Driving Under The Influence Of Drugs Research Articles

Enantiomeric profile of amphetamines in seized drug samples and in blood of impaired drivers in Iceland: the rise of (R)-methamphetamine?

Amphetamine (AMP) and methamphetamine (METH) use is increasing globally. Illegal AMP is generally a racemic mixture, whereas AMP-containing attention-deficit hyperactivity disorder drugs prescribed in Iceland consist of S-AMP. AMP is also a main metabolite of interest after METH intake. Distinguishing between legal and illegal AMP intake is vital in forensic toxicology. A chiral UPLC-MS/MS method was used to determine the enantiomeric profile of AMP and METH in circulation in Iceland by analysing blood samples from drivers suspected of driving under the influence of drugs (DUID) and seized drug samples from 2021 and 2022. All seized AMP samples (n = 48) were racemic, whereas all but one seized METH sample (n = 26) were enantiopure. Surprisingly, a large portion of the enantiopure METH samples was R-METH. DUID blood samples positive for AMP (n = 564) had a median blood concentration of 180 ng/mL (range 20-2770 ng/mL) and a median enantiomeric fraction (EFR) of 0.54 (range 0-0.73), whereas samples positive for METH (n = 236) had a median blood concentration of 185 ng/mL (range 20-2300 ng/mL) and a median EFR of 0.23 (range 0-1). The findings of this study show a significantly lower blood concentration in drivers with only S-AMP detected compared with when the R-isomer is also detected. No significant difference in blood concentration was detected between the sample groups containing S-METH, R-METH, or both enantiomers. The occurrence of R-METH in both seized drug samples and DUID cases indicates a change in drug supply and a need for better scientific knowledge on R-METH abuse.

Quantitation of hexahydrocannabinol (HHC) and metabolites in blood from DUID cases.

Hexahydrocannabinol (HHC) was first reported in the EU in May 2022. HHC has three chiral carbon atoms, but only (6aR,9R,10aR)-HHC (9R-HHC) and (6aR,9S,10aR)-HHC (9S-HHC) have been encountered in HHC products. The aim of this study was to develop and validate a method for the quantitative analysis of 9R-HHC, 9S-HHC, 11-OH-9R-HHC, 9R-HHC-COOH, 9S-HHC-COOH and 8-OH-9R-HHC. In addition, an objective was to investigate the immunochemical cross-reactivity. Blood samples from driving under the influence of drugs (DUID) cases screened positive for cannabis using enzyme-linked immunoadsorbent assay (ELISA) and confirmed negative for tetrahydrocannabinol (THC), 11-hydroxy-THC and THC-COOH were reanalyzed with a newly validated HHC method to investigate the presence of HHC and metabolites. The LC-MS-MS method was validated for matrix effects, lower limit of quantification (LLOQ), calibration model, precision, bias and autosampler stability. Cross-reactivity on an ELISA method was investigated separately for 9R-HHC-COOH and 9S-HHC-COOH at a concentration range between 5 and 200 ng/mL. The cross-reactivity was found to be 120% for 9R-HHC-COOH and 48% for 9S-HHC-COOH. In the LC-MS-MS method, 9R-HHC-COOH, 9S-HHC-COOH and 11-OH-9R-HHC showed matrix effects <25% at both concentrations, while 8-OH-9R-HHC, 9R-HHC and 9S-HHC matrix effects exceeded 25% at both concentrations but showed good precision (<10% for both inter and intra day) and low bias (<6%) in the further validation. The LLOQ was investigated and established at 0.2 ng/mL for all analytes except the carboxylated metabolites that had an LLOQ of 2.0 ng/mL. The upper LOQ was 20 and 200 ng/mL, respectively. Reanalysis of cases (n = 145) confirmed HHC and metabolites in 32 cases (22%). It was determined that the major metabolite in blood after administration of HHC was 9R-HHC-COOH followed by 11-OH-9R-HHC and that presumptive positive cases are caught by the routine ELISA screening for cannabis.

Gabapentin in drugged driving investigations.

The increasing use and misuse of gabapentin pose a major risk to public health and traffic safety. Gabapentin has been approved by the Food and Drug Administration (FDA) since 1993 for adjunctive therapy in the treatment of epilepsy and neuralgia but is increasingly being prescribed for numerous off-label uses including insomnia, anxiety, depression, and migraine. Reported side effects include blurred vision, drowsiness, and loss of coordination. Driving behaviors such as exiting the lane of travel and crashes have been reported in connection to drugged driving investigations concerning gabapentin. To further assist with the toxicological interpretation of gabapentin in driving under the influence of drugs (DUID) scenarios, a review of approximately 108,000 gabapentin-positive DUID cases was conducted. Of those, 858 cases met inclusion criteria and underwent additional evaluation. Blood specimens were screened via enzyme-linked immunosorbent assay (ELISA) and confirmed by liquid chromatography tandem mass spectrometry (LC-MS/MS) for quantitation of gabapentin. This review found an overall DUID gabapentin positivity of 7.9% between January 2020 and December 2022; 17 states from various geographical regions had at least one positive gabapentin DUID case. Observations in six driving and human performance cases where gabapentin was the only drug reported were consistent with the known adverse effects of the medication. Half of the case histories reviewed involved crashes where the driver was determined to be at fault. Additionally, 94% of the cases in this review involved gabapentin in combination with other drugs. The most prevalent drug combinations were opioids and gabapentin present in 64% of cases.

