固相支撑液液萃取-液相色谱-串联质谱测定尿液中10种双酚类化合物和5种对羟基苯甲酸酯

Determination of ten bisphenols and five parabens in urine by solid supported liquid-liquid extraction and liquid chromatography-tandem mass spectrometry

尿液中双酚类化合物(BPs)和对羟基苯甲酸酯类化合物(PBs)的浓度水平监测为考察其在人体内的暴露提供基础数据,是准确评估其健康风险的前提。本研究使用基于固相支撑液液萃取(SLE)原理的新型萃取柱,建立了新的BPs和PBs快速前处理技术,在此基础上利用液相色谱-串联质谱法(LC-MS/MS)同时测定人体尿液中10种BPs和5种PBs。尿样先酶解,然后经SLE柱富集,使用15 mL乙酸乙酯-正己烷(3∶7,v/v)混合溶液进行洗脱;通过引进水、甲醇和乙腈的三元流动相梯度洗脱系统,实现了15种目标化合物的准确定性和定量分析。在混合尿液基质中,低、中、高3个水平的加标回收率为843%~1198%;除双酚S外,其余14种化合物的基质效应均在20%以下,表明具有良好的回收率和较低的生物基质干扰。15种目标化合物在各自的线性范围内线性关系良好,相关系数均大于0995;方法定量限为003~030μg/L;精密度测试结果显示,日内和日间连续进样仪器响应的相对标准偏差分别为14%~84%和57%~146%,证明具有良好的稳定性和重复性。该方法成功应用于10个普通人群尿样中10种BPs和5种PBs的测定。结果表明,检出率最高的化合物为MeP、EtP、PrP和BPA,其中值质量浓度分别为110、060、021和055μg/L,其余化合物检出率低于50%,这可能与化合物的生产使用量、生物可利用性以及在人体内的生物代谢能力相关。

Bisphenols(BPs)and parabens(PBs)are of great concern for environmental pollu-tion and human health because of their endocrine-disrupting effects and potential health haz-ards.Urinary biomonitoring of BPs and PBs can provide basic data for human internal exposure evaluation,which is a prerequisite for accurately assessing their health risks.In this study,we developed a new pretreatment procedure based on solid supported liquid-liquid extraction(SLE)for the simultaneous separation of ten BPs and five PBs in human urine,followed by high performance liquid chromatography-tandem mass spectrometry(HPLC-MS/MS)analysis.In the instrumental analysis,the HPLC conditions and MS/MS parameters were comprehensive-ly optimized.Accurate qualitative and quantitative determination of ten BPs and five PBs was achieved by introducing a ternary gradient elution system of water,methanol,and acetonitrile for LC separation.During sample pretreatment,the extraction solvent and elution volume were optimized.Specifically,urine samples were held at room temperature and centrifuged at 3000 r/min for 10 min.The supernatant(2 mL)was then transferred to a glass tube,and the pH was adjusted to 50 using HCl(05 mL;01 mol/L)and NaAc-HAc buffer(15 mL).Thereafter,β-glucuronidase-arylsulfatase(20μL)and surrogate standard solutions(10 ng;13C12-BPS,13C12-BPAF,13C6-MeP,and 13C6-BuP)were added,and the mixture was incubated in a shaker bath in the dark at 37℃for 16 h.After incubation,the hydrolyzed sample(4 mL)was loaded onto an SLE cartridge and equilibrated for a minimum of 5 min to ensure the solution was completely absorbed by the packing material.Subsequently,the target chemicals were eluted with a mixed ethyl acetate/n-hexane solution(3∶7,v/v;15 mL).Separation of the targets was performed on a ZORBAX SB-C18 reversed-phase column(250 mm×46 mm,5μm)using an acetonitrile-methanol-water system as the mobile phase.The method was verified by spiking mixed urine samples at three levels(1,5,and 50μg/L),with the recoveries ranging from 843%to 1198%.Except for bisphenols(BPS),whose matrix effect was calculated as-218%,the ma-trix effects of other analytes were lower than 20%,indicating low matrix interference.The line-ar ranges of the analytes varied from 01-500μg/L to 1-500μg/L,with correlation coefficients higher than 0995.The method limits of quantification for target chemicals ranged from 003 to 030μg/L,and the relative standard deviations of intra-and inter-day experiments were 14%-84%and 57%-146%,respectively,suggesting high stability and reproducibility.The method was successfully applied to the determination of ten BPs and five PBs in 10 urine samples from a general population.The concentrations of target chemicals in the human urine samples varied.Methylparaben(MeP),ethylparaben(EtP),propylparaben(PrP),and bisphenol A(BPA)were detected in all samples,with median mass concentrations of 110,060,021,and 055μg/L,respectively.The detection rates of the other chemicals were less than 50%,which may be related to the production and use of specific chemicals,their bioavailability,and biological metabolism in humans.