Chinese Medicine Leech H.nipponia Whitman, The Anticoagulant Activity Of Parts Of The Pre-clinical Studies

Abstract The anticoagulation segment of the Chinese traditional medicine Leech had been extracted and isolated. Furthermore, the agent preparation and administration pathway of the segment was decided. In order to the request of preclinical study technology of the Chinese traditional medicine, the preparation technology, quality control method of the raw material and the end product were also set up. On the basis of all the work, the safety, anticoagulant and antithrombotic effective of the end product were preliminary evaluated. The anticoagulation segment was extracted with phosphate buffer (0.2M, PH 7.0) at room temperature, after removal of insoluble material by centrifugation. The EtOH concentration of the extract was adjusted to 60% by addition of cold EtOH and the precipitate was discarded. The supernatant was concentrated at 40℃ under decompression. The anticoagulation segment was obtained from the concentrated extract by precipitation with 80% EtOH after the concentration extract was adjusted to PH 4.1 by the addition of hydrochloric and by drying. The solution of the segment in phosphate buffer( 0.2M, PH 7.0)and 20mg/ml Manicol was filtrated and lyophilizated, then the end product was obtained. As to the assess of the antithrombin activity of the raw material, the anticoagulation activity of 1g raw material was decided no less than 70 antithrombin unit, at the same time, a method with little interference, good linearity, high precision and accuracy, assessing the protein content and the antithrombin activity of the end product was provided. The end product showed no haemolysis in 2% rabbit blood saline solution, no blood vessel stimulating effective after rabbit ear injection by vein and no allergic after cavy injection by muscle. The antithrombin activity of the end product cound not be tested by pour stomach at 600AT-u/kg on mice. The end product could elongate the clotting time, prothrombin time and kaolin partial thromboplastin time at 120AT-u/kg dose injection by candal vein of the mice, the clotting time was correlating with the dose. The product could also elongate the thrombin time in invitro, while the traditional anticoagulation drug heparin couldn't. The anticoagulation activity of the end product could measure quickly after injection by vein, the high anticoagulation effective lasted about thirty minutes, then decline, two hours after injection, the anticoagulation effective of the end product was similar to the saline's. At 120 AT-u/kg dose injection by candal vein on the rat ten minutes before operation, the end product reduced thrombus in the inferior vena cava. At 480AT-u/kg, the end product can completely prevent the thrombus in the inferior vena cava.