赤水丹霞土壤链霉菌CSDX 001来源的新苯邻二甲酰亚胺的分离及结构鉴定

利用改良的GYM培养基对从赤水丹霞山土壤样品中分离的链霉菌分离菌株Streptomyces sp.CSDX 001进行固体发酵,发酵产物利用乙酸乙酯萃取,减压浓缩、富集获得其粗提取物。利用反复硅胶柱层析对粗提物中的次级代谢产物进行分离纯化,通过质谱、核磁共振等波谱方法对分离产物进行结构鉴定。从链霉菌Streptomyces sp.CSDX 001中分离得到2个化合物,分别为苯邻二甲酰亚胺4-羟基-N,7-二甲基苯邻酰亚胺(1)和2-甲基-2,5-莰二醇(2),其中化合物1为新化合物。     

白色链霉菌DSM 41398中洋橄榄菌素和放线吡喃酮的发现及其生物合成途径分析

【目的】从菌株Streptomyces albus DSM 41398的发酵产物中发掘结构多样的由I型聚酮合酶催化形成的化合物,以期找到具有新颖结构或强生物活性的化合物。在结构鉴定的基础上,对其生物合成途径进行分析。【方法】利用HPLC分析方法,通过系统比较野生型菌株S.albus DSM 41398与I型聚酮合酶编码基因簇失活突变株的发酵产物差异,实现目标化合物的定向分离。然后,利用~1H-和~(13)C-NMR以及HR-ESI-MS进行化合物的结构鉴定。最后,利用生物信息学等方法对化合物的生物合成途径进行推测和分析。【结果】从5 L的S.albus DSM 41398发酵产物中,分离得到了2个具有抗肿瘤活性的聚酮类化合物放线吡喃酮和洋橄榄菌素,分别定位了它们的生物合成基因簇,并分别对其生物合成途径进行了推导。其中,放线吡喃酮的生物合成基因簇为首次报道。【结论】本研究一方面为基因组发掘S.albus DSM 41398中其他由I型聚酮合酶催化形成的化合物提供参考,另一方面也为相关化合物的结构修饰改造奠定了良好的基础。     

广西北部湾放线菌的分离筛选及活性产物的鉴定

为从广西北部湾的泥样和植物中分离海洋放线菌,筛选具有抑菌活性的菌株,分离活性化合物。研究采用普通稀释法分离菌株,对发酵产物进行抑菌活性测试,利用活性追踪分离活性化合物,并通过波谱方法确定化合物结构。结果表明从6个海泥样品和3个植物样品中共分离73株放线菌,筛选得到具有抑香蕉枯萎病和金黄色葡萄球菌活性的菌株7株,并从其中的1株链霉菌 Streptomyces sp.MDCW-126的次级代谢产物中分离鉴定了星形孢菌素。从广西北部湾分离的药用活性菌株资源具有开发和深入研究价值。     

海洋真菌Aspergillus jensenii SS5中化学成分研究

利用多种柱色谱分离技术从海洋真菌Aspergillus jensenii SS5的液体发酵产物中分离获得4个已知化合物,并通过NMR、HR-ESI-MS、X单晶衍射等技术鉴定了它们的结构,分别是epigriseofulvin(1)、sterigmatocystin(2)、brevianamide M(3)、meleagrin(4),其中化合物1～3是首次从曲霉属真菌A.jensenii中分离获得。采用SRB法对分离获得的天然产物进行体外细胞毒性测试,结果表明化合物4对人肺癌A549细胞和人肝癌Bel-7402细胞均有较好抑制作用,化合物2对A549人肺癌细胞株有较好抑制作用。本研究作为海洋真菌A.jensenii SS5的化学成分的补充研究,发现了2个具有较好细胞毒活性的化合物,为曲霉属海洋真菌活性天然产物的开发提供了理论依据。     

石磺来源海洋链霉菌Streptomyces ardesiacus SCSIO LO23中germicidin类化合物的分离鉴定及其生物合成分析

海洋共附生放线菌具有生产结构特殊和活性显著化合物的潜力。【目的】以海洋软体动物石磺Onchidium sp.来源共附生海洋链霉菌Streptomyces ardesiacus SCSIO LO23为研究对象,分离并鉴定其次级代谢产物结构,分析化合物的生物合成基因簇及其生物合成途径。【方法】采用多种培养基对该菌株进行发酵优化并放大发酵,利用多种柱色谱方法分离得到germicidin类化合物,并利用波谱学手段和X-Ray单晶衍射技术完成化合物的结构鉴定;利用Illumina HiSeq对目标菌株进行全基因组测序,结合antiSMASH和BLAST在线分析软件实现菌株中germicidin类化合物生物合成基因功能注释和生物合成途径分析,并通过异源表达进一步确定其生物合成基因。【结果】从AM3培养基发酵产物中分离鉴定了6个吡喃酮类化合物germicidinA(1)、germicidinB(2)、germicidin D (3)、germicidin H (4)、isogermicidin A (5)和isogermicidin B (6),其中化合物2和6对斑马鱼神经行为有显著兴奋作用。通过对...     

