Liquid-core waveguide technology for coupling column liquid chromatographyand Raman spectroscopy

The on-line coupling of liquid chromatography (LC) and Raman spectroscopy ( RS) via an entirely plastic liquid-core waveguide (LCW) was optimized in te rms of excitation wavelength of the laser, especially in relation to the fl uorescence background, and the length of the LCW. Excitation at 632.8 nm (H e-Ne laser) was found to be a good compromise between a wavelength long eno ugh to strongly reduce the fluorescence background and, on the other hand, short enough to avoid (re)-absorption of laser light and Raman signals by H 2O in LCWs of considerable length. This conclusion is supported by a theore tical discussion on the optimization of LCW lengths as function of the exci tation wavelength for H2O and (H2O)-H-2. When using the He-Ne laser the opt imum length is similar to 50 cm for H2O; this corresponds to a detection ce ll volume of 19 mul for an LCW of 220 mum I.D., which is fully compatible w ith conventional-size LC. The influence of an organic modifier, usually nec essary for reversed-phase LC, on the free spectral window was evaluated. Th e potential applicability of LC-LCW-RS was shown for a mixture of adenosine 5 ' -monophosphate (AMP), guanosine 5 ' -monophosphate (CMP) and uridine 5 ' -monophosphate (UMP), utilizing an aqueous eluent without the addition o f a modifier. Improved detectability was achieved by using the stopped-flow mode and applying a large-volume-injection procedure (injection volume: 20 0 mul). Under these conditions, the limit of identification for AMP, GMP an d UMP was in the 0.1-0.5-mg/ml range. (C) 2001 Elsevier Science B.V. All ri ghts reserved.