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目的建立一种同时测定血清中5-羟色胺(5-HT)和色氨酸(Trp)浓度的高效液相色谱-荧光检测法(HPLC-FD)。方法采用的色谱柱为Hypersil C18柱(250mm×4.6mmi.d.,7μm),流动相为0.1mol/L磷酸二氢钾/甲醇(85:15,V/V)溶液,流速1.0ml/min,荧光检测激发波长和发射波长在0～6min分别为278和338nm,6～10min转换为280和340nm。结果5-HT保留时间约为4.9min,线性方程为Y=4223461X-35303.9,最低检出浓度为0.001μmol/L,回收率为82.70%～95.40%,日内和日间RSD分别为2.03%和3.42%;Trp保留时间约为8.4min,线性方程为Y=421314X+38292.9,最低检出浓度为0.007μmol/L,回收率为83.67%～94.98%,日内和日间RSD分别为1.32%和3.31%;苯丙氨酸、酪氨酸、犬尿氨酸和犬尿喹啉酸等物质对本方法无干扰;健康成人(n=71)血清中5-HT和Trp的浓度分别为(544.8±156.8)nmol/L和(53.08±8.29)μmol/L。结论该方法简便、快速、灵敏,适于临床和科研应用。
Objective To establish a HPLC-FD method for the simultaneous determination of 5-hydroxytryptamine (5-HT) and tryptophan (Trp) in serum. Methods The column was Hypersil C18 (250mm × 4.6mmi.d., 7μm). The mobile phase was 0.1mol / L potassium dihydrogen phosphate / methanol (85:15, V / V) solution and the flow rate was 1.0ml / min The fluorescence excitation and emission wavelengths were 278 and 338 nm at 0-6 min and 280 and 340 nm at 6-10 min, respectively. Results The retention time of 5-HT was about 4.9min, the linear equation was Y = 4223461X-35303.9, the minimum detectable concentration was 0.001μmol / L, the recovery was 82.70% -95.40%, and the intraday and interday RSD were 2.03% and 3.42, respectively %; The retention time of Trp was about 8.4min, the linear equation was Y = 421314X + 38292.9, the minimum detectable concentration was 0.007μmol / L, the recoveries were 83.67% ~ 94.98%, RSD were 1.32% and 3.31% ; Phenylalanine, tyrosine, kynurenine and kynurenic acid did not interfere with this method. The concentrations of 5-HT and Trp in serum of healthy adults (n = 71) were (544.8 ± 156.8) nmol / L and (53.08 ± 8.29) μmol / L. Conclusion The method is simple, rapid, sensitive and suitable for clinical and research applications.