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[目的]研究苦丁茶不同粗提物的抑菌效果。[方法]制备了3种苦丁苯粗提物。粗提物1:称取苦丁茶50 g,加500 ml去离子水浸泡10 h,加热至沸腾,文火2h,过滤。药渣再加3～5倍去离子水煎煮,方法同上,重复3次。合并3次提取液,用旋转蒸发仪浓缩成50 ml,即每1 ml相当于含生药1.0 g,粗提物2:称取50 g干粉(用中药粉碎机粉碎后过80目),加500 ml去离子水,水浴80℃恒温振荡3 h,离心,取上清液,浓缩,加3倍体积无水乙醇,离心取沉淀,用去离子水配成50 ml溶液。粗提物3:称取50 g干粉,加500 ml 90%乙醇,采用回流提取法,同流时间3 h,回流次数3次,合并3次提取液,过滤,用旋转蒸发仪浓缩成50 ml。粗提物1、粗提物2、粗提物3分装于棕色试剂瓶,封口,于-20℃冰箱中保存,使用前121℃高压灭菌20 min采用试管二倍稀释法和纸片扩散法,对金黄色葡萄球菌与大肠埃希菌进行体外抑菌试验,并测定其最小抑菌浓度(MIC),最小杀菌浓度(MBC)和抑菌环直径。[结果]粗提物1抑菌效果较强,高于粗提物2和粗提物3,对金黄色葡萄球菌与大肠埃希菌的MIC分别为3.91 mg/ml和31.25 mg/ml。粗提物1对金黄色葡萄球菌的抑菌作用效果最好,抑菌能力最强,抑菌作用为高敏,抑菌环直径达(17.46±0.40)mm,对大肠埃希菌的抑菌作用为中敏,但比粗提物2、粗提物3好;粗提物2对这2种菌的抑菌能力都较差,均为低敏,特别是对大肠埃希菌,抑菌环直径只有(6.30±0.16)mm。粗提物3对金黄色葡萄球菌、大肠埃希菌的抑菌作用相差不大,都为中敏。粗提物1不但抑菌效果好,且其提取方法、步骤简单,仪器设备价格低,同时不产生废气,可为临床应用上提供支持并带来方便。粗提物3需使用有机溶剂,大量使用容易产生三废,给环境带来一定的危害。考虑到抑菌能力及成本、环保和简便等因素,采用提取粗提物1的方法在生产上比较实用。[结论]苦丁茶的不同粗提物均有不同程度的抑菌作用,为苦丁茶开发成新型、安全的抑菌剂提供了参考。
[Objective] The research aimed to study the antibacterial effect of different extracts of Kudingcha. [Method] Three crude extracts of Butylbenzene were prepared. Crude extract 1: Weigh 50 g Kudingcha, add 500 ml of deionized water soak 10 h, heated to boiling, slow fire 2h, filtered. Dregs plus 3 to 5 times deionized water boiling, the same way, repeated 3 times. The combined 3 extracts, concentrated by rotary evaporator to 50 ml, equivalent to 1 ml per 1 ml crude drug containing crude extract 2: Weigh 50 g of dry powder (crushed with a Chinese medicine mill over 80 mesh), add 500 ml deionized water, the water bath was shaken at 80 ° C for 3 h at constant temperature, centrifuged, the supernatant was concentrated, concentrated, three times the volume of absolute ethanol, centrifuged to take a precipitate, and deionized water to form a solution of 50 ml. Crude extract 3: Weigh 50 g of dry powder, add 500 ml of 90% ethanol, using reflux extraction method, the same flow time 3 h, reflux times 3 times, the combined extracts 3 times, filtered, concentrated by rotary evaporator to 50 ml . Crude extract 1, crude extract 2, crude extract 3 minutes packed in brown reagent bottle, sealed, stored at -20 ℃ freezer, 121 ℃ before using autoclave sterilization 20min by two-fold dilution of the test tube and diffusion of the paper Method, in vitro Staphylococcus aureus and Escherichia coli bacteriostasis test and determination of its minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and bacteriostatic ring diameter. [Result] The antibacterial effect of crude extract 1 was stronger than that of crude extract 2 and crude extract 3, and the MIC values of Staphylococcus aureus and Escherichia coli were 3.91 mg / ml and 31.25 mg / ml, respectively. Crude extract 1 has the best antibacterial effect on Staphylococcus aureus, the strongest antibacterial activity, the high antibacterial activity and the diameter of antibacterial loop (17.46 ± 0.40) mm, which has the antibacterial activity against Escherichia coli For the sensitive, but crude extract 2, crude extract 3 is good; crude extract 2 of these two kinds of bacteria are poor antibacterial capacity, are low sensitivity, especially for Escherichia coli, antibacterial ring Diameter only (6.30 ± 0.16) mm. Crude extract 3 of Staphylococcus aureus, Escherichia coli antibacterial effect is not much difference, are sensitive. Crude extract 1 not only has good bacteriostatic effect, but also has the advantages of simple extraction method, simple steps, low price of apparatus and equipment and no exhaust gas, which can provide support for clinical application and bring convenience. Crude extract 3 requires the use of organic solvents, heavy use of easily produced three wastes, to the environment bring some harm. Taking into account the ability of antibacterial and cost, environmental protection and simple and other factors, the extraction of crude extract 1 method is more practical in the production. [Conclusion] The different crude extracts of Kudingcha had different degree of antibacterial activity, which provided a reference for the development of new and safe antimicrobial agents.