UPLC-MS/MS法同时检测小花鬼针草中6种单体成分的含量

建立了一种应用UPLC-MS/MS同时测定小花鬼针草中6种单体成分原儿茶酸、芦丁、金丝桃苷、槲皮苷、槲皮素和山奈酚含量的方法。小花鬼针草药材粉碎后，经甲醇超声提取稀释后进样，采用负离子多反应监测（MRM）模式同时测定提取物中6种单体成分的含量，并以芍药苷为内标。色谱柱为Agilent SB C18 （50 mm × 2.1 mm, 1.8 μm），流动相为乙腈-0.1%甲酸水溶液，梯度洗脱，流速为0.4 mL/min，柱温为35 ℃。结果表明，该方法所测6种成分在相应的质量浓度范围内，峰面积与浓度线性关系良好，r＞0.999 0；平均加样回收率（n=6）为92.1% ～95.5%，相对标准偏差≤4.50%。该方法简单快捷、灵敏度高、专属性强，可同时测定小花鬼针草中6种单体成分。     

Anti-Inflammatory Activity and Mechanism of Isookanin,Isolated by Bioassay-Guided Fractionation from Bidens pilosa L.

Bidens pilosa L. (Asteraceae) has been used historically in traditional Asian medicine and is known to have a variety of biological effects. However, the specific active compounds responsible for the individual pharmacological effects of Bidens pilosa L. (B. pilosa) extract have not yet been made clear. This study aimed to investigate the anti-inflammatory phytochemicals obtained from B. pilosa. We isolated a flavonoids-type phytochemical, isookanin, from B. pilosa through bioassay-guided fractionation based on its capacity to inhibit inflammation. Some of isookanin’s biological properties have been reported; however, the anti-inflammatory mechanism of isookanin has not yet been studied. In the present study, we evaluated the anti-inflammatory activities of isookanin using lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. We have shown that isookanin reduces the production of proinflammatory mediators (nitric oxide, prostaglandin E₂) by inhibiting the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated macrophages. Isookanin also inhibited the expression of activator protein 1 (AP-1) and downregulated the LPS-induced phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-jun NH₂-terminal kinase (JNK) in the MAPK signaling pathway. Additionally, isookanin inhibited proinflammatory cytokines (tumor necrosis factor-a (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8), and interleukin-1β (IL-1β)) in LPS-induced THP-1 cells. These results demonstrate that isookanin could be a potential therapeutic candidate for inflammatory disease.     

Isolation and Identification of Bioactive Compounds from Bidens spp. Using HPLC-DAD and GC-MS Analysis and Their Biological Activity as Anticancer Molecules

The genus Bidens a member of family Compositae, is widely documented as an ethno-medicinally important genus of plants. In the present study, anticancer potential of three ethno-medicinally important species i.e., B. bipinnata, B. biternata and B. pilosa were tested. For in-vitro evaluation, an MTT (Thiazolyl blue tetrazolium bromide) assay was performed against cervical cancer cells (HeLa), hepatocellular carcinoma (HepG), and adenocarcinoma human alveolar basal epithelial cells (A549). For in vivo evaluation, Artemia salina, Danio rerio, and Caenorhabditis elegans were used. Among all the tested extracts, the ethanol extract of B. biternata appeared to have highest anticancer activity, and the compounds responsible for this activity were identified to be Tris (2,4-di-tert-butylphenyl), 4-hydroxy-2,4′-dimethoxychalcone, and 2,4-di-tert-butylphenol. This is the first report of the isolation of Tris (2,4-di-tert-butylphenyl) phosphate from the genus Bidens and the first report of 4-hydroxy-2,4′-dimethoxychalcone and 2,4-di-tert-butylphenol from B. biternata. Among the isolated compounds, 4-hydroxy-2,4′-dimethoxychalcone showed the highest anticancer activity with an LD50 value of 236.7 µg/mL. Therefore, this compound carries promising potential for being established as a pharmaceutical for chemoprevention and chemotherapy.     

HPLC method development and validation of cytotoxic agent phenyl‐heptatriyne in Bidens pilosa with ultrasonic‐assisted cloud point extraction and preconcentration

Extraction and pre‐concentration of a bioactive marker compound, phenyl‐1,3,5‐heptatriyne from Bidens pilosa, prior to HPLC has been demonstrated using both organic and ecofriendly solvents. Non‐ionic surfactants, viz. Triton X‐100, Triton X‐114 and Genapol X‐80, were used for extraction. No back‐extraction or liquid chromatographic steps were required to remove the target phytochemical from the surfactant‐rich extractant phase. The optimized cloud point extraction procedure has been shown to be a potentially useful methodology for the preconcentration of the target analyte, with a preconcentration factor of 4–99. Moreover, the method is simple, sensitive, rapid and consumes lesser solvent than traditional methods. An isocratic chromatographic separation and quantitation was accomplished on a C₁₈ column with acetonitrile–acidified aqueous as mobile phase at a flow rate of 1.0 mL/min, UV detection at 254 nm and specificity with photo diode‐array detector (PDA) and MS. Under the optimum experimental conditions recovery was satisfactory (99.18–100.33%) without interference from the surfactant. The method seems to be reliable with intraday precision and interday precision below 2.0%. Good linearity was obtained in the working range from 7.5 to 30 µg/mL with correlation coefficient >0.99. The limits of detection and quantitation were 1.84 and 6.13 µg/mL, respectively. The method was validated following international guidelines and successfully applied for quantitative assays of cytotoxic compound phenyl‐1,3,5‐heptatriyne in Bidens pilosa. Copyright © 2010 John Wiley & Sons, Ltd.     

