HPLC法测定穿心莲中穿心莲内酯和脱水穿心莲内酯的含量

[目的]建立对穿心莲中穿心连内酯和脱水穿心莲内酯进行同时测定的HPLC法。[方法]采用色谱柱Alltech C18（4．6mm×250mm,5μm）;以甲醇-水（54：46）为流动相;流速1．0mL／min;检测波长225nm;柱温35℃。[结果]穿心莲内酯及脱水穿心莲内酯的线性范围分别为0．0338～2．16μg（r=0．9992）和0．0325～2．08μg（r=0．9996）,平均回收率分别为98．7％、96．5％。[结论]该方法准确、简便、灵敏度高、重复性好,适用于穿心莲中穿心莲内酯和脱水穿心莲内酯的同时测定。     

Effect of andrographolide on osteosarcoma 143B cells and its mechanism

AIM To investigate the inhibitory effect of andrographolide （AG） on human osteosarcoma 143B cells and its underlying molecular mechanism. METHODS Osteosarcoma 143B cells were cultured in vitro and treated with AG at different concentrations (0~20 μmol/L), and the effect of AG on the proliferation of 143B cells was determined by crystal violet staining, MTT assay and colony formation assay. The wound-healing assay was performed to detect the migration ability of osteosarcoma 143B cells. Transwell assay was performed to analyze the invasive capacity of osteosarcoma 143B cells. The effect of AG on the apoptosis of osteosarcoma 143B cells was detected by Hoechst 33258 staining and flow cytometry. After treatment with of AG at different concentrations, the protein levels of the molecules related to proliferation, migration, invasion and apoptosis of osteosarcoma 143B cells were determined by Western blot. The expression of β-catenin and its related molecule c-Myc in the Wnt signaling pathway was analyzed by Western blot. RESULTS Compared with blank group, the proliferation, migration and invasion of osteosarcoma 143B cells in AG treatment group were significantly inhibited (P<0.05) in a concentration-dependent manner. The expression levels of invasion- and migration-related proteins matrix metalloproteinase-9 (MMP-9), vimentin and Snail were all down-regulated (P<0.05). AG also increased the expression of antiapoptotic protein Bcl-2, and the levels of apoptosis-related protein caspase-3 was decreased but cleaved caspase-3 was increased. At the same time, the expression levels of Wnt/β-catenin signaling pathway related proteins β-catenin and c-Myc were significantly decreased (P<0.05). CONCLUSION Andrographolide may inhibit the proliferation, migration, and invasion of osteosarcoma 143B cells and promote their apoptosis by inhibiting the activity of Wnt signaling pathway.     

Growth-inhibitory and apoptosis-inducing effects of andrographolide on acute lymphoblastic leukemia cells

AIM To explore the effect of andrographolide (AND) on the growth and apoptosis of acute lymphoblastic leukemia （ALL） cells. METHODS CCK-8 assay was used to assess the viability of human acute lymphoblastic leukemia CEM-C1 cells treated with AND for 12 h, 24 h and 48 h. The cell morphological changes were observed by Wright-Giemsa staining. The cell apoptosis was detected by flow cytometry with annexin V-FITC/propidium iodide (PI) staining, and the cell cycle was examined by flow cytometry with PI staining. The expression levels of apoptosis-related proteins were examined by Western blot. The mitochondrial membrane potential (MMP) of the cells was determined by JC-1 assay. RESULTS The results of CCK-8 assay indicated that AND inhibited the viability of CEM-C1 cells in a dose- and time-dependent manner. After administration of AND for 24 h, CEM-C1 cells shrank, the cytoplasm turned red and the cell numbers were significantly reduced. Incubation of AND for 24 h resulted in G₂-phase arrest and apoptosis. Treatment with AND for 24 h increased the protein levels of cleaved caspase-3, cleaved caspase-7 and Bax, and down-regulated Bcl-2 in the CEM-C1 cells (P<0.05). The ratios of cleaved caspase-3/caspase-3, cleaved caspase-7/caspase-7 and Bax/Bcl-2 were elevated with the increase in the concentration of AND. Collapsed MMP in CEM-C1 cells was observed after AND administration for 24 h. Treatment with AND in vivo suppressed the growth of the xenograft tumor and increased the protein level of cleaved caspase-3. CONCLUSION Andrographolide exerts growth-inhibitory and apoptosis-inducing effects on ALL cells both in vitro and in vivo.     

