Development of the sensitive determination method for the endogenous NO synthetase inhibitor and analysis of homeostasis of blood pressure.

内源性NO合成酶抑制剂灵敏测定方法的建立及血压稳态分析。

• 批准号: 13672250
• 负责人: SANTA Tomofumi
• 金额: $ 2.37万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13672250/
• 关键词: nitrogen oxide; HPLC; fluorescence; dimethylarginine; OPA; derivatization

A fully automated analyzer for methylated L-arginine metabolites (N,N-dimethyl-L-arginine (ADMA), N-methylarginine (NMMA) and N, N'-dimethyl-L-arginine (SDMA)) by high-performance liquid chromatography with post-column fluorescence derivatization was developed. This system consists of an in-line extraction, a separation on a reversed phase ion-pair chromatography, a post-column derivatization by o-phthaladehyde (OPA) and thiol reaction, and fluorescence detection. NMMA, ADMA and SDMA were separated in 40 min with isocratic elution by a combination of octanoate and cyclohexanecarboxylate as ion-pair reagents. The eluate was monitored at 450 nm with the excitation at 337 nm. The calibration curves for NMMA, ADMA and SDMA showed linearity over the range from 0.05 μmol 1^<-1> (0.5 pmol on column) to 5.0 μmol 1^<-1> (50 pmol on column). This method does not require any time-consuming pre-treatment and requires only 10-μl of plasma sample for assay.

建立了一种全自动高效液相色谱柱后荧光衍生化分析甲基化L-精氨酸代谢产物（N，N-二甲基-L-精氨酸（ADMA）、N-甲基精氨酸（NMMA）和N，N '-二甲基-L-精氨酸（SDMA））的方法。该系统由在线萃取、反相离子对色谱分离、邻苯二甲醛（OPA）柱后衍生和巯基反应、荧光检测组成。以辛酸盐和环己烷羧酸盐为离子对试剂，采用等度洗脱，NMMA、ADMA和SDMA在40 min内得到分离。在450 nm处监测荧光强度，在337 nm处激发。NMMA、ADMA和SDMA的校准曲线在0.05 μmol 1^<-1>（柱上0.5 pmol）至5.0 μmol 1^<-1>（柱上50 pmol）范围内呈线性。该方法不需要任何耗时的前处理，仅需要10 μl血浆样品进行测定。