A retrospective study of traffic offenses associated with driving under the influence of drugs in China

Objective This study aimed to investigate the prevalence and pattern of traffic offenses associated with driving under the influence of drugs (DUID) in 31 provinces of mainland China. Methods Traffic offenses were extracted from the nationally representative court file disclosure website. Drivers convicted of traffic offenses associated with DUID were included. The monthly percentage change (MPC) in rates was estimated by fitting negative binomial regression models to assess the time trends. Moran’s I was used to determine the correlations of DUID-related rates among surrounding provinces. Results In total, 896 individuals convicted of DUID offenses were identified (April 2007 to March 2022). Of these, 769 individuals were involved in traffic crashes, and they killed 629 people. Regarding the types of drugs, 638 used amphetamine-type stimulants (ATS; methamphetamine, 3,4-methylenedioxymethamphetamine, amphetamine, and tenamfetamine), 104 used ketamine, 104 used opioids (heroin, morphine, methadone, codeine, and tramadol), 20 used caffeine, 4 used tetrahydrocannabinol, and 1 used gamma-hydroxybutyric acid. The offense trends among overall drug users (MPC, −1.1; p < .001) declined from 2014 to 2020. The trends also decreased for DUID-related traffic crashes (−1.5; p < .001) and deaths (−1.7; p < .001) during the same period. Except in three provinces, ATS-related offense rates were higher than opioid-related offense rates. Ketamine-related offense rates showed a significant positive correlation among the surrounding provinces (Moran’s I, 0.22; p < .01), and South China had the highest ketamine-related offense rates. Conclusions In the sample, the case fatality rate was 70.7%, which was a strong indicator of underreporting, and most minor DUID-related offenses could not be captured by Chinese court files. The frequency of drug screening tests for people should be increased. The traffic police in Qinghai, Tibet, and Xinjiang should always be equipped with fast screening test kits for drugs, which could increase the possibility of identifying DUID offenders.

Ketamine in DUID cases in the greater Cologne area

IntroductionKetamine is primarily used as an anaesthetic or for analgesics in medical treatment, but due to its dissociative and hallucinogenic effects, abuse has increased in the past years leading to several drug impaired driving cases. MethodsEight DUID (driving under the influence of drugs) cases involving ketamine from two institutes of legal medicine over a period from January 2021 to January 2023 were evaluated. The cases were compared with regard to psychomotor impairments, adverse effects on driving performance and co-consumption of drugs. Analyses of ketamine were carried out by high performance liquid chromatography with diode array detection (HPLC-DAD). Other drugs of abuse were either detected via liquid chromatography with tandem mass spectromety (LC–MS/MS) and/or gas chromatography with (tandem) mass spectrometry (GC–MS(/MS)). ResultsKetamine plasma concentrations in a range of approx. 100–1200 ng/mL (mean: 510 ng/mL, median: 370 ng/mL) were detected. Co-consumption of at least one substance was ascertained in all cases. Besides driving impairments, recorded psychomotor impairments of the drivers comprised e.g. dilated pupils, missing or delayed pupil reactions, a slurred or decelerated speech, delayed reaction, lack of concentration, vertigo or agitation. DiscussionThe observed peculiarities were in-line with literature data. However, the assessment and differentiation of ketamine-induced impairments was aggravated due to co-consumption of other drugs of abuse or pharmaceuticals in the herein investigated cases. Nevertheless, in two cases impairments can be attributed mainly to ketamine consumption since the co-consumed substances were only detected in low concentrations. ConclusionThe presented cases provide additional data on psychomotor impairments observed in ketamine-related DUID cases. Limiting factors are co-consumption of substances, unknown habituation to drugs and the limited case number. Nevertheless, the results of this study are comparable with existing literature data. Since the abuse of ketamine has increased in the past years, these data will support forensic casework.

Comprehensive Toxicological Screening of Common Drugs of Abuse, New Psychoactive Substances, and Cannabinoids in Blood Using Supported Liquid Extraction and Liquid Chromatography-Quadrupole Time-of-Flight Mass Spectrometry (LC-QTOF-MS).

Immunoassay (IA) is currently the most widely used technique for toxicological screening in drug impaired driving investigations. However, practical limitations in the scope of testing, and the emergence of new psychoactive substances (NPS), have highlighted the need for alternative approaches, particularly mass spectrometry-based screening. High resolution mass spectrometry (HRMS) broadens the scope of testing to include NPS, and increases analyte specificity compared to IA. In addition, it provides a platform with increased flexibility and adaptability to incorporate emerging drugs of interest due to the transient drug market. In this study, a comprehensive screening procedure was developed to identify more than two hundred drugs of interest, including cannabinoids and NPS in whole blood. Supported liquid extraction (SLE) and liquid chromatography-quadrupole time of flight mass spectrometry (LC-QTOF-MS) using All Ions data acquisition was employed. The method was validated in accordance with published recommendations, and all compounds of interest were identified at recommended cutoffs for driving under the influence of drugs (DUID) investigations. Cannabinoids, including 11-nor-9-carboxy-Δ9-THC (THC-COOH), fentanyl analogs, buprenorphine, novel synthetic opioids (NSOs), and synthetic cannabinoids were identified at low to sub-ng/mL concentrations in whole blood using both positive and negative electrospray ionization (ESI) acquisition methods.

Mitragynine in Orange County DUID Population.

