一种草药氯仿萃取水相提取物止血作用的实验研究

Hemostatic properties of chloroform-water phase extract from a Chinese herb

目的:观察一种促凝血草药氯仿萃取水相提取物HB的止血作用。方法:实验于2003-12/2004-12在解放军第三军医大学西南医院全军烧伤研究所国家重点实验室完成。健康家兔18只,体质量2.5～3.0kg;健康雄性Wistar大鼠70只,体质量(250±10)g;雄性昆明种小鼠78只,体质量(25±5)g。制备HB氯仿萃取-水相提取物后给动物腹腔注射。①将70只大鼠随机分为空白对照组(n=10,30mg/kg腹腔注射生理盐水)、阳性对照组(n=10,云南白药按2mg/(kg·d)灌胃)、HB高剂量给药组(n=30,分别选取10只动物按5,10,20mg/kg剂量腹腔注射)、HB低剂量给药组(n=20,分别选取10只动物按0.5,1mg/kg剂量腹腔注射)。观察给药10d后实验动物的血生化指标(凝血酶原时间、活化部分凝血活酶时间、纤维蛋白原含量、凝血酶时间)及血小板计数;②将39只小鼠随机分为空白对照组、云南白药对照组(云南白药0.5mg/(kg·d)灌胃)和用药组(HB0.5mg/kg),每组13只。观察给药10d后小鼠的出血时间、同样的分组和干预方法观察小鼠的凝血时间。③将待试药设氯仿萃取水相提取物用药组、云南白药阳性对照组、生理盐水阴性对照组。18只家兔各取6只分别用于观察体表、肝脏及股动脉创面的止血实验,观察局部创面(皮肤、肝脏、股动脉)止血时间。结果:纳入健康家兔18只、健康雄性Wistar大鼠70只、雄性昆明种小鼠78只,均进入结果分析。①氯仿萃取水相提取物对大鼠血凝四项及血小板计数的影响:与空白对照组相比,HB氯仿萃取水相提取物腹腔注射可显著提高大鼠的血小板计数;HB高剂量给药组大鼠的凝血酶原时间明显缩短。②对小鼠出血时间及凝血时间的影响:与云南白药对照组及空白对照组相比,HB氯仿萃取水相提取物可显著缩短小鼠的出血时间[(250.2±53.3),(234.5±61.3),(100.2±27.9)s,P<0.01]及凝血时间[(202.5±50.2),(255.5±27.8),(141.8±37.6)s,P<0.01]。③对家兔局部创面的止血作用:家兔创面止血时间显著短于生理盐水阴性对照组及云南白药阳性对照组[皮肤:(41.20±6.21),(84.10±5.32),(74.80±4.73)s;肝脏:(50.20±3.39),(87.60±6.24),(74.50±5.19)s;股动脉:(62.60±4.50),(103.60±4.35),(98.20±6.07)s;P<0.01)。结论:HB氯仿萃取水相提取物具有良好的体内、外促凝血及创面止血作用。

AIM： To investigate the hemostatic property of chloroform-water extract HB from a kind of herb. METHODS： The experiment was conducted at the State Key Laboratory, Institute of Burns of Southwest Hospital, Third Military Medical University of Chinese PLA from December 2003 to December 2004. Totally 18 healthy rabbits with the body mass of （2.5-3.0） kg, 70 healthy male Wistar rats with the body mass of （250±10） g and 78 male Kunming mice with the body mass of （25±5） g were involved. Chloroform-water extract HB was prepared and then injected into animals intraperitoneally. ①Totally 70 rats were divided into blank control group （n =10,30 mg/kg saline intraperitoneally）, positive control group （n =10, 2 mg/kg per day Yunan white powder by gastric perfusion）, high-dose HB group （n =30, 5,10, 20 mg/kg in 10 animals intraperitoneally） and low-dose HB group （n =20, 0.5,1 mg/kg in 10 animals intraperitoneally）. Blood biochemical indexes such as prothrombin time （PT）, activated partial thromboplastin time （APTT）, flbrinogen （Fg）, thrombin time （TT） and platelet count （PLT） were measured 10 days after injection. ②Totally 39 mice were randomly assigned into blank control group, Yunan white powder control group （0.5 mg/kg per day Yunan white powder by gastric perfusion） and medication group （HB 0.5 mg/kg） with 13 in each group. Bleeding time and clotting time of mice by the same grouping and intervention were observed 10 days after administration. ③There were HB chloroform-water extract group, Yunnan white powder positive control group and saline negative control group. Hemostatic experiments were conducted in wound surfaces （skin, liver and femoral artery） with 6 rabbits in each experiment, and hemostatic time of partial wound （skin, liver and femoral artery） was observed. RESULTS： A total of 18 healthy rabbits, 70 healthy male Wistar rats and 78 male Kunming mice were involved in the result analysis.①Influence of HB chloroform-water extract on blood biochemical indexes and PLT： As compared with the blank control group, HB chloroform-water extract by intrapedtoneal injection could dramatically increase PLT for rats. PT could obviously shorten in the high-dose HB group. ②Influence on bleeding time and clotting time in mice; As compared with the Yunnan white powder control group and blank control group, HB chloroform-water extract could distinctly shorten the bleeding time of mice [（250.2±53.3）, （234.5±61.3）, （100.2±7.9） s,P 〈 0.01] and clotting time [（202.5±50.2）, （255.5±27.8）, （141.8±37.6） s,P 〈 0.01]. ③Hemostatic effect on partial raw surface in rabbits： Hemostatic time of rabbits was significantly shorter than that in the saline negative control group and Yunnan white powder positive control group [skin：（41.20±6.21）, （84.10±5.32）, （74.80±4.73） s;liver：（50.20±3.39）, （87.60±6.24）, （74.50±5.19） s;femoral artery： （62.60±4.50）, （103.60±4.35）, （98.20±6.07） s; P 〈 0.01]. CONCLUSION： The chloroform-water extract HB possesses good effects in promoting blood coagulation in vivo and ex vivo and has hemostatic property in wound healing.