Monitoring psychoactive substance use at six European festivals through wastewater and pooled urine analysis.

The consumption of psychoactive substances is considered a growing problem in many communities. Moreover, new psychoactive substances (NPS) designed as (legal) substitutes to traditional illicit drugs are relatively easily available to the public through e-commerce and retail shops, but there is little knowledge regarding the extent and actual use of these substances. This study aims to gain new and complementary information on NPS and traditional illicit drug use at six music festivals across Europe by investigating wastewater and pooled urine. Samples were collected, between 2015 and 2018, at six music festivals across Europe with approximately 465.000 attendees. Wastewater samples were also collected during a period not coinciding with festivals. A wide-scope screening for 197 NPS, six illicit drugs and known metabolites was applied using different chromatography-mass spectrometric strategies. Several illicit drugs and in total 21 different NPS, mainly synthetic cathinones, phenethylamines and tryptamines, were identified in the samples. Ketamine and the traditional illicit drugs, such as amphetamine-type stimulants, cannabis and cocaine were most abundant and/or frequently detected in the samples collected, suggesting a higher use compared to NPS. The analyses of urine and wastewater is quick and a high number of attendees may be monitored anonymously by analysing only a few samples which allows identifying the local profiles of use of different drugs within a wide panel of psychoactive substances. This approach contributes to the development of an efficient surveillance system which can provide timely insight in the trends of NPS and illicit drugs use.

A high-throughput solid-phase microextraction and post-loop mixing large volume injection method for water samples.

This article presents a novel approach for the analysis of 13 drugs in wastewater for use in wastewater-based epidemiology (WBE) studies. Sample preparation remains one of the principal bottlenecks in modern high-throughput analysis by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The proposed methodology is based on the micro-extraction of small volumes (1 ml) of wastewater using a HLB 96-well microplate and both large volume injection (LVI) and post-loop mixing injection (PLM). With this configuration, the limits of quantification (LOQ) were below the reported environmental concentrations of the target compounds in wastewater. Furthermore, both the complexity of collecting, transporting and storing the wastewater sample, sample preparation time, cost and amount of solvent used are all diminished, enhancing the suitability of this methodology for future WBE studies. A new workflow is also proposed in order to create a virtual specimen library bank for WBE by using high-resolution mass spectrometry (HRMS). The method was validated and the limits of quantification were between 0.2 and 6.3 ng L-1. The relative standard deviations (RSD) for a standard mixture at 200 ng L-1 (n = 6) was between 3.4 and 14.4% while the recoveries for the 13 drug target residues (DTR) were between 92 and 110%. The developed and validated method was finally successfully applied to 10 wastewater samples collected from Oslo, Norway.

Passive sampling of wastewater as a tool for the long-term monitoring of community exposure: Illicit and prescription drug trends as a proof of concept.

A passive sampling device, the Polar Organic Chemical Integrative Sampler (POCIS), was calibrated in-situ over a 4-week period in Oslo (Norway) for 10 illicit drugs and pharmaceuticals with the goal of developing an approach for monitoring long-term wastewater drug loads. The calibrations were performed in triplicate using three different overlapping calibration sets under changing environmental conditions that allowed the uncertainty of the sampling rates to be evaluated. All 10 compounds exhibited linear uptake kinetics and provided sampling rates of between 0.023 and 0.192 L d-1. POCIS were deployed for consecutive 2-week periods during 2012 and 2013 and the calculated time-weighted average (TWA) concentrations used to define different drug use trends. The relative uncertainty related to the POCIS data was approximately 40% and, except for citalopram, 85% of all the long-term measurements of pharmaceuticals were within the confidence interval levels calculated to evaluate the effects of changing environmental conditions on the TWA estimations. POCIS was demonstrated to be sufficiently robust to provide reliable annual drug use estimates with a smaller number of samplers (n = 24) than recommended for active sampling (n = 56) within an acceptable level of sample size related uncertainty < 10%. POCIS is demonstrated to be a valuable and reliable tool for the long-term monitoring of certain drugs and pharmaceuticals within a defined population.

Increased levels of the oxidative stress biomarker 8-iso-prostaglandin F2α in wastewater associated with tobacco use.

