本文選題：冬凌草 + 內(nèi)生真菌��； 參考：《鄭州大學(xué)》2017年碩士論文

【摘要】：背景:從天然產(chǎn)物中篩選生物活性成分是開發(fā)新藥的有效途徑之一,但是從藥用植物中發(fā)現(xiàn)活性化合物的幾率一直在降低。而植物內(nèi)生真菌與宿主植物可能產(chǎn)生類似活性化合物,且種類繁多,具有巨大的開發(fā)潛力。因此,從可持續(xù)發(fā)展的層面上看,從植物內(nèi)生真菌的代謝產(chǎn)物中尋找活性化合物是開發(fā)新藥的一條有效的途徑。冬凌草系唇形科香茶菜屬多年生草本植物碎米椏(Rabdosia rubescens)的干燥地上部分,主要化學(xué)成分包括貝殼杉烷二萜類、黃酮類、揮發(fā)油及多糖類等,其中以冬凌草甲素為代表的二萜類化合物對(duì)食管癌、結(jié)腸癌、賁門癌均有一定療效。我們將河南道地藥材冬凌草作為研究對(duì)象,對(duì)其內(nèi)生真菌進(jìn)行分離純化得到球毛殼菌。球毛殼菌經(jīng)過發(fā)酵的次級(jí)代謝產(chǎn)物乙醇提取物有一定的抗癌活性。且文獻(xiàn)報(bào)道的從球毛殼菌次級(jí)代謝產(chǎn)物里分離出的化合物已達(dá)200多個(gè),具有不同程度的抗菌抗癌活性。因此我們選取冬凌草內(nèi)生真菌中的球毛殼菌進(jìn)行研究以期發(fā)現(xiàn)具有抗腫瘤活性的化學(xué)成分。紅樹林(Mangrove),查閱文獻(xiàn)報(bào)道表明,擴(kuò)展青霉菌(Penicillium expansum)次級(jí)代謝產(chǎn)物中所分離得到的倍半萜類化合物和苯酚三聚體類化合物,具有良好的逆轉(zhuǎn)多藥耐藥及細(xì)胞毒活性。因此我們選取擴(kuò)展青霉菌進(jìn)行發(fā)酵以期從次級(jí)代謝產(chǎn)物中發(fā)現(xiàn)具有抗腫瘤活性的化學(xué)成分。目的意義:本文分別對(duì)球毛殼菌(chaetomium globosum)和擴(kuò)展青霉菌(Penicillium expansum)的次級(jí)代謝產(chǎn)物進(jìn)行化學(xué)成分研究,以期得到具有較好抗癌活性和抗菌活性且結(jié)構(gòu)新穎的單體化合物,這對(duì)發(fā)現(xiàn)結(jié)構(gòu)新穎生物活性化合物以及對(duì)植物內(nèi)生真菌的研究具有一定意義。方法:本文分別采用固體和液體發(fā)酵法對(duì)球毛殼菌(chaetomium globosum)和擴(kuò)展青霉菌(Penicillium expansum)進(jìn)行發(fā)酵,發(fā)酵所得次級(jí)代謝產(chǎn)物用正相硅膠柱、MCI、大孔吸附樹脂、HPLC和Sephadex LH-20等方法進(jìn)行化學(xué)成分的分離和純化,通過IR,UV,1H-NMR,13C-NMR等現(xiàn)代技術(shù)對(duì)所分離得到的單體化合物進(jìn)行鑒定并確認(rèn)結(jié)構(gòu)。用MTT法對(duì)經(jīng)分離鑒定的化合物進(jìn)行抗癌活性測(cè)定。用紙片法、倍比稀釋法和瓊脂稀釋法進(jìn)行抗菌活性研究。結(jié)論:(1)從冬凌草(Rabdosia rubescens)內(nèi)分離出16種內(nèi)生真菌,其中1-8號(hào)菌屬于暗孢科,9號(hào)菌、10號(hào)菌屬于瘤座孢菌科,11號(hào)菌屬于炭棒科,12號(hào)菌屬于黑盤孢菌科,13號(hào)菌、14號(hào)菌屬于球殼孢菌科,15號(hào)菌屬于鮮殼孢菌科,16號(hào)菌屬于半殼孢菌科。針對(duì)PC-3(前列腺癌細(xì)胞)、MGC-803(胃癌細(xì)胞)對(duì)其發(fā)酵粗提物進(jìn)行抗癌活性測(cè)試,5號(hào)菌和9號(hào)菌的發(fā)酵粗體物對(duì)PC-3的IC50分別為302μg/ml,136μg/ml,11號(hào)菌和15號(hào)菌的發(fā)酵粗體物對(duì)MGC-803的IC50值分別為181μg/ml,123μg/ml。(2)從球毛殼菌(chaetomium globosum)的次級(jí)代謝產(chǎn)物中分離鑒定出10個(gè)單體化合物,分別鑒定為:stigmasta-4,6,8(14),22-tetraen-3-one(1)、4-methyl-5,6-dihydro-2H-pyran-2-one(2)、(E)-6-hydroaxy-4,6-dimethyl-3-heptene-2-one(3)、glyceroyl monopaimitate(4)、、4-羥基-2′-脫氧嘧啶核苷(5)、胞腺嘧啶脫氧核苷(6)、麥角甾醇過氧化物(7)、bungein A(8)、β-谷甾醇(9)、鄰苯二甲酸二丁酯(10)。(3)從擴(kuò)展青霉菌(Penicillium expansum)的次級(jí)代謝產(chǎn)物中分離得到10個(gè)化合物,過氧麥角甾醇(11)、環(huán)(甘氨酸-亮氨酸)(12)、anthraquinone aversin(13)、5-methoxysterigmatocystin(14)、6,8-di-O-methyl averufin(15)、環(huán)(甘-苯丙)二肽(16)、環(huán)(L-苯丙-反-4-羥基-L-脯)二肽(17)、Brevianamide K(18)Brevianamides Q(19)、brevianamides W(20)。(4)從擴(kuò)展青霉菌內(nèi)分離出來的單體化合物Brevianamide W對(duì)MGC-803的IC50值為43.4μg/ml,5-methoxysterigmatocystin對(duì)PC細(xì)胞的IC50值為56.7μg/ml。
[Abstract]:Background: screening bioactive ingredients from natural products is one of the most effective ways to develop new drugs, but the probability of discovery of active compounds from medicinal plants has been decreasing. Plant endophyte and host plants may produce similar active compounds with great potential for development. Looking for active compounds from the metabolites of endophytic fungi is an effective way to develop new drugs. Rabdosia rabdosia is the dry ground part of the perennial herb (Rabdosia rubescens) of the perennial herb of the genus lipido. The main chemical components include two terpenoids, flavonoids, volatile oils and polysaccharides, and so on. The two terpenoids, represented by oridonin, have a certain effect on esophageal, colon and cardia cancer. We have isolated and purified the endophytic fungi from Henan oridonin, which