Crash statistics and hospital data show that injured e-scooter riders arrive at hospitals often at night and on weekends. Subsequently, the crash risk at night is higher compared to the daytime. A possible explanation might be increased rule violations, safety-critical behaviors, and changes in the user group at night compared to daytime. Therefore, we aimed to conduct an observational study analyzing the interrelationships of risky behaviors, rule violations, and user characteristics of e-scooter riders in two German cities. A total of 732 observations were analyzed with Chi-Squared tests and Generalized Estimating Equations. The results show increased rates of tandem riding at night compared to the daytime and increased rule violations of adolescents compared to older e-scooter riders regardless of the time of day. Rates of helmet use, wrong-way riding, headphone use, smartphone use, and luggage transport were comparable for daytime and night observations. The results suggest that educational campaigns should focus on tandem riding, especially targeting the user group of teenage riders. This study brings e-scooter riding at night into the light and emphasizes riders' nightly behaviors for policymakers and traffic safety.