Screening of Microbial Metabolites Affecting Cell Membrane Function (An Attempt to Effectuation of Ineffective Inhibitors for Fungal Growth by Permeability Changes by the Metabolites)

据报道，当某些酵母菌弱暴露于细胞膜损伤剂如蓼二醛和黄曲霉素B时，它们对各种抗生素和抑制剂变得过敏。在本研究中，我们试图寻找能够特异性作用于酵母细胞膜屏障的微生物代谢产物，从而使不起作用的非渗透性抑制剂能够进入酵母细胞并发挥其活性.制备用于筛选试验的酵母：用NTG处理酿酒酵母IFO 0203细胞，以产生（1）arg-p3（精氨酸通透酶缺陷型突变体）和（2）ura-p17（尿嘧啶通透酶缺陷型突变体），它们分别对精氨酸类似物刀豆氨酸和尿嘧啶类似物5-氟尿嘧啶不敏感。此外，（3）白色念珠菌IFO 1061被发现缺乏尿嘧啶透过酶。仅当在刀豆氨酸或5-氟尿嘧啶存在下进行测试时，才对约3100种培养滤液进行筛选，是否显示出针对（1）-（3）的抗酵母活性。结果，4种微生物（一种未鉴定的丝状真菌，青霉属，一种未鉴定的放线菌和链霉菌属）。被认为是这种活动的有前途的生产者。从它们的培养液中分离得到4种活性成分（影响细胞膜的代谢产物），分别鉴定为chrysodin、anhydrofulvic acid、chainin和tetramycin A.除刀豆氨酸和5-氟尿嘧啶外，当与这些影响细胞膜的代谢物联合使用时，还可有效抑制真菌生长. Chrysodin抑制酵母细胞亚麻酸和亚油酸的生物合成。由此可见，多烯脂肪酸在细胞膜对药物的渗透屏障中起着重要作用。需要进一步研究以阐明其他筛选代谢物的作用模式。

It has been reported that some yeasts become hypersensitive to various antibiotics and inhibitors when they are weakly exposed to cell membrane-damaging agents such as polygodial and amphotericin B. In this study we tried to search for microbial metabolites which can act specifically on the membrane barrier of yeast cells, so that impermeable inhibitors showing no effect would be able to enter the cells and show their activities.1. Yeasts for screening test were prepared : Cells of Saccharomyces cerevisiae IFO 0203 were treated with NTG to give (1) arg-p3 (arginine permease-deficient mutant) and (2) ura-p17 (uracil permease-deficient mutant) which are insensitive to the arginine analogue canavanine and the uracil analogue 5-fluorouracil, respectively. Also, (3) Candida albicans IFO 1061 was found to be deficient in uracil permease.2. About 3100 kinds of culture filtrates were subjected to the screening whether they showed anti-yeast activity against (1) - (3) only when they were tested in the presence of canavanine or 5-fluorouracil. As a result, 4 microorganisms (an unidentified filamentous fungus, Penicillium sp., an unidentified actinomycetes and Streptomyces sp.) were found to be promising producers of such activities.3. From their culture broths, 4 active principles (cell membrane-affecting metabolites) were isolated and identified as chrysodin, anhydrofulvic acid, chainin and tetramycin A, respectively.4. Besides canavanine and 5-fluorouracil, some other ineffective inhibitors for fungal growth were effectuated when combined with these cell membrane-affecting metabolites.5. Chrysodin inhibited the biosynthesis of linolenic acid and linoleic acid in yeast cells. Thus, it seems that polyenoic fatty acids play an important role in the permeability barrier of cell membrane to drugs. Further studies are needed to clarify the mode of action of other screened metabolites.