AbstractBackgroundExperimental and clinical evidence suggest that stress can lead to ill‐health through the disregulation of the hypothalamic–pituitary–adrenal (HPA) axis. Studies to date have produced equivocal results likely due to different methodologies and failure to account for confounding factors. This investigation aimed to assess the relation between self‐reported work‐related stressors and salivary cortisol and to clarify the role of the potential confounders.MethodsThirty‐six call‐handlers completed a self‐administered job content questionnaire and collected seven daily salivary samples on two workdays and a weekend. The diurnal salivary cortisol output was expressed as cortisol awakening response (CAR), and cortisol output in the rest of the day. Salivary cortisol data were normalized by means of square root transformation. The generalized estimating equations method was used to assess the relation between job strain and cortisol levels after adjusting for gender, weekdays and adherence to the sampling schedule.ResultsJob strain significantly influenced the total amount of cortisol response to waking (high strain vs. low strain: 1.1 (0.3–2.0) nmol/L). The cortisol response to waking showed gender‐specific differences [women excreting greater cortisol than men: 1.1 (0.3–1.9) nmol/L], and weekday differences [workdays vs. weekend: 1.0 (0.3–1.6) nmol/L]. Non‐compliance with the sampling protocol was associated with lower salivary cortisol than in adherent subjects.ConclusionsOur results provide further evidence for the HPA axis involvement in the physiological response to work stress. The measure of the CAR showed to be the sensitive index to assess the physiological response to psychosocial factors. Gender, weekday, and protocol compliance were confounding factors. Am. J. Ind. Med. 52:251–260, 2009. © 2008 Wiley‐Liss, Inc.