葛根中异黄酮含量的薄层光密度法测定

本文报道了葛根中异黄酮成分含量的薄层光密度测定法。用甲苯—甲醇—10%甲酸(7:3:0.02)和乙酸乙酯—甲醇—50%甲酸(8:2:0.2)为展开剂,在硅胶G薄层上分离了大豆甙元、大豆甙、葛根素和大豆甙元-4′,7-二葡萄糖甙,并用CS-910双波长薄层扫描仪进行了定量测定。变异系数为1.5～1.6%,采用本法测定了生药和片剂样品的含量。     

野葛中的一个新化学成分

应用色谱技术研究豆科植物野葛(Pueraria lobata (Willd.) Ohw)的化学成分，从中分离到5个化合物，通过波谱方法解析它们的结构，分别为：葛根苷D(1)，4′,8-二甲氧基-7-O-β-D-葡糖基异黄酮(2)，二十烷酸(3)，十六烷酸(4)和二十四烷酸-α-甘油酯(5)。化合物1为新化合物，化合物2、3和4为首次从葛根中分离得到。     

Chemistry of the Chinese herbal medicine Puerariae Radix (Ge-Gen): A review

Puerariae Radix (Ge-Gen in Chinese) is a popular traditional Chinese herbal medicine derived from the dried roots of Pueraria lobata (Willd.) Ohwi or P. thomsonii Benth. It shows significant pharmacological activities in the treatment of cardiovascular diseases, hypertension, and diabetes. This review summarizes progress in the chemistry study of Pueraria species, mainly covering the classes of flavonoids, puerosides, coumarins, triterpenoids, and triterpene saponins. Qualitative and quantitative chemical analyses are also covered.     

Mechanism of phytoestrogen puerarin-mediated cytoprotection following oxidative injury: estrogen receptor-dependent up-regulation of PI3K/Akt and HO-1

Phytoestrogens are polyphenolic non-steroidal plant compounds with estrogen-like biological activity. The phytoestrogen puerarin, the main isoflavone glycoside found in the root of Pueraria lobata, has been used for various medicinal purposes in traditional Chinese medicines for thousands of years. Recent studies have indicated that the estrogen receptor (ER), through interaction with p85, regulates phosphoinositide 3-kinase (PI3K) activity, revealing a physiologic, non-nuclear function of ER that may be relevant in cytoprotection. In this study, we demonstrate that the phytoestrogen puerarin inhibits tert-butyl hydroperoxide (t-BHP)-induced oxidative injury via an ER-dependent Gβ1/PI3K/Akt and heme oxygenase-1 (HO-1) pathway. Pretreatment of Hepa1c1c7 and HepG2 cells with puerarin significantly reduced t-BHP-induced caspase-3 activation and subsequent cell death. Also, puerarin up-regulated HO-1 expression and this expression conferred cytoprotection against oxidative injury induced by t-BHP. Moreover, puerarin induced Nrf2 nuclear translocation, which is upstream of puerarin-induced HO-1 expression, and PI3K activation, a pathway that is involved in induced Nrf2 nuclear translocation, HO-1 expression and cytoprotection. Puerarin-induced up-regulation of HO-1 and cytoprotection against t-BHP were abolished by silencing Nrf2 expression with specific siRNA. Also, puerarin-mediated increases in PI3K activation and HO-1 induction were reversed by co-treatment with ICI 182,780 and pertussis toxin. Taken together, these results suggest that puerarin augments cellular antioxidant defense capacity through ER-dependent HO-1 induction via the Gβ1/PI3K/Akt-Nrf2 signaling pathway, thereby protecting cells from oxidative stress.     

葛根素干预血管内膜胰岛素抵抗的应用前景

葛根（Radix Puerariae）为豆科植物野葛Pueraria lobata（Willd.）Ohwi的干燥根。葛根素（Puerarin）是葛根的主要活性成分,现代药理研究表明葛根素具有调节血管张力,抗炎,抗氧化,调节糖脂代谢等药理作用。而葛根素对心血管缺血性损害,糖尿病及其并发症等疾病的治疗作用已经有一定的临床研究基础,本文讨论了葛根素对血管内膜的保护作用,以此提示葛根素干预血管内膜胰岛素抵抗的可能途径及应用前景。     

HPLC测定丙戊酸钠注射液中丙戊酸钠的含量

目的建立测定丙戊酸钠注射液中丙戊酸钠含量的高效液相色谱法。方法采用ZORBAX Extent-C18（4.6 mm×250 mm,5 μm)色谱柱,甲醇-0.05 mol·L-1磷酸二氢钾缓冲液（80∶20）用10%磷酸调pH值至5.0±0.05为流动相,检测波长为210 nm。结果丙戊酸钠在0.932 2～9.322 3 mg·mL-1内浓度与峰面积呈良好的线性关系,r=0.999 9(n=6);平均回收率为100.37%,RSD为0.8%（n=9)。结论该方法准确可靠,重复性好,操作简便快速,可用于丙戊酸钠注射液中丙戊酸钠的含量测定。     

