产紫青霉G59的庆大霉素抗性突变株新产抗肿瘤活性产物研究

目的阐明真菌无活性野生株产紫青霉G59的庆大霉素抗性突变株2-5-3-1新产抗肿瘤活性产物。方法在活性跟踪和薄层检测指导下,通过与原始菌样品直接对照,利用液液萃取、柱层析、制备薄层层析、重结晶等技术,分离纯化活性产物。根据理化常数和波谱数据鉴定化合物结构。用MTT法测试样品对K562细胞的抑制活性。结果从突变株2-5-3-1发酵物中分离鉴定了麦角甾醇（1）、fructigenine A（2）、rugulosuvine A（3）、大黄素（4）和ω-羟基大黄素（5）。化合物1~5在100μg/ml浓度下对K562细胞的抑制率分别为52.8%、60.8%、41.2%、84.6%和37.1%,其中1、2和4的IC50分别为93.1、58.4和30.0μg/ml。结论化合物1~5均为产紫青霉G59不生产而突变株新产抗肿瘤活性产物。用DMSO介导的庆大霉素抗性筛选方法可以将无活性真菌野生株转化成活性突变株并供新产活性产物研究。

Antitumor metabolites newly produced by a gentamicin-resistant mutant of Penicillium purpurogenum G59

Objective To investigate the antitumor metabolites newly produced by the fungal strain,2-5-3-1,a gentamicin-resistant mutant of marine-derived wild-type fugal strain Penicillium purpurogenum G59 that does not produce antitumor secondary metabolites.Methods The bioactive metabolites produced by the mutant 2-5-3-1 were isolated by a bioassay-guided separation procedure with the liquid-liquid extraction,chromatography and recrystallization methods through direct comparison with the sample from Penicillium purpurogenum G59.The compounds obtained were identified by physico-chemical properties and spectroscopic data.The antitumor activity was assayed by the MTT method using K562 cells.Results Five bioactive metabolites were isolated from the fermentation products of mutant 2-5-3-1 and identified as ergosterol（1）,fructigenine A（2）,rugulosuvine A（3）,emodin（4） and citreorosein（5）,respectively.Compounds 1-5 inhibited the proliferation of K562 cells with the inhibition rates of 52.8%,60.8%,41.2%,84.6% and 37.1% at the 100 μg/ml,respectively.The IC50 values of 1,2 and 4 for the inhibition of K562 cell proliferation were determined to be 93.1,58.4 and 30.0 μg/ml,respectively.Conclusion Compounds 1-5 are antitumor metabolites newly produced by the mutant 2-5-3-1 in comparison with its parent strain Penicillium purpurogenum G59,which have not been found in the metabolites of Penicillium purpurogenum yet.It has been revealed from the present result that the introduction of gentamycin-resistant mutation by treatment with gentamycin under the DMSO-mediation can alter the secondary metabolism in Penicillium purpurogenum G59 and thus would be useful in the expansion of fugal strain sources for drug screening.