|Barbiturates belong to a class of sedative hypnotics that causes mild sedation to total anesthesia. Excessive and prolonged amount of barbiturate drugs can produce chronic symptoms and even death. Here we present a method for the analysis of barbiturates in urine using LC-MS/MS. Minimal sample preparation is required; simple ‘dilute-and-shoot’ for the urine samples. This analysis was performed on a reversed phase column analyzed by LC-MS/MS with a run time under five minutes. The assay was shown to be accurate and precise with %CV and % accuracy within ±15% of nominal across the full linear range.