ABSTRACT Background Substance use (SU) among HCWs leads to reduced productivity and overall health and, moreover, is a negative role to promote SU reduction. Aim This study aimed to estimate the prevalence of SU and assess its determinants among HCWs. Methods The study analyzed data related to the first phase of the SHAHWAR Cohort study which included 1177 university employees. Results The prevalence of current tobacco use is 11.6% (2.7% in women, and 24.5% in men), and the prevalence of lifetime tobacco use was 19.8 (6.3% in women, and 39.3% in men). The ratio of hookah: cigarette use in women is about seven times that of men. The self-reported current tobacco, alcohol and opium use were 11.6% (M: F ratio > 9), 2.6% (M: F ratio > 6) and 1.4% (M: F ratio > 7) of total subjects. There was a significant difference in substance or alcohol users based on gender, job position, working schedule, employment status, marital status and education levels. There was a negative correlation between job security and stability in terms of work status among workers with substance or alcohol use. Conclusion Since implementation of preventive interventions knowing the high-risk groups and occupational risk factors, the results of the present study may meet this need.