The Activation of Bovine Factor X by Factors IXa and VIII.

The esterase activity of bovine factor xa on the synthetic substrate α-N-benzoyl-L-arginineethyl ester can be used to follow the activation of factor X by the intrinsic pathway of coagulation. Bovine factor IXa activates factor X in a reaction having an absolute requirement for calcium ions, but little affected by phospholipid. At a molar ratio of factor IXa to factor X of 1:10, 3% of the factor X is activated after five minutes incubation at 37°C; the inclusion of bovine factor VIII in the incubation mixture results in 35% activation in the same period. Factor VIII cannot activate factor X in the absence of factor IXa. There is not an absolute requirement for phospholipid in the interaction of factors X, IXa and VIII, but the addition of crude cephalin further accelerates the rate of factor X activation. Hirudin does not affect the interaction of factors IXa and X in the absence of factor VIII, but does reduce the stimulatory effect of factor VIII on the rate of formation of factor Xa by a factor of between two and four. As hirudin specifically inhibits thrombin, this effect on factor X activation is likely to reflect the presence of trace amounts of thrombin in the incubation mixtures. The incomplete inhibition by hirudin of the effect of factor VIII on factor X activation suggests that ‘activation’ of factor VIII by thrombin cannot be a prerequisite for its interaction with factors IXa and X.