Heo H J, Yang H C, Cho H Y, Hong B, Lim S T, Park H J, Kim K H, Kim H K, Shin D H
Graduate School of Biotechnology, Korea University, Seoul.
Mol Cells. 2000 Jun 30;10(3):253-62.
We screened 42 Korean traditional tea plants to determine the inhibitory effect of acetylcholinesterase and attenuation of toxicity induced by amyloid-beta peptide, which were related to the treatment of Alzheimer's disease (AD). The methanolic extract from Artemisia asiatica among tested 42 tea plants, showed the highest inhibitory effect (48%) on acetylcholinesterase in vitro. The methanolic extract was further separated with n-hexane, chloroform, and ethyl acetate of water, in order. The chloroform solubles, which were high in inhibitory effect of acetylcholinesterase, were repeatedly subjected to open column chromatography on silica gel. From the highest inhibitory fraction (78%) on acetylcholinesterase, the single compound was obtained by the Sep-Pak Cartridge (C18: reverse phase column). This compound was found to react positively on Dragendorff's reagent (potassium bismuth iodide), which typically reacted with the alkaloid. This compound was purified by HPLC (mu-bondapack C18 reverse phase column: 3.9 x 150 mm). The IC50 (the concentration of 50% enzyme inhibition) value of this compound was 23 micrograms/ml and the inhibitory pattern on acetylcholinesterase was mixed with competitive/non-competitive type. We examined the effects of this compound on toxicity induced by A beta (25-35) in rat pheochromocytoma PC12 cells. Pretreatment of the PC12 cells for 2 h with an alkaloid of Artemisia asiatica (1200 microg/ml) reduced the toxicity induced by A beta. This study demonstrated that an alkaloid of Artemisia asiatica, which was metabolized to small molecule in digestive tract and then could pass through the blood-brain barrier, appeared to be an acetylcholinesterase inhibitor with a blocker of neurotoxicity induced by A beta in human brain causing Alzheimer's disease.
我们筛选了42种韩国传统茶树，以确定其对乙酰胆碱酯酶的抑制作用以及对β-淀粉样肽诱导的毒性的减弱作用，这些都与阿尔茨海默病（AD）的治疗相关。在测试的42种茶树中，亚洲龙蒿的甲醇提取物在体外对乙酰胆碱酯酶显示出最高的抑制作用（48%）。该甲醇提取物依次用水、正己烷、氯仿和乙酸乙酯进一步分离。对乙酰胆碱酯酶抑制作用较高的氯仿可溶物反复进行硅胶柱层析。从对乙酰胆碱酯酶抑制率最高的部分（78%），通过Sep-Pak柱（C18反相柱）获得单一化合物。发现该化合物与碘化铋钾试剂（Dragendorff试剂）呈阳性反应，该试剂通常与生物碱发生反应。该化合物通过高效液相色谱（μ-bondapack C18反相柱：3.9×150mm）进行纯化。该化合物的IC50（50%酶抑制浓度）值为23微克/毫升，对乙酰胆碱酯酶的抑制模式为竞争性/非竞争性混合型。我们研究了该化合物对大鼠嗜铬细胞瘤PC12细胞中Aβ（25-35）诱导的毒性的影响。用亚洲龙蒿生物碱（1200微克/毫升）对PC12细胞预处理2小时可降低Aβ诱导的毒性。本研究表明，亚洲龙蒿生物碱在消化道代谢为小分子后可穿过血脑屏障似乎是一种乙酰胆碱酯酶抑制剂，可阻断人脑中由Aβ诱导的导致阿尔茨海默病的神经毒性。
