Background: Indoor environment, in which we are located, influences our psychological responses, like stress. Thermal comfort is one of the most important factors determining the quality of indoor environment, obtained by calculating predicted mean vote (PMV) and predicted percentage of dissatisfaction (PPD) in a steady-state approach. Several studies revealed that hot or cold temperature (°C) could increase stress. However, other climatic thermal comfort factors like relative humidity (%) and air velocity (m/s) have not been well researched in this regard. Objectives: The purpose of this study was to investigate the immediate effects of undesirable thermal comfort on stress by measuring salivary alpha-amylase levels (sAA) in female high school students. Methods: The present study was conducted in a semi-experimental design with pre-test and post-test and control group. The sample size was 390 female high school students, who were selected by multi-stage cluster random sampling, during years 2016 and 2017, in Shiraz, Iran. Students were divided to intervention and control groups. In the intervention group, thermal comfort was interfered by changing temperature, relative humidity, and air velocity for two hours. Saliva was collected by Cocorometer's strip and sAA was measured by a hand-held device, Cocorometer (Nipro Co, Osaka, Japan). The pre-test and post-test were compared with each other. Thermal comfort in classrooms was measured by a steady-state model, PMV/PPD index, and data was analyzed by Analysis of Covariance (ANCOVA). Results: Interfering in temperatures and humidity at an average of 4.8°C and 36% for two hours in the intervention group caused thermal comfort to be in an unfavorable range (PMV > +0.5) and dissatisfaction was more than desirable (PPD > 10%). In the intervention group, when the intervention took place at the temperature, relative humidity, air velocity, and the synchronous effect of these three variables (TRHAV), the mean sAA was 43.57, 42.74, 44.23, and 45.93 (KU/L) in the pre-test and 55. 91, 52.35, 44.89 and 61.99 (KU/L) in the post-test. Also, in the control group, the mean sAA was 44.73, 47.03, 43.38, 47.19, 43.83, and 43.77 (KU/L) in the post-test. The sAA was significantly increased in the intervention group when thermal comfort and its climatic variables, including temperature (P < 0.001) and relative humidity (P < 0.001), was undesirable. No Significant increase was observed in air velocity (P = 0.659). In the control group, the mean sAA did not change significantly in pre-test and post-test. Conclusions: Undesirable thermal comfort increased stress in female high school students. Stress was affected by temperature, relative humidity and the synchronous effect of temperature, relative humidity, and air velocity yet not air velocity, individually.
