Negative emotional states, such as stress, anxiety, and depression, are prevalent in university students. Personality traits have been shown to be associated with a wide range of behaviors in students, such as academic motivation, achievement, and social well-being. The aim of this study was to investigate the association between the Big Five personality traits and negative emotion states in university students in Taiwan. A cross-sectional study was conducted on 580 university students in Taiwan. Negative emotional states were evaluated using the Depression Anxiety Stress Scale-21 (DASS-21) and the Big Five personality traits were measured using the 48-item Big Five Inventory. A hierarchical linear regression analysis was used to assess the factors associated with DASS-21 scores. Neuroticism (standardized beta std. β = 0.45, < 0.001) and openness (std. β = 0.12, = 0.003) were significantly associated with DASS-21 scores, while agreeableness (std. β = -0.10, = 0.007) was significantly and inversely associated with DASS-21 scores. Personality traits could be used to identify students at risk of negative emotional states and to undertake appropriate preventive strategies.