According to cognitive activation theory, long-lasting work demands without rest or lack of coping may lead to sustained activation and pathology. Cortisol is one of the most important stress hormones in humans and increased basal levels of cortisol are considered a valid marker for sustained activation. In order to investigate this association further, we investigate the relationships between salivary cortisol profiles, job stress, work load (effort/reward, demand/control) and health (subjective health complaints and health-related quality of life) in a population of health care workers. Forty-four women filled in a questionnaire and collected five saliva samples on two consecutive working days (1: wake-up time, 2: wake-up time+30 min, 3: wake-up time+45 min, 4: 1500 h and 5: 2200 h). There was no relationship between psychosocial factors at work and cortisol levels in the morning (cortisol level at wake-up time and awakening cortisol response (ACR)). Only the confounding variable tobacco reached a significant level in the hierarchical regressions analyses. Our significant findings are limited to the afternoon decline and the evening values. The decrease during the day relates to decision authority, physical functioning, general health, and vitality in the single, unadjusted regression analyses. The decrease also relates to coffee intake, which we included originally as a confounding variable. In the final hierarchical regression of the evening values, only decision authority and coffee were significantly related to cortisol levels in the evening.