| Summary: | <i>Cannabis sativa</i> L. has been used for a long time to obtain food, fiber, and as a medicinal and psychoactive plant. Today, the nutraceutical potential of <i>C.</i><i>sativa</i> is being increasingly reappraised; however, <i>C. sativa</i> roots remain poorly studied, despite citations in the scientific literature. In this direction, we identified and quantified the presence of valuable bioactives (namely, β-sitosterol, stigmasterol, campesterol, friedelin, and epi-friedelanol) in the root extracts of <i>C. sativa,</i> a finding which might pave the way to the exploitation of the therapeutic potential of all parts of the <i>C. sativa</i> plant. To facilitate root harvesting and processing, aeroponic (AP) and aeroponic-elicited cultures (AEP) were established and compared to soil-cultivated plants (SP). Interestingly, considerably increased plant growth—particularly of the roots—and a significant increase (up to 20-fold in the case of β-sitosterol) in the total content of the aforementioned roots’ bioactive molecules were observed in AP and AEP. In conclusion, aeroponics, an easy, standardized, contaminant-free cultivation technique, facilitates the harvesting/processing of roots along with a greater production of their secondary bioactive metabolites, which could be utilized in the formulation of health-promoting and health-care products.
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