A highly purified preparation of human plasma factor VIIa was submitte
d to chromogenic assays with S-2288 factors IXa, Xa, activated protein
C and thrombin being absent. Factor VIIa alone or in the presence of
calcium, kept its activity even in the presence of high concentrations
of aprotinin, inhibition appeared only in the presence of a factor VI
Ia-tissue factor complex. A two-stage amidolytic assay using activatio
n of purified factor X and hydrolysis of S-2765 chromogenic substrate
by the generated Xa was used to show a competitive inhibition with a K
i value of 30 muM. Aprotinin had no effect on factor Xa amidolytic act
ivity per se. The factor VIIa-tissue factor complex could be adsorbed
to immobilized aprotinin and removed by a chaotropic ion like KSCN 3 M
. The assays with the DFP inactivated VIIa-tissue factor complex prove
d that the interaction involved the active site of factor VIIa. The in
hibition of the VIIa-tissue factor complex was demonstrated in a clott
ing assay using aprotinin enriched normal or factor VIII deficient pla
sma.