In recent years, mitragynine has been consistently detected in driving under the influence of drug (DUID) cases. In this paper, we evaluate three years (2017-2019) worth of DUID data from arrested drivers in Orange County, CA, USA. From the 25,398 DUID cases received in those three years, there were 60 (0.24%) cases with detectable concentrations, >10 ng/mL, of mitragynine. The majority of the drivers were male (90%), and were stopped during the week (81%), considered Monday 0000 to Friday 1159. The concentration range for all mitragynine cases was 10.5-960 ng/mL, with a mean of 109 ng/mL and a median of 58 ng/mL. Forty-four of the 60 cases were also screened for 7-hydroxymitragynine and 27 (63%) were positive. The police reports and drug recognition expert evaluations were collected and evaluated. No case contained solely mitragynine and the most common drugs detected in combination were central nervous system depressants (ethanol, followed by benzodiazepines), stimulants (methamphetamine and cocaine), and opioids (fentanyl and indication of heroin). Two cases containing only one other psychoactive substance are discussed more thoroughly to attempt to identify the contributions of mitragynine to driving impairment. Collected demographic, toxicological, and field observations are presented for all cases.

Implementation of the First Comprehensive State Oral Fluid Drug Testing Program for Roadside Screening and Laboratory Testing in DUID Cases - A Five-Year Review.

Oral fluid (OF) is a valuable specimen for Driving under the influence of drugs (DUID) applications. This study demonstrates the implementation of the first comprehensive OF drug testing program in the United States, including approved roadside screening OF devices for law enforcement and validated LC-MS-MS confirmation methods. Three roadside OF screening devices were evaluated: the Dräger DrugTest® 5000, Abbott SoToxa®, and Randox Evidence MultiSTAT™. Two qualitative LC-MS-MS confirmation methods were validated per ASB Standard 036. The first method utilized an automated Dispersive Pipette Extraction (DPX) extraction using Integra and Hamilton STARlet platforms for drugs of abuse. The second method used a liquid-liquid extraction to detect cannabinoids. The prevalence of drugs in blood and OF was monitored over five years of casework. Calibration curves were analyzed with each batch to monitor OF concentrations for research purposes. Three roadside OF screening devices were deemed fit for purpose. Devices demonstrated appropriate sensitivity, specificity, positive and negative predictive values, and accuracy above 80% for targeted drugs except for benzodiazepines (DrugTest® 5000) and amphetamine (SoToxa®). The validated LC-MS-MS OF confirmation methods met the National Safety Council recommended cutoffs for 18/21 (86%) of the targets. Over five years of casework, THC and cocaine were detected at a positivity rate of 90% and 97% in OF versus 75% and 44% in blood, respectively. OF: blood ratios exceeded unity for parent drugs. Median concentrations of THC in OF and blood were 31 and 3.5 ng/mL, respectively. OF is a viable alternative or supplemental specimen for DUID investigations. Collecting OF close to the driving event increases the opportunity to identify pharmacologically active substances, and when combined with blood analysis results, an elevated OF: blood ratio provides valuable information for DUID investigation purposes.

The impact of fentanyl on DUIDs and traffic fatalities: Blood and oral fluid data.

Fentanyl has emerged as the most prolific drug in the ongoing opioid epidemic and has greatly impacted traffic safety in recent years. This study aimed to evaluate fentanyl prevalence and concentrations in blood and oral fluid in driving under the influence of drugs (DUID) cases in three different regions (i.e., Alabama, Orange County, CA, and Houston, TX) from 2017 to 2022. Furthermore, traffic fatalities were evaluated for Alabama and Orange County, CA. Fentanyl positivity in DUID and traffic fatalities increased for most years in this study. In Alabama, the prevalence of fentanyl DUID cases increased 4-fold in 2022 compared to 2017. Orange County's increase from 2017 to 2022 was 14-fold. In Houston, the increase was approximately 2-fold from 2019 to 2022. The greatest increase for all laboratories coincided with the start of the COVID-19 pandemic. In 2022, the median fentanyl DUID blood concentrations were 4.7, 11, and 4.7 ng/mL in Alabama, Orange County, and Houston, respectively. Most fentanyl cases were polydrug cases (≥90%). Methamphetamine, THC, and alprazolam were the most frequently detected drugs in combination with fentanyl. Alabama has collected oral fluid and blood in DUID cases since 2018. The detection of fentanyl in oral fluid was comparable to blood. However, 59% and 8.7% of fentanyl-positive cases had concentrations of >20 ng/mL in oral fluid and blood, respectively. Therefore, oral fluid as an alternative or supplemental specimen to blood is an attractive approach for fentanyl in DUID cases. This study contributes to understanding recent fentanyl trends and their impact on traffic safety.

Determination of cannabinoids in 50 μL whole blood samples by online extraction using turbulent flow chromatography and LC-HRAM-Orbitrap-MS: Application on driving under the influence of drugs cases.

