改善十六烷基三甲基溴化銨於毛細電泳/電灑法質譜儀訊號抑制之研究

Alleviation of Ion Suppression in Capillary Electrophoresis / Electrospray Ionization Mass Spectrometry Using Cetyltrimethylammonium Bromide Containing Buffer

黃裕婷(Yu-Ting Huang)

指導教授 ： 何國榮

國立臺灣大學/理學院/化學研究所/碩士(2010年)

https://doi.org/10.6342/NTU.2010.03181

摘要

利用毛細電泳分析低分子量有機酸時，會在緩衝溶液中加入帶正電荷的界面活性劑（例如：cetyltrimethylammonium bromide, CTAB）作動態塗佈，使電滲流反向，縮短分析時間並確保所有分析物都可到達偵測器。然而，CTAB會降低分析物於質譜上之靈敏度並嚴重汙染質譜的離子源。本研究利用先前實驗室已開發的低流速鞘流-液體結合式介面（liquid-junction/low-flow interface）搭配PTFE材質的銜接管，能有效避免CTA正離子(cetyltrimethylammonium cation)進入質譜，提升低分子量有機酸之靈敏度。在反向電場（reverse polarity）的操作模式下，分離管因為正電荷的界面活性劑塗佈，使得電滲流反向，因此帶負電荷的分析物與電滲流都往質譜端移動。在銜接管的部份則因為沒有塗佈，電滲流的方向與分析物相反，因此我們使用PTFE（polytetrafluoroethylene）管來降低電滲流，使分析物本身的淌度大於電滲流，分析物可以藉由自己的淌度到達偵測器，而CTA正離子則會因為攜帶正電荷的關係，留在液體接合槽中。此裝置能成功避免CTA正離子進入質譜，提升分析物在質譜上的靈敏度，並且避免離子源受到汙染。實驗數據顯示，當分析物濃度為50 ppm時，低流速鞘流介面幾乎完全無法偵測到分析物，而使用低流速鞘流-液體結合式介面搭配PTFE銜接管的裝置可成功偵測到所有分析物。
　　此外，CTAB亦被應用於毛細電泳之胜肽分離，利用微胞電動毛細管層析法（micellar electrokinetic chromatography, MEKC）增加相似胜肽之分離效果。因此我們在緩衝溶液中添加高於臨界微胞濃度的CTAB，用以分離馬的肌紅蛋白之胰蛋白酶水解胜肽。實驗結果顯示，雖然CTAB的微胞可使原本無法利用毛細管區帶電泳分離的胜肽分開，然而，對於肌紅蛋白之胰蛋白酶水解胜肽的整體分離並無明顯好處。

關鍵字

CTAB ； 動態塗佈 ； 毛細電泳質譜 ； 低分子量有機酸 ； PTFE管 ； 蛋白質水解胜肽微胞電層析

並列摘要

Analyzing low-molecular organic acid by capillary electrophoresis usually adds cationic surfactant such as cetyltrimethylammonium bromide（CTAB）in the running buffer. Dynamic coating CTAB on capillary can reverse the EOF, thus reducing the analysis time and ensuring all analytes could be detected. However, CTAB will reduce the sensitivity of organic acid and pollute the ion source of the mass spectrometer seriously. This research uses liquid-junction/low-flow interface in combination with the PTFE connecting column to prevent the CTA(cetyltrimethylammonium) cation enter the ESI source as well as the signal suppression of the analyte. Under reversed polarity and CTAB dynamic coating, the EOF and the analyte with negative charge all moved toward the detection side. However, without the adding CTAB, the EOF and the analyte were moved in the opposite directions in the connecting column. Thus, we use PTFE tube to reduce the EOF to let the mobility of the analyte can be larger than that of the EOF. And the analyte can reach the detection side at the same time the CTA cation will be staied in the liquid junction because of its positive charge. The experimental results shown that while the concentration of the analyte is 50 ppm, the system with low-flow interface cannot detect the analyte, but the system with liquid-junction/low-flow interface in combination with the PTFE connecting column successfully detected all the analytes.
Moreover, CTAB can also be used in the peptide separation in capillary electrophoresis. Better separation efficiency of similar peptides was achieved under the conditions of micellar electrokinetic chromatography（MEKC）. So we added high concentration CTAB into the buffer to separate the tryptic-digested peptides of myoglobin by MEKC. The experimental results showed that even though the separation of peptides with similar m/z was improved by MEKC. However, there was still no significant benefit obtained in the separation of overall digested peptides of myoglobin.

並列關鍵字

CTAB ； dynamic coating ； CE-MS ； low-molecular-weight organic acid ； PTFE column ； tryptic-digested peptide ； MEKC

參考文獻

48. 薛運鴻，何國榮，國立台灣大學化學研究所碩士論文 2008.

28. Fenn, J. B.; Mann, M.; Meng, C. K.; Wong, S. F.; Whitehouse, C. M. Science 1989, 246, 64.

42. Li, F. A.; Huang, J. L.; Shen, S. Y.; Wang, C. W.; Her, G. R. Anal. Chem. 2009, 81, 2810.

44. Zhong, Y. Y.; Zhou, W. F.; Hu, Z. Z.; Chen, M. L.; Zhu, Y. Chin. Chem. Lett. 2010, 21, 453－456

35. Amad, M. H.; Cech, N. B.; Jackson, G. S.; Enke, C. G. J. Mass Spectrom. 2000, 35, 784-789.