Design of Anti-ischemic drug based on their effects in liver mitochondria of the turtle as an anaerobiosis model.

我们设计了以下化合物：1)水杨酰苯胺类化合物，2)硫代二唑类化合物，3)维生素K_1环氧化合物，4)类固醇和三萜的硫酸二糖苷类化合物。甘草酸),5)二糖（麦芽糖和异麦芽糖）硫酸盐，6)环糊精-类固醇复合物作为抗缺血性药物的候选物。重点研究了噻二唑类新化合物TX-101-TX-109的合成及其对龟、大鼠肝线粒体呼吸活性的影响和小鼠体内抗缺血活性的影响。我们经常使用该基金购买的高效液相色谱紫外检测器检测我们开发的合成化合物进行分离，分析和纯化。作为生物实验，我们以淡水龟（Mauremys japonica和Chineinys reevesii）为厌氧菌，建立了抗缺血性药物研究的动物模型。从冬眠的海龟肝脏中分离到肝脏线粒体，其呼吸控制指数约为3。线粒体功能（耗氧量和ATP合成）在我们新开发的细胞中通过高灵敏度传感器（如氧、pH和膜电位）进行测量。TX-109强烈解耦线粒体氧化磷酸化[25nM(Vox max: 14natoms O/min/mg protein)]。与大鼠相比，龟肝线粒体的解偶联活性浓度相似，但其在该状态下的耗氧速率较慢[TX-109: 45nM(Vox max: 340natoms O/min/mg protein)]。这些硫代二唑衍生物TX-101-109在浓度为l00mg/kg / p时，在体内表现出较弱的抗缺血活性。我们试图利用乌龟活线粒体建立动物模型，并将其应用于抗缺血药物的开发。

We designed the following compounds : 1)salicylanilides, 2)thiodiazoles, 3)vitarmin K_1 epoxides, 4)disacchatide sulfates of steroids and triterpenoids(eg. glycyrrhizinic acid), 5)disaccharide(maltose and isomaltose)sulfates, 6)cyclodextrin-steroid complex as candidates of anti-ischemic drugs. Especially we studied the syntheses of new thiadiazole compounds such as TX-101-TX-109, their efftects against the respiratory activity in liver mitochondm in turtle as well as rat, and their in vivo anti-ischemic activity in mouse. We often used the UV detector for HPLC purchased by this grant detect our developed synthetic compounds for their isolation, analysis, and purification. As bioassay, we developed the aninmal model of anti-ischemic drug research using freshwater turtles(Mauremys japonica and Chineinys reevesii)as an animal anaerobe. The liver mitochondria were isolated from liver in turtle which was under hibernation, the RCI(Respiratory Control Index)of which was ca. 3. The mitochondrial function(oxygen consumption and ATP synthesis)was measured in our newly developed cell connected with high-sensitive sensors such as oxygen, pH, and membrane potential. TX-109 uncoupled the mitochondrial oxidative phosphorylation strongly[25nM(Vox max : 14natoms O/min/mg protein)]. This concentration of uncoupling activity in turtle liver mitochondria was similar, but its rate of oxygen consumption at this state was slower, in comparison with those in rat[TX-109 : 45nM(Vox max : 340natoms O/min/mg protein)]. These thiodiazole derivatives TX-101-109 showed weak anti-ischemic activities in vivo at the concentration of l00mg/kg i. p. We approached to develop the animal model using turtle live mitochondria and apply to the development of antiischemic drugs.