海洋放线菌分离及菌株Streptomyces sp. AH17-3的次级代谢产物研究

目的对海洋放线菌进行分离及抗肿瘤活性筛选,并对一株具有抗肿瘤活性的海洋放线菌AH17-3的次级代谢产物进行研究。方法采用溶剂萃取、柱色谱层析及制备HPLC等方法对菌株AH17-3的发酵产物进行化学分离,通过理化性质及波谱学方法并参阅文献进行化合物结构鉴定,以SRB法评价化合物的抗肿瘤活性。结果从海洋样品中分离放线菌174株,从菌株AH17-3中分离得到了4个聚酮类化合物,经鉴定其结构分别为germicidin A(1)、germicidin B(2)、daidzein(3)、genistein(4)。其中化合物1具有弱的细胞毒活性,其IC50为3.5×10-7 M。结论海洋放线菌是重要的药用微生物资源,化合物1,2均为首次从海洋放线菌中分离得到。     

海洋源放线菌Streptomyces sp.SCSIO 10428中吩嗪生物碱类抗生素的分离鉴定

目的对从广西北海斜阳岛海域沉积物中分离的1株海洋源放线菌Streptomyces sp.SCSIO 10428进行次级代谢产物及活性研究。方法对海洋源放线菌Streptomyces sp.SCSIO 10428的发酵产物进行有机溶剂萃取,利用硅胶、凝胶柱层析等手段纯化次级代谢产物,通过波谱数据分析及文献比较对化合物进行结构鉴定,对化合物进行了抗菌、卤虫致死以及抗氧化活性评价。结果从海洋源放线菌Streptomyces sp.SCSIO10428发酵产物中分离得到3个生物碱类化合物,其结构分别鉴定为1-甲氧基吩嗪(1),1-羟基吩嗪(2),吩嗪-1-羧酸(3);活性结果显示化合物1~3对白色念珠...     

冷泉来源诺卡氏菌OUCLQ19-35-1次级代谢产物的研究

目的 探究深海冷泉来源微生物的次级代谢产物产生能力,从中挖掘具有抗多重耐药(multi-drug resistant, MDR)菌活性的次级代谢产物,为新药研发提供化合物实体。方法 采用稀释涂布法分离纯化深海冷泉海泥样品中的放线菌,通过琼脂扩散法筛选具有抗MDR菌活性的放线菌;基于16S rRNA基因片段序列分析和系统进化树构建初步确定目标放线菌种属;对目标放线菌进行大规模发酵,采用有机溶剂萃取、反相硅胶柱层析、半制备高效液相等分离手段对发酵产物进行分离纯化,利用NMR、MS等波谱学技术并结合文献对化合物进行结构鉴定,然后对化合物进行抗MDR菌活性测试。结果 从深海冷泉中筛选到一株具有抗MDR菌Micrococcus luteus ML01和Staphylococcus aureus CCARM3090活性的放线菌OUCLQ19-35-1,16S rRNA序列及系统进化树分析初步确定其为Nocardiopsis synnemataformans;从其发酵产物中分离得到3个化合物,分别为questiomycin A(1)、1,6-dihydroxyphenazine(2)和5a,6,11a,12-tetrahydro-5a,11a-dimethyl[1,4]benzoxazino[3,2-b][1,4]benzoxazine(3);活性结果显示,化合物1-3均无抗MDR菌活性,但其所在的组分有抑菌活性。结论 从深海冷泉筛选得到一株诺卡氏菌OUCLQ19-35-1,其能够产生抗MDR菌的活性次级代谢产物,具有潜在的应用价值,但其活性成分待进一步的确定。     

海洋放线菌WBF16次级代谢产物的分离与结构鉴定

目的分离和鉴定海洋来源的放线菌WBF16代谢产物中的抗肿瘤活性成分。方法利用大孔树脂、Sephadex LH-20凝胶柱层析、硅胶柱层析、反相柱色谱和HPLC等方法对该放线菌发酵产物进行分离,根据理化性质和波谱学方法进行化学结构的鉴定;利用MTT法来检测化合物的抗肿瘤活性。结果从海洋放线菌代谢产物中分离得到其特征代谢产物色霉素A2(1)和2-甲基-5,6,7-三甲氧基-1,4-萘醌(2)。结论化合物2为新天然产物,化合物1为首次从海洋微生物中分离得到,同时化合物1对人口腔上皮癌KB细胞、人肺癌细胞株A549、人肝癌细胞株SMMC-7721的细胞毒活性较好,IC50值分别为3.81、7.21和12.58μg/L。     