葛根及北豆根水提物对正常小鼠血液流变学作用的比较

目的比较葛根及北豆根水提物对小鼠血液流变学的作用。方法实验一小鼠分别给予浓度为0.25kg/L和1.0kg/L的葛根水提物,实验二小鼠分别给予浓度为0.25kg/L和1.0kg/L的北豆根水提物灌胃给药,连续给药3周,并设生理盐水组和维生素E组作为对照。血细胞分析仪进行常规血细胞测定,荧光偏振法测定红细胞膜流动性和微黏度。结果葛根提取物可轻微降低小鼠白细胞数量及红细胞数量,轻微增加红细胞膜脂流动性,降低微黏度,但作用不显著;北豆根提取物可使平均血细胞容积显著降低,血小板数显著升高,低浓度北豆根可显著增加红细胞数,高浓度北豆根可显著降低白细胞数;低浓度北豆根可显著增加红细胞膜流动性,降低微黏度,而高浓度北豆根的作用不明显。结论一定浓度的北豆根水提物可以增加红细胞膜流动性,从而降低血液黏度,改善血液流变学指标;葛根水提物的作用不明显。     

葛根素注射液致急性血管内溶血的临床特征和发病机制及其安全应用

葛根素是从豆科植物野葛或甘葛藤干燥根中提出的一种异黄酮类化合物,其注射液用于心脑血管疾病的治疗。近年来葛根素注射液引起急性血管内溶血的报道增多,经检索国内相关文献,自1993年至2005年间应用葛根素注射液引起血管内溶血的患者共32例,男21例,女11例,平均年龄（69±9）岁;其中有10例死亡,平均年龄为（72±5）岁。在资料完整的25例患者中,21例单用葛根素注射液,4例为联合用药;葛根素100～700mg溶于0.9%氯化钠或5%葡萄糖注射液中静脉滴注;用药时间22例为5～10d,3例〉10d。血管内溶血的临床表现主要为腰、背痛,血尿、尿少、血红蛋白尿,网织红细胞升高（0.05～0.30）,高胆红素血症（29.1～123.2μmol/L）。22例患者经停药并给予糖皮质激素或抗组胺药物以及血液透析治疗后,多于6～7d症状消失或病情稳定。葛根素注射液致血管内溶血的可能因素为葛根素注射液的质量、患者过敏体质、年龄、用药时间或联合用药等。其发生机制可能与葛根素注射液引起的免疫性或非免疫性反应及其促氧化作用等有关。预防措施为：有过敏史者不宜应用;高龄及肝肾功能不全者用药应调整剂量;与可能致溶血的药物联用应谨慎。     

罗布麻、葛根提取物对神经刺激所致猪脑动脉反应的影响

目的观察罗布麻、葛根提取物对神经刺激引起的猪脑动脉反应的影响及其与一氧化氮的关系。方法采用离体去除内皮血管方法,测定脑动脉螺旋条的等容张力变化及药物的作用。结果罗布麻、葛根提取物对电刺激或化学刺激引起的血管舒张反应具有增强作用,此作用不受酚妥拉明和哌唑嗪、心得安、阿托品、引哚美辛的影响,但可被一氧化氮合酶抑制剂L-硝基精氨酸抑制,该抑制作用又可为一氧化氮供体L-精氨酸逆转。结论罗布麻、葛根提取物对电刺激或化学刺激所致血管舒张反应的增强作用可能与一氧化氮有关。     

The Neuroprotection of puerarin against cerebral ischemia is associated with the prevention of apoptosis in rats

Previous work has shown that puerarin (Pur), extracted from the dried root of Pueraria lobata (Wild) Ohwi, increases cerebral blood flow in dogs and attenuates cerebral and spinal cord injury resulting from ischemia and reperfusion in rats and rabbits. The present study further demonstrates the neuroprotective effects of Pur on cerebral ischemic injury in rats and the mechanisms underlying the protective effects. Male Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAo) for 50 min followed by reperfusion for 24 h. Pur (50, 100 mg/kg, i.p) was administered at the onset of MCAo. Twenty-four hours after reperfusion, neurological deficits were evaluated in Pur- and vehicle-treated rats. The infarct volume and edema ratios were assessed from stained brain slices. The results showed that Pur (100 mg/kg) markedly decreased the infarct volume by 34 % ( P < 0.01) in cerebral cortex and improved the neurological functions ( P < 0.05) after MCAo. Furthermore, flow cytometric analysis of annexin-V and PI labeling cells showed that the percentages of apoptosis and necrosis in the dorsolateral cortex were significantly reduced by 38.6 % and 28.5 % ( P < 0.01 and P < 0.05) following treatment with Pur (100 mg/kg) in MCAo rats. Caspase-3 activity, a biochemical marker of apoptosis, was significantly inhibited after treatment with Pur in the dorsolateral cortex. In agreement with this result, the expression of the X-chromosome-linked inhibitor of apoptosis protein (XIAP) was obviously up-regulated after administration of Pur (100 mg/kg), while caspase-3 gene was down-regulated in the dorsolateral cortex. These results suggest that the neuroprotection of puerarin against cerebral ischemia is associated with anti-apoptosis.     

