Several specimens of Psilocybe and Copelandia species in Koh Samui, Thailand were recently collected for herbarium deposit and scientific study. This paper presents an ethnomycological and biochemical study of one of the species; P. samuiensis Guzmán, Bandala and Allen, a new psychoactive gill fungus reported from Thailand. Mycelium for the cultivation of P. samuiensis was obtained on 6% malt agar from the spores of a dried specimen. The growth of P. samuiensis was similar to that of P. tampanensis Guzmán and Pollock, but more rapid than the mycelium of P. semilanceata (Fr.: Sacc.) Kumm. Laboratory analyses indicates that the alkaloid content in cultured fruit bodies of P. samuiensis is of the same order of magnitude as that found in naturally occurring mushrooms of this species. HPLC analyses of both naturally occurring and in vitro cultivated fruit bodies of P. samuiensis revealed high concentrations of psilocybin and psilocin. Small amounts of baeocystin were also detected. Psilocybin levels varied from 0.23% up to 0.90%. The psilocybin content was highest in the caps. Psilocybin was also found in the cultured non-bluing mycelia of P. samuiensis and varied from 0.24% to 0.32% dry weight. The relative alkaloidal content of psilocybin, psilocin, and baeocystin found in P. samuiensis was similar to that measured in many other psychoactive fungi species, but completely different from that found in P. semilanceata.