Automation in wastewater-based epidemiology for efficient analysis of 123 multi-class illicit drugs: evaluating method eco-friendliness and practicality

摘要

The performance of an automated sample preparation using the modular Biomek i7 workstation was evaluated for analysis of 123 illicit drugs (ID) in wastewater. The method underwent extensive development, including optimization of UPLC-MS/MS parameters, assessment of compound stability and extraction efficiency using three types of SPE cartridges. A 60-day stability assessment demonstrated that all analytes were stable at –25 ± 3 °C; however, storage at 6 ± 2 °C resulted in reduced stability for 30 analytes, with the most pronounced effects observed for 4-Chloro methcathinone (4-CMC), Normeperidine, and Cathinone. Extraction efficiency trials revealed that the order of suitability for SPE cartridges is, Mixed-Mode Cation Exchange (MCX) > Hydrophilic-Lipophilic Balance (HLB) > Mixed-Mode Anion Exchange (MAX). The method demonstrated specificity for all targeted IDs, with the max interfering peak for Pregabalin reaching 8.01 % of the target limit of quantification (LOQ). The method met matrix effect tolerance (± 20%) for 114 IDs, while 6 IDs showed signal suppression and 5 exhibited signal enhancement. The 8-point procedural calibration curve, (six-point for 9 IDs), prepared using automated robotic extraction has demonstrated linearity across the tested range. A spiked study at low (LQC), medium (MQC), and high (HQC) quality control (QC) levels (n = 6, repeated on three occasions) showed high precision, with 91.6 % of the observations having RSD < 10 % and only 0.5 % exceeding 15 % RSD. It was observed that the 95.7% of recoveries were falling within the 90–110% range, and less than 5% outside this interval. The high-throughput method (automated extraction of 98 samples within 2 hours and 40 minutes) fulfilled all validation requirements, demonstrated reliability in intra-laboratory comparison with an acceptable z-score, and achieved strong ratings for greenness (GAPI, AGREE) and practicality (BAGI). • Eco-friendly method meets sensitivity requirement without evaporative concentration. • Suitability of three SPE cartridge and storage stability was evaluated for 123 IDs. • Method accuracy proved with validation, MU calculation, and ILC. • Automated alternative to conventional manual protocols for testing multiclass 123 IDs. • High-throughput method rated excellent by GAPI, AGREE, and BAGI sustainability tools.