The analysis of cannabinoids in whole blood is usually done by traditional mass spectrometry (MS) techniques, after offline cleanup or derivatization steps which can be lengthy, laborious, and expensive. We present a simple, fast, highly specific, and sensitive method for the determination of Δ9 -tetrahydrocannabinol (THC), cannabidiol (CBD), cannabinol (CBN), 11-hydroxy-Δ9 -tetrahydrocannabinol (11-OH-THC), and 11-nor-9-carboxy-Δ9 -tetrahydrocannabinol (THC-COOH) in 50 μL whole blood samples. After the addition of deuterated internal standards (IS) and a simple protein precipitation step, an online extraction of sample supernatants using turbulent flow chromatography (TurboFlow-Thermo Scientific) was carried out. Analytes were separated on a C18 analytical column and detected by LC-HRAM-Orbitrap-MS using a Thermo Scientific Q Exactive Focus MS system. MS detection was performed in polarity switching and selected ion monitoring (SIM) modes using five specific acquisition windows, at a resolution of 70,000 (FWHM). Total run time was about 10min including preanalytical steps. Method validation was carried out by determining limit of detection (LOD), lower limit of quantitation (LLOQ), linearity range, analytical accuracy, intra-assay and interassay precision, carry-over, matrix effect, extraction recovery, and selectivity, for all analytes. Measurement uncertainties were also evaluated, and a decision rule was set with confidence for forensic purposes. The method may become suitable for clinical and forensic toxicology applications, taking advantage of the small matrix volume required, the simple and cost-effective sample preparation procedure, and the fast analytical run time. Performances were monitored over a long-term period and tested on 7620 driving under the influence of drugs (DUID) samples, including 641 positive samples.

Increasing Prevalence of Designer Benzodiazepines in Impaired Driving; a 5 Year Analysis from 2017-2021.

Designer benzodiazepine (DBZD) use has been increasing over the past decade and poses a threat to human health and safety, particularly when involved in driving under the influence of drugs (DUID) cases. Over a five-year period between 2017 and 2021, there were 1145 reported DBZDs in 805 blood samples submitted from law enforcement agencies for DUID testing. Eleven different DBZD were detected, including three metabolite pairs: etizolam/alpha-hydroxyetizolam, clonazolam/8-aminoclonazolam, diclazepam/delorazepam, flualprazolam, flubromazolam, flubromazepam, bromazolam, and bromazepam. Etizolam/alpha hydroxyetizolam (n=485) and flualprazolam (n=149) were the most frequently detected DBZD, at 60% and 18%, respectively. Driving behavior, standardized field sobriety tests (SFST) performance, and physical observations of individuals suspected of DUID, whose blood was toxicologically confirmed for one or more DBZD, were consistent with effects caused by central nervous system (CNS) depressants. Each DBZD has its own unique timeline, and toxicology testing had to be frequently updated to reflect the state of the novel psychoactive substance (NPS) market. DBZD play a role in impaired driving and can be the sole intoxicant in DUID cases.

Ethanol enhances JWH-018-induced impairment of sensorimotor and memory functions in mice: From preclinical evidence to forensic implication in Driving Under the Influence of Drugs

BackgroundSeveral new Synthetic Cannabinoids have appeared each year since their introduction into the illicit drug market as recreational drugs. Among these, naphtalen-1-yl-(1-pentylindol-3-yl) methanone (JWH-018) is one of the most detected compounds in biological samples from patients involved in intoxication or death cases. Furthermore, consumption of JWH-018 has been linked to several cases of Driving Under the Influence of Drugs (DUID) suggesting that effects induced by this compound can affect individuals’ ability to drive. MethodsGiven the high spread of polydrug consumption and the wide number of alcohol-related traffic accidents, this study aims to investigate the acute effects induced by co-administration of JWH-018 with ethanol on sensorimotor and motor responses, grip strength and memory functions in CD-1 male mice. Acute impairments induced by JWH-018 and ethanol alone have also been investigated, in order to compare their effects with that induced by their concurrent administration. ResultsIn vivo behavioral experiments revealed a worsening of the cognitive and sensorimotor disruption after the co-administration of JWH-018 with ethanol compared to single compounds. ConclusionsThese animal-based findings suggest a potential increased impairment on psychomotor performances which could be related to driving abilities posed by poly-drug consumption involving SCs and ethanol.

Driving under the Influence of Amphetamine: Analytical Evaluation of Illegal or Prescription Drug Intake Using Chiral UHPLC-MS-MS.

Differentiation between consumption of illegal and prescription drugs remains an important aspect in forensic toxicology. While illicit amphetamine is most often racemic, the medicinal drugs marketed in Denmark for the treatment of attention-deficit hyperactivity disorder contain the pure (S)-enantiomer or a prodrug thereof. In this study, we present a simple and efficient analytical workflow to provide information about the origin of amphetamine consumed in forensic cases concerning driving under the influence of drugs (DUID). Following quantification of amphetamine and methamphetamine using our conventional multi-target ultra-high performance liquid chromatography-tandem mass spectrometry method, determination of (R)- and (S)-amphetamine was performed by reinjecting the sample extract on a Phenomenex LUX® AMP chiral column using the same analytical instrument and mobile phases. Chiral separation was performed isocratic within a run time of 6 min. The analytical workflow was applied to blood samples from 5,248 suspected DUID cases within a 2-year period. Amphetamine was detected in 18.7% of the samples, of which both enantiomers were detected in 89.5% of the cases, indicating the consumption of illegal racemic amphetamine. In 6.1% of the positive cases, both amphetamine and methamphetamine were detected, indicating either co-consumption of both amphetamines or consumption of methamphetamine. In the remaining 4.4%, only (S)-amphetamine was detected indicating the consumption of prescription drugs containing (S)-amphetamine or a prodrug thereof. Implementation of a simple and rapid chiral method in the conventional analytical workflow for routine forensic casework proved to be an efficient way to elucidate whether a positive amphetamine result originates from illegal or prescription drug consumption, without increasing turnaround time nor costs to any significant extent, as no additional sample preparation was required.

