血脑屏障药物筛选模型的建立及丹参活性成分筛选分析

ECV304细胞在C6细胞的诱导下生长,通过细胞培养时间优化、渗透系数测定和细胞形态学观察等,建立ECV304/C6共培养血脑屏障(BBB)药物筛选模型.将该模型应用于从丹参提取液中筛选可能作用于中枢神经系统(CNS)的活性成分,结合液相色谱-质谱联用技术(LC-MS)分析,发现丹参提取液中至少有16种成分能够穿越BBB模型,其中4种成分被确认为隐丹参酮、丹参酮Ⅰ、丹参素和原儿茶酸.通过定量构效关系(QASR)分析,进一步从理论上证明所确认的4种化合物均符合CNS靶向药物的特征.研究结果表明,ECV304/C6共培养BBB模型能够在模拟生理状态下从中药复杂体系中筛选分离跨越BBB的活性成分组,可用于CNS药物开发的早期快速筛选,服务于中药现代化研究.     

Effect of compatibility on the pharmacokinetic characteristics of ginsenosides

Pharmacokinetic research, which is one of the most important parts of preclinic research, plays an important role in guiding medicine compatibility and preparation improvement. In this paper, the influence of the compatibility of GGV on pharmacokinetics of ginsenosides (GS) was studied, which consisted of ginsenosides, ginkgo biloba extract (GBE) and a chemical monomer V. The result indicated that the addition of either GBE or V could influence the pharmacokinetic parameters of ginsenosides and the influence was different when different administering routes were adopted. Therefore, it could be concluded that there are interactions among GS, GBE and V, and the synergetic and inhibitory effects of the three ingredients contribute to the pharmacological effect of GGV.     

Protective effect of total flavonoids from Spirodela polyrrhiza (L.) Schleid on human umbilical vein endothelial cell damage induced by hydrogen peroxide

In this study, we determined the protective effect of total flavonoids from Spirodela polyrrhiza (L.) Schleid (STF), which is a kind of traditional Chinese medicine, on human umbilical vein endothelial cells (ECV-304) damage induced by hydrogen peroxide (H(2)O(2)). Treated with 1mmol/L H(2)O(2) for 1h, the viability of ECV-304 cells markedly decreased. However, pretreatment with 10-50mug/mL STF resulted in a significant recovery. The survival rate of ECV-304 increased from 21.98% (only treated with 1mmol/L H(2)O(2)) to 64.74% (pretreated with 50microg/mL STF), which accompanied with the amounts of malondialdenhyde (MDA) decreasing from 1.6883nmol/L to 0.9628nmol/L. Furthermore, compared with control group, the 50mumol/L STF pretreatment enhanced the total antioxidant capacity (T-AOC) by 4.49 times, increased the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX) by 85.12%, 158.94% and 94.5%,respectively, and increased the content of nitric oxide (NO) by 116.55%.Taken together, STF protect ECV-304 cells against H(2)O(2) damage by enhancing the antioxidant ability and increasing NO production.     

TNFa在血管内皮细胞ECV304中的靶向表达研究

在缺失了3'LTR U3区内病毒的启动子／增强子序列的逆转录病毒载体pLXSNd中,用血管内皮生长因子受体KDR的特异性启动子调控了TNFa在血管内细胞ECV304中的靶向表达。将构建的载体pLXSN-TNFa,pLXSNd-KDRp-TNFa和空载体pLXSN用PA317细胞包装后获得重组病毒,并用重组病毒分别感染NIH3T3细胞和ECV304细胞,培养物上清的ELISA结果证明,KDR启动子指导的TNFa在KDR阳性细胞ECV304中的表达量为在KDR阴性细胞NIH3T3中的表达量的8倍;而TR指导的TNFa在这两种细胞中的表达无明显差异,实现了TNFa在血管内皮细胞中的靶向表达,这可能为肿瘤基因治疗提供新途径。     

Investigation on the effects of diamide on NO production in vascular endothelial cells (VEC)

