Exploring the Frontiers of Psychedelics: A New Chromatographic Method for Detection and Quantification of Psilocybin and Psilocin in Psilocybe cubensis Mushrooms

Abstract

Innovative therapies, such as psilocybin-assisted psychotherapies, hold great promises for treating anxiety, depression, and various other mental health disorders, addressing some of the challenges faced by conventional psychiatric medicine. This study focuses on developing and validating an HPLC-DAD methodology for detecting psilocybin and psilocin in extracts from psychedelic mushrooms intended for medicinal use. The methodology to has been validated following the guidelines of RDC No. 166/2017 from the Brazilian National Health Surveillance Agency (ANVISA). Utilizing a PerkinElmer C18 column (150.0 mm length × 4.6 mm internal diameter × 5 μm particle size), the gradient method employed ultrapure Milli-Q water (mobile phase A) and acetonitrile (mobile phase B), both acidified to 0.3% with formic acid (starting at a ratio of 95:5 v/v), with a flow rate of 0.8 mL/min over 18 min and an injection volume of 15 μL. The column temperature was maintained at 30 °C, and the analysis was conducted at a wavelength of 266 nm. The limit of detection (LOD) and limit od quantification (LOQ) values for psilocybin were 1.58 and 4.78 mg/L, respectively, while for psilocin, they were 1.70 and 5.17 mg/L. The method's accuracy showed recovery intervals for psilocybin ranging from 80 to 120% and for psilocin from 98 to 116%. Accurately determining the content of psilocybin (2.57%) and psilocin (0.16%) is essential for their application as pharmaceutical compounds. This methodological rigor ensures that these substances can be reliably used in therapeutic contexts, underscoring the importance of precise quantification in developing safe and effective medical treatments.