黄连解毒汤有效成分的提取及其对体内白色念珠菌的清除作用

doi:10.16431/j.cnki.1671-7236.2019.11.032

中国畜牧兽医 ›› 2019, Vol. 46 ›› Issue (11): 3414-3421.doi: 10.16431/j.cnki.1671-7236.2019.11.032

黄连解毒汤有效成分的提取及其对体内白色念珠菌的清除作用

阎力君¹, 吕欣彧¹, 张多¹, 陈昌斌², 张爽³, 高原¹, 雒江菡¹, 王福龄¹, 崔闻宇¹, 张乘蚬¹

1. 哈尔滨商业大学药学院, 细胞与分子生物学研究所, 哈尔滨 150076;
2. 中国科学院上海巴斯德研究所, 中国科学院分子病毒与免疫重点实验室, 上海 200031;
3. 辽宁省凌海市翠岩动物卫生监督所, 凌海 121227

收稿日期:2019-05-13 出版日期:2019-11-20 发布日期:2019-11-25
通讯作者: 阎力君 E-mail:ylj@hrbcu.edu.cn
作者简介:阎力君(1963-),女,黑龙江哈尔滨人,博士,教授,研究方向:生物大分子结构功能及药物干预
基金资助:
国家自然科学基金项目（31370764）；2018年度省属高校科技成果研发、培育、转化支持计划项目（TSTAU-R2018011）

Extraction of Effective Fraction of Huanglian Jiedu Decoction and Its Scavenging Effect on Candida albicans in vivo

YAN Lijun¹, LYU Xinyu¹, ZHANG Duo¹, CHEN Changbin², ZHANG Shuang³, GAO Yuan¹, LUO Jianghan¹, WANG Fuling¹, CUI Wenyu¹, ZHANG Chengxian¹

1. School of Pharmacy, Institute of Cellular and Molecular Biology, Harbin University of Commerce, Harbin 150076, China;
2. CAS Key Laboratory of Molecular Virology & Immunology, Institute Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai 200031, China;
3. Cuiyan Animal Health Supervision Institute, Liaoning Province, Linghai 121227, China

Received:2019-05-13 Online:2019-11-20 Published:2019-11-25

摘要/Abstract

摘要： 试验旨在研究黄连解毒汤有效成分的提取及对其体内白色念珠菌的清除作用。分别采用水煎煮和乙醇回流提取法，随后乙醇提取物用3种不同极性的有机溶剂萃取（正丁醇、乙酸乙酯、石油醚），以最小抑菌浓度为指标优化出最佳抑菌效果的提取工艺；观察给药前后系统性感染白色念珠菌小鼠肾脏、肝脏的真菌负荷量及活性氧（ROS）含量水平的变化，并检测血清中细胞因子白细胞介素-6（IL-6）、IL-1β、白细胞介素-1β前体（pro-IL-1β）及肿瘤坏死因子-α（TNF-α）的含量。结果显示，黄连解毒汤抑菌效果最优的提取工艺为：料液比1：30，乙醇浓度60%，温度80℃，提取时间1 h，提取次数2次，测得乙酸乙酯萃取物MIC₉₀为0.3125 mg/mL。与模型组相比，黄连解毒汤乙酸乙酯提取物（EAHD）高剂量组能显著降低小鼠肾脏真菌负荷量（P<0.05），完全治愈率达40%，并明显高于氟康唑组；EAHD低、中和高剂量组（50、100及200 mg/（kg·d））均能显著降低肝脏真菌负荷量（P<0.05），其中中剂量组和氟康唑组降低肝脏真菌负荷量效果接近。同时EAHD高剂量组能显著或极显著降低感染后小鼠肾脏和肝脏ROS含量（P<0.05；P<0.01）；显著或极显著提高血清中IL-6、IL-1β和TNF-α的含量（P<0.05；P<0.01），同时降低pro-IL-1β含量。综上所述，通过优化后的提取工艺得到的EAHD具有降低模型小鼠肾脏和肝脏ROS含量和调控炎症因子的作用，进而降低炎症反应，起到保护组织和抵抗白色念珠菌的作用。

关键词: 提取工艺优化; 黄连解毒汤乙酸乙酯提取物; 白色念珠菌; 真菌负荷量

Abstract: The aim of the experiment was to study the extraction of effective fractions of Huanglian Jiedu decoction and its scavenging effect on Candida albicans in vivo.The fractions of Huanglian Jiedu decoction were extracted by water and ethanol reflux,and then the ethanol extract was extracted with three different polar organic solvents (n-butanol,ethyl acetate,petroleum ether).The extraction process was optimized with the minimum inhibitory concentration (MIC) as the index.The changes of fungal load in kidney and liver,and reactive oxygen species (ROS) levels in a mouse model with systemic Candida albicans infection (SC) before or after taking medicine were observed.The levels of cytokines IL-6,IL-1β,pro-IL-1β and TNF-α in serum were measured by ELISA.The results showed that the optimal extraction process of Huanglian Jiedu decoction was the following:The material-liquid ratio was 1:30,the ethanol concentration was 60%,the temperature was 80℃,the extraction time and frequencey were 1 h and 2 times.Under this condition,the MIC₉₀ of EAHD was 0.3125 mg/mL.Compared with the model group,Huanglian Jiedu decoction ethyl acetate extract (EAHD) high dose group significantly reduced the kidney fungal load of mice (P<0.05),and the complete cure rate was 40% which was obvious higher than that of fluconazole group.The fungal load of liver was significantly decreased in EAHD low,medium and high dose groups (50,100 and 200 mg/(kg·d))(P<0.05),and the effect of EAHD medium dose group and fluconazole group on reducing the hepatic fungal load was similar.The ROS content in kidney and liver of EAHD high dose group were significantly or extremely significantly reduced,respectively (P<0.05;P<0.01),and the levels of IL-6,IL-1β and TNF-α in the serum were significantly or extremely significantly increased,respectively (P<0.05;P<0.01).And the pro-IL-1β content in the serum was reduced.In conclusion,the EAHD obtained by the optimal extraction process could reduce ROS content and regulate inflammatory factors in kidney and liver of model mice,thereby alleviate inflammatory reaction to protect tissues and resist Candida albicans.

Key words: extraction process optimization; ethyl acetate extract of Huanglian Jiedu decoction; Candida albicans; fungal load

中图分类号:

R285

阎力君, 吕欣彧, 张多, 陈昌斌, 张爽, 高原, 雒江菡, 王福龄, 崔闻宇, 张乘蚬. 黄连解毒汤有效成分的提取及其对体内白色念珠菌的清除作用[J]. 中国畜牧兽医, 2019, 46(11): 3414-3421.

YAN Lijun, LYU Xinyu, ZHANG Duo, CHEN Changbin, ZHANG Shuang, GAO Yuan, LUO Jianghan, WANG Fuling, CUI Wenyu, ZHANG Chengxian. Extraction of Effective Fraction of Huanglian Jiedu Decoction and Its Scavenging Effect on Candida albicans in vivo[J]. China Animal Husbandry and Veterinary Medicine, 2019, 46(11): 3414-3421.

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黄连解毒汤有效成分的提取及其对体内白色念珠菌的清除作用

Extraction of Effective Fraction of Huanglian Jiedu Decoction and Its Scavenging Effect on Candida albicans in vivo

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