Illegal Substance Analysis and Environmental Risk Assessment in Küçük Menderes River, Important Basin of the Aegean Region.

Water-based studies have come into prominence for illegal substance monitoring. There are limited studies on the detection of these substances in the surface waters as opposed to wastewater. This study aims to evaluate amphetamine, benzoylecgonine, cocaine, codeine, 3,4-methylenedioxy-N-methylamphetamine, morphine, and 11-nor-Δ9- tetrahydrocannabinol-9-carboxylic acid in the Küçük Menderes (KM) River at four different stations during three different periods and assess the environmental risks. Environmental risk assessment of psychoactive substances based on the calculation of Risk Quotients (RQ) were evaluated. RQ values for amphetamine and benzoylecgonine were < 0.01 at all sampling points during all sampling periods, indicating little risk for aquatic life. Cocaine was found as environmentally high risk (RQ > 1) based on its RQ values of 1.22 and 1.37 at KM-01 and KM-02 sampling points. This research is the first report to investigate the presence of psychoactive substances and define the environmental risks of these substances in Türkiye.

Truncations and in silico docking to enhance the analytical response of aptamer-based biosensors.

Aptamers are short oligonucleotides capable of binding specifically to various targets (i.e., small molecules, proteins, and whole cells) which have been introduced in biosensors such as in the electrochemical aptamer-based (E-AB) sensing platform. E-AB sensors are comprised of a redox-reporter-modified aptamer attached to an electrode that undergoes, upon target addition, a binding-induced change in electron transfer rates. To date, E-AB sensors have faced a limitation in the translatability of aptamers into the sensing platform presumably because sequences obtained from Systematic Evolution of Ligands by Exponential Enrichment (SELEX) are typically long (>80 nucleotides) and that obtaining structural information remains time and resource consuming. In response, we explore the utility of aptamer base truncations and in silico docking to improve their translatability into E-AB sensors. Here, we first apply this to the glucose aptamer, which we characterize in solution using NMR methods to guide design and translate truncated variants in E-AB biosensors. We further investigated the applicability of the truncation and computational approaches to four other aptamer systems (vancomycin, cocaine, methotrexate and theophylline) from which we derived functional E-AB sensors. We foresee that our strategy will increase the success rate of translating aptamers into sensing platforms to afford low-cost measurements of molecules directly in undiluted complex matrices.

A pilot study on post-mortem determination of drug abuse on dental tissues.

BACKGROUND: Post-mortem toxicology constantly deals with the research of reliable alternative matrices to be applied in case of highly damaged corpses (such us carbonized, skeletonized, human remains, etc.). Teeth represent a promising alternative matrix since dental tissues are endowed by different features, resistance and stability after death. SCOPE: Since scant literature reported on the pharmacokinetics and mechanism of incorporation of xenobiotics into dental tissues, this pilot research aims to investigate whether in the pulp can be detected the same substances found in blood in drug related death cases. Secondly, the study is addressed to disclose the possible deposit of drugs in dental hard tissues (dentine and/or enamel), thus contributing to reconstruct the drug abuse history (timing, e.g.). MATERIALS AND METHODS: The study experimented with a novel method to separately analyse dental enamel, dentin, and pulp, applied to 10 teeth collected during autopsies of drug-related deaths along with blood and hair samples for classic toxicological analyses. Each tooth was prepared by "pulverization technique" and then analysed by gas chromatography paired with mass spectrometry (GC-MS) and ultra high performance liquid chromatography coupled to high resolution mass spectrometry (UHPLC/HR-MS) for searching cocaine, opiates, and metabolites. The results were then compared with those obtained from blood and hair samples. RESULTS: Preliminary results demonstrated that teeth differ from any other classic matrix (blood and hairs) since the qualitative correspondence of the detected substances between pulp and blood as well as dental hard tissues and hair suggests that they can be useful in post-mortem evaluation as a unique matrix for both acute and chronic assumptions of drugs. The mechanism of accumulation of substances in mineralized dental tissues emerged the most significant result, being influenced by the type of molecule and the method of assumption. The main limitation of this study is the limited availability of the sample and the absence of anamnestic information of the time, rates and method of drug assumption during life. Further research is necessary to systematically investigate the distribution of different substances within the different tissues of the tooth.

