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High anthocyanin-producing cell lines, which were grown in a dark or in a light-dark regime, were selected from callus cultures initiated from stem and leaf tissues of Aralia cordata Thunb. by small-cell-aggregate selection. To verify the optimum culture conditions for anthocyanin production, cells were tested by changing the various basal media, sucrose concentration and nitrogen source and concentration. Good growth was obtained in the dark on Linsmaier-Skoog's basal medium containing 1.0 mg l-1 2,4-d and 0.1 mg l-1 kinetin, 2% (w/v) sucrose and full strength of nitrogen concentration. However, the highest anthocyanin yield (10.3% dry wt) was obtained in the dark on B5 medium containing 1.0 mg l-1 2,4-d and 0.1 mg l-1 kinetin. Our results suggested that it has became feasible to find the most effective conditions for cell growth and anthocyanin production by optimizations of the nitrogen concentration and the ratio of NH4 + to NO3 - in the medium.

Good results for anthocyanin production in dark-grown calli of Aralia cordata Thunb. were obtained on the addition of sucrose as the carbon source and NH4+ and NO3− at a molar ratio of 1:4 (15 mM total nitrogen). Also, by supplementing NAA as auxin and kinetin as cytokinin, good results were obtained. The anthocyanin yield of high-selected callus cultures was 13.0% maximum dry weight on being cultured for three weeks in darkness. The major anthocyanin, about 70% of the total pigment in the cells, was identified as cyanidin 3-xylosylgalactoside.