Nitric oxide (NO) controls several physiological functions of the cardiovascular system. The study on the effect of diamide (N₂H₄·H₂O) on NO production in vascular endothelial cells (VEC) may provide significant reference for VEC’s modeling in studying cardiovascular diseases. The objective of this study was to elucidate how high concentration diamide (V_diamide/V_{culture miedium}=5 ml/l) and low concentration diamide (V_diamide/V_{culture miedium}=0.5 ml/l) affect NO production in a human endothelial cell line (ECV304). After cells were incubated with diamide (5 or 0.5 ml/l) for 4, 6, 8 or 10 h, respectively, the amounts of NO metabolites released by the cells were quantitated and the degree of damage of VEC was observed using microscope. The results showed that NO production in VEC tended to decrease with the lapse of time in the 0.5 ml/l diamide group. In the 5 ml/l diamide group, on the contrary, NO production in VEC tended to increase with the lapse of time. At the same time, from the morphologic observation, the VEC were damaged severely after treated with 5ml/l diamide. So it could be concluded that the severe damage induced by high concentration diamide would have triggered the express of inducible nitric oxide synthases (iNOS). Just for the expresssion of iNOS, NO production in VEC treated with high concentration diamide occurred abnormally in contrast to the 0.5 ml/l group.     

Molecularly imprinted layer-coated monodisperse spherical silica microparticles toward affinity-enrichment of isoflavonoid glycosides from Radix puerariae

This paper reports the preparation of puerarin (PR) imprinted layer-coated silica microparticles toward selective recognition of PR and fast affinity-enrichment of the main isoflavonoid glycosides from the crude extract of Radix puerariae. Before the preparation, quantum mechanics (QM) method was applied to identify three kinds of common functional monomers capable of interaction with PR and then predicted optimal functional monomer (acrylamide, AA) and the relative molar ratio of template to functional monomer (PR/AA, 1:4). The obtained PR-imprinted silica microparticles were evaluated by transmission electron microscope (TEM) and rebinding experiments, exhibiting good morphology and high binding affinity to PR. Meanwhile, the rebinding amount of the imprinted microparticles to PR was nearly 2.1-folds that of non-imprinted microparticles. When the PR-imprinted microspheres were used as packing materials for solid-phase extraction, the recovery yields of PR, daidzin (DD) and genistin (GS) were simultaneously up to 90% by one-step extraction from the crude extract of Radix puerariae. Additionally, the PR-imprinted microparticles could be re-used for at least 5 times without losing any extraction efficiency. These results indicate that the PR-imprinted microparticles have highly selective adsorption capabilities to PR, DD and GS from the crude extract of Radix puerariae. The method of molecularly imprinted polymers (MIPs) coupled with solid-phase extraction (SPE) provides a good solution of the enrichment and separation of active extracts from complicated traditional Chinese medicine (TCM) with certain structures.     

慢性肾衰肾虚证患者血清微量元素的变化及其临床意义探讨

讨论了慢性肾衰肾阳虚和肾阴虚见证者血清微量元素。结果显示：肾阳虚患者血清Ｃｕ、Ｃｕ／Ｚｎ明显高于健康对照组,而血清Ｚｎ和Ｆｅ低于对照组,肾阳虚患者血清Ｃｕ、Ｃｕ／Ｚｎ也明显高于肾阴虚组,而血清Ｚｎ是低的,在肾阴虚组与对照组之间Ｃｕ、Ｚｎ、Ｃｕ／Ｚｎ均无明显差异。     

Application of pyrolysis-high-resolution gas chromatography-pattern recognition to the identification of the Chinese traditional medicine mai dong

Pyrolysis-high-resolution gas chromatography-pattern recognition (Py-HRGC-PaRe) was used to develop a potential technique for identifying the Chinese traditional medicine Mai Dong. About 1 mg of crude drug powder was pyrolysed in a furnace pyrolyser and the products were directly carried into a gas chromatograph with an FSOT capillary column (30 m x 0.265 mm I.D.) coated with DB-1701 (df 0.25 micron). The Py-HRGC data were analysed by non-linear mapping PaRe. The results showed that Mai Dong samples could be classified into two categories: Ophiopogon japonicus (L.f.) Ker-Gawl (included in the Chinese Pharmacopoeia) and Liriope spicata.     