Contamination of a drug consumption room with drugs and potential risks for social health care workers.

BACKGROUND: Studies have shown that contamination of surfaces by illicit drugs frequently occurs in forensic laboratories when manipulating seized samples as well as in pharmacies and hospitals when preparing medicinal drugs. In this project, we extended these studies to a Drug Consumption Room to investigate drug levels and possible exposure of the staff members. METHODS: We investigated pre and post cleaning contamination by heroin and cocaine and their degradation products 6-monoacetylmorphine and benzoylecgonine on different surfaces (tables, counters, computers and door handles) and in the ambient air. We also collected urine and hair samples from staff members to check for potential short and long term contaminations. RESULTS: Medium to heavy contamination has been detected on most surfaces and door handles; as expected, air contamination was particularly high in the smoking room. Drug levels were < LOD to very low in the urine and the hair samples of staff members tested. CONCLUSION: The cleaning efficiency of the surfaces, carried out by staff and drug users after drug consumption, was often not satisfactory. The very low drug levels in hair indicate that acute health risks for staff members are low.

Analysis of crude wastewater from two treatment plants in South Wales for 35 new psychoactive substances and cocaine, and cannabis.

This study investigates the presence of new psychoactive substances (NPS) and their metabolites in two wastewater treatment plants (WWTPs) situated in South Wales, UK (WWTP-1 and WWTP-2). Analysis was conducted for 35 NPS and metabolites, along with the inclusion of benzoylecgonine (main cocaine metabolite) and cannabis, the most detected illicit substances. Benzoylecgonine was identified as the predominant substance in both WWTPs. Epidemiological calculations revealed the average population consumption of cocaine to be 3.88 mg/d/1000 inhabitants around WWTP-1 and 1.97 mg/d/1000 inhabitants for WWTP-2. The removal efficiency of benzoylecgonine across both WWTPs was observed at an average of 73%. Subsequent qualitative analyses on randomly selected wastewater samples detected medicinal compounds including buprenorphine, methadone, and codeine in both WWTPs. An additional experiment employing enzymatic hydrolysis revealed the presence of morphine, an increased presence of codeine, and 11-Nor-9-Carboxy-THC (THC-COOH) post-hydrolysis. These findings underscore the significant presence of illicit substances and medicinal compounds in wastewater systems with the absence of NPS within the South Wales area, highlighting the necessity for enhanced monitoring and treatment strategies to address public health and environmental concerns.

Rapid nanomolar detection of cocaine in biofluids by electrochemical aptamer-based sensor with low-temperature effect for drugged driving screening.

Cocaine is one of the most abused illicit drugs, and its abuse damages the central nervous system and can even lead directly to death. Therefore, the development of simple, rapid and highly sensitive detection methods is crucial for the prevention and control of drug abuse, traffic accidents and crime. In this work, an electrochemical aptamer-based (EAB) sensor based on the low-temperature enhancement effect was developed for the direct determination of cocaine in bio-samples. The signal gain of the sensor at 10 °C was greatly improved compared to room temperature, owing to the improved affinity between the aptamer and the target. Additionally, the electroactive area of the gold electrode used to fabricate the EAB sensor was increased 20 times by a simple electrochemical roughening method. The porous electrode possesses more efficient electron transfer and better antifouling properties after roughening. These improvements enabled the sensor to achieve rapid detection of cocaine in complex bio-samples. The low detection limits (LOD) of cocaine in undiluted urine, 50% serum and 50% saliva were 70 nM, 30 nM and 10 nM, respectively, which are below the concentration threshold in drugged driving screening. The aptasensor was simple to construct and reusable, which offers potential for drugged driving screening in the real world.

Assessing daily patterns in stimulant use during the COVID-19 pandemic in Melbourne, Australia using wastewater analysis.

