Several cytokines are considered to be important mediators in the pathophysiology of sepsis. Cyclophilins (Cyps), the main binding proteins for the immunosuppressive drug cyclosporine A, have been suggested to function as cytokines. This study was conducted to determine (i) if serum Cyp levels were elevated in critically ill patients suffering from either sepsis or other life-threatening diseases and (ii) if so, whether there was an association between Cyp levels and a certain diagnosis and/or outcome. Serum samples of 45 patients (22 severe sepsis, 23 other diagnoses) and 17 healthy controls were prospectively analyzed by an enzymatic assay using the ability of cyclophilins to catalyze cis/trans isomerisation of peptidylprolyl-peptide bonds (PPIase activity). In addition, western blotting was applied to differentiate both isoforms. PPIase activity was significantly higher in patients with severe sepsis than in patients with other diagnoses (P = 0.004) or in healthy subjects (P = 0.001). There was no difference between healthy subjects and other critically ill patients (P = 0.067). Elevated PPIase activity was associated with high mortality (P = 0.03). It is concluded that Cyps might play a role, probably as mediators in the pathophysiology of sepsis or as symptoms of diagnostic value.