Isolation and Characterization of a Bioactive Phenylpropanoid from Ocimum sanctum L. Leaves through Chromatographic and Spectroscopic Methods

Abstract

Aims: This study aims at characterization of purified active ingredient isolated from commonly available botanical plant, Ocimum sanctum L. and to assess its potential for deployment in integrated disease management strategy of a staple food crop rice (Oryzae sativa L.) and rice based crop groundnut (Arachis hypogea) as an environment friendly approach for management of serious diseases of these two crops in lieu of synthetic chemicals. Study Design: Spectrophotometric and chromatographic methods were used.  Bioassay experiments were designed as per Complete Randomized Design (CRD) and statistical analysis was done using Cropstat 7.2 developed by International Rice Research Institute (IRRI).  Place and Duration of Study: Laboratory of Natural Plant Products, Crop Protection Division, Central Rice Research Institute, Cuttack-753006 (Indian Council of Agricultural Research), Odisha, India. Methodology: Partial purification of active principle was conducted through Column Chromatography in polarity gradient solvent systems followed by single spot isolation of active compound and mass collection through Thin Layer Chromatography (TLC).For characterization, Spectroscopic techniques viz., UV-Vis, FT-IR, 1H-NMR, 13C-NMR, DEPT, Mass and elemental analysis were conducted. For screening of different partially purified fractions of column against Pyricularia grisea, Conidial Germination Test method was preferred. To assess the fungitoxic activity of active principle against mycelial growth of five phytopathogenic fungi viz., P. grisea, Helminthosporium oryzae, Rhizoctonia solani, Curvularia lunata and Aspergillus niger, Poisoned Food Technique was used. Results: FT-IR data of active compound was in conformity with FT-IR of eugenol available in literature, 1H-NMR revealed presence of 12 protons, 13C-NMR showed the presence of 10 C atoms, DEPT along with 13C-NMR corresponds to the presence of substitution at 1, 2 and 4 positions of benzene ring. Mass spectrum reveals the presence of base peak at 164 that corresponds with the molecular weight of eugenol. Elemental analysis showed that percentages of C, H and O in the isolated compound were in conformity with calculated values of those in eugenol. The compound was found to exhibit broad spectrum fungitoxicity against five fungal pathogens tested. Conclusion: The isolated and purified compound from O. sanctum (Krishna i.e. Black) was characterized as Eugenol. It may be further used as an eco-friendly organic fungicide but only after developing suitable formulation and its evaluation under in-vivo conditions.