During the COVID-19 pandemic, one of Australia's biggest cities, Melbourne, experienced three major isolation ("lockdown") periods in 2020 (160 days) and in 2021 (111 days) which makes it one of the most locked down cities world-wide. This study assessed how the pandemic affected temporal trends in methamphetamine, MDMA and cocaine consumption using wastewater-based epidemiology. Daily samples were collected for most of 2020 and 2021 (n = 660 days). Concentrations were measured using direct-injection LC-MS/MS and back-calculated to consumption estimates. Results indicate that methamphetamine use was increasing before the first lockdown and decreased after the end of the first lockdown in 2020. Methamphetamine trends appeared to have remained steady throughout the second lockdown period before increasing steeply after it ended. For most of 2020, cocaine use remained steady, with an increase after the second lockdown. MDMA use decreased after the start of the first lockdown and remained steady throughout most of 2020 and 2021. In comparison to 2020, trends in 2021 were less variable and stimulant use did not appear to be as associated with COVID-19 restrictions. Overall, this study was able to show the impact of lockdown periods and the related social restrictions on illicit stimulant use. ENVIRONMENTAL IMPLICATION: Illicit drugs are hazardous chemicals, of concern both to humans and the environment. While studies have been undertaken to understand their temporal trends, this work utilizes wastewater-based epidemiology and daily sampling to provide a comprehensive understanding of the impact of the COVID-19 pandemic on the use of methamphetamine, MDMA and cocaine on one of the most locked-down cities in the world. Understanding the consequences of this significant intervention on illicit drug use could provide valuable insights into its potential environmental impact.

"Cocaine Shark": First report on cocaine and benzoylecgonine detection in sharks.

Cocaine (COC) and benzoylecgonine (BE), the main COC metabolite, have been detected in aquatic ecosystems. Studies focusing on wild fish are, however, very limited, and no reports concerning elasmobranchs are available. This study investigated COC and BE levels in Brazilian Sharpnose sharks (Rhizoprionodon lalandii) (n = 13) using LC-MS/MS. All samples (13/13) tested positive for COC, with 92 % (12/13) testing positive for BE. COC concentrations (23.0 µg kg-1) were over 3-fold higher than BE (7.0 µg kg-1). COC levels were about three-fold significantly higher in muscle (33.8 ± 33.4 g kg-1) compared to liver (12.2 ± 14.2 µg kg-1). Females presented higher COC concentrations in muscle (40.2 ± 35.8 µg kg-1) compared to males (12.4 ± 5.9 µg kg-1). Several positive statistical correlations were noted between COC and BE (rho = 0.84) in females, indicating systemic COC transport and metabolization, as well as between BE and weight (rho = 0.62), and between COC and the Condition Factor (rho = 0.73). A strong correlation was noted between BE and COC in the muscle of non-pregnant females (rho = 1.00). This study represents the first COC and BE report in free-ranging sharks, and the findings point to the potential impacts of the presence of illicit drugs in environments.

A systematic approach to the analysis of illicit drugs for DNA with an overview of the problems encountered.

Due to the restricted nature of illicit drugs, it is difficult to conduct research surrounding the analysis of this drug material for any potential DNA in sufficient quantities acceptable for high numbers of replicates. Therefore, the current research available in peer reviewed journals thus far regarding analysing illicit drugs for DNA has been performed under varying experimental conditions, often using surrogate chemicals in place of illicit drugs. The data presented within this study originated from the analysis of genuine illicit drugs prepared both in controlled environments and those seized at the Australian border (and therefore from an uncontrolled environment) to determine if DNA can be obtained from this type of material. This study has been separated into three main parts (total n=114 samples): firstly, methamphetamine synthesised within a controlled environment was spiked with both saliva and trace DNA to determine the yield following DNA extraction; secondly, methamphetamine also synthesised in a controlled environment but on a larger scale was tested for the amount of DNA added incidentally throughout the synthesis, including the additional steps of recrystallising, homogenising and "cutting" the drug material to simulate preparation for distribution; and thirdly, the detection of human DNA within samples of cocaine and heroin seized at the Australian border. The DNA Fast Flow Microcon Device was utilised to concentrate all replicates from the same source into one combined extract to improve the DNA profiles for the samples where no DNA spiking occurred. Full STR profiles were successfully obtained from drug samples spiked with both saliva and trace DNA. Methamphetamine was present in the final DNA extracts and caused incompatibilities with the quantification of DNA using Qubit. The yields of DNA from drugs not spiked with DNA sources were much lower, resulting in 36 % of samples yielding alleles where all others did not. These results were not unexpected given these were realistic drug samples where the history of the drug material was unknown. This is the first study to obtain DNA profiles from genuine illicit drug material in both controlled and uncontrolled environments and indicates that the analysis of illicit drugs for DNA is an avenue worth pursuing to provide information which can in turn assist with disrupting the supply of these drugs. Given that DNA profiling is carried out worldwide using essentially the same systems as described within this study, the potential for impact is on a national and international scale.

