Licorice root is traditionally used in medicine due to its content of saponins and flavonoids. Licorice leaves are not used as pharmacopoeial raw materials, although their chemical composition and biological activity have been studied recently, which makes it possible to evaluate the possibilities of using this raw material as a medicinal one. Active harvesting of licorice can jeopardize its natural populations, so urgent tasks are to cultivate cells of this plant in systems in vitro and to study the composition of cell culture metabolites. In our study, the material for obtaining a callus culture of licorice leaves was plants from the collection of the Botanical Garden of the Ural Branch of the Russian Academy of Sciences. To select optimal conditions for growing calli, nine combinations of phytohormones were compared. The best growth of calli was found on Murashige-Skoog medium with a combination of phytohormones 1 mg/L BAP and 10 mg/L NAA. Under these conditions, flavonoids accumulated in callus in quantities comparable to their content in intact leaves and roots. The content of phenolic compounds was comparable to their amount in the roots. Ethanol extracts obtained from callus culture had pronounced antioxidant activity comparable to extracts from the intact plant and standards of rutin, gallic, and ascorbic acids. When assessing the effect of extracts on animal cell cultures in the MTT test, it was shown that all obtained extracts increased the metabolic activity of both normal human cells and the HeLa line. At the same time, the extract obtained from the leaves showed the maximum effect, while that from the callus showed the minimum and differed slightly from the root extract. Thus, calli from leaf explants can be considered as a new raw material for obtaining dietary supplements with antioxidant activity.