Emission wavelengths, of fluorescent

HPLC was proven to be the most important instrumental analytical method for determination of EAs. Separation on HPLC and downstream detection by fluorescence detector or tandem mass spectrometer was performed. In the early 1970s, when the semisynthetic lysergic acid diethylamide 28 (LSD) became available in the drug scene, there was the need of a suitable analytical method for the detection of LSD. Since then, many HPLC analyses have been developed for the detection of EAs. Initially, normal phase HPLC was used for the determination of 4 with subsequent fluorescence detection. Today, reversed phase with C18 column materials is used more frequently for analysis of EAs [63, 71-73]. As mentioned before, EAs differ often in the position of the double bond in ring D of the 2. Clavine-type EAs contain sometimes a double bond at C-8 and C-9, whereas 3 and 4 carry a double bond at C-9 and C-10 instead, which influences the chromophoric features and is therefore a key parameter for the choice of excitation and emission wavelength of fluorescence detection. Extract mixtures containing EAs with a double bond at different positions (C-8 and C-9 or C-9 and C-10) should be analyzed in two runs to ensure the detection and quantification of the complete EAs. Another possibility is to use two fluorescence detectors subsequently [74]. 

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