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Alcohol and psychoactive substance use represent a major burden for global public health, increasing the risk of non-communicable diseases, violence, road traffic injuries, dependence, and mental disorders, and generating impacts on productivity and social welfare. This study aimed to estimate the burden of disease attributable to alcohol and other psychoactive substances in the departments of Colombia from 2016 to 2022. A burden-of-disease study was conducted using the Disability-Adjusted Life Years (DALYs) indicator, following the methodology of the World Health Organization Global Health Estimates. Official morbidity and mortality databases were used. An estimated 236,154.42 DALYs were attributable to alcohol and psychoactive substance use in Colombia during the study period, increasing from 14,158.7 DALYs in 2016 to 40,190.7 DALYs in 2022. The burden was heterogeneous across departments, with values above 1000 DALYs in Quindío (1779.5), Nariño (1624.3), and Norte de Santander (1008.0) and below 132 DALYs in La Guajira, Casanare, and Vaupés. Men accounted for 73.5% of total DALYs. The mean age of morbidity records associated with alcohol and psychoactive substance use disorders was 30.67 years in men and 32.37 years in women. The burden associated with psychoactive substance use is increasing in Colombia, with differences by sex and department of residence.