OBJECTIVE: Demand for new antidepressants has resulted in a re-evaluation of the therapeutic potential of psychedelic drugs. Several tryptamines found in psilocybin-containing \"magic\" mushrooms share chemical similarities with psilocybin. Early work suggests they may share biological targets. However, few studies have explored their pharmacological and behavioural effects.
METHODS: We compared baeocystin, norbaeocystin and aeruginascin with psilocybin to determine if they are metabolized by the same enzymes, similarly penetrate the blood-brain barrier, serve as ligands for similar receptors and modulate behaviour in rodents similarly. We also assessed the stability and optimal storage and handling conditions for each compound.
RESULTS: In vitro enzyme kinetics assays found that all compounds had nearly identical rates of dephosphorylation via alkaline phosphatase and metabolism by monoamine oxidase. Further, we found that only the dephosphorylated products of baeocystin and norbaeocystin crossed a blood-brain barrier mimetic to a similar degree as the dephosphorylated form of psilocybin, psilocin. The dephosphorylated form of norbaeocystin was found to activate the 5-HT2A receptor with similar efficacy to psilocin and norpsilocin in in vitro cell imaging assays. Behaviourally, only psilocybin induced head twitch responses in rats, a marker of 5-HT2A-mediated psychedelic effects and hallucinogenic potential. However, like psilocybin, norbaeocystin improved outcomes in the forced swim test. All compounds caused minimal changes to metrics of renal and hepatic health, suggesting innocuous safety profiles.
CONCLUSIONS: Collectively, this work suggests that other naturally occurring tryptamines, especially norbaeocystin, may share overlapping therapeutic potential with psilocybin, but without causing hallucinations.

Since not only psilocybin (PSB) but also PSB-containing mushrooms are used for psychedelic therapy and microdosing, it is necessary to know their concentration variability in wild-grown mushrooms. This article aimed to determine the PSB, psilocin (PS), baeocystin (BA), norbaeocystin (NB), and aeruginascin (AE) concentrations in a large sample set of mushrooms belonging to genera previously reported to contain psychotropic tryptamines. Ultra-high performance liquid chromatography coupled with tandem mass spectrometry was used to quantify tryptamine alkaloids in the mushroom samples. Most mushroom collections were documented by fungarium specimens and/or ITS rDNA/LSU/EF1-α sequencing. Concentrations of five tryptamine alkaloids were determined in a large sample set of 226 fruiting bodies of 82 individual collections from seven mushroom genera. For many mushroom species, concentrations of BA, NB, and AE are reported for the first time. The highest PSB/PS concentrations were found in Psilocybe species, but no tryptamines were detected in the P. fuscofulva and P. fimetaria collections. The tryptamine concentrations in mushrooms are extremely variable, representing a problem for mushroom consumers due to the apparent risk of overdose. The varied cocktail of tryptamines in wild mushrooms could influence the medicinal effect compared to therapy with chemically pure PSB, posing a serious problem for data interpretation.

Psilocybin is a tryptamine-derived psychoactive alkaloid found mainly in the fungal genus Psilocybe, among others, and is the active ingredient in so-called \"magic mushrooms\". Although its notoriety originates from its psychotropic properties and popular use as a recreational drug, clinical trials have recently recognized psilocybin as a promising candidate for the treatment of various psychological and neurological afflictions. In this work, we demonstrate the de novo biosynthetic production of psilocybin and related tryptamine derivatives in Saccharomyces cerevisiae by expression of a heterologous biosynthesis pathway sourced from Psilocybe cubensis. Additionally, we achieve improved product titers by supplementing the pathway with a novel cytochrome P450 reductase from P. cubensis. Further rational engineering resulted in a final production strain producing 627 ± 140 mg/L of psilocybin and 580 ± 276 mg/L of the dephosphorylated degradation product psilocin in triplicate controlled fed-batch fermentations in minimal synthetic media. Pathway intermediates baeocystin, nor norbaeocystin as well the dephosphorylated baeocystin degradation product norpsilocin were also detected in strains engineered for psilocybin production. We also demonstrate the biosynthetic production of natural tryptamine derivative aeruginascin as well as the production of a new-to-nature tryptamine derivative N-acetyl-4-hydroxytryptamine. These results lay the foundation for the biotechnological production of psilocybin in a controlled environment for pharmaceutical applications, and provide a starting point for the biosynthetic production of other tryptamine derivatives of therapeutic relevance.

Psilocybin, the prodrug of the psychoactive molecule psilocin, has demonstrated promising results in clinical trials for the treatment of addiction, depression, and post-traumatic stress disorder. The development of a psilocybin production platform in a highly engineerable microbe could lead to rapid advances towards the bioproduction of psilocybin for use in ongoing clinical trials. Here, we present the development of a modular biosynthetic production platform in the model microbe, Escherichia coli. Efforts to optimize and improve pathway performance using multiple genetic optimization techniques were evaluated, resulting in a 32-fold improvement in psilocybin titer. Further enhancements to this genetically superior strain were achieved through fermentation optimization, ultimately resulting in a fed-batch fermentation study, with a production titer of 1.16 g/L of psilocybin. This is the highest psilocybin titer achieved to date from a recombinant organism and a significant step towards demonstrating the feasibility of industrial production of biologically-derived psilocybin.

Psilocybin and its direct precursor baeocystin are indole alkaloids of psychotropic Psilocybe mushrooms. The pharmaceutical interest in psilocybin as a treatment option against depression and anxiety is currently being investigated in advanced clinical trials. Here, we report a biocatalytic route to synthesize 6-methylated psilocybin and baeocystin from 4-hydroxy-6-methyl-l-tryptophan, which was decarboxylated and phosphorylated by the Psilocybe cubensis biosynthesis enzymes PsiD and PsiK. N-Methylation was catalyzed by PsiM. We further present an in silico structural model of PsiM that revealed a well-conserved SAM-binding core along with peripheral nonconserved elements that likely govern substrate preferences.