Gaussian process regression coupled with mRMR to predict adulterant concentration in cocaine.

Street cocaine is often mixed with various substances that intensify its harmful effects. This paper proposes a framework to identify attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR) intervals that best predict the concentration of adulterants in cocaine samples. Wavelengths are ranked according to their relevance through ReliefF and mRMR feature selection approaches, and an iterative process removes less relevant wavelengths based on the ranking suggested by each approach. Gaussian Process (GP) regression models are constructed after each wavelength removal and the prediction performance is evaluated using RMSE. The subset balancing a low RMSE value and a small percentage of retained wavelengths is chosen. The proposed framework was validated using a dataset consisting of 345 samples of cocaine with different amounts of levamisole, caffeine, phenacetin, and lidocaine. Averaged over the four adulterants, the GP regression coupled with the mRMR retained 1.07 % of the 662 original wavelengths, outperforming PLS and SVR regarding prediction performance.

Assessment of a Single Quadrupole Mass Spectrometer Combined with an Atmospheric Solids Analysis Probe for the On-Site Identification of Amnesty Bin Drugs.

The surging number of people who abuse drugs has a great impact on healthcare and law enforcement systems. Amnesty bin drug analysis helps monitor the "street drug market" and tailor the harm reduction advice. Therefore, rapid and accurate drug analysis methods are crucial for on-site work. An analytical method for the rapid identification of five commonly detected drugs ((3,4-methylenedioxymethamphetamine (MDMA), cocaine, ketamine, 4-bromo-2,5-dimethoxyphenethylamine, and chloromethcathinone)) at various summer festivals in the U.K. was developed and validated employing a single quadrupole mass spectrometer combined with an atmospheric pressure solids analysis probe (ASAP-MS). The results were confirmed on a benchtop gas chromatography-mass spectrometry instrument and included all samples that challenged the conventional spectroscopic techniques routinely employed on-site. Although the selectivity/specificity step of the validation assessment of the MS system proved a challenge, it still produced 93% (N = 279) and 92.5% (N = 87) correct results when tested on- and off-site, respectively. A few "partly correct" results showed some discrepancies between the results, with the MS-only unit missing some low intensity active ingredients (N-ethylpentylone, MDMA) and cutting agents (caffeine, paracetamol, and benzocaine) or detecting some when not present. The incorrect results were mainly based on library coverage. The study proved that the ASAP-MS instrument can successfully complement the spectroscopic techniques used for qualitative drug analysis on- and off-site. Although the validation testing highlighted some areas for improvement concerning selectivity/specificity for structurally similar compounds, this method has the potential to be used in trend monitoring and harm reduction.

Counting the truxillines-11 or more, the question is now.

Truxillines are a group of tropane alkaloids present in coca leaves that are formed by photochemical dimerization of cinnamoylcocaine(s). Proportion of different truxilline forms present in cocaine serves as its geographical, manufacture, and storage "fingerprint"; thus, the quantitative determination of truxilline content represents one of the powerful methods of analysis and characterization of cocaine samples. Contrary to the statements repeatedly presented in the literature, namely, that there exist exactly 11 truxillines and that every single truxilline is diastereomer of any other, here we show that, in fact, a total of 15 truxillines exist, which can be divided in two structurally isomeric groups-five mutually diastereomeric truxillates and 10 mutually diastereomeric truxinates.

