IMPORTANCE: Direct costs of substance use disorders (SUDs) in the United States are incurred primarily among the working-age population. Quantifying the medical cost of SUDs in the employer-sponsored insurance (ESI) population can improve understanding of how SUDs are affecting workplaces and inform decision-making on the value of prevention strategies. OBJECTIVE: To estimate the annual attributable medical cost of SUDs in the ESI population from the health care payer perspective. DESIGN, SETTING, AND PARTICIPANTS: In this economic evaluation, Merative MarketScan 2018 databases were weighted to represent the non-Medicare eligible ESI population. Regression and mathematical modeling of medical expenditures controlled for insurance enrollee demographic, clinical, and insurance factors to compare enrollees with and without an SUD diagnosis to identify the annual attributable medical cost of SUDs. Data analysis was conducted from January to March 2022. EXPOSURES: International Statistical Classification of Diseases, Tenth Revision, Clinical Modification SUD diagnoses on inpatient or outpatient medical records according to Clinical Classifications Software categories (alcohol-, cannabis-, hallucinogen-, inhalant-, opioid-, sedative-, stimulant-, and other substance-related disorders). MAIN OUTCOMES AND MEASURES: Annual SUD medical cost in the ESI population overall and by substance type (eg, alcohol). Number of enrollees with an SUD diagnosis and the annual mean cost per affected enrollee of SUD diagnosis (any and by substance type) are also reported. RESULTS: Among 162 million ESI enrollees, 2.3 million (1.4%) had an SUD diagnosis in 2018. The regression analysis sample included 210 225 individuals with an SUD diagnosis (121 357 [57.7%] male individuals; 68 325 [32.5%] aged 25-44 years) and 1 049 539 individuals with no SUD diagnosis. The mean annual medical cost attributable to SUD diagnosis per affected enrollee was $15 640 (95% CI, $15 340-$15 940), and the total annual medical cost in the ESI population was $35.3 billion (2018 USD). Alcohol use disorder ($10.2 billion) and opioid use disorder ($7.3 billion) were the most costly. CONCLUSIONS AND RELEVANCE: In this economic evaluation of medical expenditures in the ESI population, the per-person and total medical costs of SUDs were substantial. Strategies to support employees and their health insurance dependents to prevent and treat SUDs can be considered in terms of potentially offsetting the existing high medical cost of SUDs. Medical expenditures for SUDs represent the minimum direct cost that employers and health insurers face because not all people with SUDs have a diagnosis, and costs related to absenteeism, presenteeism, job retention, and mortality are not addressed.