刺五加叶中金丝桃苷的薄层鉴别和含量测定

目的：以金丝桃苷为指标成分,建立刺五加叶的薄层定性和含量测定的方法。方法：采用薄层色谱法对刺五加叶中金丝桃苷进行定性鉴别,并用高效液相色谱法进行含量测定,色谱柱：Kromasil 100-5C18色谱柱（4.6mm×250mm）;流动相：甲醇-0.1％冰醋酸水溶液(35：65);流速：0.8mL/min;检测波长：360nm;柱温：30℃;进样量：10μL。结果：刺五加叶金丝桃苷的薄层鉴别特征明显,金丝桃苷在0.13～0.78μg范围内具有良好线性关系,r=0.9994,平均回收率为98.17％,RSD为1.02％（n=5）。结论：此方法简便、快速、准确,可用于刺五加叶的质量控制。     

The apoptotic and autophagic properties of two natural occurring prodrugs,hyperoside and hypoxoside,against pancreatic cancer cell lines

Pancreatic cancer is only the 12th most common cancer, but the fourth leading cause of cancer-related deaths in the world. This is due to late prognosis, poor response to chemotherapy and early metastases. Natural prodrugs may play an important role in the treatment of pancreatic cancer. The main aim of this study was to determine the cytotoxicity of five natural prodrugs, namely harpagoside, hyperoside, hypoxoside, oleuropein and polydatin, by investigating apoptosis and autophagy as possible mechanism/s of action. Hypoxoside and hyperoside have shown selective cytotoxicity at IC50 values of ∼25 and 50 μM against INS-1 and MIA PaCa-2 pancreatic cancer cells, respectively. Hypoxoside and hyperoside induced G2/M phase arrest and caspase-3 activation in INS-1 and MIA PaCa-2 cells, respectively. Hoechst/phalloidin staining confirmed morphological changes, including condensed chromatin multinucleation, membrane blebbing and loss of cytoskeletal arrangement in INS-1 and MIA PaCa-2 cells. Acridine orange staining was absent in INS-1 (hypoxoside) and MIA PaCa-2 (hyperoside) treated cells, whereas LC3B expression was not significantly increased. INS-1 and MIA PaCa-2 treated cells favour the cell death pathway, apoptosis, over the cell survival pathway, autophagy.     

Ameliorating effects of constituents from Cortex Acanthopanacis Radicis on memory impairment in mice induced by scopolamine

OBJECTIVE：To investigate the effects of active con- stituents extracted from Cortex Acanthopanacis Radicison improving the impaired memory in mice models. METHODS： The mice models of memory impair- ment were established using scopolamine. Amelio- rating effects of the fractions and constituents on scopolamine-induced memory impairment in vivo were investigated using passive avoidance and Morris water-maze task tests, and their anti-acetyl- cholinesterase（AChE） and antioxidant activities in vitro examined. The isolation of constituents was performed by chromatographic methods and their structures were identified on the basis of instru- mental analysis. RESULTS： Among the fractions tested, ethylacetate fraction exhibited the anti-AChE activity（25.83%± 0.23%） properly and excellent 2,2-diphenyl-1-picryl- hydrazyl（DPPH） radical and superoxide anion scav- enging capacity（87.50% ± 0.83% and 60.22% ± 0.43%, respectively）. However, the methylene chlo- ride fraction was much more active than the ethyl- acetate fraction in the passive avoidance task test（167.5% increase of step-through latency time） and Morris water-maze task test（33.3% decrease of es-cape latency time）. Four constituents, β-sitosterol, stigmasterol, sesamin, and hyperin were isolated from the methylene chloride fraction, among them, hyperin showed anti-acetylcholinesterase and anti-oxidant activities remarkably. Moreover, hyperin exerted a potent effect（146±38） s on mem- ory improvement in terms of passive avoidance task test compared with the reference compound tacrine（162±43） s at a dose of 2.5 mg/kg. CONCLUSION： Hyperin, a flavonoid glucoside iso- lated from Cortex Acanthopanacis Radicis, inhibited AChE activity and potently ameliorated scopol- amine-induced memory impairment, and its action may be partially mediated by the acetylcholine-en- hancing cholinergic nervous system.     

