叶柄粘球菌STXZ77的分离鉴定及抗肿瘤活性  

Screening,Identification and Antitumor Activity of Myxococcus stipitatus STXZ77

作　　者：雷良欢 黄同龙[1] 魏慧[1] 廖继燕 吴雨婧[1] 周偲[1] 夏立秋[1] 张友明[1] 

机构地区：[1]湖南师范大学生命科学学院微生物分子生物学国家重点实验室培育基地,长沙410081

出　　处：《中国生物工程杂志》2016年第11期7-15,共9页China Biotechnology

基　　金：国家"973"计划(2012CB722301);国家"863"计划(2011AA10A203);湖南省生物发育工程及新产品研发协同创新中心项目(20134486)资助项目

摘　　要：目的:从土样中分离纯化粘细菌,对其进行鉴定与归类,以丰富粘细菌菌种资源,并对其进行抗肿瘤活性初步研究,为抗肿瘤药物开发奠定基础。方法:采用灭活大肠杆菌诱导法,从土样中分离纯化粘细菌,结合形态观察、生理生化特征及16S rRNA基因序列同源性分析进行菌株鉴定;向发酵液上清中加入XAD-16大孔吸附树脂提取发酵产物粗提物,CCK-8法进行体外抗肿瘤活性研究;RP-HPLC分离抗肿瘤活性组分,LC-MS/MS分析其分子质量。结果:分离并鉴定了STXZ77菌株,命名为Myxococcus stipitatus STXZ77。该菌株的发酵产物XAD-16树脂粗提物对小鼠黑色素瘤细胞B16、小鼠乳腺癌细胞4T1、人肝癌细胞SMMC-7721、人宫颈癌细胞He La、人结肠癌细胞SW480等多种肿瘤细胞具有较好的细胞毒性,作用24h的IC_(50)值分别为5.34μg/ml、13.50μg/ml、11.93μg/ml、28.70μg/ml、48.09μg/ml,而对正常细胞人脐静脉血管内皮细胞HUVEC的毒性较小,IC_(50)值为17.09μg/ml,小于B16、4T1及SMMC-7721的细胞毒性。RP-HPLC分离得到抗肿瘤活性组分AP-C,质谱分析其分子质量为422.99m/z。结论:从土样中分离得到粘细菌Myxococcus stipitatus STXZ77,从该菌中分离得到抗肿瘤活性组分AP-C,具有开发成抗肿瘤药物的潜在价值。Objective： Screening and identifying the myxobacteria from soil samples to enrich myxobacteria resources,then preliminarily study on its antitumor activity,to lay the foundation for the exploration of anti-tumor drugs. Methods： Isolating myxobacteria strains from soil samples through inactivated Escherichia coli inducing method, identifying the strain based on morphological observation, physiologically and biochemically characterizing,and the homologous analysis of 16 S rRNA sequences of nucleotides. XAD-16 macroporous adsorption resin was added to fermented supernatant to harvest crude extracting from fermented product,and evaluated its antitumor activity in vitro through CCK-8 assay. An antitumor active component was isolated from the identified strain STXZ77 by reversed phase high-performance liquid chromatography（ RP-HPLC） technique,and its molecular weight was analyzed by LC-MS / MS. Results： The strain STXZ77,which was isolated and identified,was named Myxococcus stipitatus STXZ77. The XAD-16 resin crude extract from the fermented products expressed antitumor activity against many kinds of tumor cells,such as mouse melanoma cells B16,mouse mammary cells 4T1,human hepatoma cells SMMC-7721,human cervical carcinoma cells He La,human colon cells SW480,etc. The IC_（50） values are 5. 34μg / ml,13. 50μg / ml,11. 93μg / ml,28. 70μg / ml and48. 09μg / ml,respectively. While it showed minor toxicity to HUVEC cell line,the IC_（50） value of which is17. 09μg / ml,indicating less toxicity compared with B16,4T1 and SMMC-7721 cell lines. An antitumor active component,AP-C,was successfully isolated using RP-HPLC technique,the mass-to-charge ratio of which was determined to be 422. 99 m / z through mass spectrometry analysis. Conclusion： The myxobacterium strain Myxococcus stipitatus STXZ77 was isolated from soil samples,from which an antitumor active compound AP-C was obtained,and it has a significant value for developing antitumor drugs.

关 键 词：粘细菌 STXZ77 筛选 纯化 抗肿瘤 

分 类 号：Q93-3[生物学—微生物学]