高效液相色谱法测定大鼠海马中5-羟色胺含量的方法学研究

目的 建立了高效液相荧光色谱测定大鼠海马中5-羟色胺(5-HT)含量的检测方法 .方法 采用Alltima C18色谱柱(4.6 mm×250 mm,5 μm),流动相为0.1 mol/L磷酸二氢钾溶液-甲醇(9∶1,V/V),等度洗脱,流速1.0 ml/min.激发波长(λEx)254 nm,发射波长(λEm)338 nm.结果 线性范围为0.70～8.40 ng(r=0.999 9);当信噪比为3(S/N =3)时,检出限为1.27×10-3 ng;日内测得精密度相对标准偏差(RSD)为0.7%;重复性测定相对标准偏差(RSD)为2.8%;加样回收率为88.6%～101.5%.结论 该方法 简便、准确、灵敏,特异性强,重复性好,适用于海马中5-HT的含量测定.

Study on the Determination of 5-hydroxy Tryptamine(5-HT) in Hippocampus of Rat by High Performance Liquid Chromatography

Objective The content of 5 - HT in hippocampus of rat was determined by high performance liquid chromatograph -fluorescence (HPLC-FLU). Methods The separation and determination were carried out by following conditions: Alltima C_(18) column (250 mm±4.6 mm,5 um) ,0.1mol·L~(-1) KH_2PO_4 -methanol(9:l ,V/V) as mobile phase , flow-rate was 1.0 ml· min~(-1). The fluorescence excitation and emission wavelengths were set at λ_(Ex) 254 and λ_(Em) 338 nm, respectively. Results The linear range of the method was 0.70～8.40 ng(r = 0.999 9) for 5~HT . The detection limit of 5 - HT was 1.27×10~(-3)ng, when the ratio of signal to noise was 3. The inter - assay precision was 0.7%. The recovery was within the range of 88.6%×101.5%. Conclusion The method is simple, sensitive, reliable and has strong specificity and good repetition. It is suitable for the content determination of 5-HT in hippocampus of rat, by high performance liquid chromatography.