穿心莲高产高效施肥技术研究

通过田间试验研究不同肥料配施对穿心莲生长和内酯含量的影响。结果表明：增施钾磷肥或有机肥的平衡施肥对穿心莲的生长有明显的促进作用。不同氮磷钾和有机肥配施处理较仅施氮肥的对照产量增加16.85%-26.67%,平均增产22.07%。穿心莲内酯含量增长0.70-8.21个百分点,相对提高2.82%-33.08%;经济收入提高5202-8237元/hm2,施肥产投比为4.50-8.22∶1。     

穿心莲内酯对小鼠呼吸道免疫功能的影响

通过给肺炎模型小鼠腹腔分别注射高剂量(200mg·kg-1·d-1)和低剂量(100mg·kg-1·d-1)的穿心莲内酯,7d后,测定小鼠免疫器官和肺泡灌洗液(BALF)中相关免疫指标的变化,探讨穿心莲内酯对呼吸道免疫功能的影响.结果表明,穿心莲内酯高剂量组小鼠的脾和胸腺指数、支气管肺泡灌洗液(BALF)中分泌型免疫球蛋白A(sIgA)和干扰素-γ(IFN-γ)的含量均显著低于生理盐水组(p<0.01);而BALF中巨噬细胞吞噬率则显著升高(p<0.01).说明穿心莲内酯能增强巨噬细胞吞噬功能;并对脾、胸腺等免疫器官的生长,支气管sIgA分泌及IFN-γ表达有显著的降低,起到减缓或消退肺部炎症的作用.     

微波辅助提取穿心莲内酯的研究

在单因素试验的基础上,采用正交试验法优选穿心莲内酯微波辅助提取的最佳条件.结果表明,穿心莲内酯微波辅助提取的最佳工艺参数为:采用低火功率的微波,微波时间4 min,在60℃水浴锅中保温浸提20 min后加入活性炭脱色,pH值为2.此提取条件具有应用参考价值.     

Effect of andrographolide on oral squamous cell carcinogenesis

AIM：To study the effect of andrographolide (Andro) on oral squamous cell carcinogenesis. METHODS：Chemical-induced hamster buccal pouch cancer model was used. The tumor cell proliferation, apoptosis and microvessel density (MVD) were detected by immunohistochemical staining. RESULTS：Compared with control group, the tumor volume, MVD and tumor cell proliferation in Andro group were significantly decreased. The cell apoptotic cell number in Andro group was higher than that in control group. Caspase-3 was also activated in Andro group. CONCLUSION:Andro inhibits tumor growth by suppressing proliferation, decreasing MVD and promoting apoptosis in the hamster buccal pouch. Furthermore, Andro promotes tumor cell apoptosis through caspase-3 pathway.     

Quinone reductase (QR) inducers from Andrographis paniculata and identification of molecular target of andrographolide

In the present study, it was demonstrated that the petroleum extract of Andrographis paniculata (AP) had quinone reductase (QR) inducing activity, which might be attributed to the modification of key cysteine residues in Keap1 by Michael addition acceptors (MAAs) in it. To screen MAAs in AP, glutathione (GSH) was employed, and a LC/MS/MS method was implied. Three compounds, andrographoside, andrographolide, 14-deoxy-14,15-dehydroandrographolide were revealed could well conjugated with GSH. Then, andrographolide along with 4 new and 14 known compounds were isolated to conduct QR induction evaluation, and the CD (the concentration required to double the activity of QR) value of andrographolide is 1.43 μM. The QR induce activity of andrographolide might be attributed to its targeting multiple cysteine residues in Keap1, therefore, the alkylation of Keap1 by andrographolide was further studied and the result showed that four cysteine residues: Cys77, Cys151, Cys273 and Cys368 were alkylated, which indicated that Keap1 is a potential target for the QR induce activity of andrographolide.     

酸水法及其它方法提取穿心莲总内酯的比较

为提高以水为溶媒提取穿心莲总内酯的收率，根据穿心莲内酯可以同亚硫酸氢钠起加成反应的原理，在常温条件下，用酸水法同乙醇回流法、碱水法、乙醇冷浸法、冷水浸渍法提取穿心莲总内酯，用比色法测定其含量，结果显示酸水法提取穿心莲总内酯可达乙醇冷浸法的７５％ 以上，与乙醇回流法相当，而远远超过其它方法。酸水法可为目前以水为溶媒提取穿心莲总内酯的可行方法     

Plant growth regulators induced changes in antioxidant potential and andrographolide content in Andrographis paniculata Wall.ex Nees

In this study, 5 μM ABA and GA₃ concentration were used to study the effect of these growth regulators on the andrographolide content and antioxidant potentials of Andrographis paniculata. The growth regulators were applied by means of foliar spray during morning hours. A significant enhancement in non-enzymatic antioxidant contents was observed in all sampling days in A.paniculata plants under ABA and GA₃ treatments. Ascorbic acid and α-tocopherol content was increased significantly under the growth regulator treatments in leaves, stem and roots of A. paniculata. The activities of antioxidant enzymes such as ascorbate peroxidase, superoxide dismutase and catalase were increased by ABA and GA₃ treatments in the leaves, stem and roots of A. paniculata plants. The HPLC analysis was used to quantify the andrographolide content in control and growth regulator treated plants. The growth regulators ABA and GA₃ treated plants showed increased contents of andrographolide when compared to control.