以棉線為輔助大氣壓下游離質譜法之研究

Abstract

質譜法是目前最廣為應用的分析法之一，透過待測分析物質荷比的分析可得到分析物之質量及結構。大氣壓下游離法有別於其他游離法必須在高真空下進行，在常溫常壓下即可進行樣品游離。在我們過去的研究發現，將分析物液滴放置接近外加有高電壓的質譜進樣口前端，可使分析物液滴在高電場的誘導下產生極化效應，當電荷累積至一定的程度可形成泰勒錐，進而引發電噴灑游離產生氣相離子，而可被質譜偵測，此游離過程稱之為極化誘導電噴灑游離。而本論文的重點在於開發類似的游離法，但使用一般的棉線材料為樣品取樣基材及電噴灑噴頭，由於棉線由細小的纖維組合而成，樣品溶液可從樣品進樣端透過毛細現象被持續引導至棉線另一端。當放置此裝置至質譜進樣口前端，含有樣品溶液的棉線前端在質譜進樣口由於受到高電壓的極化效應，因而誘發液滴產生極化誘導電噴灑游離而產生氣相離子，進而可被質譜偵測，我們將此游離法稱為棉線輔助大氣壓下游離質譜法(CT-API)。棉線的前處理很簡單，僅需透過乙腈及水清洗並乾燥後，再將棉線直接浸泡於樣品溶液中即可放置質譜前端(～0.5 mm)進行質譜分析，研究中發現質譜進樣口所加的高電壓在大於2500伏特，即可誘發游離效應，且電壓愈高，游離效率愈好。本論文在偵測帶正電離子時，質譜進樣口所設定的電壓為-4500伏特。使用棉線輔助進行質譜分析時有兩種方式:一種是將棉線直接浸泡於樣品溶液，藉著毛細作用將樣品溶液持續導入棉線尖端已靠近加高電壓的質譜進樣口進行質譜分析;另一種是將棉線浸泡於有機溶劑與水混合溶劑後，將數微升的分析物點樣在棉線尖端即可進行分析。結果發現前者可得到較低之偵測極限。以精氨酸樣品為例，偵測極限約在~10-9 M左右。本游離法可用於游離有機小分子及大分子之生物樣品，可測得最高分子為分子量約66,000道耳吞之蛋白質分子。本方法有數項特點，包括樣品載體(棉線)便宜且僅需簡單的前處理步驟，以及分析物離子訊號穩定且持續時間長(> 15分鐘)等等。綜合這些優點，本方法應可進一步開發成可即時監控化學反應中反應物及產物離子隨著時間離子強度消長狀況的質譜法。如將棉線表面進行進一步修飾後，也可望開發與質譜連線的線上前處理及即時質譜分析的方法。

Mass spectrometry (MS) is one of the most common analytical techniques for molecular characterization. Molecular weights and structural information can be obtained based on the mass to charge ratios of analytes. Unlike other ionization methods that are operated in a high vacuum, atmospheric pressure ionization is conducted at ambient conditions. In our previous studies, we have discovered that sample droplets, placed close to the inlet of a mass spectrometer, which is applied with a high voltage, can be readily polarized by the high electric field provided by mass spectrometer. Owing to the polarization effect, the charges on the droplet are accumulated to a certain extent. Taylor cone is formed, leading to the generation of electrospray and gas phase ions for MS analysis. Such ionization techniques are called polarization induced electrospray ionization (PI-ESI). In this study, an ionization technique is established based on the similar concept. Cotton thread, which is composed of tiny fibers, is used as the substrate for sampling samples and as the ESI emitter for generation of electrospray. Thus, when placing one end of a thread to a sample solution, the sample solution can be continuously directly to the other end through the capillary action. When placing such a setup in front of the inlet of a mass spectrometry applied with a high voltage, the sample solution on the front end of the thread is polarized and ionized through PI-ESI processes. The resultant gas phase ions can be readily detected by MS. We named this ionization technique cotton thread-assisted atmospheric pressure ionization (CT-API). The pretreatment of thread only requires few steps by simply rinsing the thread with acetonitrile and water followed by drying. The rinsed thread can be readily used for assisting ionization of analytes in our setup. When MS analysis is conducted, one end of the thread for generation of PI-ESI has to be placed very close (~0.5 mm) to the MS inlet. The voltage set on the MS inlet should be higher than 2500 V. The higher the voltage is applied on the MS inlet, the better the ionization efficiency is obtained. Thus, the voltage is set to -4500 V when conducting MS analysis in the positive ion mode. There are two ways to conduct the MS analysis using a thread as the substrate. That is, by putting one end of the thread into a sample solution, the sample solution is directed to the front end of the thread by the capillary action for MS analysis. The other way is putting one end of the thread into the solvent containing organic solvent and water followed by depositing a few microliters of analytes on the front end of the thread that is placed close to the MS inlet. Our results show that the former method can achieve a lower detection limit than the latter one. The limit of detection of arginine is estimated to be ~ 10-9 M. Small organic molecules and large biomolecules can be readily analyzed using the current approach. The detectable molecular weight is as high as 66,000 Da. The advantages of this current method include simplicity and low-cost. The ion signals are stable and last long (>15 min). Thus, this ionization technique can be further developed to monitor the ions derived from reaction species in chemical reactions in real time by MS. In addition, it may be potentially possible to conduct on-line sample pretreatment with MS by using surface modified thread as the substrate.