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目的建立以γ-氨基丁酸转氨酶(GABA-T)抑制剂类抗癫痫神经系统药物筛选模型,并运用此模型对天麻有效成分及其类似物进行体外活性筛选和构效关系分析。方法优化酶催化反应温度、反应时间、底物NAD+浓度、底物α-酮戊二酸浓度、底物γ-氨基丁酸(GABA)浓度等因素建立GABA-T酶系活性筛选模型,采用对羟基苯甲醛(HBA)及其11种结构类似物验证模型可靠性。结果成功构建GABA-T酶系抗癫痫活性筛选模型,HBA及11个结构类似物测定结果与文献报道抗癫痫作用相一致,且构效分析发现-OH及苯环对位上的-CHO为必需药效团。结论建立的模型可应用于GABA-T酶抑制剂的高通量筛选,为GABA-T酶抑制剂类抗癫痫活性成分的筛选及其作用机制研究提供参考。
OBJECTIVE: To establish a drug screening model of GABA-T antiepileptic nervous system drugs.This model was applied to screen active constituents of Gastrodia elata Blume and their analogues and their structure-activity relationship analysis. Methods The enzymatic reaction temperature, reaction time, substrate NAD + concentration, substrate α-ketoglutarate concentration, substrate γ-aminobutyric acid (GABA) concentration and other factors were used to establish a model of GABA-T activity screening. Hydroxybenzaldehyde (HBA) and its 11 structural analogs were used to validate the model. Results The screening model of GABA-T antiepileptic activity was successfully constructed. The determination results of HBA and 11 structural analogues were consistent with the reported antiepileptic effect. The structure-activity analysis showed that -OH and -CHO on the para-position of benzene ring are necessary Pharmacophore. Conclusion The established model can be applied to the high-throughput screening of GABA-T enzyme inhibitor, which can provide a reference for the screening of active ingredients of GABA-T inhibitor antiepileptic and its mechanism of action.