海洋放线菌Rubrobacter radiotolerans的活性成分研究

目的：研究海洋共附生放线菌的化学成分。方法：采用中低压液相色谱、制备高效液相色谱法及重结晶等方法对放线菌提取物进行分离纯化,根据波谱数据结合理化性质解析所得化合物的结构,并对分离得到的单体化合物进行了细胞毒活性的筛选。结果：从放线菌 Rubrobacter radiotolerans 的醋酸乙酯萃取部分分离得到7个化合物,其结构分别鉴定为[3-[2-[(indol-3-yl)methyl]indol-3-yl]methylindole, 1]、[bis[2-[(indol-3-yl)methyl]     

海洋来源放线菌Streptomyces sp. SCSIO BEMM34的次级代谢产物研究

摘 要:目的 对一株来源于南海沉积物的放线菌Streptomyces sp. SCSIO BEMM34进行菌种鉴定及次级代谢产物研究。方法 通过16S rDNA序列分析并构建系统发育进化树来鉴定菌株,利用硅胶、凝胶层析,半制备高效液相分离等手段对海洋放线菌SCSIO BEMM34的发酵产物进行分离纯化,通过波谱数据分析及文献比对的方法对分离纯化得到的化合物进行结构鉴定,采用滤纸片法进行抗菌活性测试。结果 从一株海洋来源放线菌中分离得到2个戊二酰亚胺类化合物1和2,以及一个酚酸类化合物3。抗菌活性测试结果表明,化合物1-3都没有明显的抑菌活性。     

海洋源放线菌Streptomyces sp. SCSIO 01681次级代谢产物的研究

摘 要：目的 对从我国三亚鹿回头海水沉积物中分离得到的一株海洋放线菌SCSIO 01681进行鉴定并对其次级代谢产物进行研究。方法 通过16S rDNA序列分析并构建系统发育进化树来鉴定菌株,利用有机溶剂萃取、正相和反相硅胶层析等分离手段对海洋放线菌SCSIO 01681的发酵产物进行分离纯化,通过波谱数据分析及文献比较对化合物进行结构鉴定, 并进行了卤虫致死及抗菌活性评价。结果 该放线菌被鉴定为Streptomyces sp. SCSIO 01681,并从其发酵产物中分离得到3个化合物,其结构分别鉴定为苯乙酸（1）,亚油酸甘油酯（2）,邻苯二甲酸二（2-乙基己基）酯（3）;活性结果显示化合物1对嗜水气单胞菌、耐甲氧西林金黄色葡萄球菌、藤黄微球菌、粪链球菌具有抑制作用。结论 放线菌Streptomyces sp. SCSIO 01681能够产生3个医药、工业重要中间体,具有潜在的应用价值。     

海洋放线菌代谢产物蒽环类化合物研究进展

海洋放线菌代谢产物是抗肿瘤活性物质的重要来源。近年来,从海洋放线菌中分离到很多新化合物,其中许多结构新颖的蒽环类代谢产物具有良好的抗菌抗肿瘤活性。文章对近年来从海洋放线菌中分离得到的蒽环类代谢产物进行了归纳,并展望今后海洋天然产物的发展方向。     

一株南沙群岛柳珊瑚来源真菌 Aspergillus terreus 中化合物及 mPTPB 酶抑制活性研究

目的 从1株南沙群岛柳珊瑚来源真菌 Aspergillus terreus (NS02-09)中分离鉴定海洋天然产物,对所得化合物进行结核分枝杆菌酪氨酸磷酸激酶 (mPTPB) 抑制活性评价。方法 运用多种色谱手段分离纯化化合物,利用NMR、CD等现代波谱分析方法,对化合物进行结构鉴定、,通过衍生物制备获得两个乙酰化衍生物(2a和2b);并对化合物2及其衍生物2a和2b进行mPTPB酶抑制活性测试。结果 鉴定了1个土曲霉酮(1)和1个丁烯酸内酯 (2) 的结构; 2具有较强的mPTPB酶抑制活性,而其乙酰化产物(2a和2b)的mPTPB 酶抑制活性显著降低。运用Sybyl X 1.3 软件,对2与mPTPB酶的模拟对接计算发现,丁烯酸内酯环及环上的羟基是化合物2发挥酶抑制活性的重要作用基团。结论 从柳珊瑚来源真菌 A. terreus (NS02-09) 中发现了具有mPTPB 酶抑制活性的丁烯酸内酯类化合物,并对其作用机制进行了计算研究,该类化合物的相关研究对抗结核药物先导化合物发现具有借鉴作用。     

