CN105920082B - 猴耳环提取物在制备抗产超广谱β-内酰胺酶肺炎克雷伯菌药物中的应用 - Google Patents
猴耳环提取物在制备抗产超广谱β-内酰胺酶肺炎克雷伯菌药物中的应用 Download PDFInfo
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Abstract
本发明公开了猴耳环提取物在制备抗产超广谱β‑内酰胺酶肺炎克雷伯菌药物中的应用。经实验证明，所述猴耳环提取物对产ESBL肺炎克雷伯菌具有较强的抗菌和杀菌作用。可作为天然的抗菌药物，应用于产ESBL肺炎克雷伯菌引起的疾病治疗。
Description
技术领域
本发明涉及猴耳环提取物的新用途，特别是其在制备抗耐药菌药物中的应用。
背景技术
21世纪是多重耐药菌的时代，抗生素临床应用60年后，越来越多的院内感染和多重耐药菌感染成为目前临床抗菌治疗的一大难题，1961年Jerons报告第1例临床耐甲氧西林金黄色葡萄球菌(MRSA)感染患者，目前MRSA感染已逐渐遍及全球；2011年中国细菌耐药性监测临床中主要耐药菌的分布，大肠杆菌、肺炎克雷伯菌产超广谱β-内酰胺酶(ESBL)株分别平均为50.7％、38.5％，其耐药性的变迁和现状备受关注。另外据我国卫生部耐药监测协作组2010年度报告，鲍曼不动杆菌和铜绿假单胞菌在医院ICU分离的病原菌中居首位，鲍曼不动杆菌对亚胺培南和美罗培南的耐药率分别高达60.4％和61.4％。综上所述，在MRSA、产ESBL的各类细菌、多重耐药的鲍曼不动杆菌和铜绿假单胞菌的临床占有率逐年攀升，耐药率不断攀高的压力下，抗生素治疗面临着巨大的挑战。为避免细菌耐药现象的进一步恶化，专家学者力图发现新的抑制细菌生长以及治疗细菌引起疾病的新方法。已有研究报道证明中国传统中药如黄连、黄芩和连翘等对不同耐药细菌有一定的抑制效果，进一步研究中药抑制耐药菌生长的重点在于发现新的抑菌能力更强、耐药抑菌谱更广的中药。
猴耳环(Pithecellobium clypearia Benth)，学名围涎树，是含羞草科猴耳环属植物猴耳环的干燥幼枝和叶，其性味苦涩寒，功效清热解毒、收湿敛疮，是治疗多种热毒症候独特的南方药材。
目前有文献公开猴耳环以及其提取物具有抗病毒作用。中国专利CN103385912A公开了猴耳环的提取物具有抗MRSA作用及抗生素增敏作用，但该专利未提及该猴耳环提取物对抗产超广谱β-内酰胺酶肺炎克雷伯菌(简称“产ESBL肺炎克雷伯菌的抗菌”)的作用。
发明内容
本发明公开了猴耳环提取物在制备抗产ESBL肺炎克雷伯菌药物中的应用。包括人用药物和其他动物用药物。
所述的猴耳环提取物可以是猴耳环水提取物或猴耳环乙醇提取物。
所述的猴耳环水提取物水或猴耳环乙醇提取物可以由以下方法制备：猴耳环粗粉用水或乙醇水溶液提取，所得的提取液再用乙酸乙酯萃取，所得的萃取物即为目标产品。
所述的乙醇水溶液是按体积比计浓度为10％-95％的乙醇水溶液。
所述的乙醇水溶液是按体积比计浓度为60％的乙醇水溶液。
本发明的有益效果在于：本发明首次公开了猴耳环提取物对产ESBL肺炎克雷伯菌的抗菌作用，经本实验证明，所述猴耳环提取物对产ESBL肺炎克雷伯菌具有较强的抗菌和杀菌作用。可作为天然的抗菌药物，应用于产ESBL肺炎克雷伯菌引起的疾病治疗。
具体实施方式
为实现上述目的，本发明通过下列实施方案对猴耳环水或乙醇提取物的抗产ESBL肺炎克雷伯菌的药理作用进行筛选。
以微量肉汤稀释法测定猴耳环提取物对产ESBL肺炎克雷伯菌的最低抑菌浓度(MIC)和最低杀菌浓度(MBC)。
菌株：产ESBL肺炎克雷伯菌(KPN，编号K1-K20)20株；大肠杆菌质控菌株(ECO，ATCC25922)均由中山大学附属第一医院检验医学部临床微生物检验室提供，经中山大学附属第一医院临床微生物检验室检测确认其耐药性。
MH肉汤培养基：MH肉汤干粉(英国OXOID LTD.)2.1g，定容至100ml，NAOH调节pH至7.0，高压灭菌，置4℃冰箱备用。
方法：参照美国国立临床试验标准化委员会(NCCLS)推荐的微量肉汤稀释法操作。
以下通过具体的实施例进一步说明本发明的技术方案。
实施例1
猴耳环提取物的制备方法
猴耳环(Pithecellobium clypearia Benth)由广州市花城制药厂提供。取适量猴耳环药材打成粗粉，用水或乙醇回流2次，每次2小时，过滤；合并滤液，浓缩得浸膏(水或乙醇提取物)。取浸膏用水混悬后，用乙酸乙酯萃取，每种溶剂萃取三次，合并乙酸乙酯萃取液，浓缩得乙酸乙酯萃取物。
所述的乙醇提取物可由10-95％乙醇回流制得。
实施例2
猴耳环水或乙醇提取物的乙酸乙酯萃取物抗产ESBL肺炎克雷伯菌的抑菌杀菌试验。
1.实验方法
1)最低抑菌浓度(MIC)的测定：
将猴耳环水或乙醇提取物的乙酸乙酯萃取物在MH肉汤培养基中进行一系列倍比稀释，每孔50μl，调节接种菌为1.0×106CFU/ml，每孔50μl菌液。35℃培养；24小时，无沉淀出现的最低抗菌药物的浓度为其最低抑菌浓度(MIC)。
