反相高效液相色谱-紫外光检测法测定血浆及血小板中5-羟色胺含量

Determination of 5-hydroxytryptamine in plasma and platelet of human by RP-HPLC-VD

目的建立反相高效液相色谱-紫外光检测法测定人体血浆及血小板中5-羟色胺(5-HT)的含量。方法样本经处理后,用高效液相-紫外光色谱仪进行检测。色谱柱为Agilent technologies C18(250mm×4.6mm,5μm);流动相为甲醇-0.05mol/L磷酸二氢钾(8∶92);检测波长为275nm;流速为1mL/min。结果5-HT在12.5～0.0244μg/mL浓度范围内与峰面积有很好的线性关系(r=1);高、中、低3个浓度的日内RSD分别是0.41%、0.67%和2.77%;日间RSD分别是0.74%、0.45%和4.03%;血浆中5-HT平均回收率为82.16%,血小板中5-HT平均回收率为88.04%;最低定量限为0.0244μg/mL,最低检测限为0.0122μg/mL。结论用反相高效液相色谱-紫外光检测法测定5-HT简便,快速,特异性好,灵敏度高,重复性好,适用于临床和科研工作。

Objective To establish a method for determination of 5-hydroxytryptamine（5-HT）concentration in plasma and platelet of human by RP-HPLC-VD.Methods The chromatographic column of Agilent technologies C18（250 mm×4.6 mm,5 μm）was used with the mobile phase consisted of a mixture of methanol and 0.05 mol/L KH2PO3（8：92）at the flow rate of 1 mL/min.The detection wavelengh was 275 nm.Results It showed a good linear relation between the content of 5-HT and peak area at the range of 12.5~0.024 4 μg/mL（r=1）.The inter-day RSD of high,middle,low concentration of 5-HT were 0.41%,0.67% and 2.77% respectively,and the intra-day RSD were 0.74%,0.45% and 4.03%.The quantitation limit was 0.024 4 μg/mL,and the detection limit was 0.012 2μg/mL.The average recovery rate were 82.16% and 88.04% in human plasma and platelet.Conclusion The RP-HPLC method for the determination of 5-HT is simple,accurate,highly reproducible and with good sensitivity,it is suitable for the clinical and research work.