Chemical Constituents of Tussilago farfara L.

Two new sesquiterpenes, named tussilagonone (1) and neotussilagolactone (2), together with two known sesquiterpenes (3) and (4) and two phthalic acid derivatives (5) and (6), were isolated from the flower buds of Tussilago farfara . The structures of (1) and (2) were determined on the basis of spectroscopic analyses. Compounds (1) and (2) exhibited activities against platelet aggregation caused by platelet activating factor.

GC-MS Analysis of Essential Oil Constituents from Buds of Tussilago farfara L.

Aim To analyse the constituents of the essential oils extracted from the buds of Tussilago farfara L. in the GAP Bases of Traditional Chinese Medical Materials and provide scientific basis for quality control. Methods The essential oils were extracted by water-steam distillation and separated by GC capillary column chromatography. The components were quantitatively determined by normalization, and identified by GC-MS. Results GC-MS exhibited 259 peaks and 65 compounds were identified, accounting for 84.62% of the total essential oil. Conclusion In the total essential oil contained in the buds of Tussilago farfara L., copaene （2.36%）, （ ＋ ） -Epi-bicyclosesquiphellandrene （ 3.91% ）, γ- elemene （2.18%）, fl-bisabolene （ 13.93% ）, spathulenol （ 3.44% ） as the sesquiterpenes and its derivatives, and 1-undecene （4.83%）, （ E）-cycloundecene （8.49%）, bicycle [ 10. 1.0] tridec-l-ene （ 1. 45% ）, 1-tridecene （3.44%）, （Z）-7,11-dimethyl-3-methylene-1,6,10-dodecatriene （2.66%）, 1- pentadecene （4.57%）, [ 1R-（ 1R^＊, 4Z, 9S^＊ ） ]-4,11,11-trimethyl-8-methylene-bicyclo [ 7.2.0] undec-4-ene （ 1.03% ）, 6,6-dimethyl-2-methylene-7-（ 3-oxobutylidene ）-oxepan-3-ylmethyl acetic acid ester （2.02%）, 1, E-11, Z-13-heptadecatriene （ 3.72% ）, （ Z, Z, Z） -9,12,15-octadecatrien-l-ol （ 1.85% ）, 3,7,11-trimethyl-dodeca-2,4,6,10-tetraenal （ 1.31% ）, n-hexadecanoic acid （ 3.12% ） , （Z, Z） -9,12-octadecadienoic acid （2.26%）, （ Z, Z, Z） -9,12,15-octadecatrienoic acid methyl ester （ 1.12% ） , heneicosane （ 1.82% ）, and pentacosane （ 1.03% ） are the main components.

Chemical constituents of the flower buds of Tussilago farfara

Aim To study the chemical constituents of the flower buds of Tussilago farfara L. in the China National GAP Base of Traditional Chinese Materia Medica and provide scientific basis for quality control. Methods The constituents were separated and purified by different chromatographic methods, and their structures were elucidated by IR, MS and NMR techniques. Results Twenty eight compounds were isolated from the flower buds of T. farfara. Their structures were identified as n- heptacosane （1）, bis（2-ethylhexyl）phthalate （2）, 7β-[3＇-ethylcrotonoyloxy]-1α-[2＇-methylbutyryloxy]-3,14-dehydro-Z-notonipetranone （3）, 7β-[3＇-ethylcrotonoyloxy]-1α-[2＇-methylbutyryloxy]-3,14-dehydro-E-notonipetranone （4）, tussilagone （5）, dibutyl phthalate （6）, bauer-7-ene-3β,16α-diol （7）, isobauerenol （8）, stigmasterol （9）, β-sitosterol （10）, 2,2-dimethyl-6-acetylchromanone （11）, n- hexadecanoic acid （12）, 7β-hydroxysitosterol （13）, 7α-hydroxysitosterol （14）, 7,14-bisdesacylnotonipetrone （15）, 2,3- dihydroxypropylpalmitate （16）, daucosterol （17）, 6-hydroxy-2,6-dimethylhept-2-en-4-one （18）, ferulic acid （19）, isoferulic acid （20）, caffeic acid （21）, α-D-glucose （22）, sucrose （23）, phthalic acid （24）, p-hydroxybenzoic acid （25）, gallic acid （26）, uridine （27）, and adenosine （28）. Conclusion Compounds 1, 12-16, 18 and 20 were obtained from the genus Tussilago for the first time.