Point-of-care testing of methamphetamine and cocaine utilizing wearable sensors.

The imperative for the point-of-care testing of methamphetamine and cocaine in drug abuse prevention necessitates innovative solutions. To address this need, we have introduced a multi-channel wearable sensor harnessing CRISPR/Cas12a system. A CRISPR/Cas12a based system, integrated with aptamers specific to methamphetamine and cocaine, has been engineered. These aptamers function as signal-mediated intermediaries, converting methamphetamine and cocaine into nucleic acid signals, subsequently generating single-stranded DNA to activate the Cas12 protein. Additionally, we have integrated a microfluidic system and magnetic separation technology into the CRISPR system, enabling rapid and precise detection of cocaine and methamphetamine. The proposed sensing platform demonstrated exceptional sensitivity, achieving a detection limit as low as 0.1 ng/mL. This sensor is expected to be used for on-site drug detection in the future.

Development of a dispersive liquid-liquid microextraction method for the evaluation of maternal-fetal exposure to cocaine employing human umbilical cord tissue.

Illicit drug use is a serious and complex public health problem, not only due to the severity of the health damage but also to the social implications, such as marginalization and drug trafficking. Currently, cocaine (COC) is among the most abused drugs worldwide with about 22 million users. Drug abuse has also been found in women during the pregnancy period, which has shed light on a new group for epidemiology. The diagnosis of COC use in these cases usually depends largely on the mother's reports, which in several cases omit or deny consumption. Therefore, considering physical-chemical methods of sample preparation and exposure biomarkers, the development of analytic toxicological methods can help to confirm drug use during pregnancy. Thus, the objective of the present work was to develop an analytical method based on dispersive liquid-liquid microextraction for the determination of COC analytes, using umbilical cord tissue as an alternative biological matrix, and detection by gas chromatography coupled to mass spectrometry. Therefore, after optimization, the dispersive liquid-liquid microextraction method was fully validated for quantification of COC, benzoylecgonine, cocaethylene, ecgonine, ecgonine methyl ester and norcocaine. The limits of detection were between 15 and 25 ng/g, the limits of quantification were 30 ng/g for ecgonine and 25 ng/g for the other analytes. Linearity ranged from the limits of quantification to 1,000 ng/g. Coefficients of variation for intra-assay precision were <18.5%, inter-assay was <8.75% and bias was <16.4% for all controls. The developed method was applied in 10 suspected positive samples, based on the mother's report and maternal urine screening and confirmation. COC, benzoylecgonine, ecgonine and ecgonine methyl ester were quantified in four umbilical cords with concentrations that ranged from 39.6 to 420.5 ng/g.

Determination of fentanyl contamination on United States paper currency by LC-QQQ-MS.

Previous research has evaluated the extent to which cocaine and other drugs were detectable on currency in the USA. The literature was in agreement that the majority of bills exhibited some degree of contamination. With the increase of fentanyl in the illicit drug supply, this study was designed to evaluate the extent that fentanyl, cocaine, methamphetamine and other substances were present on circulating currency in 2022. A quantitative assay using liquid chromatography-triple quadrupole mass spectrometry was developed and validated to detect six analytes: fentanyl, 4-anilino-N-phenethylpiperidine, acetylfentanyl, benzylfentanyl, cocaine and methamphetamine. One-dollar bills were collected from 13 cities across the country. Sample preparation consisted of soaking the bills in methanol followed by liquid-liquid extraction. Chromatographic separation was achieved using a C18 analytical column and gradient elution with ammonium formate in water (5 mM, pH 3) and 0.1% formic acid in acetonitrile. The quantitative working range for this assay was 0.1 µg to 1.0 µg per bill (equivalent to 1 ng/mL to 100 ng/mL of extract). Fentanyl was detected on the majority (63%) of samples, with 61% of samples having ≥0.1 µg of fentanyl and 4% of samples having ≥1.0 µg. Cocaine and methamphetamine were detected on 100% and 98% of bills, respectively, typically in amounts >1.0 µg. The remaining fentanyl-related substances were detected in 15% of samples in amounts no >0.69 µg per bill and exclusively in the presence of fentanyl. Unsurprisingly, areas of the country with higher incidence of fentanyl use yielded higher frequency of contaminated bills and higher concentrations. Human exposure to drugs on currency is unlikely to have any significant impacts toxicologically or pharmacologically; however, our research findings suggest that paper currency could serve as a useful substrate for surveillance of drug trends regionally, nationally and/or internationally.

