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目的建立一种高效液相色谱-紫外可见检测器测定大鼠肝组织中还原型谷胱甘肽含量的方法。方法肝组织在冰浴条件下匀浆后经乙腈沉淀蛋白,离心,取上清液在硼酸缓冲液中与荧光剂4-氟-7-硝基-2,1,3-苯并氧杂恶二唑(NBD-F)进行衍生化反应。流动相为乙腈-0.02mol/L磷酸二氢钾溶液(23∶77,V/V),流速1.0mL/min,检测波长470nm,进样量10μL;采用外标法定量。结果还原型谷胱甘肽在0.200～125μg/mL范围内线性关系良好,相关系数为0.9999(n=5),最低检测限为0.0400μg/mL(S/N=3);样品加标回收率为96.2%～104.5%;日内相对标准偏差为1.1%～4.6%,日间标准偏差为1.9%～7.5%。结论本方法操作简便,快速,准确,灵敏度高,稳定性好,适合肝组织中还原型谷胱甘肽的测定。
OBJECTIVE To establish a method for determination of reduced glutathione in rat liver tissue by high performance liquid chromatography-ultraviolet visible detector. Methods The liver tissue was homogenized in an ice bath and the protein was precipitated by acetonitrile and centrifuged. The supernatant was mixed with fluorescer 4-fluoro-7-nitro-2,1,3-benzoxan in boric acid buffer Oxadiazole (NBD-F) was derivatised. The mobile phase was acetonitrile-0.02mol / L potassium dihydrogen phosphate solution (23:77, V / V), the flow rate was 1.0mL / min, the detection wavelength was 470nm and the injection volume was 10μL. Results The results showed that there was a good linearity between 0.200 ~ 125μg / mL of reduced glutathione, with a correlation coefficient of 0.9999 (n = 5) and a minimum detectable limit of 0.0400μg / mL (S / N = 3) Was 96.2% ~ 104.5%. The relative standard deviations were 1.1% -4.6%, and the daytime standard deviations were 1.9% -7.5%. Conclusion The method is simple, rapid, accurate, sensitive, stable and suitable for the determination of reduced glutathione in liver tissue.