(I) 藉由抗氧化特性探討薑在治療泛抗藥性鮑氏不動桿菌之協同效應 (II) 蘭嶼肉桂作為人類Lon蛋白酶抑制劑及抗氧化劑之研究

(I) Constituents of 2-methoxyphenol from Zingiber officinale (ginger) treat clinical extensively drug resistant Acinetobacter baumannii synergistically by antioxidation properties (II) Constituents from Cinnamomum kotoense as human Lon protease inhibitors and antioxidants

陳璽安(Hsi-An Chen)

指導教授 ： 王惠民

高雄醫學大學/藥學院/香粧品學系碩士班/碩士(2009年)

摘要

(I)
病菌對抗生素的抗藥性是目前在臨床治療上主要的問題。近年來，泛抗藥性之鮑式不動桿菌(extensively drug resistant Acinetobacter baumanii, XDRAB)已造成全球性的院內感染，尤其又以加護病房更趨嚴重。相較於抗藥性菌株的增加及蔓延，新種類抗生素之發展，已不足以應付此狀況。在無適當抗生素的情況下，另闢新的治療形式是急迫需要的。且如何針對此感染疾病而發展強而有力的天然藥物治療方式為最重要的課題。近年來自文獻中得知，從薑 (Zingiber officinale) 的根部萃取之純化合物已被發現對人體具有多種不同的功效，例如，抗腫瘤生長及抗發炎反應。但是在抗菌臨床研究上並無多做著墨，故本研究之目標著重於從薑裡面找尋對泛抗藥性菌株有作用之新成分。此外這些化合物與tetracycline結合使用，對XDRAB有顯著的協同作用。這些薑的活性分子運用在抗菌特性上，對於發展治療XDRAB感染之新藥有很大的幫助。另一方面，薑的抗氧化測定，採用清除DPPH自由基的方式，這些純化合物亦有很好的抗氧化作用。本研究用MnO2當作氧化劑與薑結合使用於抗菌能力之測定，發現其效果明顯下降，因此推測薑的抗菌能力與其抗氧化作用為相輔相成。根據此研究成果對於新種抗生素之開發有著重要的影響。
(II)
Lon是一個需要ATP活化的serine蛋白酶並且在其活性位上攜帶了一組成對的Ser-Lys催化二元體(catalytic dyad)。從文獻資料中，我們得知：腫瘤的產生與人類粒線體中Lon過度表現有關，當腫瘤產生時，Lon亦會增加。又另一方面，如果Lon的活性下降，則會使細胞繁殖能力減低且造成細胞凋亡。利用此特性可當作在癌症藥物之化學治療上的一個標的。如此一來，對人類Lon有特異性的抑制劑不僅僅提供遺傳工程上的價值，且其指標性的化合物，在發展更強而有力的抗癌藥物上也扮演著不可或缺的重要角色。而不幸地，截至目前為止，針對人類Lon蛋白酶的特異性抑制劑卻少之又少，故在此類抑制劑的開發及找尋是目前非常重要的課題之一。在此篇研究中，我們已經確立了兩個新的人類Lon蛋白酶抑制劑，分別是從蘭嶼肉桂(Cinnamomum kotoense)分離出來的obtusilactone A和(-)-sesamin。實驗數據顯示obtusilactone A和(-)-sesamin對人類Lon蛋白酶有顯著性的抑制效果。且利用分子模擬的技術，得知在Lon蛋白酶Ser855和Lys898活性位上，obtusilactone A和(-)-sesamin與其有交互作用。此外，本實驗亦有對obtusilactone A和(-)-sesamin的抗氧化能力及自由基清除率作測定。結果顯示obtusilactone A有良好的1,1-diphenyl-2-picrylhydrazyl (DPPH)自由基清除能力及還原力。而(-)-sesamin則發現有良好的還原力及金屬離子螯合力。經由上述結果得知，蘭嶼肉桂中的這些成分是天然的抗氧化劑且對於癌症的預防是有發展潛力的。同時這些成分對於人類Lon的抑制劑在癌症化療上，相信將會扮演著領導的角色。

關鍵字

(I)薑 ； 鮑式不動桿菌 ； 抗氧化劑 ； 抗菌 ； 2-methoxyphenols (II)Lon蛋白酶抑制劑 ； obtusilactone A ； (-)-sesamin ； 活性氧，抗氧化劑

並列摘要

(I)
Bacterial resistance to antibiotics is a major therapeutic problem in decades. The extensively drug resistant Acinetobacter baumanii (XDRAB) causes serious nosocomial infections worldwide, particularly at intensive care units (ICUs). Several pure compounds from the rhizomes of Zingiber officinale have been shown the various effects on human such as anti-tumor and anti-inflammation. However, their antimicrobial activities have not been reported on clinical bacterial isolates. The aim of this study was to find the components from ginger with novel anti-extensively drug resistant microbial effects. Furthermore, these compounds combined with tetracycline showed good synergic inhibition effects to XDRAB. This antimicrobial properties of ginger rendered active molecules used in the development of novel compounds for the treatment of extensively drug resistant bacterial infections.
Moreover, four ginger compounds demonstrated antioxidant properties. The antioxidant capacity was evaluated by using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay. On the other hand, the antioxidants were neutralized by MnO2, which act as an oxidizing agent. We found if the antioxidant effects were inhibited by oxidants, not only antioxidant capacity but also antimicrobial properties were decreased, either. According to the results, we considered that ginger compounds with antioxidant effects were correlated to antimicrobial bio-functions. To our knowledge, this is the first time that these bioactivity evidences of ginger were presented to date.
(II)
Lon is an ATP-dependent protease and carries a Ser-Lys catalytic dyad in the active site. Overexpression and increased human mitochondrial Lon protease activity are related to tumorigenesis. On the other hand, loss of Lon activity leads to a reduced cell proliferation activity and apoptosis. The up-regulation of Lon protease in tumorigenesis has raised its potential as a drug target in the development of cancer chemotherapy. Meanwhile, the specific human Lon inhibitors provide not only valuable chemical genetic tools for identifying the substrates but also lead compounds for the development of a powerful anti-cancer drug. Unfortunately, very limited specific inhibitors of human Lon have been found. So we have identified two novel specific inhibitors of human Lon protease, obtusilactone A and (-)-sesamin from the Cinnamomum kotoense. The results showed that the inhibition effects of obtusilactone A and (-)-sesamin on human Lon are significant and the constituents interact with Ser855 and Lys898 residues in the active site by using a model of molecular docking technique. Furthermore, the antioxidant activities and free radical scavenging capacity of obtusilactone A and (-)-sesamin were also investigated in this study. Obtusilactone A showed effective 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and reducing power compared with Vitamin C and BHA, respectively. (-)-Sesamin was found to be effective in reducing power and metal chelating power activities. These results suggest that the constituents from C. kotoense act as natural antioxidants and play a potential role in cancer prevention. Meanwhile, these pure compounds may used as lead compounds in the development of specific human Lon inhibitors for cancer chemotherapy in future research.

並列關鍵字

(I)Zingiber officinale ； Acinetobacter baumannii ； antioxidant ； antimicrobial ； 2-methoxyphenols (II)Lon protease ； inhibitors ； obtusilactone A ； (-)-sesamin ； ROS ； antioxidant

參考文獻

(I)

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