Antioxidative enzyme activities in calli during in vitro morphogenesis of Sarpagandha (Rauwolfia serpentina L.)

Subhadra R. Mallick, Priyadarsani Samal, Ram C. Jena, Kailash C. Samal


Background: A reproducible and efficient regeneration system for production of uniform, elite, true to the type plantlets for large
scale in situ and ex situ plantation is required to conserve the endangered medicinal plant Sarpagandha (Rauwolfia serpentina L. Benth. ex. Kurz). Aims: The aim of the present investigation was to assess antioxidative enzyme activities during the differentiation and regeneration of Sarpagandha and its role in catalytic scavenging of the reactive oxygen species for free radicals generated during in vitro morphogenesis. Settings and Design: The experiment was formulated using complete randomized design having 20 replicates per each treatment. Each experiment was repeated three times. Materials and Methods: The different explants of Sarpagandha
were cultured aseptically in the Murashige and Skoog (MS) basal medium containing 3% sucrose (w/v), 0.7% agar and different
concentration and combination of plant growth regulators. The antioxidative enzymes: catalase (EC, ascorbate peroxidase (EC, superoxide dismutase (EC and guaiacol peroxidase (EC were assayed employing standard biochemical procedure. Statistical Analysis: Data recorded on the different parameters were subjected to suitable statistical analysis based on the sample means of the various characters using SAS statistical software (SAS version 9.1, SAS Institute Inc. [Analysis of variance (ANOVA) was used to test statistical significance. The difference between significant treatments means and interactions were tested
against C.D. at 5% (P ≤ 0.05). Results: The maximum catalase (1.7 nkat/g fresh callus tissues) and superoxide dismutase activity (3.5 nkat/g fresh callus tissues) were observed in the shoot forming calli at the 4th week of culture. Shoot forming calli recorded the highest guaiacol peroxidase enzyme activity (2.6 nkat/g fresh callus tissues) at the 8th week of culture period whereas ascorbate peroxidase activity in the shoot forming calli at 1st week of culture recorded the maximum (0.3 nkat/g fresh callus tissues). Higher antioxidative enzyme activity was observed in shoot forming callus as compared to the non-shoot forming callus irrespective of enzymes studied. Conclusions: Higher enzyme activity was responsible for catalytic scavenging of the reactive oxygen species for free radicals and this could have led to a wholesome transformation of the component callus cells from an oxidized state to a more reduced state, the latter eventually become accountable for triggering the onset of embryogenesis.
Key words: Antioxidative enzyme activities, Callus, in vitro regeneration, R. Serpentina

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