Since human physiology has biologically adapted itself to synchronization with the light-dark cycle over thousands of years of evolution, shift work and working in atypical shifts is associated with a number of potentially negative physical and psychological effects. Many studies have shown that shift work includes a higher rate of psychophysiological disorders, sleep problems, psychoneurotic disorders, bad eating habits, gastrointestinal problems, cardiovascular problems, higher mortality, a higher frequency of accidents at work and more absenteeism. However, in relation to the aforementioned outcomes of night shifts, relatively less attention has been devoted to the study of the psychological health and well-being of healthcare workers. The aim of the research was to examine the difference in the level of subjective psychological well-being between the group that works in rotating shifts and the group that works in non-rotating shifts, and the difference between groups whose rotating shifts include night work and whose rotating shifts do not include night work. Respondents in this paper were divided into health workers working rotating shifts (Group 1; n=264) and health workers working non-rotating shifts (Group 2; n=89). The first group was further divided into a group where rotating shifts included night work (Group 1A; n=183) and where rotating shifts were without night work (Group 1B; n=81). All subjects completed the WEMWBS scale for measuring subjective psychological well-being, which was later evaluated with scores from 14 to 70. Statistical data processing did not record a statistically significant difference between group 1 and group 2, but a statistically significant difference was recorded between groups 1A and 1B. The obtained results indicate that health workers who work rotating shifts do not have a lower level of psychological well-being than health workers who work non-rotating shifts, but health workers whose rotating shifts include night work have a lower level of psychological well-being than colleagues whose rotating shifts do not include night work.