Abstract A systematic Analytical Quality by Design (AQbD) approach was applied to the development and validation of a UHPLC–MS/MS method for the determination of hallucinogenic phenethylamine-derived novel psychoactive substances in urine. Owing to the continuous emergence of structurally related compounds on the illicit drug market, AQbD enables rapid and knowledge-based development of robust analytical methods. An Analytical Target Profile was defined to ensure adequate separation, sensitivity, robustness, and compliance with white analytical chemistry principles. Method development followed a structured AQbD workflow, including chromatographic scouting of five stationary phases and different mobile-phase compositions, followed by multivariate screening and response surface optimization of critical method parameters. Optimal performance was achieved on a biphenyl column using an initial mobile-phase composition of 20 % phase B and a column temperature of 40 °C. The method operable design region was defined for a flow rate of 0.375–0.435 mL min −1 and a formic acid concentration of 0.06–0.085 %. The method was fully validated, demonstrating excellent linearity (R 2 ≥ 0.99), satisfactory precision and accuracy, and limits of detection of 0.1–1 ng mL −1 . The White Analytical Chemistry evaluation yielded an overall score of 91.9, indicating balanced analytical performance and sustainability for routine forensic and clinical toxicological analyses.
Kostić et al. (Thu,) studied this question.