海洋放线菌次级代谢产物中大环内酯化合物的研究进展

目的综述海洋放线菌次级代谢产物中大环内酯化合物的最新研究进展,为进一步开展海洋放线菌次级代谢产物中大环内酯化合物的研究打下前期基础。方法查阅文献,进行整理、分析和归纳。结果与结论从海洋放线菌中分离得到很多结构新颖,活性多样的大环内酯类次级代谢产物,与陆栖放线菌一样,很多具有潜在的药用价值,表明海洋放线菌次级代谢产物中的大环内酯化合物具有良好的研究价值和开发前景。     

海洋真菌Y26-02的灰黄霉素类化学成分

目的研究海洋真菌Y26-02发酵液的乙酸乙酯萃取部分,对其中的化学成分进行分离鉴定。方法采用硅胶、Sephadex LH-20柱色谱以及制备型高效液相色谱等手段进行分离纯化,通过理化性质和各种波谱数据对这些化合物进行结构鉴定。结果从海洋真菌Y26-02菌液的乙酸乙酯萃取部分分离得到7个化合物,分别鉴定为灰黄霉素(griseofulvin,1)、表灰黄霉素(epigriseofulvin,2)、异灰黄霉素(isogriseofulvin,3)、脱氯表灰黄霉素(dechloroepigriseofulvin,4)、脱氢灰黄霉素(de-hydrogriseofulvin,5)、4′-丁氧基异灰黄霉素(4′-butoxyisogriseofulvin,6)、2′-丁氧基表灰黄霉素(2′-butoxyepigriseofulvin,7)。结论这7个化合物均为首次从该真菌的代谢产物中分离得到,其中化合物6、7是新天然产物。     

The influence of P2Y12 receptor deficiency on the platelet inhibitory activities of prasugrel in a mouse model: evidence for specific inhibition of P2Y12 receptors by prasugrel

Prasugrel is a novel orally active thienopyridine with faster, higher and more reliable inhibition of platelet aggregation than clopidogrel reflecting its metabolism in vivo to an active metabolite with selective P2Y(12) antagonistic activity. Several lines of evidence support the contention that prasugrel provides selective P2Y(12) receptor antagonistic activity. To date, however, direct evidence of P2Y(12) specific action by prasugrel in vivo is limited. In the present study, effects of prasugrel on ex vivo platelet aggregation were examined in wild type (WT) and P2Y(12)(-/-) mice. In WT mice, prasugrel showed platelet inhibition that was 8.2 times more potent than clopidogrel. In P2Y(12)(-/-) mice, ADP induced platelet aggregation was minimal, and its extent was similar to that in prasugrel-treated WT mice. In addition, no further inhibition of platelet aggregation was observed after administration of prasugrel to P2Y(12)(-/-) mice. Furthermore, prasugrel-treated WT mice showed similar aggregation patterns using collagen- and murine PAR-4 agonist peptide to those of P2Y(12)(-/-) mice treated with vehicle or prasugrel. Overall, these results clearly provide additional in vivo evidence that prasugrel has selective P2Y(12) antagonistic activity.     

Identification of a potent inverse agonist at a constitutively active mutant of human P2Y12 receptor

Human platelets express two P2Y receptors: G(q)-coupled P2Y(1), and G(i)-coupled P2Y(12). Both P2Y(1) and P2Y(12) are ADP receptors on human platelets and are essential for ADP-induced platelet aggregation that plays pivotal roles in thrombosis and hemostasis. Numerous constitutively active G protein-coupled receptors have been described in natural or recombinant systems, but in the P2Y receptors, to date, no constitutive activity has been reported. In our effort to identify G protein coupling domains of the human platelet ADP receptor, we constructed a chimeric hemagglutinin-tagged human P2Y(12) receptor with its C terminus replaced by the corresponding part of human P2Y(1) receptor and stably expressed it in Chinese hamster ovary-K1 cells. It is interesting that the chimeric P2Y(12) mutant exhibited a high level of constitutive activity, as evidenced by decreased cAMP levels in the absence of agonists. The constitutive activation of the chimeric P2Y(12) mutant was dramatically inhibited by pertussis toxin, a G(i) inhibitor. The constitutively active P2Y(12) mutant retained normal responses to 2-methylthio-ADP, with an EC(50) of 0.15 +/- 0.04 nM. The constitutively active P2Y(12) mutant caused Akt phosphorylation that was abolished by the addition of pertussis toxin. Pharmacological evaluation of several P2Y(12) antagonists revealed (E)-N-[1-[7-(hexylamino)-5-(propylthio)-3H-1,2,3-triazolo-[4,5-d]-pyrimidin-3-yl]-1,5,6-trideoxy-beta-d-ribo-hept-5-enofuranuronoyl]-l-aspartic acid (AR-C78511) as a potent P2Y(12) inverse agonist and 5'-adenylic acid, N-[2-(methylthio)ethyl]-2-[(3,3,3-trifluoropropyl)thio]-, monoanhydride with (dichloromethylene)bis[phosphonic acid] (AR-C69931MX) as a neutral antagonist. In conclusion, this is the first report of a cell line stably expressing a constitutively active mutant of human platelet P2Y(12) receptor and the identification of potent inverse agonist.     

