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当前位置: 首页 > 文章 > 一株野生粗毛纤孔菌的分类鉴定及其发酵上清液的抗肿瘤活性 南方农业学报 2019,50 (8) 1671-1679
Position: Home > Articles > Classification and identification of a wild Inonotus hispidus and the antitumor activity of the fermentation supernatant Journal of Southern Agriculture 2019,50 (8) 1671-1679

一株野生粗毛纤孔菌的分类鉴定及其发酵上清液的抗肿瘤活性

作  者：
唐少军;陈定安;邵晨霞;许隽;杨祎;靳磊;吴胜莲
单  位：
湖南省微生物研究院;常德市鼎城区农业科学研究所
关键词：
粗毛纤孔菌;鉴定;抗肿瘤活性;液相色谱分离;半致死浓度(IC50)
摘  要：
【目的】对一株野生粗毛纤孔菌进行分类学鉴定,并分析其发酵上清液抗肿瘤活性,为粗毛纤孔菌的开发利用及抗肿瘤活性机制研究提供理论依据。【方法】通过形态特性观察、ITS序列分析及系统发育进化树构建等方法对一株野生粗毛纤孔菌(简称IH3菌株)进行分类学鉴定,并利用B16、Hep-3B、Hela、MCF-7、HCT-116和4T1 6种肿瘤细胞及正常细胞HUVEC分析该菌株发酵上清液的抗肿瘤谱。从石油醚、乙酸乙酯和正丁醇中筛选出萃取抗肿瘤活性物质的最佳有机溶剂,利用液相色谱分离技术分离抗肿瘤活性组分,并利用MTT(四唑盐)法分析活性组分对不同肿瘤细胞的半致死浓度(IC50)。【结果】IH3菌株的形态特性与粗毛纤孔菌相似,且其ITS序列与粗毛纤孔菌(MF183947.1)的相似度最高,为98%,鉴定为粗毛纤孔菌。系统发育进化树分析结果也显示,IH3菌株属于纤层孔菌属类群,与3个粗毛纤孔菌菌株的亲缘关系较近。IH3菌株的发酵上清液对不同肿瘤细胞的抑制活性排序为:B16>Hep-3B>Hela>MCF-7>4T1=HCT-116。正丁醇萃取的IH3菌株发酵上清液抗肿瘤活性组分对B16肿瘤细胞的抑制率最高,达80%,说明正丁醇对发酵上清液中抗肿瘤活性组分的萃取效果最佳。利用液相色谱分离方法可有效获得纯度较高的单一抗肿瘤活性组分(WIH3),该活性组分对B16、Hep-3B、Hela和MCF-7肿瘤细胞的半致死浓度(IC50)分别为29.322、37.387、47.029和58.009μg/mL。【结论】IH3菌株被鉴定为粗毛纤孔菌,与已发表的粗毛纤孔菌菌株存在遗传差异,可能为粗毛纤孔菌的新亚种,其发酵上清液具有广谱的抗肿瘤活性。
译  名：
Classification and identification of a wild Inonotus hispidus and the antitumor activity of the fermentation supernatant
作  者：
TANG Shao-jun;CHEN Ding-an;SHAO Chen-xia;XU Jun;YANG Yi;JIN Lei;WU Sheng-lian;Hunan Microbiology Institute;Agricultural Science Research Institute of Dingcheng District;
关键词：
Inonotus hispidus;;identification;;antitumor activity;;liquid chromatography;;medial lethal concentration
摘  要：
【Objective】The wild Inonotus hispidus was classified and identified,and the antitumor activity of its fermentation supernatant was analyzed,which provided a basis for the development and utilization of this strain as well as research of antitumor activity mechanism.【Method】A wild I. hispidus(IH3 fungi)was classified and identified through the morphology observation,ITS sequence and phylogenetic tree. The antitumor spectrum of the fermentation supernatant was analyzed by six kinds of tumor cells including B16,Hep-3 B,Hela,MCF-7,HCT-116 and 4 T1 and normal cell HUVEC.The optimal organic reagent was selected from petroleum ether, acetic ether and normal butanol. The antitumor active component was isolated by liquid chromatography,and medial lethal concentration(IC50)of different tumor cells was analyzed by MTT(tetrazolium salt).【Result】IH3 fungi was similar to I. hispidus in morphology,its ITS sequence was the closest to the I. hispidus strain MF183947.1 with the similarity of 98%. It was preliminarily identified as I. hispidus.The phylogenetic tree analysis indicated that IH3 fungi belonged to Inonotus,and had close relationship with three I. hispidus strains. Inhibition activity of IH3 fungi fermentation supernatant to the tumor cells was:B16>Hep-3 B>Hela>MCF-7>4 T1=HCT-116. The inhibition rate of antitumor active component of IH3 strain fermentation supernatant extracted from normal butanol to B16 tumoe cell was the highest(80%),indicating that normal butanol had the best extraction effect on antitumor active substances in fermentation supernatant. A single antitumor active component(WIH3)with high purity was obtained by liquid chromatography. Its IC50 to tumor cells B16,Hep-3 B,Hela and MCF-7 were 29.322,37.387,47.029,58.009 μg/mL,respectively.【Conclusion】The IH3 strain was identified as I. hispidus,which is different from the published I. hispidus strains genetically,it may be a new subspecies of I. hispidus,and its fermentation supernatant has a broad spectrum of antitumor activity.

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