Effects of different factors on friable callus induction and establishment of cell suspension culture of Hovenia dulcis (Rhamnaceae)

Abstract

Medicinal plants are an important therapeutic option for a large share of the world’s population. To establish an in vitro culture system for the production of secondary metabolites from Hovenia dulcis, we studied the effect of auxins, cytokinins, absence of light, and silver nitrate on the development of friable callus. Callus cultures were established for the first time and used to obtain cell suspension cultures. Supplementation with KIN (Kinetin) produced calli with both compact and friable areas, while the addition of TDZ (Thidiazuron) only produced compact callus. The maintenance of cultures in the dark induced a slight enhancement on friability when the auxin PIC (Picloram) was present in the culture medium. The addition of silver nitrate promoted the formation of friable calli. Dry weight analysis showed no significant differences in biomass growth, and, therefore, 2.0 mg.L^-1 was considered the most suitable treatment. The presence of silver nitrate was not required for the establishment of cell suspension cultures. Dry weight analysis of cell suspensions showed higher biomass production in the absence of silver nitrate. PIC promoted 100% of cell suspension culture formation in the absence of silver nitrate, and higher biomass production was observed with the lowest concentration (0.625 mg.L^-1). No morphological differences were observed among the different concentrations of PIC. Phytochemical screening showed the presence of saponins, flavonoids, flavonols and catechins in the extracts obtained from H. dulcis calli. These results show that the cell cultures herein established are potential sources for the production of H. dulcis secondary metabolites of medicinal interest.

Key words
cytokinins; medicinal plant; plant tissue culture; saponins; silver nitrate