Administration with hyperoside sensitizes breast cancer cells to paclitaxel by blocking the TLR4 signaling

Breast cancer is a malignancy and one of the most frequent causes of cancer death among women worldwide. Paclitaxel is a common chemotherapeutic drug and has recently been shown to facilitate tumor cell escape during cytotoxic chemotherapy by inducing inflammatory mediators and pro-survival protein expression. Hyperoside is a flavonoid glycoside compound and exerts anti-inflammation, and anti-tumor growth properties. However, its function in breast cancer chemosensitivity remains poorly elucidated. In this study, hyperoside exhibited little cytotoxicity to normal human breast mammary epithelial cell lines, and also protected against paclitaxel-induced cytotoxicity in MCF-10A. Importantly, treatment with hyperoside engendered not only inhibition of cell viability, but also potentiated cancer cell sensitivity to paclitaxel in TLR4-positive breast cancer MDA-MB-231 cells by suppressing cell viability, and increasing cell apoptosis and caspase-3 activity. Nevertheless, although hyperoside exposure restrained cell viability, its treatment presented little effects to paclitaxel sensitivity in TLR4-null HCC1806 cells. Intriguingly, paclitaxel stimulation activated the TLR4-NF-κB signaling, which was reversed after hyperoside administration. Concomitantly, hyperoside also attenuated paclitaxel-mediated anti-apoptotic Bcl-2 expression, but enhanced the effects of paclitaxel on pro-apoptotic Bax expression, and pro-inflammatory cytokine IL-6 and IL-6 levels in MDA-MB-231 cells. Importantly, restoring the TLR4 pathway overturned hyperoside-evoked chemosensitivity to paclitaxel in MDA-MB-231 cells. Thus, hyperoside may elevate breast cancer cell sensitivity to paclitaxel by blocking TLR4 activation-mediated pro-inflammatory and pro-survival approaches, thereby endorsing its usefulness as a promising therapeutic combination to overcome chemosensitivity in breast cancer.     

复方臭灵丹合剂质量标准研究

目的建立复方臭灵丹合剂的质量标准。方法采用TLC对桔梗和鱼腥草进行定性鉴别,依据《云南省中药材(民族药材)质量标准研究技术指导原则(试行)》对其进行分析方法验证及重现性考察。采用HPLC对合剂中的洋艾素和金丝桃苷进行定量分析,同时进行相关方法学的考察。结果桔梗和鱼腥草TLC鉴别色谱图中,供试品与对照药材相应位置上均显相同颜色的特征斑点,阴性无干扰,且分析方法验证与重现性考察均合格。洋艾素和金丝桃苷分别在2.59~55.47μg/mL和4.02~51.68μg/mL范围内线性关系良好,回归方程分别为Y₁=43.24X-12.39(R=0.9996)和Y₂=63.82X-30.04(R=0.9994),准确度试验中,平均加样回收率分别为100.06%和100.14%,RSD值分别为2.36%和2.74%(n=9),其余方法学考察均合格,洋艾素和金丝桃苷的平均含量分别为2.41μg/mL和9.95μg/mL。结论建立的质量标准稳定可行。     

金丝桃苷、淫羊藿苷在大鼠附睾精子冷冻保存中的作用

目的：本研究旨在观察金丝桃苷、淫羊藿苷在大鼠精液冷冻保存过程中的应用效果,并对大鼠精液冷冻保存体系进行优化。方法：将金丝桃苷、淫羊藿苷加入大鼠精液冷冻保护剂中,冷冻保存大鼠附睾尾部的精子,并评估冷冻前后精子活率指数,采用伊红染色评估精子畸形率。结果：添加金丝桃苷、淫羊藿苷的冷冻保护剂可以提高解冻后的精子活率指数。结论：金丝桃苷、淫羊藿苷在大鼠精液冷冻保存过程中有一定的保护作用。     

