On-line fourier transform infrared detection in capillary electrophoresis.

Abstract

The coupling of Fourier transform infrared (FT-IR) spectroscopy as a new on-line detection principle in capillary electrophoresis (CE) is presented. To overcome the problem of total IR absorption by the fused-silica capillaries that are normally employed in CE separations, a micromachined IR-transparent flow cell was constructed. The cell consists of two IR-transparent CaF2 plates separated by a polymer coating and a titanium layer producing an IR detection window, 150 microm wide and 2 mm long, with a path length of 15 microm. The IR beam was focused on the detection window using an off-axis parabolic mirror in an optical device (made in-house) attached to an external optical port of the spectrometer. The connections between the fused-silica capillaries and the flow cell were made by a small O-ring of UV-curing epoxy adhesive on the sharply cut ends of the capillaries, allowing the capillaries to be easily replaced. Aqueous solutions comprising mixtures of adenosine, guanosine, and adenosine monophosphate were used to test the system's performance. Conventional on-line UV detection was employed to obtain reference measurements of analytes after the IR detection flow cell. The limit of FT-IR detection for all analytes (in absolute amounts) was in the nano- to picogram range corresponding to concentrations in the low-millimolar range.