Combined Use of Flubromazepam and Stimulants: Blood and Oral Fluid Concentrations and Impact on Driving Ability.

"Designer" benzodiazepines (DBZDs) are becoming increasingly available in Europe, with the European Monitoring Centre of Drugs and Drug Addiction currently monitoring ∼30 new benzodiazepines. The following driving under the influence of drug (DUID) case describes the oral fluid (OF) and blood concentrations, as well as the observed effects after the combined use of stimulants and flubromazepam. Both OF, collected via the Intercept i2 collector (Immunalysis, Pomona, CA, USA), and blood (collected in containers with various stabilizers) were screened using a liquid chromatographic (LC) time-of-flight (TOF) mass spectrometric (MS-MS) method. In addition, various LC-MS-MS methods in multi-reaction monitoring mode were applied for confirmation and quantification. The OF and blood samples were taken 2 h 25 min and 9 h 19 min after the accident, respectively. OF contained 789 ng/mL amphetamine, 5,173 ng/mL MDMA, 168 ng/mL benzoylecgonine, 492 ng/mL cocaine, 134 ng/mL 4-methylmethcathinone (4-MMC) and traces of flubromazepam (less than limit of quantification (LLOQ); 2 ng/mL). The sodium-fluoride blood samples contained 19 ng/mL amphetamine, 284 ng/mL MDMA, 20 ng/mL MDA, 38 ng/mL benzoylecgonine, 4 ng/mL methylecgonine, 161 ng/mL flubromazepam and traces of 4-MMC (<LLOQ; 2.5 ng/mL). The driver was observed to have an irregular speed driving pattern and could not keep his lane. He demonstrated the following effects after the accident: bloodshot eyes, red face, sweating, fatigue, disorientation in time and space and mental confusion. Even 24 h after the accident, the driver was confused, disoriented, had red spots on his face and could not keep his balance. The effects of flubromazepam combined with several stimulants are demonstrated. Moreover, this case illustrates well the pros and cons of the different biological matrices applied in a DUID context. Differences between the biological matrices are not only observed concerning the ease/practicality of (on-site) collection, but also in the final drug detectability due to the large variations in OF/blood drug concentration ratios and metabolism/elimination rates as a result of the different chemical entities of the compounds.

Tetrahydrozoline: Death by Eyedrops

Tetrahydrozoline (THZ) is an alpha-adrenergic receptor agonist most commonly used in eyedrops to reduce redness. Reported plasma concentrations are less than 1 ng/mL after normal use. Adverse effects following oral ingestion include drowsiness, tremors, irregular heartbeat and respiratory depression. Several cases of individuals surviving accidental THZ exposure and drug facilitated crimes victims rendered unconscious by the drug have been reported. The purpose of this study is to summarize THZ concentrations from cases submitted to a reference laboratory and examine its role in death cases. Data were extracted from NMS Labs’ (Horsham, PA, USA) Laboratory Information Management System for all cases received from December 2010 to December 2021. Cases with a reported blood THZ concentration were classified as postmortem (PM), driving under the influence of drugs (DUID), drug facilitated crime (DFC) or unknown. The mean, median and concentration ranges for each group were determined. The number and types of other drugs reported were also examined in cases which underwent comprehensive testing. 129 cases were positive for THZ. The reported concentrations are summarized below: – Case Type N Mean (ng/mL) Median (ng/mL) Range (ng/mL) – PM 112 20 1.7 0.19–290, – DUID 1 0.64 0.64 -, – DFC 3 0.35 0.35 <0.10–0.52, – UNKNOWN 13 18 1.2 0.27–130. Comprehensive testing was performed on approximately 100 cases. The most commonly detected other drugs were ethanol, Delta-9 THC), diphenhydramine, fentanyl, amphetamine, and morphine. THZ concentrations ≥100 ng/mL were reported in 10 cases. One case (THZ = 100 ng/mL) was from a living individual and the status of the patient could not be determined in another case (THZ = 130 ng/mL). The remaining cases included a dual suicide, one sudden death, one “suspected poisoning”, one homicide and one case still under active investigation. A woman admitted to lacing her boyfriend's water with tetrahydrozoline on multiple occasions from 2009–2012 to get him to pay more attention to her. The man, who survived, sought medical attention for trouble breathing, irregular blood pressure and nausea and had a blood THZ concentration of 49 ng/mL. The THZ concentration in cardiac blood from a decedent whose death was originally believed to be due to natural causes was 68 ng/mL. THZ, an uncommon finding in toxicology testing, has been reported in multiple homicide, suicide and drug facilitated crimes highlighting the importance of its inclusion in comprehensive post-mortem toxicology testing.