红旱莲不同部位金丝桃苷含量测定

目的 比较了红旱莲药材不同部位（茎、叶、花、果）中金丝桃苷的含量差异。方法 采用高效液相色谱法进行测定,采用C₁₈（4.6 mm×250 mm,5 μm）色谱柱,乙腈-0.1%磷酸为流动相,360 nm为检测波长,流速为1.0 mL·min^-1,柱温30 ℃。结果 金丝桃苷的线性范围为2.904～58.08 μg·mL^-1,相关系数r=0.999 8,平均回收率为 95.1%,RSD为1.3%;红旱莲药材不同部位（茎、叶、花、果）中金丝桃苷的含量分别为0.405、3.221、2.260、0.371 mg·g^-1。结论 红旱莲中不同部位中金丝桃苷含量具有显著差异,叶中含量最高。     

金丝桃苷下调微小 RNA-199a对脂多糖诱导的人肺泡上皮细胞的影响

目的 探讨金丝桃苷对脂多糖(LPS)诱导的人肺泡上皮细胞(HPAEpiC)的影响及分子机制.方法 将HPAEpiC分为对照组、模型组、金丝桃苷低、中、高剂量药物组、anti-miR-NC组、anti-miR-199a组、高剂量药物组+miR-NC组、高剂量药物组+miR-199a组.四甲基偶氮唑盐比色法(MTT)检测细胞增殖;流式细胞术检测细胞周期和细胞凋亡情况;酶联免疫吸附试验(ELISA)检测肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)水平;实时荧光定量PCR(RT-qPCR)检测miR-199a表达水平;蛋白质印迹(Western blotting)法检测裂解的半胱天冬蛋白酶3(Cleaved-caspase3)、半胱天冬蛋白酶3前体(pro-caspase3)的蛋白表达.结果 金丝桃苷处理后,LPS诱导的HPAEpiC中细胞OD值升高,金丝桃苷低、中、高剂量药物组OD值分别为(0.49±0.02)、(0.60±0.03)、(0.72±0.03),G0/G1期细胞所占比例降低,S期细胞所占比例升高,细胞凋亡率降低,TNF-α、IL-1β水平降低,miR-199a表达水平降低,Cleaved-caspase3表达水平降低,pro-caspase3表达水平升高,且呈剂量依赖性(P<0.05);抑制miR-199a表达可抑制LPS诱导的HPAEpiC细胞凋亡和炎症因子的释放.且miR-199a过表达逆转了金丝桃苷提取物对LPS诱导的HPAEpiC细胞凋亡和炎症因子的作用.结论 金丝桃苷可能通过下调miR-199a抑制LPS诱导的HPAEpiC细胞凋亡和炎症反应.     

不同采收期和部位黔产千里光中总黄酮、绿原酸和金丝桃苷含量动态比较

[目的]研究不同采收期及部位黔产千里光中总黄酮、绿原酸和金丝桃苷含量的动态变化规律,结合生物产量,为确定黔产千里光的采收期和药用部位提供科学理论依据。[方法]采用紫外分光光度法测定总黄酮,高效液相色谱法测定绿原酸和金丝桃苷的含量。[结果]不同采收期黔产千里光总黄酮的含量以6月份含量最高,绿原酸和金丝桃苷含量均以8月份最高,不同部位千里光叶中总黄酮、绿原酸和金丝桃苷的含量最高;8月份千里光的生物产量最大,茎、叶的产量占较大比重。[结论]综合考虑3种活性成分的含量和生物产量,建议黔产千里光在7—9月份采收地上部位为宜。     

金莲花不同植株部位的金丝桃苷含量比较

目的:测定金莲花中金丝桃苷的含量。方法:采用80%乙醇提取总黄酮,HPLC法测定金莲花不同部位中金丝桃苷的含量。色谱柱为Phenomenex Gemini C18柱(4.6 mm×250 mm,5μm);流动相为乙腈-0.05%磷酸溶液(V/V,18∶82);流速为1.0 ml/min;柱温为25℃;检测波长为360 nm;进样量为10μl。结果:金丝桃苷在0.04~0.56μg范围内与峰面积呈良好的线性关系,相关系数为0.9997(n=6);其中茎、叶、籽中金丝桃苷的含量分别为0.0279,0.3868,0.0016 mg/g。结论:金莲花不同部位中金丝桃苷的含量分布为:叶﹥茎﹥籽,上述结果可为金莲花的综合利用提供科学依据。