The present study aims to identify the relationship between workplace spirituality, compassion, relationship with others at work, spiritual orientation, organizational value and alignment of personal values, and psychological well-being among universities' teachers. Further, the mediating effect of job stress was also identified between workplace spirituality and psychological well-being. Similarly, the mediation of environmental mastery between organizational values and alignment of personal values and psychological well-being was examined. Finally, we examined the mediation of personal growth between spiritual orientation and psychological well-being. The data were collected through a structured and adapted five-point Likert scale using a purposive sampling technique, with a total sample of 873 male and female university teachers working in the private and government universities. We employed structural equation modeling to check the relationship among the considered variables for analysis purposes. The results show a strong positive relationship between the independent and dependent variables. The findings further demonstrated that the mediation analysis confirms that job stress mediates the relationship between workplace spirituality and psychological well-being, and environmental mastery mediates between organizational values and alignment of personal values and psychological well-being. Finally, personal growth mediates between spiritual orientation and psychological well-being. To maintain the excellent quality of education, educational institutes need to identify and imply the practice of workplace spirituality that will help to reduce job stress and improve the psychological well-being of universities' teachers, thus resulting in better educational output.