多功能离子液体分散液液微萃取结合高效液相色谱法检测人尿中5种邻苯二甲酸酯代谢物

将多功能离子液体与分散液液微萃取(DLLME)技术相结合,建立了测定尿液中5种邻苯二甲酸酯类(PAEs)物质代谢产物的高灵敏度新方法。对影响DLLME效率的各单因素进行了优化,包括萃取剂的种类及体积、分散剂的种类及体积、萃取温度、超声时间、冷却时间、离心时间和盐效应等条件,经过严格的优化,最佳的萃取条件分别为:萃取剂C8MIM]PF6]35μL,分散剂BSO3HMIm]OTf]30μL和C4MIM]BF6]120μL,萃取温度为35℃,超声时间5 min,冷却时间5 min,离心时间5 min,盐析剂NH4PF60.1 g。在最佳的萃取条件下,5种PAEs代谢物在0.5~1000μg/L范围内具有良好的线性关系,决定系数(R2)均大于0.9955,方法检出限为0.16~0.19μg/L,尿液中添加低中高水平(5、20、100μg/L)的PAEs代谢物,其回收率为92.9%~105.0%,日内精密度及日间精密度的相对标准偏差(RSD)均小于5.96%,方法学验证各指标及稳定性均符合分析要求。对所采集的10份糖尿病患者的尿液进行检测,并对该人群PAEs代谢物的暴露水平进行评价。结果表明,各PAEs代谢物均有检出,其中邻苯二甲酸单(2-乙基己基)酯(MEHP)的检出率为100%。总之,该方法萃取过程中未添加有毒的有机试剂,均使用多功能离子液体作为萃取剂、分散剂和盐析剂,萃取过程绿色环保,简单高效;方法的灵敏度较高,稳定性较好,适用于人体尿液中痕量PAEs代谢物的检测。

Combination of dispersive liquid-liquid microextraction using multifunctional ionic liquids with high performance chromatography for determination of phthalate ester metabolites in human urine sample

A novel sensitive method for the determination of the five primary metabolites of phthalate esters (PAEs) in urine was developed by combining dispersive liquid-liquid microextraction (DLLME) using ionic liquids with high performance liquid chromatography (HPLC). The factors affecting the efficiency of DLLME were optimized. The types and proportions of extraction solvent and dispersants, as well the ultrasonic extraction time, cooling time, and centrifugal time, were determined. The optimal conditions were as follows:extraction solventC₈MIM]PF₆] 35 μL; dispersantsBSO₃HMIm]OTf] 30 μL, C₄MIM]BF₆] 120 μL; NH₄PF₆ 0.1 g, extraction at 35℃, ultrasonic dispersion for 5 min, cooling in ice water for 5 min, centrifugation at 4000 r/min for 5 min. After optimization, the five primary metabolites of PAEs were determined. The method showed a good linear relationship within the concentration range of 0.5-1000 μg/L. The determination coefficients (R²) were greater than 0.9955. The detection limit was in the range of 0.16-0.19 μg/L. Under the optimized conditions, the extraction recoveries for the PAEs were 92.9%-105.0%, and the relative standard deviations (RSDs) of the intra- and inter-day precisions were < 5.96%. Urine samples collected from 10 diabetic patients were tested, and the exposure level of the population to the PAE metabolites was evaluated. All the PAE metabolites were detected in these samples, and the detection rate of 2-ethylhexyl hydrogen phthalate (MEHP) was 100%. In conclusion, no toxic organic reagents were added during the extraction process in this method, and multifunctional ionic liquids were used as the extraction agent, dispersant, and salting-out agent. In other words, the extraction process was demonstrated to be green, simple, and efficient. The developed method has high sensitivity and stability, and it is suitable for the determination of trace PAE metabolites in human urine.