不同采收及加工方式对穿心莲药材质量的影响

通过考察穿心莲的生长期、产地加工、营养器官和全草分段等因素对药材质量的影响,为穿心莲优质药材的生产提供科学依据。采用"一测多评"法,同步测定78份样品中穿心莲内酯(andrographolide,AD)、新穿心莲内酯(neoandrographolide,NAD)、14-去氧穿心莲内酯(14-deoxyandrographolide,14-DAD)和脱水穿心莲内酯(dehydroandrographolide,DHAD)的含量,并分别采用直观分析、聚类分析和偏最小二乘法判别分析(PLS-DA)等统计学方法进行数据分析和挖掘。结果表明:穿心莲按照快速生长期(Ⅰ期)、枝叶茂盛期(Ⅱ期)、花蕾期(Ⅲ期)、花盛期(Ⅳ期)、盛果期(Ⅴ期)和果期后期(Ⅵ期)6个生长期来划分,4种二萜内酯总量(TC)在Ⅰ~Ⅲ期内较为稳定,Ⅳ期开始下降,Ⅴ期下降最为迅速,Ⅵ期降至最低,因此穿心莲药材的采收期以花蕾期为最佳。产地加工时,尽管晒干和阴干对TC的影响较小,但与晒干比较,阴干样品中AD含量降低,DHAD和NAD含量升高,即阴干过程可以导致AD的分解和转化。因此,在天气条件允许的情况下,产地加工尽量采用晒干法进行干燥。AD,DHAD和TC在穿心莲不同全草分段和营养器官的分布分别为:全草上部>全草中部≈全草>全草下部,叶>全草>茎,且4种二萜内酯在叶和全草中的含量远高于茎,建议药材采收时以全草的上部和中部或含叶以上部位为佳,以保证其叶的占比要求。     

引种不同来源穿心莲叶中穿心莲内酯及脱水穿心莲内酯积累分析

目的观察不同来源穿心莲叶中穿心莲内酯和脱水穿心莲内酯的积累,以期为品种选育提供依据。方法收集同一引种地各生长阶段的不同来源地穿心莲叶,以HPLC测定叶中穿心莲内酯和脱水穿心莲内酯的含量。结果经数据分析,穿心莲内酯和脱水穿心莲内酯与生长期呈现明显的相关性。穿心莲内酯含量无论大叶型还是小叶型与始花期均成正相关,即开花期越晚,生长期越长,含量越高,大叶型相关系数的平均值为0.951,小叶型为0.899;脱水穿心莲内酯含量与始花期均成负相关,即开花期越晚,脱水穿心莲内酯含量越低,大叶型相关系数均值为-0.930,小叶型为-0.979。结论建议穿心莲品种应从大叶型中选育始花期晚的品种。     

In vivo and in vitro anti-inflammatory activities of neoandrographolide

Neoandrographolide, one of the principal diterpene lactones, isolated from a medicinal herb Andrographis paniculata Nees, was tested in vivo and in vitro for its anti-inflammatory activities and mechanism. Oral administration of neoandrographolide (150 mg/kg) significantly suppressed ear edema induced by dimethyl benzene in mice. Oral administration of neoandrographolide (100-150 mg/kg) also reduced the increase in vascular permeability induced by acetic acid in mice. In vitro studies were performed using the macrophage cell line RAW264.7 to study the effect of neoandrographolide on suppressing phorbol-12-myristate-13-acetate (PMA)-stimulated respiratory bursts and lipopolysaccharide (LPS)-induced production of nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha). Respiratory bursts were quantified by chemiluminescence (CL) measurements.Results showed that neoandrographolide suppressed PMA-stimulated respiratory bursts dose-dependently from 30 muM to 150 muM. Neoandrographolide also inhibited NO and TNF-alpha production in LPS-induced macrophages, contributing to the anti-inflammatory activity of A. paniculata. These results indicate that neoandrographolide possesses significant anti-inflammatory effects, which implies that it would be one of the major contributing components to participate in the anti-inflammatory effect of A. paniculata. and a potential candidate for further clinical trial.     

