In Vitro Production of Echioidinin, 7-O-Methywogonin from Callus Cultures of Andrographis lineata and Their Cytotoxicity on Cancer Cells.

Arifullah Mohammed,Yerra Koteswara Rao,Kishore K Chiruvella,Sathees C Raghavan,Madamanchi Geethangili,Rama Gopal Ghanta,Brett Neilan

doi:10.1371/journal.pone.0141154

Abstract

Andrographis lineata is an herbal medicinal plant used in traditional medicine as a substitute for Andrographis paniculata. Here, using mature leaf explants of A. lineata we demonstrate for the first time the callus induction established on MS medium containing 1.0 mg l–1 IAA. Dried callus was subjected to solvent extraction with acetone. Further the acetone residue was separated by silica gel column chromatography, crystallized and characterized on the basis of nuclear magnetic resonance (proton and c13) and liquid chromatographic mass spectroscopy. This analysis revealed the occurrence of two known flavones namely, 7-O-methylwogonin (MW) and Echioidinin (ED). Furthermore, these compounds were tested for their cytotoxicity against leukemic cell line, CEM. We identify that ED and MW induced cytotoxicity in a time- and concentration-dependent manner. Further increase in the LDH release upon treatment with ED and MW further confirmed our cytotoxicity results against leukemic cell line. Strikingly, MW was more potent than ED when compared by trypan blue and MTT assays. Our results recapitulate the utility of callus cultures for the production of plant specific bioactive secondary metabolites instead of using wild plants. Together, our in vitro studies provide new insights of A. lineata callus cultures serving as a source for cancer chemotherapeutic agents.

Highlights

We show isolation and structural characterization of echioidinin (ED) and 7-O-methywogonin (MW) by silica gel column chromatography followed by nuclear magnetic resonance and liquid chromatographic mass spectroscopy
We established callus cultures from mature leaf explants on Mass spectra (MS) medium fortified with auxins like naphthalene acetic acid (NAA), 2,4-D and indole acetic acid (IAA)
Organogenic callus was increased in its volume by subculturing callus segments on MS medium fortified with 1.0 mg l–1 IAA for every twenty days

Summary

Introduction

Previous studies with A. paniculata have demonstrated the utility of callus cultures for the production of secondary metabolites instead of using wild plants. We report for the first time establishment of callus cultures from leaf explants of A. lineata. Prior made silica gel coated plates was developed in different solvent systems with hexane, ethyl acetate and methanol extracts. Echioidinin (ED) and 7-O-methywogonin (MW) used in the present study are natural flavonoids purified from leaf callus of A. lineata.

Results

Conclusion