In the context of severe coronavirus disease 2019 (COVID-19) illness, we examined endogenous glucocorticoid concentrations, steroidogenic enzyme activity, and their correlation with inflammation and patient outcomes. This observational study included 125 hospitalized COVID-19 patients and 101 healthy individuals as a reference group. We utilized LC-MS to assess serum concentrations of 11-deoxycortisol, cortisol, and cortisone, as well as activities of steroidogenic enzymes (11β-hydroxylase and 11β-hydroxysteroid-dehydrogenase type 1). Cox proportional hazards regression analysis and competing risk analysis were employed to analyze associations between glucocorticoid concentrations and outcomes, adjusting for relevant factors. In patients with COVID-19, cortisol concentrations were higher and cortisone concentrations were lower compared to the reference group, while 11-deoxycortisol concentrations were similar. Steroidogenic enzyme activity favored cortisol production. Correlations between glucocorticoid concentrations and inflammatory markers were low. A doubling in concentrations cortisol, was associated with increased 90-day mortality and mechanical ventilation (HR: 2.40 95% CI: (1.03–5.59) , P = 0.042 and HR: 3.83 (1.19–12.31), P = 0.024). A doubling in concentrations of 11-deoxycortisol was also associated to mortality (HR: 1.32 (1.05–1.67), P = 0.018), whereas concentrations of cortisone were associated with mechanical ventilation (HR: 5.09 (1.49–17.40), P = 0.009). In conclusion, serum concentrations of glucocorticoid metabolites were altered in patients hospitalized with severe COVID-19, and steroidogenic enzyme activity resulting in the conversion of cortisone to biologically active cortisol was preserved, thus not favoring critical-illness-related corticosteroid insufficiency at the enzymatic level. Glucocorticoid release did not counterbalance the hyperinflammatory state in patients with severe COVID-19. High serum concentrations of 11-deoxycortisol and cortisol were associated with 90-day mortality, and high serum concentrations of cortisol and cortisone were associated with mechanical ventilation.