Application of Precision-Cut Rat Liver Slice to Study the Influence of Monocrotaline, Tussilago farfara Alkaloids on the Expression of Cytochrome P450 Enzymes

Precision-cut liver slice has been successfully used to study the mechanism of drug-induced hepatotoxicity, the prediction of liver toxicity, the discovery of early hepatic toxicity biomarker and the metabolism of drug in liver. We detected the expression of CYP3A4, CYP2B1 + CYP2B2 and CYP2E1 in precision-cut liver slice after co-cultured with monocrotaline or Tussilago farfara alkaloids to investigate the hepatotoxicity mechanism of those drugs. After co-culturing with monocrotaline or Tussilago farfara alkaloids for 6 hours, the expression of CYP3A4 in the microsome of precision-cut liver slices was detected by Western blot, and the expressions of CYP2B1 + CYP2B2 and CYP2E1 were detected by immunofluorescence. The results showed that monocrotaline induced the expression of CYP3A4 and CYP2B1 + CYP2B2, and Tussilago farfara alkaloids obviously up-regulated the expression of CYP2E1 and CYP3A4. Thus, we conclude that the up-regulation of CYP3A4, CYP2B1 + CYP2B2 and CYP2E1 may be one of the toxic mechanisms of liver injury of those drugs.

Effects of Flower Buds Extract of Tussilago farfara on Focal Cerebral Ischemia in Rats and Inflammatory Response in BV2 Microglia

Objective： To investigate the effects of the flower buds extract of Tussilago farfara Linné（Farfarae Flos; FF） on focal cerebral ischemia through regulation of inflammatory responses in activated microglia. Methods： Brain ischemia was induced in Sprague-Dawley rats by a transient middle cerebral artery occlusion（t MCAO） for 90 min and reperfusion for 24 h. Twenty rats were randomly divided into 4 groups（n=5 per group）： normal, t MCAO-induced ischemic control, t MCAO plus FF extract 300 mg/kg-treated, and t MCAO plus MK-801 1 mg/kg-treated as reference drug. FF extract（300 mg/kg, p.o.） or MK-801（1 mg/kg, i.p.） was administered after reperfusion. Brain infarction was measured by 2,3,5,-triphenyltetrazolium chloride staining. Neuronal damage was observed by haematoxylin eosin, Nissl staining and immunohistochemistry using anti-neuronal nuclei（Neu N）, anti-glial fibril ary acidic protein（GFAP）, and antiCD11 b/c（OX42） antibodies in ischemic brain. The expressions of inducible nitric oxide synthase（i NOS）, tumor necrosis factor（TNF-α）, and hypoxia-inducible factor-1 a（HIF-1α） were determined by Western blot. BV2 microglial cel s were treated with FF extract or its main bioactive compound, tussilagone with or without lipopolysaccharide（LPS）. Nitric oxide（NO） production was measured in culture medium by Griess assay. The expressions of i NOS, COX-2 and pro-inflammatory cytokines mRNA were analyzed by reverse transcription-polymerase chain reaction. The expression of i NOS, and COX-2 proteins, the phosphorylation of ERK1/2, JNK, and p38 MAPK and the nuclear expression of NF-κB p65 in BV2 cells were determined by Western blot. Results： FF extract significantly decreased brain infarctions in ischemic rats（P〈0.01）. The neuronal death and the microglia/astrocytes activation in ischemic brains were inhibited by FF extract. FF extract also suppressed i NOS, TNF-α, and HIF-1α expression in ischemic brains. FF extract（0.2 and 0.5 mg/m L, P〈0.01） and tussilagone 20 and 50 μmol/L, P〈0.01） significantly decreased LPS-induced NO production in BV2 microglia through downregulation of i NOS m RNA and protein expression. FF extract and tussilagone significantly inhibited LPS-induced expression of TNF-α, IL-1β, and IL-6 m RNA, and also suppressed the phosphorylation of ERK1/2, JNK and p38 MAPK and the nuclear expression of NF-κB in a dosedependent manner. Conclusion： FF extract has a neuroprotective effect in ischemic stroke by the decrease of brain infarction, and the inhibition of neuronal death and microglial activation-mediated inflammatory responses.

