Studies on improvement of multiwavelength detection high performance liquid chromatography for diagnostic

本研究开发了一种新型的高效液相色谱多波长检测系统。为了克服二极管阵列探测器在探测波长范围和灵敏度方面的局限性，该系统采用了光电倍增管作为光学探测器，因为它的灵敏度比二极管阵列大得多，而且它的探测波长范围比二极管阵列宽得多，特别是在紫外光小于200 nm的区域。开发了一种新的机械扫描系统，其中带有螺旋形狭缝的圆盘在光谱仪的像平面上旋转。当圆盘旋转时，狭缝的位置在光谱仪衍射的光谱上从左向右移动。通过狭缝的光被收集在光电倍增管上，并将光电倍增管的输出信号输入到基于微型计算机的信号处理单元。将该系统应用于多波长检测高效液相色谱系统中，分离出的化合物在190 nm和250 nm的紫外区有吸收光谱。利用所研制的多波长检测系统，对氨基酸、糖和羧酸进行了分析。在190~250 nm范围内获得了这些组分的三维色谱图，具有良好的信噪比。该系统还应用于多波长荧光检测。虽然作为荧光检测器的灵敏度还不够高，但证实了该检测系统作为多波长荧光检测系统用于高效液相色谱检测的可行性。

In this study, a new type of a multiwavelength detection system for high performance liquid chromatography was developed. To overcome the limitations of diode array detectors about their detectable wavelength range and sensitivity, the system adopted a photomultipliers as an optical detector since its sensitivity is much greater than that of a diode array and its detectable wavelength range is much wider than that of a diode array, especially in the region of ultra-violet light shorter than 200nm. A new mechanical scanning system was developed where a disk with a spiral shaped slit is rotating at the image plane of the spectrometer. The position of the slit moves from left to right on the spectrum diffracted by the spectrometer as the disk rotates. Light passing through the slit is collected on the PMT and PMT output signal is introduced to a microcomputer based signal processing unit. The developed system was applied to the multiwavelength detection HPLC system where the absorption spectra of separated compounds in the region from 190nm and 250nm in ultraviolet region. By the developed system, amino acids, sugars and carboxylic acids were analyzed by the developed multiwavelength detection HPLC system. Three dimensional chromatograpms of these components in the region from 190nm to 250 nm were obtained with good SIN ratio. The developed system was also applied to multiwavelength fluorescence detection. Although the sensitivity was not enough as a fluorescence detector, the feasibility of the detection system as an multiwavelength fluorescenece detection system for HPLC was confirmed.