Studies on anti-tumor activity of crude drugs. II. The effects of aqueous extracts of some crude drugs in shortterm screening test. (2)

The selective toxicity was often shown by experiments with in vitro screening test in human cancer cell (JTC-26) and human normal embryonic cell (HE-1) on the crude extract of crude drugs. The active fractions with selective toxicity were obtained by column chromatography. However, the selective toxicity disappeared in the simple constituent in which the activity remained. The selective toxicity of crude drugs was presumed to appear with the complex constituents.     

西双版纳粗榧内生真菌菌株CM112中分离到的两个新bisabolane型倍半萜

目的 从西双版纳粗榧内生真菌菌株Aspergillus sp. CM112中分离和鉴定抗菌活性产物。方法 采用反相硅胶色谱、凝胶色谱和HPLC等分离技术,对菌株CM112的发酵产物进行分离纯化;通过1D、2D NMR、HR-ESI-MS以及旋光数据进行结构测定。结果 鉴定了5个bisabolane型倍半萜的结构,包括两个新化合物(7S,11S)-(+)-11-hydroxyl-sydonol(1)和(7S,11S)-(+)-11-hydroxyl aspergiterpenoid A(2)以及3个已知化合物(7S,11S)-(+)-11-hydroxysydonic acid(3), (7S,11S)-(+)-11,12-dihydroxysydonic acid (4)和(7S,11S)-(+)-12-hydroxysydonic acid(5);并通过纸片扩散法测试了它们对3株病原细菌的抗菌活性。结论 化合物1和2是新的bisabolane型倍半萜,2和3为抗菌活性成分。     

Centella asiatica accelerates nerve regeneration upon oral administration and contains multiple active fractions increasing neurite elongation in-vitro

Axonal regeneration is important for functional recovery following nerve damage. Centella asiatica Urban herb, also known as Hydrocotyle asiatica L., has been used in Ayurvedic medicine for centuries as a nerve tonic. Here, we show that Centella asiatica ethanolic extract (100 microg mL-1) elicits a marked increase in neurite outgrowth in human SH-SY5Y cells in the presence of nerve growth factor (NGF). However, a water extract of Centella was ineffective at 100 microg mL-1. Sub-fractions of Centella ethanolic extract, obtained through silica-gel chromatography, were tested (100 microg mL-1) for neurite elongation in the presence of NGF. Greatest activity was found with a non-polar fraction (GKF4). Relatively polar fractions (GKF10 to GKF13) also showed activity, albeit less than GKF4. Thus, Centella contains more than one active component. Asiatic acid (AA), a triterpenoid compound found in Centella ethanolic extract and GKF4, showed marked activity at 1 microM (microg mL-1). AA was not present in GKF10 to GKF13, further indicating that other active components must be present. Neurite elongation by AA was completely blocked by the extracellular-signal-regulated kinase (ERK) pathway inhibitor PD 098059 (10 microM). Male Sprague-Dawley rats given Centella ethanolic extract in their drinking water (300-330 mg kg-1 daily) demonstrated more rapid functional recovery and increased axonal regeneration (larger calibre axons and greater numbers of myelinated axons) compared with controls, indicating that the axons grew at a faster rate. Taken together, our findings indicate that components in Centella ethanolic extract may be useful for accelerating repair of damaged neurons.     

