Effect of Plant Growth Regulators and Subculture Frequency on Callus Culture and the Establishment of Melastoma malabathricum Cell Suspension Cultures for the Production of Pigments

Friable callus of Melastoma malabathricum could be induced from the leaf explants on MS medium supplemented with 1 mg L super(-1) N6-benzylaminopurine (BA) + 6 mg L super(-1) 1-naphthaleneacetic acid (NAA). Continuous subculturing of the friable calluses on the callus induction medium could increase the callus biomass at every 4 weeks subculture cycle. The production of callus biomass became stable with a growth index of 10 or more after the tenth subculture cycles. The cell suspension culture of M. malabathricum was initially established by culturing the leaf-derived friable callus in the liquid callus induction medium. The cells subsequently grew healthily and maintained well in MS liquid medium supplemented with 0.25 mg L super(-1) BA and 0.5 mg L super(-1) NAA. The growth kinetics of M. malabathricum cells followed a general growth pattern of a sigmoid curve. However, browning occurred when the cultures reached the highest fresh cell mass. In order to maintain healthy cultures, subculturing had to be done before browning occurred. After nine days of culture, cell inoculums of 0.75 and 1.0 g could produce fresh cell mass of 5.560 and 5.147 g, respectively. On the 15th day, cell cultures with initial inoculums of 0.25 and 0.5 g produced fresh cell mass of 5.416 and 6.150 g, respectively with cell mass increased of 22 and 15 fold, respectively. To maintain the cultures, an initial inoculum of 0.25 to 0.5 g of cells could be used and subcultured between 9 and 15 days after initial inoculation.