Trazodone is a Serotonin receptor Antagonist and Reuptake Inhibitor (SARI) that is used for its anxiolytic properties in human and small animal veterinary medicine. Studies in horses have shown that it has sedative effects and can be used to modify behavioural issues following oral dosing, but it has a high level of side effects following intravenous administrations. To ensure effective control, pharmacokinetic studies are required to understand its metabolism and excretion. Therefore, an oral administration of 2.5 mg/kg of trazodone was performed to six Thoroughbred horses to assess detection in equine plasma and urine. Initially, LC-HRMS was utilised to assess phase I and II metabolism of trazodone in unhydrolysed and hydrolysed plasma and urine samples. Subsequently, extraction and LC-MS/MS methods were developed and quantitatively validated for detection of trazodone, its active metabolite m-chrlorophenylpiperazine (m-CPP) and 4’-hydroxytrazodone in unhydrolysed plasma and hydrolysed urine. Sample analysis results will be presented herein.