The Action of a Proteolytic Enzyme From Aspergillus Oryzae on Components of the Blood Clotting System

Abstract

@r1.7, a highly purified proteolytic eilzyrne fr-on1 ~l s p e r g i l l~~v orysae, acts 011 a 11nrnbea of the c~:~yulatiori iactors. I n low c-oncentrrstio~~s, inlrncclinte action is to decrease the rec.alcifleti clottirrg tirne. 'The c\idcc~ce, as furrlish~d by exg~erirnientl-011 a prothroinbin fractio~i, is t h a t it C~) C S thi:, tiy co~iv~'rhi~lg pruthronltin to thronr~bii~. In this regard its actio11 is similar to that of thr-onil~r~plastii~. Ilurialg inc-uI,ation, espec*irnl%y mith ialcreasing concentraticirrii, it destroys prothrombi11 and fibrinogen, dud plasirla beco~i~es inc-ongul,ible. It aicstroys ilc~l-~nal pl,sm'~ ailtithronlbirr hut, paradoxically, generates ;t. thrortibiii-fibrinogen inhibiting dctivity by its proteoiytic action on f~briiiogcii in plasma. 111 :t search for better tllerapeaatic-agents to aid the treatraaent of thro~l-nbotic disorders, Stefai-aiali and illari~a (li described the preparation of a crrlde proteolytic extract frorn i1,sf)ergiii'us oryznc. Connaught hIedicaI Research 1,aboratories iinprovecl the t.xtr:~ction armd obtained a Inore l~urified product (lot 142). Tising this anaterial, Stefanini and his colleregucs studied its action on blood coagulation in vitro (2) and in rnlan (3).More recently Tosol-ai and his colleagues a t the Connaught Lal~sratorics have produced a still more acaive inaterial (Ci1.7). 'I'lais r~matcrial has the characteristics of a single enz3me. Roschlau has recently (4) evaluated the use of this eilzynle in the renlov:\l of experirncntal arterial arscl venous tlmrombi and devised : I technique for evaluatiisg indiviclual variations in protcase resistance. Our paper descritxs the action of Ci1.7 CHI different con~ponents of the blood clottirlg system. Methods and MaterialsRecnlci,fied clolting tis7zes were carried out at 37' ' C. 'To 0.1 1111 aliquots of citrated or oxalated plasrrla 0.1 rlnl of 0.0:3 , I f 'I'ris buffer and 0.1 ml of calcium chloride were added. The buffer was used as a mlet1iur-n ira which to add the proteolytic enzyrile to the test system.

Influence of Brinase on Fibrinogen: Fibrin Transition in in vitro and in vivo Studies

“…The results of Monkhouse et al [ 18] are in favor of the first explanation: Addition of brinase to a bovine prothrombin solution caused in an isolated in vitro system acceleration of prothrombin to thrombin, in the presence of calcium. The authors concluded from these findings that brinase converted prothrombin to thrombin.…”

Possibilities of Development of Fibrinolytic Activity with New and Projected Agents

“…However, it is a very toxic compound which has profound effects on the coagulation mechanism and the effective doses are so close to the toxic or even fatal dose that it seems almost certain it will not be clinically useful (22).…”

Systemic Toxicity of Ca-7 (Fibrinolytic Enzyme From Aspergillus Oryzae) in the Dog

“…Insufficient dosage, although being without toxicity, was ineffective in lysing thrombi. (Very low dosage carried the additional hazard of being thrombogenic.3~ 6,11) Owing to this narrow range of therapeutic effectiveness (i.e., the need for attainment of a delicate balance between the lowering of CA-7 inhibitors to the level of fibrinolysis, and their total depletion), a closer examination of the occurrence of systemic toxic sequelae within this range appeared essential. EXPERIMENTAL Thirty-six mongrel dogs between 12 and 20 kg in weight were identically treated with a variety of CA-7 preparations.…”