浅紫灰链霉菌CQ01819及其次级代谢产物的定向发掘

【目的】采用特征次级代谢产物生物合成的保守功能基因探针,定向分离土壤中产生特征次级代谢产物的菌种资源,借助基因转录分析为导向的培养基优化方法,获得目标次生代谢产物。【方法】首先,根据5种特征次级代谢产物保守的合成功能基因设计简并引物,定向从土壤样品中筛选、分离并纯化菌株。然后,以RT-qPCR为指导开展目标产物的发酵培养基优化;最后,对菌株进行发酵,利用多种色谱技术分离纯化目标天然产物,并结合高分辨质谱与核磁共振等技术对所获得的化合物进行结构鉴定。【结果】从土壤中筛选得到了一株AHBA合酶基因和环氧化酶基因均为阳性的链霉菌菌株(编号为CQ01819),根据转录分析优化发酵培养基,最终从该菌株分离纯化得到了含有AHBA结构单元的丝裂霉素C、聚醚类抗生素莫能霉素A和缬吲霉素。【结论】本研究通过菌株的定向分离纯化,筛选得到了产生预期抗生素的浅紫灰链霉菌菌株CQ01819;基于RT-qPCR指导的发酵培养基优化,确定了菌株的发酵条件;获得发酵粗提物后,采用多种色谱整合技术和光谱分析策略,快速分离并鉴定了目标产物。该研究为目标菌种资源的定向筛选、菌株的发酵条件的快速优化和化合物的定向分离提供了较好的参考。     

南海珊瑚来源真菌Aspergillus sp.SCSIO 40435的分离鉴定及其次级代谢产物研究

【目的】南海珊瑚共附生真菌Aspergillus sp. SCSIO 40435次级代谢产物分离鉴定及抑菌活性筛选。【方法】利用稀释涂布平板法分离珊瑚共附生真菌。采用单菌多代谢产物方法(one strain many compounds,OSMAC)对分离菌株进行化学多样性筛选,并采用滤纸片扩散法对真菌发酵产物进行抑菌活性分析。通过ITS测序鉴定活性菌株SCSIO 40435的分类地位,运用多种色谱手段从其粗提物中分离纯化单体化合物,并利用各种波谱手段(HRESIMS、1D和2D NMR、单晶X-ray衍射法等)确定化合物的结构。最后,采用微量肉汤稀释法对单体化合物的抑菌活性进行评估。【结果】从南海珊瑚样品中分离得到19株共附生真菌,结合化学多样性和抑菌活性分析,筛选出1株产物丰富且具有多种抑菌活性的菌株SCSIO40435。利用ITS测序分析将其鉴定为曲霉属真菌(Aspergillus sp.),进一步从其发酵产物中分离鉴定了4个对三联苯类化合物：dicandidusin A(1)、candidusin A(2)、terphenyllin(3)和4″-deoxyterphenylli...     

红树林放线菌Stremptmeces costaricanus SCSIO ZS0073抗菌活性次级代谢产物的研究

从红沙公园的红树林泥土中分离得到一株生产较强抑菌活性次级代谢产物的放线菌Strepomyces costaricanus SCSIO ZS0073,采用滤纸片法进行抑菌活性追踪,并利用硅胶、凝胶柱层析和HPLC等分离手段对其发酵产物进行分离纯化,得到3个化合物,通过NMR、MS等波谱数据分析并参阅文献将3个化合物分别鉴定为放线菌素D、放线菌素X_(Oβ)和制霉色基素(fungichromin)。其中放线菌素D在发酵条件未优化的情况下产量就达到300 mg/L以上。     

