Psilocybin ( 1 ) is the major alkaloid found in psychedelic mushrooms and acts as a prodrug to psilocin ( 2 , 4-hydroxy- N , N -dimethyltryptamine), a potent psychedelic that exerts remarkable alteration of human consciousness. In contrast, the positional isomer bufotenin ( 7 , 5-hydroxy- N , N -dimethyltryptamine) differs significantly in its reported pharmacology. A series of experiments were designed to explore chemical differences between 2 and 7 and specifically to test the hypothesis that the C-4 hydroxy group of 2 significantly infuences the observed physical and chemical properties through pseudo-ring formation via an intramolecular hydrogen bond (IMHB). NMR spectroscopy accompanied by quantum chemical calculations was employed to compare hydrogen bond behaviour in 4- and 5-hydroxylated tryptamines. The results provide evidence for a pseudo-ring in 2 and that sidechain/hydroxyl interactions in 4-hydroxytryptamines influence their oxidation kinetics. We conclude that the propensity to form IMHBs leads to a higher number of uncharged species that easily cross the blood-brain barrier, compared to 7 and other 5-hydroxytryptamines, which cannot form IMHBs. Our work helps understand a fundamental aspect of the pharmacology of 2 and should support efforts to introduce it (via the prodrug 1 ) as an urgently needed therapeutic against major depressive disorder.