Occurrence of GHB in blood and urine specimens from victims of alleged sexual assault or blackout in Western Switzerland

Although much has been written on the subject of drug facilitated sexual assault (DFSA) or victim of blackout, no epidemiological study has yet been conducted in Switzerland to evaluate the occurrence of GHB and other drugs in this context. GHB is frequently mentioned on social networks and in the media as the most regularly used substance in DFSA, but often without being based on scientific studies. In order to have an assessment of the situation, political authorities asked our laboratory to systematically screen for GHB in samples obtained in case of DSFA or blackout, and driving under the influence of drugs (DUID), with the aim to assess the occurrence of voluntary or involuntary GHB use in Western Switzerland. All DFSA, blackout and DUID cases received in 2021 in our laboratory were routinely tested for the presence of GHB at non-physiological concentrations by enzymatic assay with a urine cut-off of 10 mg equivalent GHB/L. All positive cases were confirmed by gas chromatography coupled to mass spectrometry (GC-MS) using a cut-off of 6 mg/L in urine and 1.5 mg/L in whole blood. All DFSA and blackout cases were also analyzed by GC-MS, to ensure the absence of false negative cases. When requested by the justice, other substances such as ethanol, benzodiazepines, cannabinoids, cocaine, amphetamines, opioids, antidepressant, and other psychoactive substances were screening in urine and/or blood, by immunoassays, head-space gas chromatography coupled to flame ionization detector (HS-GC-FID), GC-MS, and/or liquid chromatography coupled to tandem mass spectrometry or high resolution mass spectrometry (LC-MS/MS; LC-HRMS (orbitrap)). 60 DFSA and blackout cases and 633 DUID cases were included in the study. Women predominated over men in the DFSA and blackout group (77%), while men predominated over women in the DUID group (90%). Median age in the DFSA and blackout group was 23, and median age in the DUID group was 30. The median delay to obtain urine after the event was 12 hours in case of DFSA and blackout, and 2 hours for DUID cases. The median delay to obtain whole blood after the event was 12 hours in case of DFSA and blackout, and 1.5 hours for DUID cases. The presence of GHB at concentration above the cut-off was detected in 1 DFSA and blackout case (130 mg/L in blood and 730 mg/L in urine), and in 3 DUID cases (78 mg/L, not detected, and not detected in blood, and 660 mg/L, 8.6 mg/L, and 25 mg/L in urine, respectively). These cases concerned only men. For these 4 cases, other substances were also detected. Thus, for the DFSA and blackout case, methamphetamine, methylphenidate, zolpidem and pregabaline were detected, and in the DUID cases, methamphetamine was detected in one case, ethanol, cocaine and cannabis were detected in another case, and 3-MMC was detected in the last case. When analyzed, ethanol was detected in 48% of DFSA and blackout cases, cannabinoids in 17%, benzodiazepines and Z-drugs in 10%, antidepressant in 9.8%, cocaine in 6.5%, amphetamines in 6.5%, methylphenidate in 4.8%, and opioids in 2.1%. Median concentration of ethanol after back calculation was 2.07 g/kg in DFSA and blackout cases. The results obtained, showing a very limited voluntary or involuntary use of GHB, are comparable to those obtained in other international studies. For the first time in Switzerland, a study was conducted for the evaluation of the occurrence of GHB in blood and urine specimens from victims of alleged sexual assault or blackout, and DUID cases. The results showed that GHB is very rarely consumed and used in DSFA or blackout cases, in comparison to other psychoactive drugs.

Driving under the influence of drugs (DUID) profiling in Spain: Data from oral fluid results between 2016 and 2022, including the effect of COVID-19 lockdown

To provide information about driving under the influence of drugs (DUID) cases in Spain during the last 6 years and to evaluate the impact of the Spanish lockdown (March 14th to May 6th 2020) on drivers’ consumption profile. In total, 16,461 oral fluid samples were collected from drivers in Spain that tested positive on the roadside between January 1st 2016 and April 15th 2022. Samples were collected at different locations throughout the country and analyzed by LC-MS/MS at the Toxicology Service of the Institute of Forensic Sciences of the University of Santiago de Compostela. For data analysis, drugs and their metabolites were classified in 8 different groups (cannabis, cocaine, amphetamines, opioids, benzodiazepines, ketamine, methadone and zolpidem). Demographic characteristics and drug profile were evaluated during time. The impact of COVID-19 pandemic lockdown was also considered, applying Chi 2 test. Drivers were mainly young adults (33.3 ± 9.9 years old) and men (94.2% male). In 97.6% of cases, LC-MS/MS results confirmed at least one of the positive results detected on the roadside. Cannabis (72.6%) and cocaine (64.4%) were the most detected drugs, followed by amphetamines (13.8%), opioids (12.9%), benzodiazepines (7.7%), and methadone (7.4%). Polydrug use was observed in 55.1% of drivers, being the combination of substances from two different groups the most prevalent. Cannabis and cocaine were the most common combination (either only the two of them or mixed with more substances). Changes in drivers’ consumption profile was noticed when comparing data from the specimens collected during the Spanish lockdown period ( n = 151) vs. specimens obtained during 2020 before (“pre-lockdown”, n = 557) and after (“post-lockdown”, n = 1974) that period. During the lockdown period drivers were 3 years older in average, and polyconsumers of 4 and 5 different groups of substances doubled (in average, 4.5% vs. 9.9% for 4 groups; 0.7% vs. 1.3% for 5 groups). Nevertheless, results were only statistically different for consumers of 4 groups of substances during lockdown, either compared with pre- and post-lockdown results. Also, for polyconsumers, the combination of cannabis and cocaine with amphetamines, with high prevalence before lockdown, was substituted by the combination of cannabis and cocaine with methadone. Furthermore, opiates use was two times higher (in average, 14.0% vs. 24.5%), being this difference statistically significant when comparing the 3 periods. On the other hand, amphetamines and ketamine were less consumed by drivers during lockdown (in average, 11.2% vs. 9.9% for amphetamines; 3.2% vs. 0.7% for ketamine). Nevertheless, these differences were only statistically significant when considering ketamine consumption post-lockdown (either compared with pre-lockdown or lockdown results). The use of oral fluid samples was able to confirm the vast majority of the screening results found in roadside screening tests. In Spain, cannabis and cocaine remain the most detected drugs on the roadside, as previously reported from our group in the period 2013–2015 (Lema-Atán JA. Drug Alcohol Depend 2019;195:106–113). During the lockdown period in Spain, due to stay-at-home orders, mobility was only allowed for “essential workers”. Samples collected in those months showed an increase in polydrug use, especially for opiates and methadone, as well as a decrease in the use of “party drugs” (amphetamines and ketamine). LC-MS/MS confirmation result was negative in only 2.4% of the cases. The DUID driver profile was a young man, consuming either cannabis or a combination of cannabis and cocaine, alone or combined with more substances. During lockdown, drivers consumed less “party drugs” and more opiates and methadone. Moreover, there was a higher rate of polyconsumers (4 or 5 groups of substances).