穿心莲UPLC-PDA指纹图谱研究及4种内酯类成分含量测定

分析研究不同部位及产地穿心莲药材,为穿心莲相关制剂工艺提供依据。以穿心莲内酯峰为参照,确定了27个共有指纹峰,建立了23批穿心莲药材的UPLC-PDA数字化定量指纹图谱,特征提取后对其进行主成分分析(PCA);采用外标法测定23批穿心莲药材中穿心莲内酯、新穿心莲内酯、去氧穿心莲内酯、脱水穿心莲内酯成分含量,并采用Plackett-Burman试验设计结合pareto图研究耐用性因素对方法的影响。结果发现,穿心莲药材部位比产地对药材质量的影响更显著;药材不同部位4种内酯含量差异大;流动相p H是影响方法耐用性较大的因素。     

穿心莲内酯及其衍生物药理活性研究

穿心莲内酯为二萜内酯类化合物,由于其具有广泛的药理作用而受到关注。文章简要阐述了穿心莲内酯的理化性质以及提取分离方法。重点综述了穿心莲内酯的药理活性以及穿心莲内酯进行分离纯化和以穿心莲内酯为母体进行不同的取代侧链的结构改造后,生成的穿心莲内酯衍生物或类似物在抗菌、抗炎、心血管系统、免疫调节、抗肿瘤这些方面的药理活性的改善。随着对其药理活性及结构改造的深入研究,穿心莲内酯类新药表现出广阔的研究和开发前景。     

穿心莲甲羟戊酸5-焦磷酸脱羧酶基因的克隆与表达

穿心莲内酯是穿心莲的主要药效成分,广泛用于抗炎。为了对它的生物合成进行遗传调控,提高内酯类成分的合成量,作者在转录组数据分析的基础上,获得了穿心莲内酯生物合成途径中甲羟戊酸5-焦磷酸脱羧酶基因的3个克隆,包含的ORF均长1 260 bp,分别在4个位点具有单碱基的差异。它们编码由419个残基组成的氨基酸序列;保守结构域中均具有11个高度保守的氨基酸,分别决定催化反应的特异性和活性;N端不含有质体定位的信号肽;与丹参的MVD蛋白(Gen Bank号AEZ55675.1)一致性较高。在穿心莲茎和叶片中,MVD基因的表达量分别在花蕾期和初花期最高,但各时期表达量的倍数差异不大。该研究获得的MVD基因,为后续进行详细的功能解析,并进一步应用于穿心莲内酯的遗传调控,奠定了基础。     

穿心莲香叶基香叶基焦磷酸合成酶基因的克隆与表达

目的从穿心莲中克隆穿心莲内酯合成途径中的香叶基香叶基焦磷酸合成酶(geranylgeranyl diphosphate synthase,GGPS)基因,并进行组织表达等特征研究。方法采用CTAB-LiC1法从穿心莲茎和叶中提取总RNA,简并引物扩增获得保守区段,RACE法获得基因全长,ProtParam等软件分析基因特征,实时荧光PCR法检测不同发育时期穿心莲茎和叶中基因的表达。结果获得了一个长1 047 bp的GGPS基因,编码一条由348个氨基酸组成的蛋白质序列,与长春花的GGPS蛋白亲缘关系较近,N端含有一个由47个氨基酸构成的质体信号肽。在穿心莲的茎和叶中,GGPS基因在花蕾期表达量较高,初花期降低;到了花果混合期表达量升高,青果期下降。表明穿心莲花蕾期和花果混合期相关代谢成分的合成可能更活跃。结论从穿心莲中克隆了GGPS基因,为开展穿心莲内酯合成途径的调控奠定了基础。     

