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I nvestigation of the in vitro ability of plasma from pregnant women to inhibit exogenous thrombin (25 nM) demonstrated that heparin cofactor II inhibited more thrombin (3.0 +/- 0.7 nM, mean +/- SD) than plasma from women 3-5 d postpartum (1.9 +/- 0.5 nM) or plasma from nonpregnant adults (1.5 +/- 0.4 nM). Levels of heparin cofactor II were only slightly increased over normal in both pregnant and postpartum women and did not account for the observed increase in thrombin bound to heparin cofactor II. Assay of pregnancy plasma for dermatan sulfate anticoagulant activity demonstrated the presence of activity equivalent to 0.23 +/- 0.02 micrograms/ml of porcine mucosal dermatan sulfate. This activity could not be demonstrated in normal adult plasma or plasma from women on the contraceptive pill. The mass of dermatan sulfate in pregnancy and umbilical cord plasmas was increased over adult control plasma by 0.20 micrograms/ml (53%) and 0.29 micrograms/ml (76%), respectively. The glycosaminoglycan-containing fraction of plasma was isolated and an assay for anticoagulant dermatan sulfate confirmed its presence in both pregnancy and cord plasmas but minimal activity in adult plasma. Gel chromatography of isolated fractions from both pregnancy and cord plasmas revealed a polydisperse, active species with apparent Mr 150,000 D. Reductive elimination decreased the apparent Mr of the active species on gel chromatography to 31,000 D for cord and 21,000 D for pregnancy products. This confirmed the presence of an anticoagulant active dermatan sulfate proteoglycan circulating in the plasmas of pregnant women at term and fetuses at delivery.