Micropropagation for mass multiplication and enriched production of ascorbic acid in tissue culture foliage of roselle (Hibiscus sabdariffa L.)

An efficient protocol was developed for the micropropagation of Hibiscus sabdariffa through nodal explants (auxillary bud cultures) of in vitro seedlings. Nodal explants were able to induce multiple shoots on Murashige and Skoog (MS) basal medium supplemented with 2.0 mg L-1 6-benzylaminopurine, 0.5 mg L-1 indole-3-acetic acid, and 10 mu M silver nitrate (AgNO3; 7-8 shoots/explant) or 20 mu g L-1 triacontanol (TRIA; 4-5 shoots/explant). Medium devoid of AgNO3 that was supplemented with 0.1-0.5 mg L-1 gibberellic acid (GA(3)) induced the proliferation of up to two shoots/explant, but supplementation with 1-5 mg L-1 GA(3) induced a single shoot. The highest frequency of shoot elongation and rooting was obtained on 0.5-strength MS medium supplemented with 0.1 or 0.5 mg L-1 GA(3), and on this medium, further proliferation of shoots was also evident. Micropropagated plants were hardened in the greenhouse and successfully established in soil. Ascorbic acid level in leaves of micropropagated plants from AgNO3- and TRIA-supplemented media was highest (2.24 +/- A 0.14 and 2.18 +/- A 0.13 mg g(-1) fresh weight, respectively), which was 2.8-fold more than normal ex vitro leaves of the same age (0.79 +/- A 0.03 mg g(-1) fresh weight).