Polyhydroxylated eudesmane sesquiterpenoids and sesquiterpenoid glucoside from the flower buds of Tussilago farfara

Chemical fractionation of the n-BuOH partition,which was generated from the EtOH extract of the flower buds of Tussilago farfara,afforded a series of polar constituents including four new sesquiterpenoids(1-4),one new sesquiterpenoid glucoside(5)and one known analogue(6)of the eudesmane type,as well as five known quinic acid derivatives(7-11).Structures of the new compounds were unambiguously characterized by detailed spectroscopic analyses,with their absolute configurations being established by A-ray crystallography,electronic circular dichroism(ECD)calculation and induced ECD experiments.The inhibitory effect of all the isolates against LPS-induced NO production in murine RAW264.7 macrophages was evaluated,with isochlorogenic acid A(7)showing significant inhibitory activity.

款冬花多糖调控miR-889-3p/TLR4对IL-6诱导的乳腺癌细胞增殖、迁移及侵袭的影响

[目的]探讨款冬花多糖对白细胞介素-6(IL-6)诱导的乳腺癌细胞恶性行为作用机制。[方法] IL-6诱导人乳腺癌细胞MCF-7后,不同浓度的款冬花多糖处理。转染miR-NC、miR-889-3p mimics的MCF-7细胞经IL-6处理;转染NC-inhibitor、miR-889-3p-inhibitors的MCF-7细胞经款冬花多糖与IL-6处理;噻唑蓝(MTT)、平板克隆形成以及TranswellTM实验分析细胞增殖、迁移及侵袭;实时荧光定量聚合酶链反应(qRT-PCR)检测miR-889-3p、Toll样受体4(TLR4)的表达量;蛋白免疫印迹(Western blot)法检测基质金属蛋白酶(MMP)-2、MMP-9、TLR4蛋白表达量;双荧光素酶报告实验验证miR-889-3p与TLR4的靶向关系。[结果]款冬花多糖可降低IL-6诱导的MCF-7细胞增殖(P<0.05)、迁移及侵袭能力(P<0.05),而提高miR-889-3p的表达(P<0.05),呈剂量依赖性;TLR4是miR-889-3p的靶基因,miR-889-3p可靶向调控TLR4的表达;转染miR-889-3p mimics可降低IL-6诱导的MCF-7细胞增殖、迁移及侵袭(P<0.05);转染miR-889-3p-inhibitors可恢复款冬花多糖对IL-6诱导的MCF-7细胞增殖、迁移及侵袭。[结论]款冬花多糖可通过调节miR-889-3p/TLR4表达而抑制IL-6诱导的乳腺癌细胞恶性行为。

基于“法象思维”探颐药对“紫菀-款冬”治疗肺胀

法象思维是古人以效法取象的方式将物象维度上升至意象维度,来探讨分析药物功效的思维模式。中医药学的发展在“法象思维”的指导下应用广泛,源远流长,在临床遣方用药中起着不可替代的作用。紫菀、款冬作为临床常用经典药对且二者常相须为用,属于以“降”气为主的宣肺平喘的配伍。本文从“法象思维”的角度,对紫菀、款冬药对的药“象”作以分析,探寻在治疗肺胀病“象”用药的作用途径,以期为临床辨证论治、用药处方提供参考。

款冬花的化学成分及药理活性研究进展

款冬花含有萜类、黄酮、生物碱等化学成分,具有止咳、祛痰、平喘的作用,以及升压作用,抗血小板活化因子作用,抗炎作用等药理活性。作者阐述了款冬花的化学成分和药理活性研究的最新进展,并针对其毒性提出了安全评价问题,指出了款冬花研究存在的主要问题和今后研究的主要方向。

中药款冬花的化学成分研究

目的研究中药款冬花的化学成分。方法采用硅胶、Sephadex LH-20柱色谱等方法分离化合物,根据波谱数据和理化常数确定其结构。结果从款冬花中分离得到11个化合物,鉴定为:克氏千里光碱(senkirkine,1)、款冬酮(tussilagone,2)、款冬巴耳二醇(bauer-7-ene-3β,16α-diol,3)、sitoindosideΙ(4)、pedatisectineE(5)、β-carboline(6)、苯甲酰胺(benzamide,7)、腺嘌呤核苷(adenosine,8)、正二十七酸(heptacosanoic acid,9)、β-谷甾醇(β-sitoterol,10)和胡萝卜苷(daucosterol,11)。结论化合物1,4,5,6,7,9,10为首次从款冬中分离得到,其中4,5,6为首次从菊科植物中分离得到。化合物1和2对组胺和乙酰胆碱诱导的豚鼠离体气管条痉挛有抑制作用。

