Callus Production Protocol for Aegle marmelos (L.) Corr.: As a Tool for Extraction of Secondary Metabolites

Abstract

Background: Aegle marmelos (L.) Corr. is an important medicinal and a fruit tree belongs to the family Rutaceae possessing numerous valuable secondary metabolites. The growing commercial importance for secondary metabolites has led to a great demand in the pharmaceutical industry in recent years. Therefore, an efficient callus production protocol was developed as a tool for extracting valuable secondary metabolites from Aegle marmelos.Methods: For seeds, callus induction was observed under three conditions as with seed coat, after removing seed coat and split into two halves after removing seed coat. For callus multiplication, 1cm2 pieces of initiated calli were used. These explants were established in MS medium supplemented with combinations of 2, 4 D either with BAP or Kinetin. All experiments were arranged according to the completely randomized design (CRD) with 20 replicates at the Tissue Culture Laboratory, Department of Crop Science, Faculty of Agriculture, University of Ruhuna, Sri Lanka, for a period of 1 year. Percentage of fungal and bacterial contaminations and percentage of bleached explants were observed to select the best explant/s. Percentage of responded explants were observed to select the best condition for callus induction and quality of callus. Growth of callus was observed visually by giving a score. Best hormonal combination for callus multiplication was observed as fresh weight and dry weight of callus produced under each treatment.Result: High quality callus with higher growth was observed in all combinations of BAP and 2, 4 D tested: ranging from 0.5 mgL-1 to 1.5 mgL-1 BAP and 1.0 mgL-1 to 2.0 mgL-1 2,4 D in Murashige and Skoog (MS) medium. Initiated calli were further multiplied in MS medium supplemented with 2,4 D combined with either BAP or Kinetin. Highest amount of callus biomass was recorded in the MS medium with 0.5 mgL-1 2, 4 D and 1.0 mgL-1 Kinetin (132.58 gL-1 fresh weight). The optimized protocol could be used to produce higher amount of callus in order to extract secondary metabolites from Aegle marmelos (L).