一株红树林曲霉菌Aspergillus sp.WHUF0343的次级代谢产物研究

【目的】研究分离自三亚亚龙湾红树林根系土壤的真菌Aspergillus sp. WHUF0343的次级代谢产物及其生物活性。【方法】利用硅胶柱层析、Sephadex LH-20凝胶柱层析和半制备高效液相色谱等技术对该菌的发酵产物进行分离纯化;综合利用核磁共振波谱(NMR)和质谱(MS)等现代波谱技术以及与文献数据对照确定化合物的结构;分别采用肉汤微量稀释检测法和MTS法对化合物进行抗菌和肿瘤细胞毒活性测试。【结果】从真菌Aspergillus sp.WHUF0343的发酵产物共分离并鉴定了10个化合物,分别为isoechinulin A (1)、neoechinulin A (2)、neoechinulin E (3)、preechinulin (4)、neoechinulin D (5)、variecolorin J (6)、dehydroechinulin (7)、questinol (8)、emodin (9)和catenarin (10)。活性评价显示化合物2、9和10对幽门螺杆菌(Helicobacter pylori)和金黄色葡萄球菌(Staphylococcus aure...     

耐盐菊芋块茎内生真菌次生代谢产物的分离与结构鉴定

盐生植物菊芋(Helianthus tuberosus)是一种新型经济作物,特殊生境可能会使其内生真菌产生特殊的次生代谢产物,如结构新颖或活性显著的化合物.为研究这种耐盐植物内生真菌的次生代谢产物以及植物宿主和内生真菌之间可能存在的关系,本实验室从耐盐菊芋块茎中分离纯化出10株内生真菌,对这10株内生真菌进行抗卤虫活性筛选,初步筛选出1株抗卤虫活性良好的菌株Mucor sp.ht-7,规模发酵后通过提取分离技术得到7个化合物,用波谱手段鉴定了7个化合物的结构,对这株内生真菌和宿主次生代谢产物之间的关系进行分析探讨,并对化合物生物活性加以分析.     

Anti-plasmodial and antioxidant activities of constituents of the seed shells of Symphonia globulifera Linn f

A xanthone derivative, named gaboxanthone (1), has been isolated from the seed shells of Symphonia globulifera, together with known compounds, symphonin (2), globuliferin (3), guttiferone A (4), sistosterol, oleanolic acid and methyl citrate. The structure of the compound was assigned as 5,10-dihydroxy-8,9-dimethoxy-2,2-dimethyl-12-(3-methylbut-2-enyl) pyrano [3,2-b]xanthen-6(2H)-one, by means of spectroscopic analysis. The anti-plasmodial and antioxidant activities of the phenolic compounds were evaluated, respectively, in culture against W2 strain of Plasmodium falciparum and using the free radical scavenging activity of the DPPH radical, respectively. Compounds 1-4 were found to be active against the Plasmodium parasites (IC(50) of 3.53, 1.29, 3.86 and 3.17 microM, respectively). Guttiferone A (4) showed a potent free radical scavenging activity compared to the well-known antioxidant caffeic acid.     

DPPH free radical scavenger components from the fruits of Alpinia rafflesiana Wall. ex. Bak. (Zingiberaceae)

The methanol extract of the dried ripe fruits of Alpinia rafflesiana was investigated for its DPPH free radical scavenger constituents. 2',3',4',6'-Tetrahydroxychalcone (7), which has never been isolated from natural sources was found to be most active as a DPPH free radical scavenger with the IC50 value of 55 microM. Other known compounds isolated from this species include 5,6-dehydrokawain (1), flavokawin B (2). 1,7-diphenyl-5-hydroxy-6-hepten-3-one (3), (-)-pinocembrin (4), cardamonin (5) and (-)-pinostrobin (6). The DPPH free radical scavenger compounds were detected using TLC autographic analysis. The percentage inhibition of DPPH free radical scavenging activity was measured on isolates (5-7) using colorimetric analysis.     

Chemical constituents of the root bark of Erythrina droogmansiana

Two new compounds namely 7,4′-dihydroxy-2′-methoxy-3′-(3-methylbut-2-enyl)isoflavanone (1) and 5,7,3′-trihydroxy-8-(3-methylbut-2-enyl)-[6′′,6′′-dimethylpyrano(2′′,3′′:4′,5′)]isoflavone (4) were isolated from the root bark of Erythrina droogmansiana together with eight known compounds. Their structures were elucidated on the basis of spectroscopic analyses (1D- and 2D-NMR and ESI-HRMS) and by comparison with literature data. In addition, the correct ¹³C NMR of 5,7,2′,4′-tertahydroxy-8,5′-di-(3-metylbut-2-enyl)isoflavone (6) was assigned. The DPPH free radical scavenging properties of the isolated compounds were evaluated. Compounds 4–7 showed weak to moderate DPPH free radical scavenging activities.     

