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Multiple blood samples were collected over the first 15 min after an i.v. injection of 25 units/kg of heparin in four healthy subjects. Plasma heparin activity in each sample was determined by a chemical neutralization assay using polybrene and a bioassay based on activated partial thromboplastin time. Chemically assayed heparin declined much more rapidly than bioassayed heparin over the first 5-7 min after the dose. Subsequently, both heparin activity vs time curves declined with a similar terminal half-life. Slopes of the in vivo 1n APTT vs plasma heparin activity (determined by chemical neutralization) relationships were significantly greater than those of the corresponding in vitro relationships between 1n APTT vs heparin activity added to plasma. A possible explanation for these heparin assay differences is a rapid in vivo enhancement of the anticoagulant effect of heparin. How such enhancement may occur, however, is presently unclear.