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THROMBINASE
The search for new and more efficacious thrombolytic
agents is a rapidly advancing field of research. Several products are currently
in
the market, e.g., streptokinase, urokinase and tissue plasminogen activator
(TPA). The new product is a novel thrombin dissolving agent containing a fibrinolytic
enzyme named as thrombinase, isolated, identified and purified for the first
time from a Bacillus species. This product has potential use for the treatment
of cerebral thrombosis (stroke), myocardial infraction and deep vein thrombosis
and in the prevention of post surgical adhesions. The preparation can be used
on its own or in combination with specific antibodies. Though, the product is
not a plasminogen activator, it acts very specifically on fibrin clots. The
fibrinolytic action is greater than streptokinase or urokinase. It has optimum
activity in the range of 7-9 pH and is stable over the pH range 5-7.5. Molecular
weight (gel filtration) was estimated to be around 20,000. It is stable below
40oCand retains 80 % or more activity on standing for 24 hours at
30oC.
BIOLOGICAL ACTIVITY
Fibrinolytic activity of the purified enzyme was evaluated in comparison to standard streptokinase according to the modified method of Astrup and Mullertz (1952). Fibrinolytic potency was estimated by measuring the extent of liquefaction (zone diameter) produced by the enzyme on fibrin clot of fibrinogen, plasminogen and thrombin. The reference standard was diluted to contain 500 units per ml. Thrombinse was dissolved at pH 7.5 to contain 1 mg protein per ml. Quantities of 20 and 40 m l of streptokinase and the test preparation were placed as a droplet on the surface of the fibrin clot and incubated. The diameter of the digested area was measured after 30 minutes of incubation.
IDENTITY AND PURITY
Identity and purity of the final product was checked by HPLC analysis and as found quite encouraging.
PRESENT STATUS OF THE TECHNOLOGY
Scale of work: Pilot Plant Scale (100 litre capacity).
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