Metabolic activation of a drug leading to reactive metabolite(s) that can 
covalently modify proteins is considered an initial step that may lead to 
drug-induced organ toxicities. Characterization of reactive metabolites is 
critical to designing new drug candidates with an improved toxicological 
profile. High performance liquid chromatography (HPLC) coupled with mass 
spectrometry (MS) predominates over all analytical tools used for screening and 
characterization of reactive metabolites. In this review, a brief description of 
experimental approaches employed for assessing reactive metabolites is followed 
by a discussion on the reactivity of acyl glucuronides and acyl coenzyme A 
thioesters. Techniques for high-throughput screening and quantitation of 
reactive metabolite formation are also described, along with proteomic 
approaches used to identify protein targets and modification sites by reactive 
metabolites. Strategies for dealing with reactive metabolites are reviewed. In 
conclusion, we discuss the challenges and future needs in this field of 
research.