Objective: To analyze whether the results on quality assurance and safety culture in a healthcare organization are related to and affected by the actions implemented. Setting: Health Insurance of Work-related Accidents and Occupational Diseases. Methods: The study was conducted as a longitudinal observational study that analyzed the relationship of the Safety Culture and Quality Assurance measurements. Participants who were involved came from small centers with less than eight workers (N = 52), big centers (eight and more workers) (N = 707), and those centers with quality coordinators (N = 91). Data were collected during the years 2015 and 2016. Results: A total of 595 healthcare professionals responded in 2015 and 491 in 2016. The scores showed a positive progression both in Quality Assurance (T-test = 3.5, p = 0.001) and in Safety Culture (T-test = 5.6, p < 0.0001). Hence, the gradient of improvement in quality (average 5.5%) was greater compared to that of the safety culture (2.1%). Conclusions: The assessments of the quality assurance goals were consistent with the safety culture assessment. Hence, the results on Safety Culture were observed to be more stable over time.