Daily life has changed due to COVID-19. This has affected nursing education and caused a shift in online learning. This study examined the effects of online learning on nursing students’ knowledge, self-regulation, and learning flow. We used a quasi-experimental design on a sample comprising 164 senior nursing students. We compared pre- and post-test scores to examine the educational effects. The pre-test was conducted a week before the educational intervention, and the post-test was conducted a week after it. We found a significant increase in knowledge (t = −14.85, p < 0.001) and learning flow (t = −2.15, p = 0.033) in the post-test. We also found an increase in self-regulation (t = −1.57, p = 0.119) from the pre- to the post-test that was not statistically significant. The results could help instructors to provide additional information in online learning. They highlight the need to assess learners’ readiness for online learning and to prepare the learning environment with systematic educational planning, design, development, and evaluation for improving the effectiveness of online learning outcomes.