HPLC with UV/Vis diode array detector
The most important combination in high performance liquid chromatography (HPLC) is the combination of chromatographic system and UV/Vis diode array detector and is the method of choice for routine qualitative and quantitative substance determinations. Beside retention time the UV/Vis-spectrum provides a second parameter for identification which allows a rapid assignment to a known reference compound by means of automated comparison with a spectra data base.
LC-MS (mass selective detection)
In LC-MS the output of an HPLC is fed into a mass spectrometer. Linkage of both methods results in a mass spectrum for each point of a chromatogram. In this way it is possible to deduce the chemical structure of individual compounds in a mixture. The use of high-resolution mass spectrometer allows the direct determination of the empirical formula of all components.
Linking liquid chromatography and NMR spectroscopy is one of the most effective analytical methods for structure determination of compounds in mixtures. In principle three forms of LC-NMR linkage are distinguished: continuous-flow-, stopped-flow- and the state of the art LC-SPE-NMR-technique. This enables directed enrichment of individual substance peaks by post-column peak-trapping and thus permits structure determination of secondary components in complex mixtures like extracts of natural compounds or samples from stress tests of active pharmaceutical ingredients.
Only for long time projects
Linkage of gas chromatograph and mass spectrometer is one of the most effective analytical techniques available for volatile substances with relatively low molecule mass (<1000 Daltons). As with LC-MS a mass spectrum is generated for each point of the chromatogram. The automatic calibration with commercial databases which contain more than 100.000 compounds allows for an effective identification of the spectra of unknown probes. This makes GC-MS an effective technique in the analyses of metabolites.