8味补阴药补阳药中微量元素Fe、Mg、Zn、Ca、Mn、Cu的测定

采用火焰原子吸收光谱法对补阴药石斛、玉竹、北沙参、女贞子和补阳药肉苁蓉、杜仲、菟丝子、补骨脂8味中药进行Fe、Mg、Zn、Ca、Mn、Cu含量测定。结果表明,以平均值而论Mg、Cu含量补阴药与补阳药相当,Fe、Ca含量补阴药低于补阳药,Zn、Mn含量补阴药高于补阳药。以每味中药而论,补阳药菟丝子Fe含量最高,是其他几味中药的近10倍,补阴药石斛Zn、Mn含量最高,是其他几味中药的3~10倍。补阴药、补阳药药效是相对的,但本实验测定各微量元素含量并没有呈现相对性,因此,补阴药、补阳药的药效与Fe、Mg、Zn、Ca、Mn、Cu微量元素的相关性有待于进一步研究。     

中药注射剂荧光光谱法的快速鉴别和热稳定性研究

红外光谱指纹图谱技术应用中药注射剂的快速鉴别中，某些试样因难于成膜造成制样困难，为此，直接或仅经水稀释后测定了12种21批次的常用中药注射剂的荧光光谱。结果表明：由于中药注射剂常含荧光性物质，但因不同注射剂所含具体荧光性物质不同，同种注射剂又因厂家的具体配方的差异或制备工艺条件的波动，均会使其特征的荧光激发、发射光谱不同或其强度呈现差异；从而可充分利用荧光分析的高灵敏度，使其作为中药整体红外指纹识别的一种辅助手段，根据注射剂的荧光图谱的差异达到快速鉴别、认定和控制配方、工艺的目的。比较而言，荧光法表现出来的差异更为一目了然，易于判断。此外，荧光光谱法还可用于中药注射液的热稳定性的研究。     

中国的中药微量元素研究Ⅱ.微量元素:中药有效药成分的核心组分

在中国开展的中药微量元素研究证明,中药有效药成分由有机药成分和无机药成分组成,两者共生共存。从含水络合离子、天然有机金属配合物、中药有机药成分-微量元素配合物,以及中药有机药成分-微量元素复合物四个方面,论述了微量元素在中药有效药成分中的核心地位和核心作用。     

关于中药质量控制与体内代谢研究的思考

质量控制是保证中药安全、有效的重要基础。体内代谢研究可以为阐明中药的治病机制提供依据。本文以蟾酥为例,讨论了当前中药质量控制及体内代谢研究存在的主要问题和解决办法:认为把中药指纹图谱的全面指认与多指标成分定量相结合是有效控制中药质量的可行途径;液相色谱-质谱联用技术(LC/MS)将在中药化学研究、体内代谢研究等方面发挥重要作用;中药的化学指纹图谱应与药理活性相关联,从而建立合理的中药质量评价体系;结构修饰对基于天然产物的新药发现具有重要意义。     

Determination of Paeoniflorin in Total Glucosides of Peony Root by HPLC

The crude drug of Radix Paeoniae Alba has been used extensively in traditional Chinese medicine as an analgesic, an antispasmodic,an astringent, and a sedative for the treatment of a variety of painful afflictions. A modern way of using Radix Paeoniae Alba is the application of Total Glucosides of Peony Root (TGPR), which is the ethanol extract from the crude drug. Our interest in this work is to develop a method to determine one of the main effective components, Paeoniflorin, in TGPR by HPLC. Although several HPLC methods for the determination of Paeoniflorin were reported before,our method has the advantages of simplicity, rapidity and accuracy.     

矿物中药自然铜的组成与热稳定性研究

采用X射线粉末衍射、热重-差热分析、等离子体发射光谱等分析手段, 对不同产地自然铜和煅自然铜的结构组成、热稳定性和微量元素进行测定和分析. 结果表明: 自然铜的主要物相为FeS2, 煅自然铜则因产地而异, 出现了FeS2(南京中医药大学样品中约86%, 湖南89%, 四川32%, 山西19%), FeS(湖南11%, 安徽24%, 四川24%), Fe3O4(安徽29%), Fe2O3(安徽14%)和FeO(OH) (南京中医药大学14%, 山西74%)等复杂物相. 自然铜在加热到450～800 ℃时, 逐渐发生了由FeS2转变为FeS的相变. 在这些药材中含有丰富的与人体健康密切相关的微量元素, 既含有对人体有益的Ca, Fe, Zn等微量元素, 也含有As, Cd, Pb等有害微量元素. 实验结果为该矿物药的鉴定和评价提供了科学数据.     

