LED激發高效液相層析螢光偵測法之相關研究

Research about HPLC fluorescence detector with LED as a light source

楊馥維(Fu-Wei Yang)

指導教授 ： 陳建銘

國立臺北科技大學/電資學院/光電工程系研究所/碩士(2011年)

https://doi.org/10.6841/NTUT.2011.00384

摘要

本研究使用高亮度發光二極體(LED)做為高效液相層析儀(HPLC)螢光偵測器的光源。雖然LED的窄頻譜特性(FWHM 30 nm)無法包含多種螢光分子所需的激發波長，卻適用於研究單一種螢光分子。本研究使用單顆中心波長在505 nm，1W的高功率青色光LED，激發待測物：3-羥基丁酸(3-hydroxybutyrate, 3-HB)。完成3-HB的同日間(Intra-day)、異日間(Inter-day)檢量線，濃度分別是7.93，15.86，39.65，79.30，158.60和237.90 (μM)。由檢量線得知，相同濃度待測樣品的螢光訊號再現性良好，相對標準偏差(RSD)<13%，在此濃度範圍內線形狀況良好(R2 = 0.997)，可媲美傳統光源氙燈的線性度。
為了提升偵測螢光訊號的訊雜比(S/N)，使用光阻斷器(chopper)將LED改為閃爍式光源，並使用鎖相放大器從螢光偵測器的輸出訊號中挑選出與閃爍頻率相同的訊號，得到訊號處理過的層析圖，並對結果進行討論。

關鍵字

發光二極體 ； 高效液相層析儀 ； 螢光偵測法 ； 3-羥基丁酸 ； 檢量線 ； 光阻斷器 ； 鎖相放大器

並列摘要

This research use high power cyan LED as a light source to excite fluorescence in HPLC. Although LED’s narrow spectrum couldn’t excite all kinds of fluorochrome, but is useful for a specific kind of analyst. This research use a single LED (1W, center wavelength 505 nm) to excite 3-hydroxybutyrate(3-HB). After completed Inter-day and Intra-day calibration curve, concentration at 7.93，15.86，39.65，79.30，158.60 and 237.90 (μM), we found that the calibration curve has good precision (RSD <13%) and good linearity (R2=0.997) .
To enhance the S/N ratio of fluorescence signal, we use chopper to make LED a pulse light source and use lock-in amplifier to choose signal which frequency is equal to the pulse LED from the output of fluorescence detector. And get a signal processed chromatogram, and gave some discussion about the result.

並列關鍵字

LED ； HPLC ； fluorescence detector ； 3-HB ； calibration curve ； chopper ； Lock-in Amplifier

參考文獻

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