Development of Optimal Milling Procedures for Accurate Quantification of Psilocybin and Psilocin in Psilocybin Mushrooms Utilizing Fritsch Instrumentation.

Abstract: Psilocybin and psilocin, two psychoactive compounds found in various species of mushrooms, have recently attracted significant scientific and regulatory attention due to their therapeutic potential. Sample preparation should be performed in a way that provides a homogeneous sample, without introduction of artifacts or sample degradation, in an efficient manner. The Fritsch Pulverisette 11 meets these needs and future proofs sample analysis as matrices change. Problem Statement: Accurate quantitation requires homogenization and mushroom samples may change as they are incorporated into to a variety of food matrices e.g. chocolates, gummies, etc. A lack of sample uniformity i.e. heterogeneity can lead to inaccurate quantification and introduce an unacceptable level of error into analytical measurements thereby negatively impacting clinical trials. Methods: Dry and cryomilling of psychedelic mushrooms was investigated as well as refinement of RPM and duration settings. A comparison to alternative methods of milling was also performed. Flexible SOP programmability in the Pulverisette 11 allows for reproducible results independent of user or technique, a key requirement for GLP. Results: Wet, dry, and cryomilling techniques that provide process controls mitigate causes of sample degradation, e.g. heat buildup. In this case, dry milling offers several benefits including speed, reduced cost, greater reproducibility, and higher particle size uniformity as compared to rudimentary grinding methods. Conclusions/Implications: When relying on precise analytical methods to guide accurate dosing in clinical settings, reproducible and reliable sample preparation is paramount. The Pulverisette 11 not only meets these needs, but does so in a simple, straight forward software interface and is versatile for future needs as sample matrices change.