Besides its clinical application, the anaesthetic agent propofol is being increasingly misused, mostly by healthcare professionals, and its abuse potential gained worldwide attention after the tragic death of Michael Jackson in 2009. Due to the short duration of its narcotic effects, propofol abuse is especially easy to hide compared with the use of other recreational drugs. However, propofol possesses a very narrow therapeutic window between the desired effect and potentially fatal toxicity, making abuse of the drug extremely dangerous even in experienced physicians. Consequently, it is important that forensic laboratories possess a sensitive and specific method for the detection of chronic propofol abuse. We present a simple, fast and reliable method to simultaneously extract propofol and its main metabolite propofol glucuronide from hair, followed by sensitive LC–MS/MS analyses, allowing to determine a chronic propofol abuse. Difficulties regarding the detection of propofol using LC–MS/MS were solved by using a derivatization reaction with 2-fluoro-1-methylpyridinium-p-toluene-sulfonate and triethylamine. Reliability of extraction method and subsequent LC–MS/MS analyses was confirmed under consideration of the validation parameters selectivity, linearity, accuracy and precision, analytical limits, processed sample stability, matrix effects and recovery. Appropriate quantification (LLOQ = 10 pg/mg hair) and detection limits (3.6 pg/mg hair for propofol and 7.8 pg/mg hair for propofol glucuronide) could be achieved, enabling to detect even small amounts of both analytes. Applicability of the method was confirmed by analysis of three human hair samples from deceased with suspicion of chronic propofol abuse.