had a certain anticancer activity through the ethanol extracts of the secondary metabolites of the fermentation. And more than 200 compounds have been separated from the secondary metabolites of S. oridote, which have different degrees of antiseptic and anticancer activity. Therefore, we have selected the fungus in the endophytic fungi of oridonin to find out the chemical constituents of the antitumor activity. Mangrove. The sesquiterpene and phenol triad compounds isolated from the secondary metabolites of Penicillium expansum have a good reversal of multidrug resistance and cytotoxic activity. Therefore, we choose to ferment the Penicillium to discover antitumor chemical constituents from secondary metabolites. In this paper, the chemical composition of secondary metabolites of Chaetomium globosum and Penicillium expansum was studied in order to obtain the monomers with good anti-cancer activity and antibacterial activity with novel structure. The study of bacteria is of certain significance. Methods: in this paper, solid and liquid fermentation methods were used to ferment Chaetomium globosum and Penicillium expansum respectively. The secondary metabolites obtained by fermentation were separated by normal silica gel column, MCI, macroporous adsorption tree fat, HPLC and Sephadex LH-20 to separate chemical components. And purification, through the IR, UV, 1H-NMR, 13C-NMR and other modern technology to identify the isolated monomers and identify the structure. MTT method was used to determine the anticancer activity of the isolated compounds. The antibacterial activity was studied by paper method, double dilution method and agar dilution method. Conclusion: (1) from Rabdosia rubescens (Rabdosia rubescens) 16 endophytic fungi, of which No. 1-8 bacteria belong to the dark spores, 9 bacteria and 10 bacteria belong to the family cyclosporaceae, 11 bacteria belong to the charcoal stick family, the 12 bacteria belong to the spporaceae, 13 and 14 belong to the family sporospora, and the 15 bacteria belong to the fresh shell spores, and 16 bacteria belong to the hemicarporaceae. The antitumor activity of the fermented crude extracts was tested. The IC50 of PC-3 was 302 mu g/ml and 136 mu g/ml respectively. The IC50 value of MGC-803 was 181 mu g/ml, and 123 mu g/ml. (2) was separated from the secondary metabolites of S. ducrum (Chaetomium globosum) for 10 monomers. Compounds were identified as stigmasta-4,6,8 (14), 22-tetraen-3-one (1), 4-methyl-5,6-dihydro-2H-pyran-2-one (2), (E) -6-hydroaxy-4,6-dimethyl-3-heptene-2-one (3), glyceroyl monopaimitate (4), 4- hydroxyl -2 '- deoxy pyrimidine (5), cytosine pyrimidine deoxynucleoside (6), ergosterol peroxide (7), bungein A (8), and beta valerol (9). ), dibutyl phthalate (10). (3) separation of 10 compounds from secondary metabolites of Penicillium extended (Penicillium expansum), peroxy ergosterol (11), ring (Glycine leucine) (12), anthraquinone aversin (13), 5-methoxysterigmatocystin (14), 6,8-di-O-methyl averufin (15), ring (Gan - phenylpropyl) two peptide (16), ring (L- benzene). Two peptide (17), Brevianamide K (18) Brevianamides Q (19), brevianamides W (20), brevianamides W (20). (4) Brevianamide W of the monomer separated from the Penicillium extended to MGC-803 IC50 value is 43.4 micron.
【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R915

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