2)最低杀菌浓度(MBC)的测定：
采用平板涂布计数法，从1)项无菌生长的孔中吸取50ul菌悬液至血平板上，均匀涂布，35℃培养24小时，菌落计数，使最初的实验活菌数减少99.9％或以上所需要的最低抗菌药物的浓度为其最低杀菌浓度(MBC)。
通过测定药物的最低抑菌浓度(MIC)和最低杀菌浓度(MBC)，并对数据进行统计得出MIC50、MIC90、MBC50、MBC90，来评价药物抗产ESBL肺炎克雷伯菌的作用。
2.实验结果
猴耳环水提取物的乙酸乙酯萃取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果见表1。经过统计分析，猴耳环水提取物的乙酸乙酯萃取物对产ESBLs肺炎克雷伯菌的体外抑菌和杀菌的MIC50、MIC90、MBC50、MBC90见表2、表3。
猴耳环10％乙醇提取物的乙酸乙酯萃取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果见表4。经过统计分析，猴耳环10％乙醇提取物的乙酸乙酯萃取物对产ESBLs肺炎克雷伯菌的体外抑菌和杀菌的MIC50、MIC90、MBC50、MBC90见表5、表6。
猴耳环60％乙醇提取物的乙酸乙酯萃取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果见表7。经过统计分析，猴耳环60％乙醇提取物的乙酸乙酯萃取物对产ESBLs肺炎克雷伯菌的体外抑菌和杀菌的MIC50、MIC90、MBC50、MBC90见表8、表9。
猴耳环95％乙醇提取物的乙酸乙酯萃取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果见表10。经过统计分析，猴耳环95％乙醇提取物的乙酸乙酯萃取物对产ESBLs肺炎克雷伯菌的体外抑菌和杀菌的MIC50、MIC90、MBC50、MBC90见表11、表12。
表1
猴耳环水提取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果
表2
猴耳环水提取物对产ESBL肺炎克雷伯菌的体外抑菌的MIC50、MIC90
表3
猴耳环水提取物对产ESBL肺炎克雷伯菌的体外抑菌的MBC50、MBC90
表4
猴耳环10％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果
表5
猴耳环10％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌的MIC50、MIC90
表6
猴耳环10％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌的MBC50、MBC90
表7
猴耳环60％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果
表8
猴耳环60％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌的MIC50、MIC90
表9
猴耳环60％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌的MBC50、MBC90
表10
猴耳环95％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌杀菌试验结果
表11
猴耳环95％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌的MIC50、MIC90
表12
猴耳环95％乙醇提取物对产ESBL肺炎克雷伯菌的体外抑菌的MBC50、MBC90
猴耳环水提取物的乙酸乙酯萃取物单用对产ESBL肺炎克雷伯菌的MIC50＝1600μg/ml，MIC90＝1600μg/ml，MBC50及MBC90均大于1600μg/ml。
猴耳环10％乙醇提取物的乙酸乙酯萃取物单用对产ESBL肺炎克雷伯菌的MIC50＝1600μg/ml，MIC90＝1600μg/ml，MBC50及MBC90均大于1600μg/ml。
猴耳环60％乙醇提取物的乙酸乙酯萃取物单用对产ESBL肺炎克雷伯菌的MIC50＝800μg/ml，MIC90＝1600μg/ml，MBC50及MBC90均大于1600μg/ml。
猴耳环95％乙醇提取物的乙酸乙酯萃取物单用对产ESBL肺炎克雷伯菌的MIC50＝1600μg/ml，MIC90＝1600μg/ml，MBC50及MBC90均大于1600μg/ml。
Claims (2)
1.一种猴耳环提取物在制备抗产超广谱β-内酰胺酶肺炎克雷伯菌药物中的应用，其特征在于，所述猴耳环提取物由以下方法制备：猴耳环粗粉用浓度为10％-95％乙醇水溶液提取，所得的提取液再用乙酸乙酯萃取，所得的萃取物即为目标产品。