天麻中香草醇对灰树花菌丝体生物量和胞外多糖合成的影响

为探讨天麻醇提物特征成分香草醇对灰树花菌丝体生物量和胞外多糖(exopolysaccharide, EPS)合成的影响,在灰树花发酵体系中添加不同质量浓度的香草醇,采用苯酚-硫酸法测定多糖含量并记录菌丝体质量。研究表明:一定质量浓度的香草醇对灰树花菌丝体生长和EPS合成有促进作用;香草醇质量浓度为200mg/L时,促进灰树花菌丝体生长和EPS合成的效果最佳,相比于空白组(未添加香草醇)分别增加了21.1%和19.9%,均显著高于空白组(P＜0.05)。此外,在香草醇参与的灰树花整个发酵过程中,发酵第13天开始生物量和EPS产量趋于稳定,第14天时灰树花菌丝体生物量和EPS产量为最大值,分别为(1.42±0.01)g/L和(2.10±0.03)g/L,但低于天麻醇提物和对羟基苯甲醛实验组;且第14天同第0天相比,香草醇含量降低了68.3%。香草醇作为天麻醇提物中的特征成分可以被灰树花用于自身菌丝体生长和EPS的合成。     

民族药鬼针草的生药学鉴定

采用原植物、性状、显微、薄层鉴别方法对民族药鬼针草进行系统的生药学鉴定,为其鉴别及应用提供科学依据.结果表明:通过原植物、性状鉴别、显微鉴别、薄层色谱鉴别能够很好地鉴定该药材.     

根癌病土壤杆菌诱导转化毛状根能力的评价

以鸟头(Aconitum carmichaeli)、Bidens sulphureus和洋金花Datura metal)三个植物种为材料(用A.rhizogenes的九个菌株和A tumefaciens的一个突变系为感染源.诱导出转化毛状根.根据所诱导出的转化毛状根数量来评价各菌株诱导产生转化毛状根的能力.结果表明同一菌株对不同植物和同一植物接受不同菌株的感染力有明显差异.A. rhizogenes ATCC15834、ATCC 25818、A 42017和A.tumefaciens、TMR 338诱导产生转化毛状根的能力最强,A.rhizogenes R1000、TR 105次之.ATCC 11325、ATCC 13332、ATCC 13333和Melon;2—1较差 作者认为,目前仍可依照Dawso (1942)、在烟草Nicotiana tabacum离体根培养 得出的结论 按生长率、干重和分枝数作为建立转化毛状根无性系的初选形态学标准.     

高大鬼针草的形态与显微鉴定

运用分子、植物形态以及显微鉴别的方法对高大鬼针草进行研究。通过测定DNA的ITS序列确定高大鬼针草为鬼针草属,并对其形态学以及茎、叶的显微鉴别特征进行详细描述和记录。该研究建立的方法为高大鬼针草的鉴定以及开发利用提供了参考依据。     

鬼针草和肿柄菊提取物对小白菜种子萌发的化感作用

采用培养皿滤纸法,研究入侵植物鬼针草和肿柄菊的水浸液和菌浸液对小白菜种子萌发的化感作用。研究结果表明:不同浓度的鬼针草、肿柄菊浸提液对同种受体植物种子萌发的化感效应不同,随着浸提液浓度的增大,其抑制作用越明显,其中鬼针草菌提液抑制效应最强。在根系生长方面,2种植物浸提液表现出低浓度促进、高浓度抑制效应,最适促进质量浓度为5 g·L-1,化感指数为047～058;最大抑制质量浓度为100 g·L-1,化感指数为-1。在株高生长方面,处理质量浓度25～50 g·L-1对小白菜株高的促进作用随浓度的增大而递增,其中肿柄菊菌浸液对株高的促进作用最强。可见,2种入侵菊科植物不同浸提液处理对同种植物不同器官的化感作用存在差异。小白菜发芽指数结果表明,4种浸提液化感抑制的强弱依次为:鬼针草菌浸液>肿柄菊水浸液>肿柄菊菌浸液>鬼针草水浸液。     

固体菌质中灰树花多糖含量的测定

目的：测定固体菌质中灰树花多糖的含量．方法：用苯酚-硫酸法，于488nm波长处测定其含量，结果：固体菌质中灰树花多糖含量平均为17．50％．结论：本方法简便、准确，易于推广。