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目的:对测定宽体金线蛭抗凝血活性的凝血酶滴定方法的影响因素进行分析。方法:用白瓷点滴板代替试管进行滴定反应(简称白瓷板法),考察白瓷板法和常规的试管法、不同凝血酶滴定间隔时间和凝血酶浓度对抗凝血活性测定的影响。结果:在同样条件下,白瓷板法测定抗凝血活性的精确度和稳定性较试管法高。用白瓷板法测定,宽体金线蛭样品浓度在一定范围内(0.125~0.333 g·ml-1)、凝血酶浓度为20 u·ml-1或10 u·ml-1时,每隔1 min滴加1次,每次5μl,样品浓度与消耗的凝血酶体积呈线性相关,r值分别为0.961和0.992;所测得样品的抗凝血活性分别为(33.08±2.64)和(31.24±1.32)u·g-1,RSD分别为8.0%和4.2%;对于0.333 g·ml-1的样品,其测量产生的理论误差不大于10%和5%。结论:白瓷板法可用于测定宽体金线蛭样品的抗凝血活性,用10 u·ml-1的凝血酶溶液,每分钟滴加一次,每次5μl进行测定,其线性和精确度均较好。
OBJECTIVE: To analyze the influencing factors of thrombin titration method for measuring anticoagulant activity of Hirudo dabryanus. Methods: The titration reaction was performed with white porcelain plate instead of test tube (white porcelain plate method). The effects of white porcelain plate method and conventional test tube method, different titration intervals of thrombin and thrombin concentration on anticoagulant activity were investigated. Results: Under the same conditions, the accuracy and stability of the white porcelain plate method in determining anticoagulant activity were higher than those in the test tube method. The white porcelain plate method was used to determine the concentration of leeches in a wide range of gold wire samples (0.125-0.333 g · ml -1) and thrombin concentration of 20 u · ml -1 or 10 u · ml -1. The concentration of thrombin was linearly correlated with the volume of thrombin consumed, with r values of 0.961 and 0.992, respectively. The anticoagulant activities of the samples were (33.08 ± 2.64) and (31.24 ± 1.32) u · g-1 with RSDs of 8.0% and 4.2%, respectively. For the 0.333 g · ml-1 samples, the theoretical error was less than 10% and 5%, respectively. CONCLUSION: The white porcelain plate method can be used to determine the anticoagulant activity of Hirudo haematobium samples. The linearity and precision of the samples were measured with 10 μl · ml-1 thrombin solution, once every 5 μl. Are better.