高效液相色谱法同时测定妊娠合并乙型肝炎患者血浆中的吲哚与3-甲基吲哚

Simultaneous determination of plasma indole and skatole in pregnant women with hepatitis B virus infection by high performance liquid chromatography

建立了一种高效液相色谱-荧光检测法用于同时测定血浆中的吲哚与3-甲基吲哚。样本经液液萃取法提取,采用Shim-Pack VP-ODS柱(150 mm×4.6 mm,4.6μm),以15 mmol/L磷酸二氢钠溶液-甲醇(40∶60,v/v)为流动相,甲奈酚为内标,荧光激发和发射波长分别为274 nm和340 nm。吲哚和3-甲基吲哚的线性范围分别为2.22~88.89μg/L和1.11~44.44μg/L;检出限分别为0.11μg/L(吲哚)和0.06μg/L(3-甲基吲哚);平均回收率为95.5%~112.3%,日内与日间相对标准偏差均小于6.8%。利用该方法对妊娠合并乙肝患者(n=29)和正常孕妇(n=46)的血浆进行了测定,结果表明妊娠合并乙肝患者血浆中吲哚和3-甲基吲哚水平均显著高于正常对照组,且与肝损伤指标转氨酶水平呈正相关。

A high performance liquid chromatographic （ HPLC） method with fluorescence detection （FD） was established to simultaneously determine plasma indole and skatole. Plasma samples were pretreated by liquid-liquid extraction. Chromatographic separation was accomplished on a Shim-Pack VP-ODS column （150 mm×4.6 mm,4.6 μm） using an isocratic mixture of 15 mmol/L sodium dihydrogen phosphate solution and methanol （40：60, v/v ）. The fluorescence excitation and emission wavelengths were 274 nm and 340 nm, respectively. The linear ranges were 2. 22-88. 89μg/L for indole and 1. 11-44. 44 μg/L for skatole. The detection limits were 0. 11 μg/L and 0. 06 μg/L for indole and skatole, respectively. The recoveries were in the range of 95. 5% -112. 3% with the relative standard deviations less than 6. 8%. The method was successfully applied to the analysis of plasma from healthy pregnant women （ n = 46） and pregnant women with hepatitis B virus （HBV） infection （ n = 29）. The results showed that plasma indole and skatole levels were significantly different between two groups. In pregnant women with HBV infection,the concentrations of indolic compounds were positively associated with transaminase levels.