新型三液相萃取裝置結合液相層析串聯質譜儀偵測尿液中甲苯暴露指標成分

Analysis of Urinary Markers for Toluene Exposure by LC-MS/MS with a New Liquid-Liquid-Liquid Extraction Device

黃豊鈞(Li-Jun Huang) ； 謝仲允(Jhong-Yun Sie) ； 沈振峯(Jenn-Feng Sheen)

《化學》 74卷1期 (2016/03) Pp. 33-43

https://doi.org/10.6623/chem.2016004

摘要

甲苯為國內工業界常用之溶劑，甲苯暴露過量會造成神經系統、肝和腎臟危害。因此對甲苯暴露的監控可藉由分析尿中甲苯代謝物進行對偵測。本研究主要利用一自行組裝之三液相萃取裝置結合液相層析串聯質譜儀分析尿液樣品中甲苯暴露代謝物芐基硫醇酸(Benzylmercapturic acid, BMA)與鄰-甲酚(o-cresol)，此裝置所使用的材料皆為實驗室易取得的離心管及樣品瓶且十分容易操作。經最佳化後之萃取條件為：萃取溶劑為1.5 mL之toluene與diethyl ether (1:1; v/v)混和溶劑、萃取相體積400 μL、6 mL之尿液樣品(pH = 2)，萃取時間為150分鐘。經過裝置萃取後，基於萃取相含緩衝鹽，我們使用自行修飾之放電套頭化學游離界面來進行液相層析質譜儀分析，其尿液樣品萃取前後BMA訊號增加將近100倍，而o-cresol訊號增加將近8倍。以本方法分析BMA及o-cresol之尿液添加樣品，在濃度範圍0.5-200 ng/mL及12.5-400 ng/mL分別可得到良好的線性關係(r^2 ＞ 0.99)。為減低不同來源尿樣的相對基質效應，尿樣需經十倍稀釋後再進行萃取分析，取七種不同來源尿樣進行萃取分析，測得各指標物訊號峰面積之相對變異(RSD)皆小於15%，同時分析靈敏度亦可清楚偵測未添加尿樣中BMA及o-cresol之背景訊號。

關鍵字

三液相萃取 ； 放電套頭界面 ； 芐基硫醇酸 ； 鄰-甲酚

並列摘要

Bio-monitoring of low level toluene exposure is commonly performed by determining the urinary benzylmercapturic acid (BMA) and o-cresol. In this study, a new liquid-liquid-liquid extraction (LLLE) device was developed and applied in the analysis of the urinary markers of toluene exposure. The new device was simply constructed with centrifuge tubes and sample bottle which is very easily to operate. Because of the high buffer contents of the acceptor phase, the APCI mode performed with a patented discharge adaptor interface was chosen for the LC-MS/MS analysis. The optimized extraction parameters were 6 mL donor phase (pH = 2), 1.5 mL mixed organic phase of toluene and diethyl ether (1:1, v/v) and 400 μL acceptor phase of 0.1 M NaOH_((aq)) (pH~13). In spiked urine, after 150 min extraction, the urinary markers were selectively enriched in the donor phase, the ion signals were enhanced 100 times for BMA and 8 times for o-cresol. To minimize the sample lots effect to less 15%, the urine samples were 10-fold diluted before the LLLE. Good linearity (r^2 ＞ 0.99) was obtained in the range of 0.5-200 ng/mL and 12.5-400 ng/mL for urinary BMA and o-cresol, respectively. The analytical sensitivity of the method was able to clearly detect the BMA and o-cresol signals in the un-spiked urine sample.

並列關鍵字

Liquid-liquid-liquid extraction (LLLE) ； Discharge-adaptor interface ； Benzylmercapturic acid ； o-Cresol