放线菌1356抗真菌次级代谢产物分离鉴定及活性研究

发布时间：2018-01-09 03:02

本文关键词：放线菌1356抗真菌次级代谢产物分离鉴定及活性研究　出处：《沈阳药科大学》2008年硕士论文　论文类型：学位论文

【摘要】： 放线菌1356菌株是本实验室从土壤放线菌中分离得到的一株具有抗真菌活性的菌株,系griseochromogenes的一个变种。其抗真菌次级代谢产物KZJ-1,在实验室条件下对多种农业致病真菌有很好的抑菌杀菌活性,具有很好的应用前景。本论文利用有机溶媒萃取、大孔吸附树脂洗脱和制备型高效液相等分离提纯方法,成功地从菌株1356发酵液中分离纯化了其抗真菌成分—KZJ-1。同时考察了KZJ-1的理化性质：进行了稳定性试验及纸色谱、pH色谱实验。采用核磁共振波谱(NMR)、质谱(MS)、紫外(UV)、红外(IR)吸收光谱技术确证菌种1356抗真菌代谢产物——KZJ-1为鲁斯霉素；解释该结构在NMT、MS、UV、IR等谱图中的特征吸收峰、共振峰及特征离子的一一对应归属；首次利用二维核磁共振技术确定了鲁斯霉素分子中各个C原子的归属。摸索出KZJ-1液相检测条件为：流速为0.8 mL·min-1,进样量20μL,检测波长303nm,流动相为甲醇：水=75：25,为后续工作奠定了基础。通过胞内外含量测定实验得知,KZJ-1主要存在于菌丝体当中。胞内与胞外活性成分比为7：1,这为发酵液预处理时弃去上清液提供了依据。光热稳定性实验表明,KZJ-1对光热不稳定,特别是光照条件下,KZJ-1极不稳定。KZJ-1甲醇溶液,室温避光条件下放置两个月,活性约降低为原来一半(47.7%)；但自然日光下,一个星期活性降低为原来9.4%,两个星期基本失活。采用制备型高效液相等手段,成功的从菌株1356发酵液中分离提纯出抗真菌活性物质KZJ-1纯度达98.3%。本文利用核磁共振波谱(NMR)、质谱(MS)、紫外(UV)、红外(IR)吸收光谱技术确证KZJ-1为鲁斯霉素,并用二维核磁共振技术重新对共轭烯烃部分C进行归属。
[Abstract]:1356 Actinomyces Strains with antifungal activity strains from the soil actinomycetes isolated a strain, a variant of griseochromogenes. Its anti fungal secondary metabolite KZJ-1, has a good antifungal activity to many agricultural pathogenic fungi under laboratory conditions, has good application prospects..
In this paper, using organic solvent extraction, macroporous resin elution and preparative HPLC separation and purification method, successfully from 1356 strains in fermentation liquid separation and purification of its antifungal component KZJ-1. also investigated the physicochemical properties of KZJ-1: the stability test and paper color spectrum, pH chromatography by NMR experiments. Resonance spectroscopy (NMR), mass spectrometry (MS), ultraviolet (UV), infrared (IR) absorption spectroscopy confirmed 1356 strains of antifungal metabolites - KZJ-1 lucimycin; interpretation of the structure in the NMT, MS, UV, IR and other spectral features in absorption peaks, corresponding to the resonance peak and belonging the characteristics of ion; for the first time by using 2D NMR techniques to determine the Ruth by each C atoms in molecules belonging. Find out KZJ-1 detection in liquid phase conditions: the flow rate was 0.8 mL / min-1, sample volume 20 L, the detection wavelength of 303nm, the mobile phase was methanol: water =75:25, for The continuation of the work laid the foundation.
Through intracellular and extracellular content determination experiments, KZJ-1 was mainly found in mycelia. The ratio between intracellular and extracellular active components was 7:1, which provided a basis for abandoning the supernatant during pretreatment of fermentation broth.
The experiment shows that KZJ-1 is unstable to light and heat stability, heat, especially light conditions, KZJ-1 is extremely unstable.KZJ-1 methanol solution at room temperature for two months to avoid light conditions, the activity was reduced to about half (47.7%); but in natural daylight, a week reduced activity for the original 9.4%, two weeks basic deactivation.
The purity of the anti fungal active substance KZJ-1 was purified to 98.3%. by the preparation of the equal method of preparation of high performance liquid, and the isolation and purification of the antifungal active substance KZJ-1 from the fermentation broth of the strain.
In this paper, using nuclear magnetic resonance spectroscopy (NMR), mass spectrometry (MS), ultraviolet (UV), infrared (IR) absorption spectroscopy confirmed KZJ-1 as lucensomycin, and 2D NMR techniques to C conjugated alkenes were assigned.

【学位授予单位】：沈阳药科大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R379

【相似文献】