Wastewater analysis has been demonstrated to be a complementary approach for assessing the overall patterns of drug use by a population while the full potential of wastewater-based epidemiology has yet to be explored. F2-isoprostanes are a prototype wastewater biomarker to study the cumulative oxidative stress at a community level. In this work, 8-iso-prostaglandin F2α (8-iso-PGF2α) was analysed in raw 24 h-composite wastewater samples collected from 4 Norwegian and 7 other European cities in 2014 and 2015. Using the same samples, biomarkers of alcohol (ethyl sulfate) and tobacco (trans-3'-hydroxycotinine) use were also analysed to investigate any possible correlation between 8-iso-PGF2α and the consumption of the two drugs. The estimated per capita daily loads of 8-iso-PGF2α in the 11 cities ranged between 2.5 and 9.9 mg/day/1000 inhabitants with a population-weighted mean of 4.8 mg/day/1000 inhabitants. There were no temporal trends observed in the levels of 8-iso-PGF2α, however, spatial differences were found at the inter-city level correlating to the degree of urbanisation. The 8-iso-PGF2α mass load was found to be strongly associated with that of trans-3'-hydroxycotinine while it showed no correlation with ethyl sulfate. The present study shows the potential for 8-iso-PGF2α as a wastewater biomarker for the assessment of community public health.

Target and suspect screening of psychoactive substances in sewage-based samples by UHPLC-QTOF.

The quantification of illicit drug and pharmaceutical residues in sewage has been shown to be a valuable tool that complements existing approaches in monitoring the patterns and trends of drug use. The present work delineates the development of a novel analytical tool and dynamic workflow for the analysis of a wide range of substances in sewage-based samples. The validated method can simultaneously quantify 51 target psychoactive substances and pharmaceuticals in sewage-based samples using an off-line automated solid phase extraction (SPE-DEX) method, using Oasis HLB disks, followed by ultra-high performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC-QTOF) in MS(e). Quantification and matrix effect corrections were overcome with the use of 25 isotopic labeled internal standards (ILIS). Recoveries were generally greater than 60% and the limits of quantification were in the low nanogram-per-liter range (0.4-187 ng L(-1)). The emergence of new psychoactive substances (NPS) on the drug scene poses a specific analytical challenge since their market is highly dynamic with new compounds continuously entering the market. Suspect screening using high-resolution mass spectrometry (HRMS) simultaneously allowed the unequivocal identification of NPS based on a mass accuracy criteria of 5 ppm (of the molecular ion and at least two fragments) and retention time (2.5% tolerance) using the UNIFI screening platform. Applying MS(e) data against a suspect screening database of over 1000 drugs and metabolites, this method becomes a broad and reliable tool to detect and confirm NPS occurrence. This was demonstrated through the HRMS analysis of three different sewage-based sample types; influent wastewater, passive sampler extracts and pooled urine samples resulting in the concurrent quantification of known psychoactive substances and the identification of NPS and pharmaceuticals.

Using biomarkers in wastewater to monitor community drug use: a conceptual approach for dealing with new psychoactive substances.

Data obtained from the analysis of wastewater from large-scale sewage treatment plants has been successfully applied to study trends in the use of classical illicit drugs such as cocaine, but the dynamic nature of the new psychoactive substances (NPS) market presents a unique set of challenges to epidemiologists. In an attempt to overcome some of the challenges, this paper presents a framework whereby a collection of tools and alternative data-sources can be used to support the design and implementation of wastewater-based studies on NPS use. Within this framework the most likely and most suitable biomarkers for a given NPS are predicted via in-silico metabolism, biotransformation and sorption models. Subsequent detection and confirmation of the biomarkers in samples of wastewater are addressed via high-resolution mass spectrometry (HRMS). The proposed framework is applied to a set of test substances including synthetic cannabinoids and cathinones. In general, the in-silico models predict that transformation via N-dealkylation and hydroxylation is likely for these compounds, and that adsorption is expected to be significant for cannabinoids in wastewater. Screening via HRMS is discussed with examples from the literature, and common-fragment searching and mass-defect filtering are successfully performed on test samples such that spectral noise is removed to leave only the information that is most likely to be related to the NPS biomarkers. HRMS screening is also applied to a set of pissoir-sourced wastewater samples and a total of 48 pharmaceuticals and drugs including 1-(2-methoxyphenyl)piperazine (oMeOPP) are identified. The framework outlined in this paper can provide an excellent means of maximizing the chances of success when identifying and detecting biomarkers of NPS in wastewater.