NPS-EQA PART I: Four years' experience in external quality assessment program in Italy for classical and new psychoactive substances analysis in oral fluid.

In 2019, Italian National Institute of Health established an external quality assessment program (EQA) to evaluate the performance of oral fluid testing for classical and new psychoactive substances by laboratories participating in the National Early Warning System collaborative centres. This report presents the results of four rounds between 2019 and 2023. Eleven oral fluid specimens, including 3 blank samples, were prepared by adding different classes of and new psychoactive drugs at known concentrations to pre-screened drug-free oral fluid. False-negative and false-positive results were calculated for the qualitative data evaluation. The quantitative evaluation measured the imprecision and accuracy of the results, in terms of coefficient of variation (CV%) and percent error (ERR%), respectively, with respect to a mean value obtained by reference laboratories. Z-score values were then calculated. Over the years, there has been a significant improvement in false-negative results (from 42.7% in the first year to 19.4% in the last year), but not in false-positive results (from 33.3% in the first year to 22.2% in the last one). In addition to the classic drugs of abuse (e.g. cocaine, amphetamine, methadone), the substances found in false positive samples belonged to the class of synthetic cannabinoids (e.g 5-fluoro CUMYL-PINACA and 5-fluoro-EDMB-PICA), synthetic opioids (e.g butyrylfentanyl) and tryptamines (e.g. 5-methoxy-N-methyl-N-isopropyltryptamine). The four rounds yielded a mean ERR% of approximately 22.1% and a mean CV% of around 41.5%. The participating laboratories demonstrated variable performances in relation to the class of analysed psychoactive substances, as evidenced by the calculated Z-scores. Between 25% and 60% of the reported results in all rounds should be considered satisfactory. EQA is a crucial element of laboratory quality management systems. It promotes continuous improvement and maintains high standards in the field of forensic and clinical drug testing.

Hair toxicological analysis of infants and their mothers: a 5-year retrospective study focusing on cocaine.

Prenatal and infant exposure to drugs of abuse is an emerging social and public health problem affecting children health and which may relate to child abuse and neglect. Exposure to drugs of abuse may occur through different routes, including intrauterine, breastfeeding, accidental intake, passive inhalation, and intentional administration. Currently, cases of suspected exposure can be investigated by hair toxicological analysis, the interpretation of which is, however, often difficult, leading to consequent difficulties in the management of such cases. In order to provide a contribution in terms of interpretation of the analytical results, this study aimed to search for the possible existence of elements, from a toxicological point of view, indicative towards the route of exposure. A retrospective study was performed on cases of suspected exposure to drugs of abuse in children aged 0-1 year, evaluated at a University Hospital between 2018 and 2022. Data of children hair toxicological analysis were analyzed and then compared with those of their mothers, when available; 41.6% children tested positive for cocaine. The study found a significant correlation between cocaine and benzoylecgonine concentrations, and a benzoylecgonine/cocaine ratio that tends to decrease as the age of children increases. From the comparison with mothers, a child/mother cocaine concentration ratio lower than 1 was found in all cases of hair sampled within the first week of life, and a ratio greater than or equal to 1 in all cases in which the sampling was performed later. These results, if confirmed in a larger cohort, could represent a contribution in the interpretation of cases of infant exposure to drugs of abuse and be integrated in the context of their multidisciplinary evaluation.

Mn-Co-Ce/biochar based particles electrodes for removal of COD from coking wastewater by 3D/HEFL system: Characteristics, optimization, and mechanism.

