Isolation, quantification and anti‑microbial screening of flavonoids from callus of Vitex negundo Linn

Abstract

Context: Natural habitats for medicinal plant are disappearing fast and together with environmental instabilities, it is increasingly difficult to acquire plant‑derived compounds. This has prompted scientists to consider the possibilities of using cell cultures as an alternative supply for the production of plant natural products. Aim: The present investigation deals with the establishment of callus cultures from leaf explants of Vitex negundo, to extract flavonoids from the raised callus and to assess their anti‑microbial potential. Materials and Methods: Callus culture of Vitex negundo has been established on Murashige and Skoog’s medium supplemented with 2, 4‑D using leaf explants. Growth index (GI) was calculated for the tissues obtained after 2, 4, 6 and 8 weeks, respectively. Calli of 2, 4, 6 and 8 weeks old were used for extraction of total flavonoids (free and bound) using standard method. Flavonoid extract of 6 weeks old callus with maximum GI and maximum flavonoid content was then tested against Staphylococcus aureus, Agrobacterium tumefaciens (bacteria), Candida albicans (yeast) for their anti‑microbial potential. Minimum inhibitory concentration, minimum bactericidal/fungicidal concentrations and total activity were also evaluated. Results: GI was found maximum in 6 weeks old tissue (0.76). Total flavonoids from 6 week old tissue showed maximum zone of inhibition against A. tumefaciens. Kaempferol was identified from total flavonoids of 6 week old callus through, thin layer chromatography (TLC), preparative TLC, Melting point and Infra red spectral studies. Content of kaempferol in 6‑week‑old calli was also estimated using standard Spectrophotometric method, which came out to be 54.25 μg/mg d.wt (dry weight) of calli. Conclusion: The protocol of callus induction of V. negundo opens new vistas that could facilitate phytochemical production, extraction of pharmaceuticals and their screening against different human pathogens from the callus without harvesting the plant itself. Key words: Anti‑microbial potential, callus, flavonoids, Vitex negundo