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4. Discussion The global antifungal market was estimated at $9.4 billion in 2010 and is expected to grow at a rate of 1.9% during 2010-2017. The major class of antifungal compound includes azoles, polyenes, pyrimidines, allyamines and echinodians. The antifungal therapies intend eradicating fungi through several action mechanisms, mainly involving destruction of cell wall and inhibition of cell division. There is a significant unmet need of novel anti- fungal compounds, because the present portfolio of treatments interact unfavorably with other medication, have resistance problem, low spectrum of activity, are fungistatic as opposed to fungicidal and are often toxic. Hence research has been focused on development of potent and safe antifungal compounds, either synthetic or from microbial sources [24]. In the present study, we exploited the marine resources for isolation of actinomycetes strains producing antifungal compounds. In the course of study, the marine invertebrate associated active acti- nomycetes strain PM0525875 was isolated and identified as Actinoalloteichus cyanogriseus. The antifungal principle from this strain was characterized as Caerulomycin A that showed potent in-vitro activity against pathogenic and drug resistant Candida strains. Caerulomycin A was first isolated from S. caeruleus [25] [26] as an antibiotic. Later it was reported to have anti-cancer activity [27]. In-vitro and in-vivo activity of Caerulomycin A against Entamoeba histolytica has been reported [28]. Natural derivatives of Caerulomycin A has also been reported [29]. Caerulomycin B and C, a new 2,2′-dipyridyl derivatives were isolated from Streptomyces caeruleus supplemented with 1 mM L-tryptophan. This group also reported the isolation of Caerulomycin D [19]. Additional natural derivatives of Caerulomycin F-K were isolated from marine derived actinomycetes (Actinoalloteichus cyanogriseus) that showed anticancer and antibacterial activity [29]. Lin et al. identified the biosynthetic gene cluster for Caerulomycin A from ma- rine actinomycetes Actinoalloteichus cyanogriseus. It mainly consists of unusual hybrid polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) [30]. The biosynthesis of Caerulomycin A based on enzy- matic modifications leading to structural changes, have been studied [31]. Chemical synthesis of Caerulomycin A-C has also been reported [32] [33]. The immunosuppressive activity of Caerulomycin A and its natural ana- logues has been reported [34]. Although Caerulomycin A has not been entered in clinic so far, may be due to its limitations, it could be the ideal molecule for generating semi synthetic druggable derivatives having potential application as an anticancer, antifungal or immunosuppressive drug. The strain and the optimized fermentation process described here could be useful for large scale production of this molecule for further drug development studies. 5. Conclusion In our in-house screening program in the quest of novel and superior antifungal compounds, an actinomycetes strain PM0525875 was isolated from a marine invertebrate. The active principle was characterized to be Caeru- lomycin A. Minimum inhibitory concentration (MIC) of the compound was found in the range of 0.39 - 1.56 μg/ml against pathogenic fungal test strains. The phylogenetic analysis of producer strain using 16S rRNA se- quence showed closest match with Actinoalloateichus cyanogriseus. Overall, Caerulomycin A was isolated from marine invertebrate-associated Actinoalloteichus sp. using optimized medium and fermentation conditions. |
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4 討論 2010年全球抗真菌市場(chǎng)估計(jì)值為94億美元，且在2010至2017年間有望以1.9%的增長(zhǎng)率持續(xù)增長(zhǎng)。 抗菌劑的主要成分含唑類(lèi)、多烯類(lèi)、嘧啶類(lèi)、以及烯胺類(lèi) �？拐婢煼ㄔ噲D通過(guò)多種作用機(jī)制—主要是破壞細(xì)胞壁及抑制細(xì)胞的分解—來(lái)根除真菌。新型抗真菌化合物的需求量遠(yuǎn)未得到滿足，其原因是現(xiàn)有的組合療法內(nèi)部機(jī)制間存在相互抵制，且作用范圍小，其與抗菌劑的作用相悖，甚至?xí)箍咕鷦┦А?br /> 因而有研究致力于開(kāi)發(fā)高效、安全的抗真菌化合物，包括合成及從細(xì)菌中提取[24]。在本研究中，我們開(kāi)發(fā)利用了細(xì)菌源進(jìn)行放線菌株的隔離，來(lái)制取抗真菌化合物。我們?cè)谠囼?yàn)中將海洋無(wú)脊椎動(dòng)物的相關(guān)抗菌性菌株進(jìn)行了隔離，并確定其為海洋來(lái)源放線菌。該菌株的抗菌性原理類(lèi)似于淺藍(lán)霉素 A，其對(duì)致病性和耐藥性念珠菌菌株表現(xiàn)出很強(qiáng)的體外活性。 淺藍(lán)霉素 A是一種首次由S. caeruleus [25] [26]分離的抗生素。之后有報(bào)道稱其有抗癌作用[27]。淺藍(lán)霉素 A的體內(nèi)抗痢疾變形蟲(chóng)作用已經(jīng)報(bào)道[28]。還有報(bào)道指出淺藍(lán)霉素 A的人工合成方法[29]。 淺藍(lán)霉素B與C是由青藍(lán)鏈霉菌補(bǔ)充以1mM左旋色氨酸分離而得的一種新型聯(lián)吡啶衍生物。該課題組還報(bào)道了淺藍(lán)霉素D 的分離提取[19]。其他自然衍生物淺藍(lán)霉素F-K均分離自海洋來(lái)源放線菌(稱為Actinoalloteichus cyanogriseus)，具有抗癌性及抗菌性[29]。 Lin等鑒定了分離自海洋放線菌Actinoalloteichus cyanogriseus的淺藍(lán)霉素A的生物基因組。結(jié)果發(fā)現(xiàn)該淺藍(lán)霉素A主要含不常見(jiàn)的雜聚酮合酶（PKS）及無(wú)核糖體型肽合成酶（NRPS）[30]. 淺藍(lán)霉素A 的生物合成以對(duì)酶的修飾為基礎(chǔ)實(shí)現(xiàn)其結(jié)構(gòu)的改變，已有相關(guān)研究[31].淺藍(lán)霉素A-C的化學(xué)合成法已有報(bào)道[32][33]. 淺藍(lán)霉素A免疫抑制性及其自然界類(lèi)似物已有報(bào)道[34]。雖然淺藍(lán)霉素A至今還未用于臨床——可能由于其限制性——但其仍可作為制取半合成藥物衍生物的理想分子，將成為一種有應(yīng)用潛力的抗癌、殺菌或免疫抑制性的藥物。 5 結(jié)論 我們?yōu)榱双@得新型、優(yōu)化的抗真菌化合物設(shè)定了室內(nèi)篩選程序，利用該程序從一種海洋無(wú)脊椎動(dòng)物中分離出一段放線菌主鏈PM0525875。其活性成分為淺藍(lán)霉素A。該化合物經(jīng)對(duì)抗致病真菌的測(cè)試，得出其最低抑菌濃度為0.39-1.56μg/ml。利用16S rRNA對(duì)產(chǎn)生菌進(jìn)行系統(tǒng)分析，結(jié)果表明其與Actinoalloteichus cyanogriseus（海洋來(lái)源放線菌）有相關(guān)匹配性。綜上所述，本研究利用最優(yōu)方法及最佳發(fā)酵條件，從海洋無(wú)脊椎聯(lián)合放線菌淺藍(lán)霉素A中分離得到了淺藍(lán)霉素A。 |
2樓2014-10-22 10:52:43
bxzc1233樓2014-10-22 19:33:49
4樓2014-10-22 20:39:03
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