Highlights ■ We validated the UHPLC/UV method for accurate busulfan quantification in plasma. ■ Inaccuracy and imprecision were below 15%, ensuring reliable therapeutic drug monitoring results. ■ This enables effective pharmacokinetic studies with rapid turnaround times in patient samples. ABSTRACT Objective: This study aimed to validate a sensitive, accurate, and precise bioanalytical ultrahigh- performance liquid chromatography coupled with ultraviolet (UHPLC/UV) method for the determination of busulfan in human plasma using 1,6-bis-(methanesulfonyloxy) hexane as an internal standard for therapeutic drug monitoring. […]