穿心莲二萜内酯类化合物的合成及抗肿瘤构效关系研究

 目的合成穿心莲二萜内酯类化合物,并进行抗肿瘤活性研究。方法以穿心莲内酯为底物合成14-去氧-11,12-二去氢穿心莲内酯、14-去氧穿心莲内酯、异穿心莲内酯和穿心莲酸等穿心莲二萜内酯类化合物。以人白血病HL-60为体外模型,小鼠H22肝癌皮下移植肿瘤为体内模型观察其抗肿瘤活性,同时观察对小鼠迟发性变态反应及碳粒廓清功能的影响。结果目标物结构经红外、质谱、核磁共振氢谱确证。体外实验,异穿心莲内酯和穿心莲内酯对HL-60细胞的IC₅₀分别为7.15和28.34μmol·L^-1,其余化合物均大于50μmol·L^-1;体内实验,筛选剂量为100mg·kg^-1时,穿心莲内酯和异穿心莲内酯有明显的抗肿瘤活性,抑制率分别为64.4%和61.2%,其余化合物抗肿瘤活性不明显;相同剂量下,穿心莲内酯,穿心莲酸对迟发性变态反应有促进作用,穿心莲内酯,穿心莲酸,14-去氧穿心莲内酯,14-去氧-11,12-二去氢穿心莲内酯能降低小鼠碳粒廓清指数,而异穿心莲内酯对迟发性变态反应和炭粒廓清指数均无影响。结论初步生物活性实验表明,穿心莲二萜内酯类化合物的内酯环是抗肿瘤的必须活性基团;该类化合物的抗肿瘤作用可能与内酯环部分(包括C₁₁和C₁₂)双键的个数及位置和14-位羟基均有一定关系,具有环外12,13-位双键和14-位羟基的抗肿瘤活性较强;异穿心莲内酯的四氢呋喃环有增强抗肿瘤活性之作用。异穿心莲内酯有非常显著的抗肿瘤活性,且对小鼠免疫功能无影响,提示它可能是一类有发展前景的抗肿瘤药物。     

Andrographolide inhibits the tumour necrosis factor-α-induced upregulation of ICAM-1 expression and endothelial-monocyte adhesion

Andrographolide, a diterpene isolated from Andrographis paniculata, has been shown to have several biological activities including analgesic, antipyretic and antiinflammatory effects. Since the upregulation of adhesion molecules expression and endothelial-leucocytes adhesion are key steps in the development of inflammation, the present study examines whether andrographolide modulates these biological processes in vitro. Incubation of endothelial cells with non-toxic concentrations (0.16–16.7 υg/mL) of andrographolide attenuated the tumour necrosis factor-α (TNF)-induced intercellular adhesion molecule-1 (ICAM-1) expression. Similar concentration ranges of andrographolide also inhibited the TNF-induced endothelial-monocyte adhesion in a concentration-dependent manner. These effects of andrographolide may account for its reported in vivoantiinflammatory activity. © 1998 John Wiley & Sons, Ltd.     

UPLC快速分析穿心莲药材及制剂中内酯成分

目的:建立UPLC-PDA技术快速测定穿心莲药材及制剂中主要内酯类成分含量的方法.方法:采用超高效液相色谱仪,用Acquity BEH Shield RP C18色谱柱(2.1 mm×50 mm,1.7 μm);流动相甲醇(A)-水(B),梯度洗脱(0～2 min,40％A;2 ～4 min,40％ ～45％ A;4 ～ 10 min,45％ ～50％ A;10 ～15 min,50％A);流速0.5 mL· min -1,检测波长226 nm,柱温30℃.结果:在15 min内可以完成药材和制剂中穿心莲内酯、脱水穿心莲内酯的分析,分离度良好,线性范围分别为0.0286 ～0.286 g·L-1(r=0.9996,n=6),0.0940～0.940 g·L-1(r =0.9993,n=6);平均加样回收率分别为97.32％,97.11％.结论:该方法快速、准确、灵敏度高,可用于穿心莲药材及制剂的质量控制.     

Anticancer and immunostimulatory compounds from Andrographis paniculata

Andrographis paniculata extract is traditionally used as a medicine to treat different diseases in India, China and Southeast Asia. In the present study, we evaluated the anticancer and immunomodulatory activity of the methanolic extract of Andrographis paniculata in human cancer and immune cells. The methanolic extract of Andrographis paniculata was fractionated into dichloromethane, petroleum ether and aqueous extracts and screened for bioactivity. Our results indicate that the dichloromethane fraction of the methanolic extract retains the active compounds contributing for both the anticancer and immunostimulatory activity. Dichloromethane fraction significantly inhibits the proliferation of HT-29 (colon cancer) cells and augments the proliferation human peripheral blood lymphocytes (HPBLs) at low concentrations. On further fractionation of the dichloromethane extract we could isolate three diterpene compounds, i.e. [1] andrographolide, [2] 14-deoxyandrographolide and [3] 14-deoxy-11,12-didehydroandrographolide. Andrographolide showed anticancer activity on diverse cancer cells representing different types of human cancers. Whereas all the three molecules showed enhanced proliferation and interleukin-2 (IL-2) induction in HPBLs.     