Ketamine in DUID cases in the greater Cologne area. A case series

Cases of ketamine consumption in driving under the influence of drugs (DUID) were evaluated with regard to recorded psychomotor impairments and possible co-consumption of drugs of abuse or other substances. Ketamine is a substance that is used e.g. as an anaesthetic or for analgesics in medical treatment. Due to its dissociative effects, ketamine abuse has increased in the past years leading to several drug-impaired driving cases. According to literature, ketamine plasma concentrations above 50 ng/mL are associated with significant psychomotor impairments, whereby mind-altering effects can be expected at plasma concentrations of ketamine between 50 and 200 ng/mL. They seem to be concentration dependent. The most commonly described impairments comprise a slurred speech, drowsiness, confusion and vomiting as well as lack of coordination and dystonia, blurred vision or visual field narrowing, dissociation, referential thinking, euphoria, depersonalization and anxiety. Moreover, pupil dilatation, a lack of convergence, horizontal gaze nystagmus, elevated pulse rate and failure of divided attention tests like walk-and-turn and one-leg stand test were observed. DUID cases from two institutes of legal medicine from the years 2021 to early 2022 were evaluated with regard to drug-impaired drivers who consumed ketamine. The cases were compared with regard to psychomotor impairments, salience in driving and co-consumption of drugs. Technical devices for the analyses of drug concentrations in blood were drug-dependent and therefore either performed by GC-MS/MS, LC-MS/MS or HPLC-DAD. Five cases of drivers (all male, between 21 and 44 years old) were collected. Ketamine consumption was admitted in two out of five cases. Indications for a medical application were not given in the other three cases without confessed consumption. Ketamine concentrations of approx. 165 ng/mL, 245 ng/mL, 610 ng/mL, 676 ng/mL and 3800 ng/mL were detected in the serum samples. Moreover, co-consumption of either one or multiple substances like cannabis ( n = 2), amphetamine ( n = 4), 3,4-methylenedioxymethamphetamine (MDMA) ( n = 1), cocaine ( n = 2) and clonazepam ( n = 1) could be ascertained in all cases. The recorded psychomotor impairments of the drivers comprised e.g. dilated pupils ( n = 2), missing or delayed pupil reactions ( n = 3), as well as aggressive behaviour ( n = 3), a slurred or decelerated speech ( n = 2), a delayed reaction ( n = 2), lack of concentration ( n = 2) or vertigo ( n = 3). The observed psychomotor impairments correlated with literature data. However, the assessment and differentiation of impairments with regard to ketamine consumption is aggravated due to co-consumption of other drugs of abuse or substances. In literature, co-consumption of e.g. alcohol, cannabis, MDMA, amphetamine, benzodiazepines or cocaine is also described when ketamine is abused, which is also in line with the collected data. The extremely high concentration of ketamine (approx. 3800 ng/mL) in one case from the present case series, might allow the supposition that the recorded impairments are most likely due to ketamine consumption, although cocaine and its metabolite benzoylecgonine (BE) were also detected in moderate concentrations (cocaine = 32 ng/mL, BE = 729 ng/mL) as well. However, a decent distinction of the impairments is not possible. The presented case series provides additional data on psychomotor impairments observed in DUID cases after the consumption of ketamine, although the results have to be regarded critically due to co-consumption of other substances and the limited case number. Nevertheless, correlation between existing literature data could be observed. Since the abuse of ketamine has increased in the past years, those data can support the assessment of cases, where ketamine abuse is in question.

Automating DUID methods using robotics for rapid high throughput sample preparation – Section 4 and Section 5A UK Road Traffic Act Compounds in Whole Blood