瓦宁木层孔菌中多酚和黄酮类成分分离及清除自由基活性的研究

采用硅胶和反相C18柱层析方法,首次从瓦宁木层孔菌中分离得到了5个化合物,运用NMR波谱法分析和鉴定为樱花亭、7-甲氧基二氢莰非素、二氢莰非素、4-(3,4-二羟苯基)-3-丁烯-2-酮、hispolon。并通过建立体外二苯基苦味酰基苯肼自由基（·DPPH）、超氧阴离子自由基（·O2?）以及羟自由基（·OH）发生体系,研究了5个化合物对·DPPH、·OH和·O2?的清除作用。结果表明当浓度达到100μg/mL时,化合物4-(3,4-二羟苯基)-3-丁烯-2-酮和hispolon对·DPPH清除率分别为92%和93%,对·OH的清除率分别为90%和95%,而对·O2?的清除率分别为70%和77%,略低于清除·DPPH和·OH的能力;二氢莰非素对·O2?自由基的清除率为39%,强于清除·OH和·DPPH的能力;而樱花亭和7-甲氧基二氢莰非素对3种自由基的清除率均低于30%。2个多酚类化合物清除自由基的能力均强于3个黄酮类化合物。5个化合物清除自由基能力均表现出一定的浓度依赖性。     

色钉菇子实体化学成分及其生物活性初探

从色钉菇Chroogomphus rutilus子实体中分离纯化出11种化合物,经核磁等方法鉴定为：（1）麦角甾-7,22-二烯-3-酮;（2）麦角甾-4,6,8(14), 22-四烯-3-酮;（3）邻苯二甲酸二丁酯;（4）顺式-3-己烯醇;（5）3β,5α,6β-三羟基麦角甾-7,22-二烯;（6）α-甜没药醇;（7）2-甲氧基腺嘌呤核苷;（8）(4E,8E)-2-N-(2-羟基棕榈酰)-1-O-B-D-吡喃葡萄糖基-9-甲基-4, 8-sphingadienine;（9）5-羟基尿嘧啶核苷;（10）2-氨基-3-醛基-6-甲氧基吡啶;（11）Polyozellin。对分离化合物的体外抗氧化、抗肿瘤活性的初步测定结果显示,2-甲氧基腺嘌呤核苷、顺式-3-己烯醇表现出较强的清除DPPH自由基活性,EC50值分别为0.06128mg/mL和0.08253mg/mL。5-羟基尿嘧啶核苷对人胃癌细胞株BGC的增殖有较强的抑制作用,IC50值为7.92μg/mL。研究结果为进一步开发利用色钉菇提供了科学依据。     

朱红栓菌提取物抗氧化、抗肿瘤活性评价及相关化学成分分析

采用石油醚、乙酸乙酯、乙醇浸提朱红栓菌 Trametes cinnabarina 子实体干粉,得到不同极性提取物;采用清除DPPH 自由基、羟自由基、超氧阴离子自由基能力,测定提取物的体外抗氧化活性;MTT法检测提取物对人肝癌细胞株HepG2细胞增殖的抑制作用。结果表明,朱红栓菌石油醚、乙酸乙酯、乙醇提取物均具有一定的抗氧化、抗肿瘤活性;各提取物在浓度为4-5mg/mL时,对DPPH自由基、羟自由基和超氧阴离子自由基清除能力大小依次为乙酸乙酯提取物>乙醇提取物>石油醚提取物;乙酸乙酯提取物对3种自由基的最高清除率分别为60.23%、74.49%、63.84%。各提取物对人肝癌细胞株HepG2细胞增殖抑制作用大小依次为乙酸乙酯提取物>乙醇提取物>石油醚提取物;乙酸乙酯提取物的抑制率最高达55.93%。采用硅胶和凝胶等柱色谱方法结合核磁、波谱和质谱等技术对乙酸乙酯提取物的化学组分进行分析,共分离纯化出11种化合物,分别鉴定为：麦角甾醇（1）,邻苯二甲酸二丁酯（2）,对羟基苯甲酸甲酯（3）,麦角甾-7,22,二烯-3-酮（4）,1-[(12E,16E)-12,16-二十碳二烯酰基]-2-[(E,E)-7,11-十八碳二烯酰基]-3-硬脂酰基甘油（5）,cinnabarin（6）,过氧麦角甾醇（7）,尿嘧啶（8）,甘露醇（9）,腺嘌呤核苷（10）,豆甾-7,22-二烯-3β,5α,6β-三醇（11）。除化合物6外均为首次从朱红栓菌子实体中分离得到。研究结果为开发利用朱红栓菌子实体提供了科学依据。     