款冬花硒多糖的制备及抗氧化性研究

本文对款冬花多糖(TFPs)进行硒化修饰工艺优化研究,并初步探讨了其抗氧化活性。研究采用亚硒酸钠硒化法,制备款冬花硒化多糖(STFPs);以多糖中硒含量为指标,采用响应面法对制备工艺条件进行优化;并进行STFPs清除DPPH·、O-2·和OH·的实验,研究其体外抗氧化活性。得到最佳硒化条件为:硒化试剂质量比(m(亚硒酸钠)∶m(款冬花多糖))=1.06∶1,反应温度70℃,时间11h,硝酸体积分数0.5%。此时STFPs中的平均硒含量为3.84mg/g,与TFPs相比,清除DPPH自由基的能力显著提高,对O-2,OH·的清除能力也有一定程度的提高。

超声提取款冬花多糖的响应面法工艺优化

用响应面法考察了超声法提取款冬花多糖的最佳工艺。以多糖提取率为评价指标,考察了提取温度、超声时间、m(水)/m(款冬花)比值、提取次数对多糖提取率的影响。通过单因素实验和响应面法确定的最佳工艺条件为:超声时间36 min,提取温度68℃,m(水)/m(款冬花)=27,提取次数3次,在该条件下,多糖提取率为1.97%。超声法与传统工艺比较,提取时间缩短了71%。

款冬花多糖对荷瘤小鼠的抑瘤率及对白血病小鼠生存期的影响

目的研究款冬花多糖对荷瘤小鼠的抑瘤率和对白血病小鼠生存期的影响,并探讨其抗肿瘤机制。方法以小鼠肉瘤(S180)、小鼠肝癌(H22)瘤株及小鼠网状细胞白血病(L615)瘤株造模,设正常对照组、阳性药物组、款冬花高剂量组和款冬花低剂量组4组,考察款冬花多糖对各组小鼠抑瘤率和对白血病生存期的影响。结果款冬花多糖对S180、H22小鼠的抑瘤率分别为55.76%、45.61%;对L615白血病小鼠生命延长率>55.76%。结论款冬花多糖是抗肿瘤、抗白血病的有效药物,可干扰肿瘤细胞的有丝分裂过程和提高机体免疫力。

生品款冬花不同化学成分的镇咳祛痰作用

目的研究生品款冬花不同化学成分的镇咳、祛痰作用。方法镇咳实验:昆明种小鼠,随机分成空白组、阳性枸橼酸喷托维林对照组(0.01 g·kg^(-1))、生品款冬花(按生药量计,下同)水煎液组、总萜组、总皂苷组、总黄酮组、总生物碱组(6.0g·kg^(-1)),给药体积20ml·kg^(-1),连续给药5 d,然后观察25%浓氨水定量喷雾后出现咳嗽的潜伏期及2 min内的咳嗽次数;祛痰实验:分组及给药量同上,连续给药3d,气管段酚红法测定末次给药后45min小鼠痰液分泌量。结果与模型对照组相比,总萜组、总皂苷组、总黄酮组、总生物碱组镇咳效果显著,咳嗽次数明显减少(P<0.001);与水煎液组相比,总萜组、总黄酮组、总生物碱组能明显延长咳嗽潜伏期(P<0.05)。此外,与模型对照组及款冬花水煎液组相比,总萜组、总皂苷组、总黄酮组、总生物碱组均不能增加小鼠气管酚红排泌量。结论款冬花中生物碱、黄酮、萜、皂苷类化合物具有镇咳作用,是款冬花的药效成分。此外,萜类化合物具有毒性。

