CN115381841A - 一种异石胆酸的用途 - Google Patents
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Abstract
本发明公开一种已知化合物―异石胆酸的用途，即在制备抗缓症链球菌ATCC49456药物中的应用。本发明首次发现了异石胆酸具有抑制缓症链球菌ATCC49456的活性，实验表明异石胆酸具有良好的抑制缓症链球菌ATCC49456的活性，是一种潜在的可代替现有抗生素抑制缓症链球菌ATCC49456的药物。
Description
技术领域
本发明涉及一种已知化合物―异石胆酸的用途。
背景技术
胆汁酸是胆汁里的一个有效成分，是参与胆汁消化功能的重要成分之一。其含量变化在肝脏病诊断中具有重要的参考价值，已公开的文献中披露胆汁酸具有抑制肿瘤的效果。另，中国发明专利申请202111019726.6公开胆石酸具有抗金葡菌MRSA和MSSA、大肠埃希菌Esbls+、大肠埃希菌Esbls-和铜绿假单胞菌的抑菌活性。异石酸分子式为C24H40O3,结构式如下：
发明内容
本发明涉及异石胆酸的一种用途，即在制备抗缓症链球菌ATCC49456药物中的应用。
缓症链球菌属于α-溶血性链球菌(亦称甲型溶血性链球菌)，通常是人体口腔、消化道、生殖道等部位的正常菌群之一，该菌是一种条件致病菌，可致中毒性休克综合征，亚急性心内膜炎，前列腺炎等，此外，还可致肺炎、心包炎、腹膜炎、唾腺炎、口面部感染、牙源性感染、中耳炎、鼻窦炎等。据报道在发病猪体内分离到此病原菌，并且导致仔猪发病和死亡严重，说明缓症链球菌对猪也是有致病性。在儿科常用抗生素中，缓症链球菌对氨曲南、头孢曲松、哌拉西林/他唑巴坦、阿莫西林/克拉维酸钾均有不同程度耐药，耐药率达到42.3％-48.1％。这些抗生素目前仍然是儿童肺炎的常用药物。细菌耐药是全球问题，但相应的抗生素的开发则进展缓慢，形式非常严峻，因此迫切需要一种新的抗菌药物或抗菌产品来抑制细菌。
本发明首次发现了异石胆酸具有抑制缓症链球菌ATCC49456的活性，实验表明异石胆酸具有良好的抑制缓症链球菌ATCC49456的活性，是一种潜在的可代替现有抗生素抑制缓症链球菌ATCC49456的药物。
附图说明
图1：异石胆酸浓度与缓症链球菌生长抑制率关系图。
图2：氯霉素浓度与缓症链球菌生长抑制率关系图。
具体实施方式
本发明以下结合相关实验详细说明，实验中采用微量肉汤二倍稀释法对异石胆酸进行了体外抗菌活性测试，考察其对缓症链球菌的抗菌活性，以对照品氯霉素进行实验系统的评价。
(一)供试品及对照
1.1供试品
名称：异石胆酸，市售商品
供试品临用前用乙醇配制成最大浓度16.67mg/mL，依次对倍稀释。
1.2对照品
名称：氯霉素，市售品。
(二)测试菌株及其培养方法
2.1菌株：缓症链球菌ATCC49456
2.2菌种来源：购于北京百欧博伟生物技术有限公司。
2.3培养条件：Todd-Hewitt Broth，37℃孵育24h-48h。
2.4菌种开启接种并培养
用75％酒精棉擦拭西林瓶外部，在安全柜内使用尖嘴钳去除塑料盖及铝盖。缓慢开启胶塞，用75％酒精棉消毒瓶口部分，使用无菌吸管注入0.5ml液体培养基复溶冻干粉末，将悬液接种在培养基上，37℃培养24小时后，挑取单个菌落于液体培养基中在相同的培养条件下培养增菌。
2.5菌液工作液的准备
当OD600值达到0.5左右(约108CFU/ml)，经THB肉汤梯度稀释(0、10、100、1000、10000、100000、10000倍)后，取100μL菌悬液接种于固体培养基上，培养24h后计数活菌数，计算菌悬液浓度，再将此混悬菌液进行稀释，使菌悬液最终浓度约为2×106CFU/ml。菌种操作均在无菌条件下进行，实验完毕，所有耗材经灭菌再做丢弃处理。
(三)抑制50％受试菌所需最低抑菌浓度的测定(MIC50)
3.1测试方法：采取微量肉汤稀释法，进行MIC50的测定。
实验分为阳性对照组(即不含药物的菌悬液)、阴性对照组(含溶媒乙醇的培养基)和实验组(含不同浓度的异石胆酸菌悬液)。以上各组均设置3个平行组。将浓度为100μg/ml的异石胆酸实验液按对倍稀释溶于液体培养基中，使其终浓度为100μg/ml、50μg/ml、25μg/ml、12.5μg/ml、6.25μg/ml、3.125μg/ml、1.5625μg/ml、0μg/ml。浓度为2×106CFU·mL-1菌悬液与各组药液按比例1∶1各100μL，分别接种于无菌96孔板中，使菌悬液的终浓度为1×106CFU/ml(含105CFU/孔)，置于适当的培养条件下培养20h。同时设置对照品氯霉素，使其终浓度分别为32μg/ml、16μg/ml、8μg/ml、4μg/ml、2μg/ml、1μg/ml、0μg/ml。培养结束后，在黑色背景下进行肉眼观察，平皿内未见细菌生长的最低药物浓度即为异石胆酸的最低抑菌浓度(MIC)。所有菌种的MIC测定实验重复3次。悬浮菌体测OD600值，不同浓度的供试品及对照抑菌率的计算方法：抑菌率(％)＝1-(供试品OD值—阴性对照OD值)/(阳性对照OD值—阴性对照OD值)×100％。通过曲线拟合得函数计算而得MIC50。
3.2结果判断
本次试验结果见表1，抑制率图见图1。当阳性对照孔(即不含药物)内细菌明显生长，试验才有意义。结果显示，阳性对照孔(即不含药物)内细菌正常生长，溶媒对照培养基孔无菌生长。对照品氯霉素对缓症链球菌的MIC值为4μg/ml，参见表2和图2，符合美国临床实验室标准化委员会(National committee for clinical library standardization,NCCLS)2001年的《抗微生物药物敏感试验的执行标准：第十一版信息增刊》M100-S11中氯霉素对缓症链球菌≤4μg/ml的标准，说明本实验方法是可靠的。
统计结果表明：异石胆酸对受试的缓症链球菌的MIC50为12.52μg/ml。综上所述，异石胆酸有良好的抑制缓症链球菌活性。
表1：不同浓度异石胆酸对缓症链球菌生长影响的吸光值测定
表2：不同浓度氯霉素对缓症链球菌生长影响的吸光值测定
Claims (1)
1.异石胆酸在制备抗缓症链球菌ATCC49456药物中的应用。
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| US20190070200A1 (en) * | 2016-02-29 | 2019-03-07 | Tel Hashomer Medical Research Infrastructure And Services Ltd. | Use of bile acids and bile salts as anti bacterial agents for inhibition of bacterial conjugation and horizontal gene transfer |
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