. A hundred and one different substances were detected. Overall, ethanol was the most prevalent substance, detected in 30.9% of the cases (309 cases), followed by cannabinoids (THC/THCCOOH/11-OH-THC) (28.8% of cases), amphetamine/methamphetamine (16.5% of cases), cocaine/metabolites (10.4% of cases), and clonazepam/metabolite (7.6% of cases). The mean, median and range concentrations of ethanol in blood (n = 309) were 98.6 mg/dL, 82.0 mg/dL and 9.2-366 mg/dL, respectively. Ethanol and cannabinoids were the most frequent combination found. The absence of alcohol and drugs in some cases may represent delay in collecting samples.

An Experimental Study of Diazepam and Alprazolam Kinetics in Urine and Oral Fluid Following Single Oral Doses.

Benzodiazepines are commonly seen in samples submitted for drug testing of patients, people involved in child welfare cases, work-place drug testing, as well as in drug-facilitated assaults. Limited previous experimental studies are available regarding the excretion of benzodiazepines in urine and oral fluid. The aim of this study was to investigate the concentrations of diazepam and alprazolam in oral fluid and urine for up to 2 weeks after ingestion of a single oral dose in healthy volunteers. A total of 11 healthy volunteers ingested 10 mg diazepam at the start of the study and 0.5 mg alprazolam on Day 3 of the study. A total number of 10 oral fluid samples and 17 urine samples were collected from each participant. The samples were analyzed by liquid chromatography with tandem mass spectroscopy and ultra-high performance liquid chromatography tandem mass spectrometry methods. The median detection time was 252 h for the longest detected diazepam metabolite in urine (oxazepam, range 203-322) and 132 h in oral fluid (N-desmethyldiazepam, range 109-136). For alprazolam, the median detection time was 36 h (metabolite α-OH-alprazolam, range 26-61) in urine and 26 h (alprazolam, range 4-37) in oral fluid. These results show that detection times are only 36 h for alprazolam in urine after intake of a single therapeutic oral dose. For diazepam in urine, detection times were 11 days. Detection times were generally shorter in oral fluid compared to urine. The results could be helpful in the interpretation of diazepam or alprazolam findings in drug testing cases involving urine or oral fluid.

Persistence of alprazolam in river water according to forced and non-forced degradation assays: adsorption to sediment and long-term degradation products.

Alprazolam is a pharmaceutical compound that it is detected in surface waters. Some degradation studies in aqueous solutions and pharmaceutical products are available, but there is no reliable information about its stability in river water. Here, assays have been conducted under forced biological, photochemical, and thermal conditions, and under non-forced conditions, to estimate the fate of alprazolam in river water and know its degradation products. The forced assays indicated that the biological and photochemical degradation of alprazolam was negligible; heating at 70°C for a long time barely affected it. The degradation of alprazolam in river water at 100 µg/L was about 5% after 36 weeks, keeping the water under a natural day-night cycle at room temperature and limiting partially the exposure to sunlight as happens inside a body of water; no change in concentration was observed when the monitoring was performed at 2 µg/L. The results suggest the persistence of alprazolam in surface water and a possible accumulation over time. Residues were monitored by ultra-pressure liquid chromatography/quadrupole time-of-flight/mass spectrometry after solid-phase extraction; nine degradation products were found and the structures for most of them were proposed from the molecular formulae and fragmentation observed in high-resolution tandem mass spectra. (5-chloro-2-(3-methyl-4H-1,2,4-triazol-4-yl)phenyl)(phenyl)methanol was the main long-term transformation product in conditions that simulate those in a mass of water. The degradation rate in presence of sediment was equally very low under non-forced conditions; adsorption coefficients of alprazolam and major degradation products were calculated. Copyright © 2016 John Wiley & Sons, Ltd.

PLS-LS-SVM based modeling of ATR-IR as a robust method in detection and qualification of alprazolam.

According to the United States pharmacopeia (USP), Gold standard technique for Alprazolam determination in dosage forms is HPLC, an expensive and time-consuming method that is not easy to approach. In this study chemometrics assisted ATR-IR was introduced as an alternative method that produce similar results in fewer time and energy consumed manner. Fifty-eight samples containing different concentrations of commercial alprazolam were evaluated by HPLC and ATR-IR method. A preprocessing approach was applied to convert raw data obtained from ATR-IR spectra to normal matrix. Finally, a relationship between alprazolam concentrations achieved by HPLC and ATR-IR data was established using PLS-LS-SVM (partial least squares least squares support vector machines). Consequently, validity of the method was verified to yield a model with low error values (root mean square error of cross validation equal to 0.98). The model was able to predict about 99% of the samples according to R2 of prediction set. Response permutation test was also applied to affirm that the model was not assessed by chance correlations. At conclusion, ATR-IR can be a reliable method in manufacturing process in detection and qualification of alprazolam content.

