|M.Sc Student||Milay Looz|
|Subject||Evaluation of the Optimal Storage Conditions for|
|Department||Department of Biology||Supervisor||DR. David Meiri|
|Full Thesis text|
The number of patients using medical cannabis is consistently increasing worldwide. The therapeutic effects of cannabis largely depend on the content of its pharmacologically active secondary metabolites and their interactions, mainly terpenoids and phytocannabinoids. Terpenoids constitute an extremely diverse family of compounds that are present in many species including plants, animals and microorganisms. On the other hand, phytocannabinoids are unique compounds which have been almost exclusively identified in cannabis plants. Terpenoids and phytocannabinoids are biosynthesized by specific metabolic enzymes in the glandular trichomes, which are abundant on the surface of female inflorescences. Once harvested and during storage, these natural compounds may decarboxylate, oxidize, isomerize, photochemically react, evaporate or more. Hundreds of these degradation compounds have been identified in cannabis inflorescences at different concentrations. However, despite the widespread and increasing use of medical cannabis, there is little data on the stability of most of the plant's terpenoids and phytocannabinoids during storage. Therefore, in this study, we aimed to determine the optimal storage conditions for preserving the composition of the naturally biosynthesized secondary metabolites in cannabis inflorescences and cannabis extracts. To this end, cannabis inflorescences (whole versus ground inflorescences) and cannabis extracts (in DMSO, EtOH and olive oil) from two major groups, were stored at different temperatures (25, 4, -30 and -80 °C), and their phytocannabinoid and terpenoid profiles were analyzed for a whole year. Changes in phytocannabinoid compositions were evaluated by ultra-high-performance liquid chromatography ultraviolet detection or liquid chromatography mass spectrometry (UHPLC/UV or LC/MS). Changes in terpenoid compositions were measured with a static headspace sampler, followed by gas chromatography-mass spectrometry (SHS-GC/MS/MS). We found that storage at 25°C resulted in a statistically significant decrease in the concentrations of naturally occurring phytocannabinoids (and an increase in the concentration of degradation products) over time as compared to the lower temperature storage conditions (4, -30 and -80 °C). Moreover, storage at 25°C led to faster degradation of ground samples, as opposed to the whole samples. All terpenoid concentrations decreased over time. The most significant difference in the terpenoid profile occurred for ground inflorescences stored at -80 ºC for 4 months. According to these results, we concluded that storing whole inflorescences at 4 °C is optimal to maintain the original chemical composition of terpenoids and phytocannabinoids in cannabis inflorescences. For cannabis extracts, the optimal storage conditions were in olive oil and 4 °C. Moreover, we found that although CBN had the same trend of increasing during time in both types in cannabis inflorescences, the concertation at the type III (high CBD) chemovar was very low and therefore CBN cannot be used as an aging indicator, as it been use today all over the world. This study provides important information for cannabis growers, clinicians, manufacturers and distributors of medical cannabis, who need to provide standardized, pharmaceutical-grade cannabis inflorescences for patients.