| 摘要: | 人造皮製造業勞工尿液中代謝物之固相微萃取氣相層析質譜分析方法研究 目的：二甲基甲醯胺(N,N-dimethylformamide ; DMF)為工業中常使用的溶劑，特別在於人造皮製造業中。目前對於DMF的所常用生物暴露指標為尿中的NMF(N,N-methylformamide)。本研究擬開發一個以頂空固相微萃取吸附的方式，再以氣相層析質譜儀來分析尿液中DMF代謝物。 方法：本研究以Carbowax-Divinylbenzene(CW/DVB)纖維作為吸附介質。因DMF代謝物揮發性低，故執行分析前以加熱的方式增加其頂空濃度，再以頂空法萃取尿液中DMF代謝物。完成採集後的固相微萃取纖維以氣相層析質譜儀分析。實驗室技術建立時，係將DMF代謝物標準品添加於健康非暴露者尿液中，製備標準溶液進行分析。本研究將探討的參數包括：尿液樣本之固相微萃取pH值、加入鹽類(NaCl、KNO3、NaNO3、(NH4)2SO4)、溫度、攪拌(包括攪拌時的轉速、攪拌時間)對尿液中DMF代謝物萃取效率的影響，以及氣相層析質譜儀器分析條件設定。 結果： 1.SPME方法為：將pH值為7±0.2的4 mL尿液樣本置於15 mL的樣本瓶中，添加0.8 g的氯化鈉混合均勻，在60 ℃、轉速為1000 rpm的條件下，同時用CW/DVB 纖維進行頂空固相微萃取25分鐘，最後在氣相層析質譜儀注射口以240 ℃熱脫附 8分鐘。 2.在此萃取分析條件下，所求得的丙酮檢量線濃度範圍介於0.1582-31.64 µg/mL，DMF檢量線濃度範圍介於0.944-188.8 µg/mL，NMF檢量線濃度範圍介於1.992-398.4 µg/mL，其相關係數(r)皆在0.995 以上。此外，丙酮的方法偵測極限（MDL）為0.0177 µg/mL，DMF的方法偵測極限為0.432 µg/mL；NMF的方法偵測極限為0.446 µg/mL。 3. 本分析方法發現，丙酮、DMF、NMF 分別在15.82、0.791 µg/mL；94.4、4.72 µg/mL；199.2、9.96 µg/mL之濃度值的尿液樣本密封儲存於-20℃冰箱內，在四週內的回收率皆大於86 %。 結論：以HS-SPME-GC-MS的方式來分析尿液樣本中DMF代謝物，可省去繁雜的前處理步驟，同時其具有免溶劑、快速、方法簡便、以及不受樣本基質干擾等優點。 關鍵字：二甲基甲醯胺、生物暴露指標、固相微萃取、頂空法; SPME-GC/MS method development for synthetic leather worker urinary metabolites Objective：N,N-dimethylformamide (DMF) has been used as a solvent in many industries, especially in artificial leather industry. Urinary N,N-methylformamide (U-NMF) is commonly used as the biomarker of DMF exposure. The purpose of this study is to develop an analytical method for determining the DMF metabolism using solid phase microextraction followed by gas chromatography/mass spectrometry analysis. Method：The Carbowax-Divinylbenzene(CW/DVB) fiber was tested to be the most suitable material for sample extraction. Due to the characteristic of low vaporization of DMF metabolism, and to increasing the DMF metabolism consistency, heating procedure was executed prior to the analysis. The pretreated procedure was then used to extract DMF metabolism in urine by headspace extraction. The SPME fiber was inserted into the injection port of GC/MS and analyzed. The standard solution was prepared by spiking DMF metabolism standard into the blank pooled urine of unexposed person. The optimum extraction parameters investigated were: pH, salt (NaCl, KNO3, NaNO3, (NH4)2SO4), temperature, stirring speed, extraction time, as well as measurement condition of GC/MS. Results： 1.The SPME method was placing 0.8 g NaCl and 4mL urine sample of pH=7±0.2 in an 15mL vial, stirring at 1000 rpm 60 ℃stirring plant, CW/DVB fiber was used to perform headspace solid-phase microextraction for 25 min and thermal desorption 240 ℃for 8 min. 2. For the quality control calibration, the concentration ranges of Acetone, DMF and NMF in urine were 0.1582-31.64 µg/mL, 0.944-188.8 µg/mL, and 1.992-398.4 µg/mL. The correlation coefficients(r) were all above 0.995. The method detection limit（MDL）for Acetone, DMF and NMF were 0.0177 µg/mL, 0.432 µg/mL and 0.446 µg/mL, respectively. 3. In the storage stability tests at -20 ℃, we found the recovery rates for the urine spiked with Acetone at 15.82、0.791 µg/mL, with DMF at 94.4、4.72 µg/mL, and with NMF at 199.2、9.96 µg/mL were all above 86 % within 4 weeks. Conclusion：The analytical method of DMF metabolism by HS-SPME-GC-MS can be reduced complicated pretreated procedure, as well as having the advantage of solvent-free, rapidity, simplicity, and avoid interfering with a complex matrix. Key word：N,N-dimethylformamide, biomarker, solid phase microextraction, headspace |
