無電極式/無翹流界面在毛細管電泳/電噴灑游離質譜法的開發與應用

Abstract

毛細管電泳結合電噴灑游離質譜法已經廣泛地應用於各種樣品的分析，而結合此二種方法的界面設計可決定分析結果的好壞，其中無鞘流界面的的設計常是針對電噴灑尖端塗佈的材料進行探討。一般來說通常需要將金屬、導電高分子、或石墨等導電材料塗佈於毛細管樣品出口尖端，外加第二組電壓於導電尖端以提供電噴灑時所需要的電場，但如何將導電材料穩定地塗佈於矽氧材質的尖端表面一直是此界面探討的主題之一。本論文嘗試直接利用高溫拉製及氫氟酸處理過的毛細管尖端做為電噴灑尖端，此設計不需要塗佈導電材料及施加第二組電壓於毛細管尖端處，此無鞘流界面因此能夠避免因導電材料塗佈不密合穩定而造成的易脫落及壽命短等問題。 為了得到此無鞘流界面的最佳實驗條件，論文中也針對了毛細管尖端口徑大小、毛細管樣品端施加電壓、流速、進樣角度與緩衝溶液等條件進行探討。本論文已成功地將此界面應用於蛋白質分子的分析，與商用鞘流界面相比更有樣品使用量減少之優點。另外，論文中以胜肽混合物為樣品進行毛細管電泳線上質譜分析，結果顯示出混合胜肽片段經過分離後改善了分析物同時游離時相互抑制的問題，且此無鞘流界面能夠有效地串聯毛細管電泳與電噴灑游離質譜法。 由於毛細管電泳/電噴灑游離質譜法容易受到電噴灑溶液中成份的干擾造成分析結果不理想，例如大量鹽類存在於樣品溶液中會造成分析物訊號下降。本論文最後一部份是設計了一個實驗，利用具有功能性的磁性鐵奈米粒子先專一濃縮分離樣品中的目標物-鈣離子，再配合本論文中所設計的無鞘流界面進行電噴灑質譜分析。實驗結果發現即使在複雜的生化樣品如牛奶與血清中都能利用功能性奈米粒子進行樣品的前處理後順利地在電噴灑質譜中偵測到鈣離子。 本論文所提出之界面設計相當簡單且容易製作，並和一般無鞘流界面的效果相同，可達到高靈敏度的分析結果。

Capillary electrophoresis/electrospray ionization mass spectrometry (CE/ESI MS) has been widely used in the analysis of various types of analytes. The design of the interface for combining CE and ESI MS may affect the analysis results. Generally, electric-conductivity materials such as metal, conductive polymers, and graphite are coated on the outside surface of the capillary tip followed by the application of an external high voltage on the tip to provide a sufficient electric field to generate electrospray in seathless interface. Thus, the fabrication of the electric-conductivity material on the surface of fused-silica is one of the main issues when using sheathless interfaces. In this thesis, a pulled bare capillary tip was proposed to be used as the ESI emitter. The outside surface of the tip was neither coated with electric-conductivity materials nor subjected to an external high voltage. Thus, there is no concern such as short lifetime that generally observed in conventional sheathless tips. The diameter of the capillary tip, the applied voltage on the capillary inlet, the flow rate for the sample introduction, the angles of the tip coupling to the mass spectrometer, and the composition of the ESI solution are examined in this thesis. The proposed interface has been demonstrated to an effective interface for the analysis of proteins and peptides in ESI MS. Furthermore, the sample consumption is much less than that used for commercial one. This sheathless interface is also applicable in directly coupling CE with ESI MS. When a peptide mixture is separated by CE prior to ESI MS analysis, the ion suppression problem during ionization between peptides can be avoided in ESI MS analysis. The presence of a high concentration of salts or impurities can greatly affect the ionization of the analytes and results in low ion intensities of analytes in ESI MS. Thus, in the last part of thesis, functionalized nanoparticles that are capable of selectively concentrating calcium ions from complex samples were employed for the sample treatment prior to electrodeless/sheathless ESI MS analysis. The results show that calcium in complex samples such as milk and serum can be quantitatively determined using this proposed approach. In conclusion, the sheathless interface presented herein is easy to be fabricated. Furthermore, it has a comparable sensitivity with the conventional one.