Molecular authentication of Radix Puerariae Lobatae and Radix Puerariae Thomsonii by ITS and 5S rRNA spacer sequencing

In the present study, we examined nuclear DNA sequences in an attempt to reveal the relationships between Pueraria lobata (Willd). Ohwi, P. thomsonii Benth., and P. montana (Lour.) Merr. We found that internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA are highly divergent in P. lobata and P. thomsonii, and four types of ITS with different length are found in the two species. On the other hand, DNA sequences of 5S rRNA gene spacer are highly conserved across multiple copies in P. lobata and P. thomsonii, they could be used to identify P. lobata, P. thomsonii, and P. montana of this complex, and may serve as a useful tool in medical authentication of Radix Puerariae Lobatae and Radix Puerariae Thomsonii.     

三维HPLC法同步测定犬血浆中的葛根素及阿魏酸

以对 羟基苯甲酸为内标 ,甲醇 水 醋酸 (5 6 2 5mol/L ) (39∶5 8∶3 ,v/v)为流动相 ,InertsilODS 3色谱柱 (15 0mm× 4 6mm ,5 μm)为固定相 ,流速 0 9mL/min。以二极管阵列检测器同时获取 2 48nm和 32 1nm两个波长下的数据 ,并通过三维图观察其紫外吸收定性 ,以沸水浴法处理血浆样品。结果 :葛根素和内标及阿魏酸分离完全 ,葛根素在 0 2 35 4～ 1 883μmol/L范围内线性关系良好 (r =0 9943) ,变异系数 <10 % ,平均回收率为 10 1 0 2 % ,最低检测限为 0 33ng ,最低检测浓度为 0 0 396 μmol/L;阿魏酸则在 1 0 341～ 5 35 6 μmol/L范围内线性关系良好 (r =0 9942 ) ,变异系数 <10 % ,平均回收率为 10 0 88% ,最低检测限为 0 45ng ,最低检测浓度为1 115 9μmol/L。此法可用于不同波长下同步以内标法测定血浆中两种或多种成分     

Puerarin 6″-O-xyloside suppresses growth,self-renewal and invasion of lung cancer stem-like cells derived from A549 cells via regulating Akt/c-Myc signalling

Cancer stem cells have been identified as the major cause of cancer initiation and progression. To investigate the effects of puerarin 6″-O-xyloside (PXY), derived from Pueraria lobata (Willd.) Ohwi, on lung cancer stem cells, we enriched and identified a subpopulation of lung cancer stem-like cells (LCSLCs) derived from lung adenocarcinoma A549 cells with traits including high self-renewal and invasive capability in vitro, elevated tumourigenicity in vivo, and high expression of stem cell markers CD44, CD133 and aldehyde dehydrogenase 1 (ALDH1). We found that PXY could impair cell viability, suppress self-renewal and invasive capability, and decrease CD133, CD44 and ALDH1 mRNA expression in LCSLCs in a dose-dependent manner. Furthermore, we showed that PXY suppressed the self-renewal and invasive capability of LCSLCs at least in part through suppressing the activation of Akt/c-Myc signalling. In conclusion, PXY can block the traits of LCSLCs, indicating that PXY may be a candidate compound for lung adenocarcinoma therapy via eliminating LCSLCs.     

