Impact Of Callus Induction Conditions On Metabolic Diversity Of Calli Cell Lines Of Glycyrrhiza Sp

Traditional Chinese herbal medicinces (TCM) is facing the crisis of plantresource depletion due to over-exploitation and extensive uses. Glycyrrhiza sp. as oneof the most common Chinese herbal plant is depleting and could not meet the marketdemand. Plant cell and tissue culture could provide an alternative technology for thelarge-scale production of the pharmacologically-important secondary metabolites orbioactive fractions of these endangered Chinese herbal plants.The purpose of this study was to evaluate the impact of typical callus inductionconditions on secondary metabolite diversity of the calli cell lines of traditionalChinese medicinal plant Glycyrrhiza sp.(Glycyrrhiza) by combined chemicalanalysis and HPLC fingerprint. These induction conditions included Glycyrrhizaspecies, types of explants, light conditions, combination of hormones and hormonesconcentrations. Firstly, the induction ratio of calli (the number of explants producingcalli over the number of explants inoculated) was investigated. It was found that thetype of explants had the greatest impact with100%callus induction ratio fromradicles and hypocotyls, only30-70%from cotyledon. Other induction conditions hadno significant impact on calli induction ratio. All the calli induced grew healthy,except that the color of calli grown under dark was yellow or light yellow, and thecolor of calli grown under light was green.After the calli were subcultured to become stabilised under the same conditionsof their induction the diversity of secondary metabolites was evaluated by thecombination of chemical analyses of total phenolics and total flavonids, and HPLCfingerprint. The first evaluation was to use one way ANOVA to examine the impactof different calli induction conditions on the contents of total phenolics and totalflavonoids. The contents of total phenolics and total flavonoids in calli weresignificantly (P <0.05) impacted by Glycyrrhiza species, types of explants, basicmedia, light condition, and hormone concentrations; while combinations of hormoneshad significant impacted on the content of total phenolics (P≥0.05), but not on thecontent of total flavonoids. The second evaluation was to use Diversity Factor (DF)based on the comparison of HPLC finger-prints to examine these calli lines that havesignificantly different contents of total phenolics and total flavonoids. DF varies significantly for calli induced under differentinduction conditions and the highest DFreached0.451. Light condition and combinations of hormones had the greatest impacton the diversity of secondary metabolites of Glycyrrhiza calli. It was concluded thatthe types and quantities of secondary metabolites in calli cultured are significantlyinfluenced by different induction conditions. These results provide importantknowledge for the selection of suitable calli cell lines for the production ofpharmacologically-important secondary metabolites or bioactive fractions by in vitrocell and tissue culture of Glycyrrhiza sp.