Direct Quantitation of Psilocybin and Psilocin by One-Dimensional 1H and 31P qNMR in a revived Greek specimen of Psilocybe cyanescens

The genus Psilocybe of Basidiomycota includes more than two hundred species of mushroom-forming fungi, which are widely known for the production of the secondary metabolite psilocybin, a prodrug of its in vivo dephosphorylated active metabolite psilocin [1]. Psilocybin is being currently used in numerous clinical trials including the treatment of Major Depressive Disorder (MDD) [2], existential distress of terminally ill patients [3] and Alcohol Use Disorder (AUD) [4], creating the need of investigating and quantitating these substances.

Αn experimental study was carried out in order to test the viability of basidiospores coming from dried specimens. Among the various specimens of basidiomycetes kept at the Dried Specimen Collection of the Mycetotheca ATHUM (ATHens University Mycetotheca) was a specimen identified as Psilocybe cyanescens Wakef. [5]. Surprisingly, the spores of the specimen dried almost twenty-three years ago were germinated, producing mycelium which was subsequently studied for the production of the secondary metabolites of interest.

The lyophilized mycelium was extracted with methanol via ultrasounds. In order to verificate the presence of psilocybin, a dephosphorylation step was devised and the extract was subjected to GC/MS analysis, that revealed the presence of the dephosphorylated product psilocin. The initial and dephosphorylated extracts were analyzed with 1H and 31P NMR in D2O using calcium formate and dimethyl methylphosphonate as internal standards, respectively. To our knowledge, this is the first time a qNMR method is utilized for the analysis of psilocybin and psilocin containing fungi, without the need of separation and reference standards in less than five minutes.

Fig. 1 NMR spectra of P. cyanescens: (a) and (b) ¹H NMR before and after dephosphorylation, respectively; (c) and (d) ³¹P NMR before and after dephosphorylation, respectively.

Publikationsverlauf

Artikel online veröffentlicht:
12. Dezember 2022

• References

• 1 Hofmann A, Heim R, Brack A. et al. Psilocybin und Psilocin, zwei psychotrope Wirkstoffe aus mexikanischen Rauschpilzen. Helv Chim Acta 1959; 42: 1557-1572
• 2 Davis AK, Barrett FS, May DG. et al. Effects of Psilocybin-Assisted Therapy on Major Depressive Disorder: A Randomized Clinical Trial. JAMA Psychiatry 2021; 78: 481
• 3 Agin-Liebes GI, Malone T, Yalch MM. et al. Long-term follow-up of psilocybin-assisted psychotherapy for psychiatric and existential distress in patients with life-threatening cancer. J Psychopharmacol 2020; 34: 155-166
• 4 Bogenschutz MP, Forcehimes AA, Pommy JA. et al. Psilocybin-assisted treatment for alcohol dependence: A proof-of-concept study. J Psychopharmacol 2015; 29: 289-299
• 5 Delivorias P. A systematic and ecological study of macrofungi og the mountainous region of Agrafa (Greece) [PhD Thesis]. National & Kapodistrian University of Athens; 2014