复方野葛根胶囊质量控制

建立复方野葛根胶囊的质量标准。方法：采用薄层色谱鉴别法鉴别处方中的主要药味西洋参;采用高效液相色谱法测定葛根素的含量。结果：处方中的西洋参的薄层色谱具有鉴别特征;葛根素在进样量0．102～1．020μg范围内与峰面积线性关系良好,r=0．9998,平均回收率为99．21％,RSD=0．52％。结论：复方野葛根胶囊质量可控制。     

Protective effect of Puerarin on β-amyloid-induced neurotoxicity in rat hippocampal neurons

Puerarin (CAS Number 3681-99-0), a major isoflavone glycoside purified from Pueraria lobata, was reported to possess antioxidative and estrogen-like biological activities. Recent studies showed that puerarin protects different cell types from damage caused by a variety of toxic stimuli. In the present study, we investigated the neuroprotective effect of puerarin against Aβ25-35-induced neurotoxicity in cultured hippocampal neurons, as well as the underlying mechanism(s). Following exposure of cells to Aβ25-35, cell survival and glutathione peroxidase (GSH-Px) and catalase (CAT) activities were reduced while production of reactive oxygen species (ROS) was increased. Preincubation of the cells with puerarin prior to Aβ25-35 exposure increased cell survival and GSH-Px and CAT activities and decreased ROS production. It was previously shown that overactivation of glycogen synthase kinase-3β (GSK-3β) is implicated in Aβ-induced cell death. In this study, Aβ25-35 treatment is found to increase GSK-3β activity and pretreatment with puerarin preventesAβ-induced activation of GSK-3β based on Western blot analysis. In addition, puerarin is shown to activate protein kinase B (PKB)/Akt, an important upstream kinase of GSK-3β, possibly promoting subsequent GSK-3β inhibition. Our data suggest that puerarin attenuates cell death induced by Aβ25-35 via various mechanisms, which might be beneficial for the treatment of Alzheimer's disease.     

夜香颈椎片质量标准研究

目的建立夜香颈椎片的质量标准。方法采用聚酰胺薄层色谱法对夜香颈椎片中葛根、夜香牛、透骨草进行了定性鉴别;采用分光光度法测定总黄酮的含量,为夜香颈椎片的质量控制提供依据。结果定性鉴别方法专属性强。芦丁在0.206～1.236 mg内线性良好,平均加样回收率为99.09%,RSD为1.13%。结论所建立的方法准确可行,重复性好,可有效控制夜香颈椎片的质量。     

野葛根中葛根素的提取及HPTLC测定

采用有机溶剂萃取结合酸水解的方法,直接从野葛根中分离葛根素,并用HPTLC进行纯度分析。产率为1．15％,含量为97．6％。操作简便,产品纯度和产率高,成本低。     

Emodin accelerates osteoblast differentiation through phosphatidylinositol 3-kinase activation and bone morphogenetic protein-2 gene expression

Emodin is a naturally occurring anthraquinone present in the roots and bark of numerous plants of the genus Rhamnus. Here, we identified emodin as one of compounds activating the mRNA expression of bone morphogenetic protein (BMP)-2 in the differentiation process of mouse osteoblastic MC3T3-E1 subclone 4 cells. Without any effect on cell growth, the low concentration (up to 5 microM) of emodin highly induced the mRNA expression of BMP-2, the expression of alkaline phosphatase (an early marker of osteoblast differentiation), and the mineralization. Interestingly, emodin induced the activation of phosphatidylinositol 3-kinase (PI3K), Akt and mitogen-activated protein (MAP) kinases, but those inductions by emodin were completely inhibited by the PI3K inhibitor, LY294002, suggesting that the up-regulation of BMP-2 by emodin could be mediated through the activation of both Akt and MAP kinases by activating PI3K. Additionally, emodin-induced activation of NF-kappaB suggested that NF-kappaB might be required for its anabolic activity in part. In conclusion, the use of natural compounds with anabolic activity such as emodin could have a beneficial effect on bone health and this kind of studies further elucidate the pharmacological roles of natural compounds in the prevention of osteoporosis and provide the initiative in the early drug discovery and development for osteoporosis.     

The effect of puerarin on serum nitric oxide concentration and myocardial eNOS expression in rats with myocardial infarction

Puerarin (1) is a major effective ingredient extracted from the traditional Chinese medicine Ge-gen (Radix Puerariae, RP). Recently, puerarin has been used to treat patients with coronary artery diseases (CAD). However, the mechanisms of puerarin on CAD are still not very clear. In this study, we investigated the role of puerarin on serum nitric oxide (NO) concentration, myocardial endothelial nitric oxide synthase (eNOS) gene expression, the protein expression of eNOS and inducible nitric oxide synthase (iNOS), as well as the level of protein kinase B (Akt/PKB) phosphorylation in rats with myocardial infarction. We found that puerarin (120 mg/kg/day, i.p.) could increase serum nitrite concentration in rat with myocardial ischemia (MI). It also induced the gene expression or activation of eNOS, protein expression of eNOS, and the Akt/PKB phosphorylation. From these results, we suggested that puerarin could increase serum nitric oxide level of rat with myocardial infarction, which should be one of the mechanisms of the therapeutic effect of puerarin on CAD. The increased expression of eNOS and the Akt/PKB pathway may be the underlying mechanism by which puerarin stimulates NO production.