Homoisoflavonoids from Ophiopogon japonicus Ker-Gawler

From the ethyl acetate extract of the tuberous roots of Ophiopogon japonicus (Liliaceae) eight known and five new homoisoflavonoidal compounds were isolated. The new compounds are 5,7-dihydroxy-8-methoxy-6-methyl-3-(2'-hydroxy-4'-methoxybenzyl)chroman-4-one (1), 7-hydroxy-5,8-dimethoxy-6-methyl-3-(2'-hydroxy-4'-methoxybenzyl)chroman-4-one (2), 5,7-dihydroxy-6,8-dimethyl-3-(4'-hydroxy-3'-methoxybenzyl)chroman-4-one (3), 2,5,7-trihydroxy-6,8-dimethyl-3-(3',4'-methylenedioxybenzyl)chroman-4-one (4) and 2,5,7-trihydroxy-6,8-dimethyl-3-(4'-methoxybenzyl)chroman-4-one (5). Their structures have been elucidated by mass and NMR spectroscopy. Compounds 4 and 5 are the first isolated homoisoflavonoids with a hemiacetal function at position 2.     

Free radical scavengers from Cymbopogon citratus (DC.) stapf plants cultivated in bioreactors by the temporary immersion (TIS) principle

The biomass production of Cymbopogon citratus shoots cultivated in bioreactors according to the temporary immersion (TIS) principle was assessed under different growth conditions. The effect of gassing with CO2-enriched air, reduced immersion frequency, vessel size and culture time on total phenolic and flavonoid content and free radical scavenging effect of the methanolic extracts was measured. From the TIS-culture of C. citratus, seven compounds were isolated and identified as caffeic acid (1), chlorogenic acid (2), neochlorogenic acid (3), p-hydroxybenzoic acid (4), p-hydroxybenzoic acid 3-O-beta-D-glucoside (5), glutamic acid (6) and luteolin 6-C-fucopyranoside (7). The occurrence of compounds 1-7 and their variability in C. citratus grown under different TIS conditions was determined by HPLC. The free radical scavenging effect of the methanolic extract and compounds was measured by the discoloration of the free radical 1,1-diphenyl-2-picrylhydrazyl (DPPH). The main metabolites in 6- and 8-week-old cultures, both in 5 and 10 1 vessels, were chlorogenic acid (2) (100-113 mg%) and neochlorogenic acid (3) (80-119 mg%), while in the cultures with CO2-enriched air and reduced immersion frequency the main compound detected in the extracts was glutamic acid (6) (400 and 670 mg% for the green and white biomass and 619 and 630 mg% for the green and white biomass, respectively). The most active compounds, as free radical scavengers, in the DPPH discoloration assay were caffeic acid (1), chlorogenic acid (2), neochlorogenic acid (3) and the flavonoid luteolin 6-C-fucopyranoside (7).     

Iridoids from the aerial parts of Verbena littoralis (Verbenaceae)

The iridoids, 6S-hydroxy-8S-methyl-4-methylene-hexahydro-cyclopenta[c]pyran-3-one and 6S,9S-dihydroxy-8S-methyl-4-methylene-hexahydro-cyclopenta[c]pyran-3-one, were isolated from the aerial parts of Verbena littoralis. Their structures and stereochemistry were elucidated by means of NMR spectral data analysis. Both compounds showed moderate in vitro activity against gram positive and negative bacteria as well as moderate in vivo intestinal peristaltic action in mouse. The iridoids also showed moderate free radical scavenging activity against l,l-diphenyl-2-picrylhydrazyl (DPPH) as well as antioxidant activity, the latter being evidenced by redox properties measured using E1CD-HPLC.     

Cytotoxic and antimicrobial metabolites from marine lignicolous fungi, Diaporthe sp

A new compound (1), named diaporthelactone, together with two known compounds (2 and 3) were isolated from the culture of Diaporthe sp., a marine fungus growing in the submerged rotten leaves of Kandelia candel in the mangrove nature conservation areas of Fugong, Fujian Province of China. The new compound was elucidated to be 1,3-dihydro-4-methoxy-7-methyl-3-oxo-5-isobenzofuran-carboxyaldehyde (1), which showed cytotoxic activity against KB and Raji cell lines (IC50 6.25 and 5.51 microg mL(-1), respectively). Two known compounds, 7-methoxy-4,6-dimethyl-3H-isobenzofuran-1-one (2) and mycoepoxydiene (3), were also demonstrated to exhibit cytotoxic activities for the first time. All three compounds were assessed for antimicrobial activity.