加速溶剂提取-同位素稀释-高分辨气相色谱-高分辨质谱法测定生物样品中82种多氯联苯

我国水产品中多氯联苯(PCBs)的检测方法,主要以6种指示性PCBs和12种二噁英类共平面PCBs为主,仅涵盖有限的PCBs。为更全面地获得生物体中PCBs的浓度水平,深入探讨PCBs在生物体内的代谢和富集特征,进而准确评价PCBs对人类的暴露水平及风险,以鱼和贝类作为生物样品代表,建立了加速溶剂提取-同位素稀释-高分辨气相色谱-高分辨质谱(ASE-ID-HRGC-HRMS)测定生物样品中82种PCBs的方法。比较了振荡提取和加速溶剂提取两种提取方式的回收率和重复性,最终采用正己烷-二氯甲烷(1∶1, v/v)对PCBs进行加速溶剂提取。考察了各流分淋洗液对PCBs的回收率,确定了样品提取液经8 g 44%酸性硅胶层析柱(内径15 mm), 90 mL正己烷洗脱的净化方式。样品提取液净化浓缩后进行HRGC-HRMS分析,色谱柱采用DB-5MS超低流失石英毛细管柱(60 m×0.25 mm×0.25 μm)。通过优化后的升温程序对化合物进行分离,以保留时间和两个特征离子精准定性,采用同位素内标法定量。结果表明,在0.1~200 μg/L范围内,平均相对响应因子(RRF)的相对标准偏差值(RSD, n=7)均≤20%,相关系数(r²)>0.99。生物样品中PCBs的方法检出限为0.02~3 pg/g;鱼类中PCBs平均加标回收率为71.3%~141%, RSD(n=7)为2.1%~14%;贝类中PCBs平均加标回收率为76.9%~143%, RSD为1.4%~11%。该方法灵敏、准确、可靠,可以更加全面具体地分析鱼和贝类等水产品受PCBs的污染情况,为国内外开展生物监测提供有效的技术支持,从而服务于相关生态环境管理及履行《斯德哥尔摩公约》。

Determination of 82 polychlorinated biphenyls in biological samples using accelerated solvent extraction-isotope dilution-high resolution gas chromatography-high resolution mass spectrometry

In China, the detection methods for polychlorinated biphenyls (PCBs) in aquatic products are mainly effective for 6 indicative PCBs and 12 coplanar dioxin-like PCBs, which only account for a limited proportion of PCBs in organisms. In this study, to obtain the detailed concentration levels of PCBs in organisms, elucidate the metabolism and enrichment characteristics of PCBs in organisms, and accurately evaluate the exposure level and risks of PCBs to humans, an improved method for the simultaneous determination of 82 PCBs in fish and shellfish samples was developed using isotope dilution-high resolution gas chromatography-high resolution mass spectrometry (ID-HRGC-HRMS). The recovery and reproducibility of two extraction methods, i. e., oscillatory extraction and accelerated solvent extraction (ASE), were compared. Finally, ASE was chosen for subsequent experiments. Specifically, after adding 1 ng¹³C-labeled extraction internal standards, the samples were extracted under pressure by ASE using a mixture of n-hexane-dichloromethane (1∶1, v/v). The experimental conditions employed for this were a pressure of 10.3 MPa, heating temperature of 100 ℃, heating time of 5 min, static time of 8 min, flush volume of 60%, purging time of 120 s, and 34 mL cells. Subsequently, the extracts were loaded on an 8 g acid silica gel (44%) column (inner diameter: 15 mm) and eluted with 90 mL of n-hexane. After purification and concentration, the analytes were determined by HRGC-HRMS with a fused-silica capillary column (DB-5MS, 60 m×0.25 mm×0.25 μm). The temperature program was optimized to separate the most target compounds at the baseline. Specifically, the initial oven temperature was 120 ℃, which was held for 1 min, following by heating to 180 ℃ at 30 ℃/min, heating to 210 ℃ at 2 ℃/min and holding for 1 min, and further heating to 310 ℃ at 2.5 ℃/min and holding for 1 min. The injector and ion source temperatures were 270 ℃ and 280 ℃, respectively. With a static resolution of 10000, the HRMS instrument was operated in the selected-ion monitoring mode at an electron energy of 35 eV. The 82 PCBs were qualified by their retention time and two characteristic ions, and thereafter quantified using the mean relative corresponding factor (RRF). The results showed that the relative standard deviation (RSD) of the RRF obtained from six-point calibration standard solutions was less than 20%. The linearity ranges were from 0.1 to 200 μg/L, and the correlation coefficients (r²) were greater than 0.99. Under optimum conditions, the method detection limits (MDLs) for the PCBs of biological samples were in the range of 0.02-3 pg/g. To validate the method, the fish and shellfish samples were spiked with a low level (0.4 ng) and high level (3.6 ng) of native PCB standards. The spiked recoveries using low-concentration native PCBs were 71.3%-139% in fish and 76.9%-143% in shellfish, and the RSDs (n=7) were 2.1%-14% and 4.5%-14%, respectively. The spiked recoveries using high-concentration native PCBs were 77.6%-141% and 82.2%-131%, respectively, and the RSDs (n=7) were 1.4%-9.4% and 1.7%-11%, respectively. An analysis of fresh fish and shellfish samples showed that the contents of a single PCB ranged from “not detected” to 54.1 pg/g, where 12 coplanar dioxin-like PCBs were detected in the range of 12.6 pg/g to 74.5 pg/g, six indicative PCBs in the range of 30.9 pg/g to 62.1 pg/g, and 82 PCBs in the range of 174 pg/g to 672 pg/g. It was concluded that this method could be successfully applied for the determination of PCBs in biological samples with good accuracy and precision. This comprehensive analytical method of PCBs in aquatic products provides effective technical support for biological monitoring; it will also aid in ecological and environmental management and the implementation of the Stockholm Convention policies.