分散液-液微萃取结合气相色谱质谱测定尿液中的尼古丁和可替尼

Determination of Nicotine and Cotinine in Urine by Dispersive Liquid-Liquid Microextraction Combined with Gas Chromatography-Mass Spectrometry

目的探讨建立溶剂去乳化分散液-液微萃取（dispersive liquid-liquid microextraction,DLLME）结合气相色谱-质谱法（gas chromatography-mass,GC/MS）测定尿液中尼古丁和可替尼的方法。方法考察GC/MS检测和DLLME条件。取5mL尿样,用NaOH调节pH至9.0,加入氯化钠振荡溶解后,取100μL三氯甲烷（含内标物喹啉）和1 000μL甲醇分别作为萃取剂和分散剂并混合,注入样液中使乳化分散,4 000r/min离心5min,弃上清,取1μL下层有机相进GC/MS分析。以DB-5毛细管色谱柱为分离柱,程序升温分离样液中的尼古丁和可替尼,离子监测（SIM）模式质谱检测和内标工作曲线法进行定量。结果尼古丁和可替尼在0.2~100ng/mL范围内线性良好,方法检出限分别为0.010ng/mL和0.022ng/mL,方法精密度（相对标准偏差）分别为8.2%和9.6%,加标回收率分别为92.0%~108.0%和83.0%~110.0%。结论本方法简单快速、灵敏准确,适用于尿液中尼古丁和可替尼的测定,能满足人群烟草暴露评价的需要。

Objective To develop a method for the determination of nicotine and cotinine in urine samples by dispersive liquid-liquid microextraction（DLLME）combined with gas chromatography-mass spectrometry（DLLMEGC/MS）.Methods The experimental conditions for GC-MS and DLLME were investigated in detail.DLLME was performed with the following procedure：5mL of urine sample was adjusted to pH9.0with NaOH solution;NaCl was added to increase ionic strength;100μL chloroform（containing internal standard of quinolone）as extractant was mixed with 1 000μL methanol as dispersant and then injected into the urine sample to make it emulsified and dispersed.The sample solution was centrifuged for 5 min at 4 000r/min,and 1μL of its extraction solvent was injected into the GC/MS system for analysis.GC separation was performed with DB-5column under programmed temperature.Nicotine and cotinine were quantified using selected ion monitoring（SIM）mode of mass spectrum detection and internal standard working curve.Results Good linear relationship was obtained for detecting nicotine and cotinine ranging from 0.2ng/mL to 100ng/mL,with a detection limit of 0.010ng/mL and 0.022ng/mL for nicotine and cotinine respectively.The relative standard derivation（RSD）for determination of nicotine and cotinine in urine samples were 8.2% and 9.6% respectively.The spiked recoveries ranged from 92.0% to 108.0% for nicotine,83.0%to 110.0%for cotinine.Conclusion The method is rapid,sensitive,accurate and simple,with little consumption of organic solvent.It is suitable for determination of nicotine and cotinine in urine,and can meet the requirements for evaluating human tobacco exposure.