The effect of oxygen exposure on nitrosylhemoglobin of whole human blood or its buffered solution has been determined. The amount of methemoglobin formed was determined by an anaerobic modification of the Evelyn-Malloy method; 59% of the total hemoglobin of whole blood was oxidized to methemoglobin in the first 15 min of the oxygen exposure and 78% of the total hemoglobin was oxidized after 120 min of oxygen exposure. Similar results were obtained when nitrosylhemoglobin buffered solutions were exposed to the oxygen of the air. A comparison of the present in vitro results with these obtained by injecting nitric oxide into the rat peritoneal cavity and its implications are discussed.