泽泻醇类化合物与血清白蛋白相互作用的分子机理研究

中药泽泻具有抗肿瘤作用,可能与血清中蛋白成分的改变有关.利用荧光光谱、圆二色谱结合分子模拟技术研究了模拟生理条件下泽泻有效成分泽泻醇类化合物与人血清白蛋白的相互作用.实验结果表明,23-乙酰泽泻醇B与蛋白的结合作用远强于24-乙酰泽泻醇A.分子模拟结果与实验一致,并且表明,结合强弱的差异与小分子的侧链结构有关.该结果可...     

Static headspace‐multicapillary column with gas chromatography coupled to ion mobility spectrometry as a simple approach for the discrimination of crude and processed traditional Chinese medicines

The processing procedure can alter the nature and chemical transformation of traditional Chinese medicine to accommodate different clinical dispensing and preparation requirements. In this study, static headspace‐multicapillary column with gas chromatography coupled to ion mobility spectrometry was developed for the rapid and sensitive discrimination of crude and processed traditional Chinese medicine. Using Radix Paeoniae Alba as a traditional Chinese medicine model, the combined power of this approach was illustrated by classifying the crude and processed Radix Paeoniae Alba samples into two main categories. The contents of the main components in Radix Paeoniae Alba varied significantly. The established method could promote the use of ion mobility spectrometry in intrinsic quality control and differentiation of herbal medicines from other processed products or preparations.     

Authentication of Chinese crude drug gecko by DNA barcoding

Gekko gecko, an animal used as a valued traditional Chinese medicine, has been widely used for over 2000 years. Due to localized habitat destruction, the amount of G. gecko has dramatically decreased in recent years. As a result, more and more adulterants have been detected in the traditional medicine, which has resulted in a chaotic market. Therefore, a correct identification method is badly needed. In this study, we employed a new molecular method of DNA barcoding for discriminating gecko from its adulterants. Fifty-seven specimens of gecko and its adulterants were collected as test samples. The full-barcode and mini-barcode sequences of these specimens were separately amplified and sequenced separately. Together with other published barcode sequences, we detected that the intra-specific sequence diversity was far lower than the inter-specific diversity in G. gecko and its adulterants (3% compared with 35% in full-length barcode; 4% compared with 33.5% in mini-barcode). These results showed that both the full-length and mini-barcodes were effective for identifying gecko, which suggested that the DNA barcode could be an effective and powerful tool for identifying the Chinese crude drug gecko.     

Effect of wine processing and acute blood stasis on the serum pharmacochemistry of rhubarb: A possible explanation for processing mechanism

As a specific item mentioned in traditional Chinese medicine theory, processing can fulfill different requirements of therapies. Crude and wine‐processed rhubarbs are used as drastic and mild laxatives, respectively. In this study, a practical method based on ultra‐fast liquid chromatography coupled with diode‐array detection and ion trap time‐of‐flight mass spectrometry was developed to screen and analyze multiple absorbed bioactive components and metabolites in the serum of both normal and acute blood stasis rats after oral administration of crude or wine‐processed rhubarbs. A total of 16 compounds, mainly including phase II metabolites, were tentatively identified. Possible explanations for the processing‐induced changes in pharmacological effects of traditional Chinese medicines were first explored at serum pharmacochemistry level.     

多囊卵巢综合征相关雄性激素的液相色谱-串联质谱检测

建立了液相色谱-串联质谱(LC-MS/MS)检测血清中硫酸脱氢表雄酮(DHEAS)、雄烯二酮(A4)、睾酮(T)、17-羟基孕酮(17-OHP)、双氢睾酮(DHT)5种激素的分析方法。血清样本经蛋白沉淀后,采用固相萃取,经Agela Venusil MP C₁₈色谱柱(3.0 mm×50 mm,3μm)分离,以含0.1%甲酸的甲醇和含0.02%甲酸的水为流动相进行梯度洗脱。使用正-负离子多反应监测模式进行数据采集。结果表明,5种激素在各自质量浓度范围内线性关系良好(r²>0.995),回收率为96.0%～105%,各激素的批内精密度和批间精密度均小于10%,DHEAS、A4、T、17-OHP、DHT的定量下限(LOQ)分别为5.00、0.05、0.05、0.025、0.025 ng/mL。该方法快速、准确、灵敏,适用于临床对多囊卵巢综合征(PCOS)患者血清雄性激素水平的检测。