Determination of psilocybin in hallucinogenic mushrooms by reversed-phase liquid chromatography with fluorescence detection.

The determination of psilocybin was carried out by reversed-phase liquid chromatography (HPLC) with fluorescence (FL) detection. Psilocybin was labeled with 5-dimethylaminonaphthalene-1-[N-(2-aminoethyl)]sulfonamide (DNS-ED) at 60 degrees C for 4h in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) as the activation reagent. The resulting derivative was separated on a Mightysil RP-18 GP column (150mmx4.6mm, i.d. 3mum) with the mixture of 50mM ammonium acetate (AcONH(4)) and CH(3)CN, and detected at 539nm (excitation at 321nm). The structure of the derivative was identified by HPLC-ESI-MS. A good linear relation of the calibration curve of psilocybin was observed under the proposed conditions for labeling, separation and detection. The quantification limit was 4.4ng in 1mg dried mushroom. The proposed procedure was successfully used for the determination of psilocybin in real samples. The contents of psilocybin in six magic mushrooms by the proposed HPLC-FL method were less than 20.0ng in 1mg dried samples.