When very complex mixtures need to be analysed, separation techniques can be coupled with molecular spectroscopy for detection and/or identification. For example, in recent years the following systems have been developed:
-Liquid chromatography (LC) and capillary electrophoresis (CE) coupled to laser induced fluorescence in the visible and in the UV range, including miniaturization for chip-based analysis.
- CE with quenched phosphorescence detection, offering low detection limits (also suitable for chiral, mirror-image analytes).
- Liquid flow systems coupled with time-resolved fluorescence resonance energy transfer (TR-FRET) for the screening of biological activity of enzyme inhibitors.
- LC or CE separation in combination with surface-enhanced Raman spectroscopy, using new silver-coated substrates.
- LC-Raman applying a liquid core waveguide (LCW) for increased optical pathlengths.
- LC and CE combined with deep-UV Raman. At those short wavelengths fluorescence interference is strongly reduced, and via the resonance enhancement phenomenon a strong increase in sensitivity can be obtained, as well as extra selectivity over the solvent matrix.
An exciting new development (collaboration with Atomic, Molecular & Laser
Physics) is the use of cavity ring-down spectroscopy (CRDS) in the liquid state.
A 2-mm detector cell was constructed from two concave mirrors and connected
to an LC separation system. Short, 532-nm pulses will bounce back and forth
inside the cavity; a small fraction leaks out of the cavity and is measured
by a fast detector. The resulting decays (typically of the order of 10-100 ns,
corresponding to an effective pathlength of 2-20 m) are analyzed after each
laser pulse. A shorter-than-usual decay time indicates that an extra absorbing
species passes the cavity; with this setup extremely low absorbances can be
detected (see figure).
The cavity ring-down signal as
measured on the oscilloscope for
pure ethanol (blank; black curve)
and after injection of a 10 nM CVethanol
solution (red curve). Also
displayed is the detector response
curve (green). [After Bahnev et al.,
Analytical Chemistry, Vol. 77, 1188-