Anti-inflammatory and anti-melanogenic steroidal saponin glycosides from Fenugreek (Trigonella foenum-graecum L.) seeds

Fenugreek seed ( Trigonella foenum-graecum L.) is used as an herbal medicine for treating metabolic and nutritive dysfunctions. To determine if this plant has other beneficial effects, we tested the inhibitory activities of a methanol (MeOH) extract of fenugreek seed on the production of inflammatory cytokines and melanin synthesis in cultured cell lines in vitro. The MeOH extract inhibited the production of phorbol-12-myristate-13-acetate-induced inflammatory cytokines such as tumor necrosis factor (TNF)-α in cultured THP-1 cells, and also restrained the intracellular synthesis of melanin in murine melanoma B16F1 cells. We isolated three active constituents from fenugreek seed extracts. These were identified as the steroidal saponins 26- O-β-D-glucopyranosyl-(25 R)-furost-5(6)-en-3 β,22 β,26-triol-3- O-α-L-rhamno-pyranosyl-(1' → 2')-O-[β-D-glucopyranosyl-(1' → 6')- O]-β-D-glucopyranoside 1, minutoside B 2, and pseudoprotodioscin 3. Compounds 1 and 2 strongly suppressed the production of inflammatory cytokines, whereas 3 showed a weaker suppressing effect. Melanogenesis in B16F1 cells was significantly suppressed by 1 and 3, and weakly suppressed by 2. All three compounds showed moderate cytotoxicities. These results indicate that fenugreek extract and its active constituents could protect against skin damage.     

高效液相色谱法测定灯盏细辛提取物中两种有效成分的含量

目的：探讨灯盏细辛提取物中野黄芩苷、3,4-二-O-咖啡酰奎宁酸2种有效成分含量的高效液相色谱测定方法。方法：色谱柱选择Dikma Kromasil C18（250mm×4.6mm,5.0μm）,流动相选择乙腈：0.2%磷酸溶液=30∶70,检测波长为332nm,柱温为35℃。结果：野黄芩苷的线性回归方程为Y=1.554×103-0.201×102（r=0.9997）;3,4-二-O-咖啡酰奎宁酸的线性回归方程为Y=1.724×103-0.081×102（r=0.9996）;野黄芩苷在0.0341～1.3121μg范围内,3,4-二-O-咖啡酰奎宁酸在0.0592～2.3426μg范围内呈良好的线性关系。结论：采用高效液相色谱法同时测定灯盏细辛提取物中2种有效成分的含量,操作简单,精密度高,重现性好,可以作为灯盏细辛提取物中这2种有效成分的定量分析方法。     

钩吻生物碱的提取分离及结构鉴定

中草药钩吻中具有抗癌活性生物碱单体成分的提取分离,并测定其结构。方法：药材粉碎后用3%NaOH碱化后,晾干,用氯仿浸泡,回收氯仿提取液得浸膏。加酸溶解后,用氯仿萃取,碱化水层,再用氯仿萃取,得到钩吻总生物碱。然后经硅胶柱层析、反相ODS柱层析以及制备型HPLC层析分离纯化,并经理化性质和光谱学数据的测定鉴定单体化合物的结构。结果：共分离得到12个单体化合物,经测定紫外光谱、1H-NMR数据并与文献报道相比较,初步确定两个化合物的结构分别为gelsemine和koumine。结论：通过硅胶柱层析、ODS柱层析和HPLC相结合的方法可以达到分离钩吻总生物碱的目的。     

The effect of ginger on serotonin induced hypothermia and diarrhea

One of the important medicinal properties of ginger is known to remove chills caused by common cold and to warm body. In the present study, acetone extract of ginger at 100 mg/kg p.o. significantly inhibited serotonin (5-HT) induced hypothermia. Therefore, the active constituents of ginger were further examined. The acetone extract was functioned into 4 fractions by column chromatography. Fractions 1 and 2 showed significant activity. Fraction 2 was further purified and [6]-shogaol which was obtained from fraction 2-2, at 10 mg/kg p.o. was shown to inhibit 5-HT induced hypothermia. Anticathartic activity is known to be one of the medicinal effects of ginger. In the present study, acetone extract of ginger at 75 mg/kg p.o., significantly inhibited 5-HT induced diarrhea. In order to clarify the active constituents, the acetone extract was fractionated into 4 fractions by silica gel chromatography. Fractions 2 and 3, which was especially effective, were further purified and [6]-shogoal, [6]-dehydrogingerdione, [8]- and [10]-gingerol were found to have an anticathartic action. [6]-Shogaol was more potent than [6]-dehydrogingerdione, [8]- and [10]-gingerol.