论文部分内容阅读
目的探讨溶血毒素活性测定的标准化。方法以溶血毒素产生藻——海洋卡盾藻为材料,在系统分析和考查超声波细胞破碎法和液氮研磨法、不同减压蒸馏温度、不同来源红细胞及样品保存时间和温度等对溶血毒素活性影响的基础上,提出海洋卡盾藻溶血毒素及活性的提取、测定最优化方法。结果超声波细胞破碎法和液氮研磨法得到的溶血活性分别为288.23和94.89HU/L;超声时间分别为5、10、20和30min时,测得溶血活性分别为80.57、157.45、288.23和279.17HU/L;减压蒸馏温度分别为40、60和80℃时,溶血活性为288.23、124.97和120.68HU/L。分别选择兔、昆明鼠、花身和人的红细胞作为底物,测定等量毒素的溶血活性,测得溶血活性分别为244.98、288.23、266.35和195.47HU/L;将离心后的藻细胞分别于0、20和-20℃保存1、3和7天,藻细胞的溶血活性随时间延长逐渐降低。结论超声波为最佳藻细胞破碎法;功率为200W时,超声时间以20min为宜;蒸馏温度应保持在40℃左右;兔红细胞对溶血毒素最为敏感。实际测定时,最好选择5%兔血红细胞;藻类样品在0℃下最多可保存3天。
Objective To investigate the standardization of the determination of hemolytic toxin activity. Methods Hemolytic toxin-producing marine algae-marine Cladophylla was used as material in the systematic analysis and examination of the effects of ultrasonic cell disruption and liquid nitrogen grinding, different vacuum distillation temperatures, different sources of red blood cells and sample storage time and temperature on hemolytic toxin activity On the basis of the proposed Marine Cladophora hemolytic toxin and activity of extraction, determination of optimization methods. Results The hemolytic activities of the cells were 288.23 and 94.89 HU / L, respectively. When the ultrasonic time was 5, 10, 20 and 30 min, the hemolytic activities were 80.57, 157.45, 288.23 and 279.17HU, respectively / L; when the vacuum distillation temperature was 40, 60 and 80 ℃ respectively, the hemolytic activity was 288.23, 124.97 and 120.68HU / L. The haemolytic activity of the same amount of toxins was determined by using rabbit, Kunming mouse, Hystrix pallidum and human erythrocytes as substrate respectively, and the hemolysis activities were 244.98,288.23,266.35 and 195.47HU / L, respectively; After 1, 3 and 7 days of storage at 0, 20 and -20 ° C, the hemolysis activity of algal cells gradually decreased with time. Conclusion Ultrasound is the best method of algae cell disruption. When the power is 200W, the ultrasonic time is 20min, the temperature of distillation is about 40 ℃, and the rabbit erythrocytes are the most sensitive to hemolytic toxin. In the actual measurement, it is best to choose 5% rabbit red blood cells; algae samples can be stored at 0 ℃ up to 3 days.