黄芩不同部位中15种黄酮类成分的含量测定及化学计量学研究

Objective: The aerial parts of Scutellaria baicalensis were used as Huangqin Tea for thousands of years and mainly contain flavonoids which contribute to its bioactivities. However, there is no appropriate quality evaluation method of Huangqin Tea, and three flavanones of isocarthamidin-7-O-β-D-glucuronide,carthamidin-7-O-β-D-glucuronide, and isoscutellarein-8-O-β-D-glucuronide with high contents in the aerial parts have never been defined quantitatively. Here, an HPLC-DAD method for simultaneous determination of 15 flavonoids and systematically compared their contents and distribution in the roots,stems, leaves, and flowers of S. baicalensis was established.Methods: Under the HPLC-DAD chromatographic conditions, 77 batches of samples of S. baicalensis were analyzed. Meanwhile, the chromatographic fingerprint of different parts of S. baicalensis was established.Subsequently, principal component analysis(PCA), orthogonal projections to latent structures discriminant analysis(OPLS-DA), and clustering heat map were performed based on the contents of 15 flavonoids in different parts of S. baicalensis.Results: The results showed significant differences in the contents and distributions of 15 flavonoids among the different parts of S. baicalensis. The chemical composition of stems showed some similarities to leaves, and their contents were all lower than leaves. The contents of isocarthamidin-7-O-β–glucuronide [(106.66 66 ± 22.68) mg/g], carthamidin-7-O-β-D-glucuronide [(19.82 ± 11.17) mg/g],and isoscutellarein-8-O-β-D-glucuronide [(3.10 ± 1.73) mg/g] were the highest in leaves. The content of apigenin-7-O-β-D-glucopyranoside and chrysin-7-O-β-D-glucuronide were the highest in flowers. The contents of baicalin, baicalein, wogonoside, wogonin, alpinetin, and oroxylin A were higher in roots than in other parts.Conclusion: The method was fully validated and could be effectively used to characterize the contents and distributions of main flavonoids in the different parts of S. baicalensis. It may lay a foundation to establish the quality evaluation system for Huangqin Tea.     

黄独脱毒苗快繁技术的研究

目的 以黄独脱毒苗为试材,研究不同因素对黄独带芽茎段芽增殖和生根的影响,以期对黄独脱毒苗的快繁技术进行优化.方法 采用植物组织培养的方法 进行茎尖培养和快繁研究,采用RT-PCR法对茎尖脱毒植株进行病毒检测.结果 黄独脱毒苗带芽苇段的最佳培养基是MS+KT 2 mg/L+6-BA 1 mg/L+NAA 0.5 mg/L;黄独脱毒苗带芽茎段增殖的最佳蔗糖质量浓度和琼脂质量浓度分别是30和0 g/L;黄独脱毒苗带芽茎段生根的最佳培养基是1/2 MS+IBA 0.1 mg/L+NAA 0.5 mg/L+PP_(333) 1 mg/L;黄独试管苗移栽的最好基质是珍珠岩:蛭石(2:1);黄独试管苗移栽时最佳的PP_(333)质量浓度是50 mg/L.结论 首次成功建立了黄独脱毒苗的快繁技术,为黄独脱毒苗的工厂化生产奠定了技术基础.     

妇科千金方及其拆方、单味饮片主要成分分析及10种成分测定

目的 分析妇科千金方(穿心莲、单面针、功劳木等)及其拆方(抗炎方、抑菌方、补益方)、单味饮片主要成分,并测定原儿茶酸、木兰花碱、芦丁、染料木苷、党参炔苷、小檗碱、穿心莲内酯、染料木素、脱水穿心莲内酯、藁本内酯的含量.方法 制备全方、拆方、单味饮片,其70％甲醇提取物的HPLC分析采用Kromasil 100-5-C18...     