To fully automate a 96-well plate sample preparation technique using liquid handling robotics for the purpose of significantly increasing sample throughput for whole blood samples taken under the UK Road Traffic Act (RTA). Introduction: demand for analysis of whole blood samples taken under the Road Traffic Act 1988 has increased in recent years due to UK Policing and Government policy on tackling driving under the influence of drugs (DUID). Previous extraction techniques utilised by our laboratory include manual or semi-automated approaches that are lengthy and labour intensive. Both approaches require time consuming human input, to precipitate blood and prepare calibrators and quality controls. Recent developments have sped up chromatographic methods through use of techniques such as ultra-high performance liquid chromatography. However, there has been little progress with speeding up sample preparation and extraction processes, particularly for whole blood to compliment increased LC-MS/MS capacity. Use of 96-well plate formats for protein precipitation and extraction provide a platform that can be readily coupled to automated liquid handling (ALH) instruments. This provides a solution to the increase in demand by allowing for extraction of more samples, with minimal human input, in a much shorter amount of time. Here we present development, validation and implementation of an automated robotic sample preparation method for whole blood for analysis of drugs in RTA cases. Aliquots (200 μL) of blank blood and casework samples were dispensed into a 96-well plate using a robotic pipetting system before being loaded onto a Hamilton ALH. Casework samples and blank blood were spiked with internal standard. Additional blank blood samples were further spiked to produce calibrants and quality controls. The plate was mixed with the addition of a cell lysis solution and the aliquots transferred to a protein/lipid depletion (PPT) plate. A positive pressure manifold was used to draw the solution through the PPT plate into a collection plate where the precipitate was dried. The precipitate was reconstituted and loaded by the Hamilton onto a microelution SPE plate. Following extraction, the samples were reconstituted and the collection plate placed in the LC-MS/MS autosampler for analysis. The LC-MS/MS method consists of a 2.1 mm × 30 mm, 1.7 μm, C18 column running a 4 minute gradient of water and methanol with 0.1% formic acid. Quantitation of 34 compounds (including common drugs of abuse and their metabolites) was successfully validated with CVs ranging from 3.0%–9.0% and accuracies of 85.8%–111.7%. Within batch precision had CVs of < 10.0%. The robotic platform was able to undertake sample pre-treatment and extraction of 40 case samples in 3.5 hours. The method was implemented into casework following UKAS accreditation and in 2021, 12,400 samples were analysed using the method. Introduction of this method to our laboratory has resulted in a significant increase in speed for a sample to be processed and results obtained. Each ALH is capable of extracting two batches per day; equating to 160 case sample versus 28 cases on semi-automated methods. This, along with a reduced sample volume requirement and reduced solvent consumption has had a very beneficial impact in terms of cost, turnaround times and ability to use low sample volume cases. Full automation of sample preparation of whole blood samples coupled with high speed chromatographic techniques can significantly increase throughput, decrease turnaround times and reduce solvent consumption.

Automating DUID methods using robotics for rapid high throughput sample preparation – Analysis of Delta-9-Tetrahydrocannabinol and its major metabolite Delta-9-tetrahydrocannabinol acid in whole blood

To fully automate, using liquid-handling robotics, a liquid chromatography tandem mass spectrometry method for the analysis of delta-9-tetrahydrocannabinol (THC) and delta-9-tetrahydrocannabinol acid (THC-A) in whole blood. Uptake in usage of ultra-high performance liquid chromatography (UHPLC) has led to development of methods with very fast (typically < 5 minutes) chromatographic run times used in the forensic toxicology laboratory. Nevertheless, use of whole blood for quantitative analysis continues to present challenges for delivering high throughput methods with rapid analysis times. Whole blood requires the use of solid phase extraction (SPE) for sample pre-treatment and preparation, which is time consuming and labour intensive. Recent developments in SPE technologies have produced 96-well format platforms that are used to extract greater number of samples in parallel and are ideally suitable for automated liquid handlers. With recent sensitivity increases in mass spectrometers, smaller volumes (< 250 μL per aliquot of blood) can be used for extraction in 96-well plates. This facilitates the use of robotic platforms together with rapid liquid chromatography tandem mass spectrometry (LC-MS/MS) methods to give truly high throughput and fast analytical methods, increasing method efficiencies, reducing human intervention and errors. Here we present the development, validation and implementation of a fully automated robotic sample preparation method for whole blood for analysis of THC and THC-A in UK driving under the influence of drugs (DUID) cases. Aliquots (200 μL) of blank blood and casework samples previously dispensed into 96-well plates by a robotic system (BRAND) were prepared by the liquid handling system (Hamilton). Casework samples and blank blood were spiked with internal standard and blank blood samples were further spiked to produce calibrants and quality controls. The plate is then mixed with the addition of a cell lysis solution and the aliquots transferred to a protein/lipid depletion (PPT) plate (Waters). A positive pressure manifold was used to draw the solution through the PPT plate into a collection plate where the precipitate was dried. The precipitate was reconstituted and loaded by the Hamilton onto a C18 microelution SPE plate (Waters). Following extraction, the samples are reconstituted and the collection plate is placed in the autosampler for analysis. The LC-MS/MS method consists of a 2.1 mm × 50 mm, 1.7 μm, C18 column running a 4 minute gradient of water and acetonitrile with 0.1% formic acid. The method was successfully validated, with LLOQs for THC and THC-A of 1 μg/L and 5 μg/L, respectively. Precision (%CV) for THC was < 8% and < 6% for THC-A. Accuracy was within 15% of the spiked concentration for THC and THC-A across all validation batches ( n = 11). The robotic platform was able to undertake sample pre-treatment and extraction in 3.5 hours. A batch of 40 casework samples can be completed in 13 hours including the LC-MS/MS run time. The introduction of this method to our laboratory has resulted in a significant increase in the speed for a sample to be processed and results obtained. We can prepare and extract double the amount of samples in less than half the time taken by our manual method. We have reduced the amount of sample used and decreased our turnaround times all for our customer's benefit. Full automation of the sample preparation of whole blood samples using liquid handling robots can significantly improve laboratory throughput and capacity and efficiently utilise high speed chromatography providing rapid analysis from start to finish.