Psychological inflexibility is defined as a transdiagnostic mechanism characterized by rigid efforts to control psychological reactions to painful, undesirable, unpleasant internal experiences. The Acceptance and Action Questionnaire-University Students (AAQ-US) was designed to assess students' academic context-related psychological inflexibility. This study adapted the AAQ-US to Portuguese and examined its factor structure, validity, and reliability in a college student sample (sample 1: N = 262). Exploratory factor analysis and parallel analysis suggested a two-factor structure. A confirmatory factor analysis was conducted in a second sample to cross-validate the AAQ-US factor structure (sample 2: N = 260). One higher-order factor (psychological inflexibility) with two lower-order factors (cognitive fusion and experiential avoidance) revealed a very good fit to the data. The AAQ-US total and dimensions showed good reliability, convergent and incremental validity. Overall, the Portuguese version of the AAQ-US is a reliable and valid instrument for assessing context-specific psychological inflexibility in university students.