Six new diterpenoids from Suregada multiflora

Six new diterpenoids were isolated from a CH(2)Cl(2)-MeOH extract of the bark of Suregada multiflora. The structures were established on the basis of one- and two-dimensional NMR and other spectroscopic studies and chemical derivatizations. Two compounds, suregadolides C (1) and D (2), were identified as new diterpene lactones of two antipodal series, containing a cyclopropane ring bridging C-3 and C-4 of the basic abietane skeleton. Suremulide A (3) was found to be a new abietene diterpene lactone. Bannaringaolide A (4), a diterpene lactone, based on a novel carbon skeleton with a seven-membered ring, possibly formed by the rearrangement of the exocyclic C-17 in ring C of an ent-pimarane framework, has also been isolated. A kaurane triol, suremulol A (5), and a kaurane diol, suremulol B (6), were also identified as new metabolites.     

Two diterpenes and three diterpene glucosides from Phlogacanthus curviflorus

Two new diterpene lactones, phlogacantholides B (1) and C (2), and three new diterpene lactone glucosides, phlogacanthosides A (3), B (4), and C (5), together with lupeol, beta-sitosterol, betulin, beta-daucosterol, (+)-syringaresinol, and (+)-syringaresinol-4-O-beta-D-glucopyranoside, were isolated from the roots of Phlogacanthus curviflorus. Their structures were elucidated by chemical and spectroscopic evidence. The structure, including the relative configuration of phlogacantholide B (1), was confirmed by X-ray crystallographic analysis of its diacetate (6).     

Andropanolide and isoandrographolide, minor diterpenoids from Andrographis paniculata: structure and X-ray crystallographic analysis

Phytochemical investigation of the leaves of Andrographis paniculata has led to the isolation of a new labdane type diterpenoid, andropanolide (1), along with seven known diterpenoids including isoandrographolide (2), previously reported as a rearrangement product of andrographolide. The structures and stereochemistry of compounds 1 and 2 were established by X-ray crystallographic analysis.     

栝楼的组培快繁技术研究

目的:探讨栝楼组织培养及种苗快速繁殖技术。方法:通过茎尖、带芽茎段、茎段和叶片诱导丛生芽及愈伤组织。结果:栝楼2年生块茎幼苗的茎尖和带芽茎段在MS 2 mg/L BA 0.5~0.05 mg/L NAA培养基上可诱导丛生芽,并在生根培养基MS 0.1 mg/L NAA 0.2 mg/L IBA中生根,经驯化移栽可实现快速繁殖。无芽茎段和叶片用含4 mg/L BA和0.5 mg/L NAA的MS培养基可诱导愈伤组织,由茎段来的愈伤组织经6周后分化为无根苗。结论:栝楼茎尖和带芽茎段的组织培养,可在短期内大量繁殖栝楼雌雄种苗,具有成本低,效益高的特点。     

施用硼肥对菊花生长及品质的影响

目的:研究施用硼肥对于菊花生长和品质的影响,旨在为菊花生态种植合理施肥提供科学的依据。方法:以杭菊为研究对象,采用砂培实验,设置不施硼、适硼、高硼3个施硼浓度,测定不同硼浓度下菊花经济学性状及产量、矿质元素吸收以及内在成分含量等指标。结果:相较于不施硼肥,适量施硼可以显著提高杭菊花的产量,增产幅度可达39.02%,而高硼显著降低了杭菊花产量,降低了19.43%。适量硼肥能显著提高蕾期杭菊植株茎秆干物质量和花期植株叶片干物质量,分别为12.04%和36.63%。施加硼肥对于不同时期不同部位矿质元素吸收的影响不尽相同。施加适量硼肥能提高总黄酮和3,5-O-二咖啡酰基奎宁酸含量,但会降低绿原酸和木犀草苷含量;而高硼胁迫虽然能显著提高菊花的总黄酮、3,5-O-二咖啡酰基奎宁酸、绿原酸和木犀草苷含量,但却显著降低了菊花产量,结论:在菊花种植生产上,应重视硼肥的施用。     

不同树龄银杏叶在不同季节中总黄酮和总内酯的含量变化

目的：确定银杏叶最适宜采收季节。方法：采用HPLC法（二级管阵列检测器DAD和蒸发光散射检测器ELSD）分别测定银杏叶中总黄酮和总内酯的含量。结果：不同树龄,不同季节的银杏叶总黄酮、总内酯含量不同,结论：银杏叶的总黄酮、总内酯以二、三年生的为高,总黄酮在5月份,总内酯在9月份含量为高。