本研究開發以表面增強拉曼光譜(surface-enhanced Raman spectroscopy, SERS)為基礎的快速篩選分析法。分析物包括化學結構式類似的孔雀石綠與結晶紫，兩者均為三苯基甲烷類染料。為了有效遮斷雷射(Nd:YAG第二倍頻波: 532 nm, 300 mW)造成的雷利散射光，以提高拉曼散射的偵測靈敏度，本研究首先架設了一組雙-單光器(double-monochromator)。該實驗系統除了能有效去除不必要的干擾以外，亦提供了絕佳的光譜解析度；解析度為 ± 2 cm-1 (相當於0.06 nm)。此外，本系統在實驗操作上不僅能快速更換樣品，以達快速分析的目的，並且能將樣品置入液態氮中，得到解析度更佳的超低溫SERS光譜。
一般的拉曼光譜，由於訊號微弱，難以做為快速篩選分析法之用。本研究則研發及嘗試各種不同粒徑的奈米銀。實驗發現配製直徑為~ 23 nm的奈米銀(最大吸收峰在波長408 nm)對孔雀石綠與結晶紫而言，均能提供非常明顯的SERS光譜。相較於拉曼訊號增強了~ 104倍。隨後為了探討分析物與奈米銀的作用情形，比較了拉曼與SERS光譜同範圍內各譜峰的相對強度與相對波數的改變。實驗發現孔雀石綠的表面增強拉曼光譜相較於拉曼光譜，譜峰在1221 (N-C、N(CH3)2彎曲)、989 (N-C伸縮)、941 (N(CH3)2 彎曲、N-φ伸縮) cm-1處，有明顯的藍光位移現象。已知影響藍光位移主要是因為氮原子與銀表面作用的大小不同，而相對強度的改變是因為存在垂直於銀表面的振動模式有所異動。據此，可推測出孔雀石綠分子是以垂直站立方式，矗立於奈米銀表面之上。此推論以市售電腦軟體(HyperChem 5.01)以理論計算的結果得到驗證。結晶紫則在1485、1446、1180、807 cm-1處有明顯的藍光位移，但是由於化學結構上與孔雀石綠分子不盡相同，沒有觀察到苯環的振動模式會影響相對強度。因此推論結晶紫是以平躺方式，趴臥在銀的表面上。經電腦軟體理論計算結果亦得到了驗證。這些譜峰位移與相對強度上的改變，乃是做為快速篩選定性分析上重要的依據。
孔雀石綠經常被拿來做為魚貨類的消毒劑或殺菌劑，最高含量不得超過2 ppb。一般對於孔雀石綠的檢驗方式以液相層析法最為普遍。但是層析分離法需要較長的時間，不足以應付例行性/低濃度孔雀石綠的檢驗工作。再者，這類樣品分析經常會受到螢光物質的干擾，實際上無法進行直接測量。本研究發現，在添加直徑為~23 nm奈米銀之後，可以非常有選擇性的增強孔雀石綠的SERS訊號。該訊號不僅可以有效抑制樣品中螢光性物質的螢光干擾，其特徵譜峰的存在，亦可做為快速篩選是否含有孔雀石綠。為了確認本方法的可行性，在添加了2 ppb孔雀石綠的實驗室魚缸水樣品進行偵測，可以正確無誤的指認孔雀石綠的存在。結晶紫則常以不同濃度，做為市售紅、藍、黑原子筆墨水的原料。對於刑事鑑識上的筆跡鑑定，結晶紫的黏稠性質，難以使用氣相層析質譜法，對不同濃度比例的墨水痕跡進行分析的工作。再者原子筆墨水當中除了結晶紫以外，還有許多未知的螢光性物質，實際上也無法進行直接測量。然而本研究發現，在添加直徑為~23 nm奈米銀之後，可以非常有選擇性的增強結晶紫的SERS訊號，可清楚分辯原子筆墨水的類別，藉此可達到筆跡鑑定的目的。

This study developed a rapid screening method on surface-enhanced Raman spectroscopy (SERS). A double-monochromator (resolution, 2 cm-1) was used to measure and record Raman and SERS spectra, using a doubled Nd:YAG (532 nm, 300 mW) diode laser as the light source to measure two triphenylmethane type non-fluorescent compounds, malachite green (MG) and crystal violet (CV). Besides, this system could be attended better resolution in SERS spectra by means of adding liquid Nitrogen.
The signal intensity was too weak for detection in Raman spectroscopy. Therefore, this study attempted to prepare different particle size of silver colloids. The particle size about 23 nm (λabs-max, 408 nm) could be obviously enhanced signal intensity in MG and CV cases. On the SERS mode, has been enhanced about 104 times by comparing with Raman spectra. Raman shift and relative intensity change were the main factor to influence the combination of silver colloids. In the experiment, there was apparent blue shift in MG and CV cases by comparing with Raman and SERS spectra. The correlation between Raman shift and the vibration mode of Nitrogen was demonstrated by HyperChem, a theoretical calculation software. One side, the vibration mode of benzene would change the relative intensity in MG case. According to these reasons, the combination of MG and silver colloids could be predicted to stand a way by perpendicularity. Nonetheless, the vibration mode of benzene would not been changed the relative intensity in CV case. Therefore, the combination of CV and silver colloids could be predicted with the horizontal type.
Malachite green, a synthetic dye, that is potentially dangerous to human health, has been used illegally in the treatment of certain fish diseases, mainly, against parasites in fishwater and marine fishes. Thus far, the current detection methods for MG include HPLC, LC-MS, and UV/CE-stacking methods, etc. Each method has unique advantages and disadvantages with respect to sensitivity, precision and simplicity of use. In this study, we found that MG can be easily detected and quantified using the SERS mode. On the SERS mode, the limit of detection for MG could be detected to 2 ppb. Thus, the method was also extended to the determination of MG in an actual sample. Crystal violet, an industrial dye, that is often used to be the material of ballpoint pan inks. Ballpoint pen inks were manufactured from a wide variety of materials that exhibit different chemical materials. By adding the silver colloids (diameter, ~ 23 nm), different ballpoint pen inks could be made out and the fluorescent compounds in the inks could not be detected. These proposed methods may solve problems that were frequently encountered on pen inks analysis.

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