Toxicological urinalysis is a highly sensitive and specific test that detects recent substance use. It has been established for substance misuse treatment but has not been routinely used at liver transplantation (LT) centers. Patients with a history of substance misuse are required to be abstinent from alcohol and illicit drugs before they are listed for LT. In this cross-sectional study, we sought to determine the prevalence of recent substance use in LT candidates via toxicological urinalysis. One hundred nine adults who were admitted for an LT assessment provided data, and they were categorized by the etiology of their liver disease [alcohol-related liver disease (ALD), hepatitis C virus (HCV), or other liver diseases]. Urine was toxicologically screened for drugs and their metabolites as well as the urinary alcohol metabolites ethyl glucuronide and ethyl sulfate. The prevalence of alcohol metabolites in patients with ALD was 20%. Licit and illicit substances together provided a positive toxicological result in 30% of the patients. Positive results were more common among patients with HCV (40%) and ALD (38%) versus patients with other liver diseases (18%). During the clinical assessment, 4% of the patients with ALD or HCV self-reported current alcohol or illicit drug use. These results correspond to the findings of other studies and emphasize the uncertainty of self-reported substance use data for LT candidates. Liver Transpl 17:1200–1204, 2011. © 2011 AASLD.