2.如权利要求1所述的应用，其特征在于，所述的乙醇水溶液是按体积比计，浓度为60％的乙醇水溶液。
Priority Applications (9)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610308207.4A CN105920082B (zh) | 2016-05-10 | 2016-05-10 | 猴耳环提取物在制备抗产超广谱β-内酰胺酶肺炎克雷伯菌药物中的应用 |
| PCT/CN2017/071671 WO2017193635A1 (zh) | 2016-05-10 | 2017-01-19 | 猴耳环提取物在制备抗菌药物中的应用 |
| EP17795261.1A EP3456335B1 (en) | 2016-05-10 | 2017-01-19 | Pithecellobium clypearia benth extract and application for preparing anti-microbial agent |
| US15/920,480 US11154582B2 (en) | 2016-05-10 | 2018-03-14 | Method of chinese herbal medicine extract used for treating multiple diseases caused by drug resistant bacteria infection |
| US17/129,901 US20210106641A1 (en) | 2016-05-10 | 2020-12-21 | Method of chinese herbal medicine extract used for treating multiple diseases caused by drug resistant bacteria infection |
| US17/129,885 US20210106640A1 (en) | 2016-05-10 | 2020-12-21 | Method of chinese herbal medicine extract used for treating multiple diseases caused by drug resistant bacteria infection |
| US17/129,853 US11491198B2 (en) | 2016-05-10 | 2020-12-21 | Method of chinese herbal medicine extract used for treating multiple diseases caused by drug resistant bacteria infection |
| US17/494,858 US11793849B2 (en) | 2016-05-10 | 2021-10-06 | Method of Chinese herbal medicine extract used for treating multiple diseases caused by drug resistant bacteria infection |
| US17/494,850 US11654174B2 (en) | 2016-05-10 | 2021-10-06 | Method of chinese herbal medicine extract used for treating multiple diseases caused by drug resistant bacteria infection |
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| CN107970272A (zh) * | 2017-12-22 | 2018-05-01 | 成都乾坤动物药业股份有限公司 | 猴耳环或提取物在制备治疗和/或辅助治疗畜禽支气管肺炎的药物中的用途 |
| CN113384587A (zh) * | 2021-08-06 | 2021-09-14 | 吉林大学 | 委陵菜酸在制备碳青霉烯酶抑制剂中的应用 |
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| CN101032547A (zh) * | 2006-03-07 | 2007-09-12 | 广东奇方药业有限公司 | 一种抗炎抗病毒的药物组合物 |
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| CN101032547A (zh) * | 2006-03-07 | 2007-09-12 | 广东奇方药业有限公司 | 一种抗炎抗病毒的药物组合物 |
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