In this work, the Mn, Co, Ce co-doped corn cob biochar (MCCBC) as catalytic particle electrodes in a three-dimensional heterogeneous electro-Fenton-like (3D-HEFL) system for the efficient degradation of coking wastewater was investigated. Various characterization methods such as SEM, EDS, XRD, XPS and electrochemical analysis were employed for the prepared materials. The results showed that the MCCBC particle electrodes had excellent electrochemical degradation performances of COD in coking wastewater, and the COD removal and degradation rates of the 3D/HEFL system were 85.35% and 0.0563 min-1 respectively. RSM optimized conditions revealed higher COD removal rate at 89.23% after 31.6 min of electrolysis. The efficient degradability and wide adaptability of the 3D/HEFL system were due to its beneficial coupling mechanism, including the synergistic effect between the system factors (3D and HEFL) as well as the synergistic interactions between the ROS (dominated by •OH and supplemented by O2•-) in the system. Moreover, the COD removal rate of MCCBC could still remain at 81.41% after 5 cycles with a lower ion leaching and a specific energy consumption of 11.28 kWh kg-1 COD. The superior performance of MCCBC, as catalytic particle electrodes showed a great potential for engineering applications for the advanced treatment of coking wastewater.

In-sample stability and postsampling analysis of 21 illicit drugs, their metabolites and cotinine in wastewater.

A thorough understanding of the degradation of chemical biomarkers in wastewater after the sampling is critical in the surveillance of illicit drug use based on the back-calculation technique. Herein, three temperatures, eight groups of matrices, and acidification were applied to simulate the preservation condition of 21 illicit drugs, their metabolites, and cotinine for a 240-day stability study. It was proved that the temperature, matrices, and acidification play vital roles in their stability in wastewater. Most of them demonstrated high stability (transformation rates < 20%) during room temperature for 45 days, and the transformation rates decreased while the storage temperature reduced. The stability of the target compounds such as cocaine (COC), 6-monoacetylmorphine (6-MAM), and amphetamine (AM) is influenced by matrices. Acidification prevented the majority of analytes from transforming, making it a feasible solution for preservation after sampling. A model that combined the effects of temperature and matrix was developed to back-calculate the concentration of target compounds during the postsampling process. The feasibility of this model was validated by correcting the loss of COC and 6-MAM from 24.2% and 16.2% to 2.98% and 2.77%. This study simulated a typical large-scale sampling and storage scenario. The effect of the temperature, pH, and matrix on in-sample stability and the postsampling analysis of selected target compounds was investigated for the first time in this study.

Drug driving in Italy. The results of the first roadside drug testing service utilizing on-site confirmatory analysis between 2019 and 2022.

BACKGROUND: Drug driving represents a public safety concern, and the size of this issue in Italy is not fully known. Drug testing is composed of two steps: 1) screening and 2) confirmatory analysis. The second step, and the associate medical examination to assess the state of impairment, usually are not performed right after the screening as they require specialized personnel and instrumental equipment that are not historically available at roadblocks. These pitfalls make this process both complicated and time-consuming. METHODS: A mobile laboratory was set up in 2019 by the Forensic Lab Service S.r.l. (limited liability company) to improve roadblock timing, planning, as well as to shed light on the extent of the drug driving issue in Italy. Drug screenings were performed using DrugWipe® Saliva testing. Confirmatory analysis was performed on oral fluids by liquid chromatography coupled with tandem mass spectrometry. A dedicated room of the mobile laboratory was also designed for drug driving medical assessment. RESULT: 2082 samples were collected during 88 road safety services held in different locations across Italy. In total, 9 % of the tested subjects were positive to both the screening and the confirmatory analysis. The most prevalent illicit drugs found in this study were THC (72 %), followed by cocaine (41 %). Drug drivers were mostly male (93 %) and younger than 30 years of age (58 %). CONCLUSIONS: The prevalence of drivers testing positive for illicit drugs resulted to be higher compared to the results obtained in the DRUID project and to other surveys previously performed in Italy. These data demonstrate the need for control services to improve road safety in regards to drug driving.