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研究生:簡逸栩
研究生(外文):Chien, Yi-Hsu
論文名稱:自放線菌中篩選生理活性物質之研究
論文名稱(外文):Screening of bioactive substances from actinomycetes
指導教授:蔡英傑蔡英傑引用關係
指導教授(外文):Tsai, Ying-Chieh
學位類別:碩士
校院名稱:國立陽明大學
系所名稱:生物化學研究所
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:1995
畢業學年度:83
語文別:中文
論文頁數:75
中文關鍵詞:生物化學生物學科學放線菌生理活性物質
外文關鍵詞:BIOCHEMISTRYBIOLOGYSCIENCE
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  • 被引用被引用:1
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我國製藥工業發展了數十年,始終乏善可陳,為求深植根基,篩選新型生理活性物質,以開發獨創性醫藥用或農業用產品,是絕對必要的研究投資,本研究之擬定,即基於此種深切認知,希望能建立有效之篩選系統,以用於開發 型生理活性物質。本論文的主要研究項目有三項:(1)一藥用抗細菌性物質之篩選,(2)抗真菌性物質之篩選,(3)HIV-1蛋白水解酵素抑制物質之篩選。
生理活性物質的篩選系統,可分為以下四項重要工作:(1)生產菌之分離與選擇,(2)發酵培養,(3)活性檢測,以及(4)物理化學性質鑑定。其中尤以生產菌分離與活性檢測兩項最為重要,生產菌種類繁多且特殊化,篩選方法新穎且簡便化,則篩選得新物質的可能性越高。
首先自台灣東南部採集土壤標本及木柵動物園採集動物糞便,設計組合各種微生物分離方法,自土壤及糞便中分離出各種放線菌菌種,共分離得約1847株放線菌,其中由上壤分離得約620株,而自糞便中分離得約1227株,然後對所分離之菌種,依形態初步分類整理後保存。
在活性檢測方面,利用生長抑制法及形態異常誘發法,自所分離菌種中篩選抗細菌及抗真菌物質生產菌種。另外利用HIV-1蛋白水解酵素活性偵測之ELISA系統,篩選HIV-1蛋白水解酵素抑制物質之生產菌種。結果在抗細菌活性部分篩選得98株,在抗真菌活性部分篩選到104株,但在HIV-1蛋自水解酵素抑制物質方面只篩選到2株生產菌株。
至於活性物質萃取方面,將所篩選得到之具生理活性物質生產菌,挑取5株活性較高的菌株進行液體培養,然後分菌體及菌液處理,利用乙酸乙酯萃取其活性物質,並利用bioautography檢視其活性之再現性,後來選取有活性之萃取液,進行HPLC等分析。
The pharmaceutical industry in our country has been rising for the past few decades, but there has no any real success in finding new and useful bioactive substances. In order to improve this situation, it is neccessary to screen for new bioactive substancess, moreover to develope a unique product. In our research, we intend to design an effective screening system, in order to search for new bioactive substancess. This study is divided into three major topics: Firstly, to find a bactericide for nosocomial infection; secondly, to screen for anti fungal bioactive substancess; thridly, to screen for the inhibitor HIV-1 protease.
The soil samples were collected from the south east region of Taiwan, and the animals feces from in Mu Zha zoo. We then devised methods to isolate the bacteria and actinomyces from the soil and fecal samples. In all 1847 isolates of actinomyces, 620 strains were isolated from the soil samples, and 1227 strains from the fecal samples. These strains were futher classified based on their morphological charactericitics, and were then preserved for further testing.
As to the bioassay area, the substances were tested for their anti fungal, and antibacterial activity. We identified the activity by observing the morphological changes of the tested fungi in combination with the cup assay. We also used an ELISA method for testing HIV-1 protease activity to screen for the substances that inhibit the HIV-1 protease. As a result, we found 98 strains of actinomyces that have antibacterial characteristic, and 104 strains that have anti fungal characteristic. However, only two strains were found to contain inhibitory activity to HIV-1 protease.
Wtih the method of activity guided extraction, we can find if these anti-fungal, antibacterial, or HIV-1 protease inhibitory activities are in the supernatant or in the pellet. We selected five strains to culture in medium , and then extracted with ethy1 acetate. Bioautography was then used to check if these characteristic are stable and consistent. If these characteristics are consistent, then further analysis was then carried out through HPLC.
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