款冬花化学成分的研究

目的：研究款冬花的化学成分。方法：采用硅胶柱色谱方法进行分离、纯化，NMR和MS等方法进行结构鉴定。结果：从款冬花的干燥花蕾乙醇提取物中分离鉴定了11个化合物，分别为3，4-O-二咖啡酰基奎宁酸甲酯（1），3，5-O-二咖啡酰基奎宁酸甲酯（2），4，5-O-二咖啡酰基奎宁酸甲酯（3），3，5-O-二咖啡酰基奎宁酸（4），3-O-咖啡酰基奎宁酸甲酯（5），3-O-咖啡酰基奎宁酸（6），金丝桃苷（7），芦丁（8），山柰酚3-O-吡喃葡萄糖苷（9），槲皮素（10）和山柰酚（11）。结论：化合物2-5为首次从款冬属植物中分离得到。

款冬花化学成分的研究

从中药款冬花中分出一个新萜类成分,命名为款冬花素(farfaratin,Ⅰ)经红外、紫外、质谱、碳谱分析,最后经X-ray衍射确定结构。

款冬花不同部位提取物镇咳祛痰作用研究

目的研究款冬花不同溶剂提取物的镇咳、祛痰作用。方法采用小鼠氨水引咳法、小鼠气管酚红排泌法分别研究款冬花醇提取物、石油醚提取物、氯仿提取物、乙酸乙酯提取物、正丁醇提取物、水残余物高、低剂量的镇咳祛痰作用。结果与空白对照组比较,正丁醇高剂量组、氯仿高剂量组(6g/kg)能显著减少小鼠咳嗽次数并延长小鼠咳嗽潜伏期,差异有统计学意义(P<0.05或P<0.01);氯仿高剂量组(6g/kg)的祛痰效果较好,但与空白对照组相比,差异无统计学意义。结论款冬花镇咳部位是正丁醇、氯仿部位,祛痰部位可能是氯仿部位。

款冬花挥发油的提取及其在卷烟中的应用

采用同时蒸馏萃取-GC/MS法对款冬花挥发油进行了分离鉴定,并用该挥发油进行了卷烟加香试验。结果表明:①款冬花挥发油共鉴定出70种挥发性成分,主要有β-红没药烯、十六烷酸、匙叶桉油烯醇、7,10,13-十六三烯醛、δ-杜松烯、亚油酸、亚麻酸、β-荜澄茄油烯和β-紫罗兰酮等;②款冬花挥发油具有改善和修饰卷烟香气,增加清香香韵,减轻刺激性的作用。该挥发油可用于卷烟加香。

基于色谱技术的款冬花蕾与花梗代谢组成差异分析

本研究拟对款冬花蕾与花梗的化学组成差异进行比较。采用基于气质联用的代谢组学技术对花蕾与花梗的代谢组成差异进行分析,代谢产物的结构指认通过NIST数据库检索和标准品对照确定,GC-MS数据通过XCMS预处理后导入SIMCA-P软件进行多元统计分析;采用超高效液相色谱法对花蕾和花梗中吡咯里西啶生物碱的含量进行测定。结果基于GC-MS的代谢组学分析共鉴定了款冬中54个代谢产物,主成分分析显示款冬花蕾与花梗的代谢组成明显不同。OPLS-DA分析的载荷图显示,与花蕾相比,花梗中肌醇、香橙烯、豆甾醇、棕榈酸等化合物含量较低,而蔗糖、花生四烯酸等含量较高。此外,花蕾中吡咯里西啶生物碱含量为102μg·g-1,明显高于花梗中的含量(64μg·g-1)。款冬花蕾与花梗的化学组成存在较大差异,为了保证中医临床用药的安全有效,药用款冬花中应剔除花梗,并在质量标准检查项中规定花梗限量。

药用植物款冬花芽分化过程观察

实验以不同生长发育阶段的款冬花序芽突起为材料,通过制作石蜡切片,在显微镜下观察款冬花序芽分化各阶段的形态特征。结果表明:款冬花序芽从7月上旬开始花序(盘)分化至十月初小花胚珠分化完成,分化时期可分为分化前期、花盘形成期、花原基分化期、中央花(筒状)花瓣原基分化期、中央花雄蕊原基分化期、中央花雌蕊原基分化期、边缘花(舌状)花瓣原基分化期、边缘花雌蕊原基分化期、中央花花粉分化形成期、子房胚珠分化期共10个时期,阐明了款冬花序芽分化各时期与生长时间的关系。