INVESTIGATION OF A MIXTURE CONTAINING ALPRAZOLAM, CODEINE AND PARACETAMOL USING THIN-LAYER AND HIGH PERFORMANCE LIQUID CHROMATOGRAPHY METHODS.

The increasing drug consumption in Lithuania and all over the world makes us think about the negative consequences - the risk of toxicity. Fast and accurate identification of material that caused the poisoning reduces the probability in death cases and makes easier to determine the main cause of death. The results have shown that the most appropriate systems of solvents for qualitative analysis by TLC method of the mixture consisting of alprazolam, codeine and paracetanol are: system "D" (trichloromethane : acetone : conc. ammonia = 55 : 40 : 5 (v/v/v)) and system "F" (trichloromethane : diethyl ether: isobutanol : conc. ammonia = 50 : 30 : 15 : 5 (v/v/v/v)). For qualitative analysis of the mixture consisting of alprazolam, codeine and paracetamol by HPLC method the chromatographic column ACE C18 (25 cm x 4.6 mm x 5 µm), gradient elution mode (mixture of 3% acetic acid and methanol and the flow rate 1 mL/min have been used. The injection volume was 10 pL. Photodiode array detector (210 - 240 nm range) has been used. UV absorption spectra of materials measured using photodiode array detector have been identical to those presented in the scientific literature.

New approach to the determination of contaminants of emerging concern in natural water: study of alprazolam employing adsorptive cathodic stripping voltammetry.

Contaminants of emerging concern (CECs) are chemicals, including pharmaceutical and personal care products, not commonly monitored in the aquatic environment. Pharmaceuticals are nowadays considered as an important environmental contaminant. Chromatography methods which require expensive equipment and complicated sample pretreatment are used for detection of CECs in natural water. Thus, in this study we proposed a simple, fast, and low-cost voltammetric method as a screening tool for the determination of CECs in natural water prior to chromatography. A case study was conducted with alprazolam (benzodiazepine). The method was optimized and validated in-house. The limit of quantification was 0.4 µg L(-1) for a 120 s preconcentration time. The recoveries ranged from 93 to 120 % for accuracy tests. A further proposal aim was to determine for the first time the occurrence of alprazolam in Brazilian river water and to evaluate its potential use as a marker of contamination by wastewater.

Detection Times of Diazepam, Clonazepam, and Alprazolam in Oral Fluid Collected From Patients Admitted to Detoxification, After High and Repeated Drug Intake.

BACKGROUND: Clonazepam, diazepam, and alprazolam are benzodiazepines with sedative, anticonvulsant, and anxiolytic effects, but their prevalence in drug abuse and drug overdoses has long been recognized. When detection times for psychoactive drugs in oral fluid are reported, they are most often based on therapeutic doses administered in clinical studies. Repeated ingestions of high doses, as seen after drug abuse, are however likely to cause positive samples for extended time periods. Findings of drugs of abuse in oral fluid collected from imprisoned persons might lead to negative sanctions, and the knowledge of detection times of these drugs is thus important to ensure correct interpretation. The aim of this study was to investigate the time window of detection for diazepam, clonazepam, and alprazolam in oral fluid from drug addicts admitted to detoxification. METHODS: Twenty-five patients with a history of heavy drug abuse admitted to a detoxification ward were included. Oral fluid was collected daily in the morning and the evening and urine samples every morning for 10 days, using the Intercept device. Whole blood samples were collected if the patient accepted. The cutoff levels in oral fluid were 1.3 ng/mL for diazepam, N-desmethyldiazepam, and 7-aminoclonazepam and 1 ng/mL for clonazepam and alprazolam. In urine, the cutoff levels for quantifications were 30 ng/mL for alprazolam, alpha-OH-alprazolam, and 7-aminoclonazepam, 135 ng/mL for N-desmethyldizepam, and 150 ng/mL for 3-OH-diazepam and for all the compounds, the cutoff for the screening analyses were 200 ng/mL. RESULTS: The maximum detection times for diazepam and N-desmethyldiazepam in oral fluid were 7 and 9 days, respectively. For clonazepam and 7-aminoclonazepam, the maximum detection times in oral fluid were 5 and 6 days, respectively. The maximum detection time for alprazolam in oral fluid was 2.5 days. New ingestions were not suspected in any of the cases, because the corresponding concentrations in urine were decreasing. Results from blood samples revealed that high doses of benzodiazepines had been ingested before admission, and explains the longer detection times in oral fluids than reported previously after intake of therapeutic doses of these drugs. CONCLUSIONS: This study has shown that oral fluid might be a viable alternative medium to urine when the abuse of benzodiazepines is suspected.