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Bosco Lawarence, Aswathy J. M., Greeshma G. M., Sumyya S. S., Kavitha C. H. and K. Murugan*


Bridelia retusa is traditionally used as an astringent and also for rheumatic pains. Present study reveals that 2, 4-D either alone or in combination with kinetin in MS medium showed good response in terms of callus induction from leaf explants of B. retusa. Significant amount of callus was induced with 2.5 mg/L N6-benzyladenine (BA) + 2 mg/L 2, 4-dichlorophenoxyacetic acid (2, 4-D) i.e., 98.9%. The optimal fresh and dry weights of the calli were i.e., 1.9 ± 0.04 and 0.45 ± 0. 03 g respectively. The calli incubated in light on MS medium with 4% glucose containing 2.5 mg/L BA and 2 mg/L 2, 4-D at pH 3.5 yielded 2.8 mg /g of anthocyanin. MS medium with glucose was optimal compared with sucrose for inducing anthocyanin synthesis. However addition of KIN inhibited anthocyanin accumulation. Pigmented calli transferred to half strength MS medium containing NH+/NO3 (1:4 ratio), 70g/L sucrose with BA + 2, 4-D yielded remarkable biomass with anthocyanin content. Suspension cultures in half MS medium containing 2.5 mg/L 2, 4-D + 2 mg/L BA at pH 5.8 enhanced anthocyanin synthesis. pH and temperature factors against anthocyanin extraction and stability were analyzed. Enhancement in the degradation rate constant with a corresponding decline in the t1/2 values was seen with the increasing temperature at pH 1 and 4. Fractionation of anthocyanin was carried out by HPLC coupled mass spectrometry revealed 9 fractions comprising acylated cyanidins, two peonidins, cyanidin 3-p-coumaroyl and feruloyl diglucoside-5-glucosides. Hepatoprotective experimental results revealed that the anthocyanin extract at 200 and 400 mg/kg of body weight remarkably reversed the changes in serum biochemical parameters and also histology of liver caused by CCl4 indicating their hepatoprotective efficacy. Results were comparable with silymarin. The data was further supported by antioxidant enzyme profiles.

Keywords: Bridelia retusa, in vitro culture, phytohormones, anthocyanin, hepatoprotective, antioxidant.

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