EFFECT OF C-GLYCOSYL FLAVONE FROM URGINEA INDICA ON ANTIBIOTIC INDUCED MICROBIAL CELL DEATH Original Article BEVARA GANESH BABU a , A. D. NAVEEN KUMAR a , ANIL BADANA a , SEEMA KUMARI a , ANJALI JHA b , RR MALLA a * a Cancer Biology Lab, Department of Biochemistry, GITAM University, Visakhapatnam, Andhra Pradesh, India 530045, b Department of Chemistry, GITAM University, Visakhapatnam, Andhra Pradesh, India 530045 Email: dr.rrmall@gmail.com Received: 12 Feb 2016 Revised and Accepted: 30 Mar 2016 ABSTRACT Objective: The aim of the present study was to isolate and characterize C-glycosyl flavone from Urgenia. indica bulb and to evaluate its effect on microbial cell growth. Methods: Methanolic extract of U. indica bulb was prepared using Soxhlation. Isolation was performed using silica gel column chromatography, and characterization was done based on IR, NMR and mass spectral data. The effect of C-glycosyl flavone was evaluated on microbial growth inhibition C- glycosyl flavone in terms of cytotoxicity, cell membrane damage, and cell death. Results: The results indicates that O-glycosyl flavanone (5,4 1 -di hydroxyl-3 1 -methoxy-7-O-[(α-L-rhamnosyl-(1[11] 1 -6[11])-β-D-glucopyranosyl)] flavanone), O-glycosyl flavone (5,4 1 -dihydroxy-3 1 -methoxy-7-O-β-D-glucopyranosyl flavone) and C-glycosyl flavone (5,7-dihydroxy-2-[4 1 -hydroxy-3 1 - (methoxymethyl) phenyl]-6-C-β-glucopyranosyl flavones) showed significantly sensitivity against Staphylococcus aureus, Bacillus subtilis, Rhizopus oryzae and Aspergillus niger. Synergism of C-glycosyl flavone on antibacterial activity of ciprofloxacin with the FIC index of 0.3 against S. aureus and 0.5 against B. subtilis. Antifungal activity of clotrimazole with the FIC index 0.3 against R. oryzae and 0.48 against A. niger. C-glycosyl flavone increased the ciprofloxacin-induced cytotoxicity from 63 to 91% against S. aureus and 56 to 89% against B. subtilis, whereas clotrimazole is induced cytotoxicity from 36 to 49% against R. oryzae and 23 to 41% against A. niger. C-glycosyl flavone increased the ciprofloxacin-induced cell death in S. aureus and B. subtilis and clotrimazole induced cell death in R. oryzae and A. niger as evident by propidium iodide staining, Tunel positive cells, and cytoplasmic membrane damage. Conclusion: The present investigation provides scientific and rationalism for the folkloric use of U. indica as an antimicrobial agent. Keywords: Antimicrobial, Synergism, Cytotoxicity, Cytoplasmic membrane damage and cell death © 2016 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/ ) INTRODUCTION Studies in the recent past have reported that diseases and disease- causing agents were recurring in new forms and resistant to existing antibiotics [1]. The emergence of multi-drug resistant microbial strains is limiting the effectiveness of current drugs and significantly causing treatment failure of infections [2]. To face such a challenge, there is a need to develop an alternative, approaches that can enhance the efficacy of antibiotics to target resistance. Combination therapy has been shown to be beneficial for drug- resistant infectious diseases which do not normally respond to single-drug therapy. Studies have demonstrated that the natural products enhanced the action of antibiotics against multidrug resistant microorganisms [3-4]. However, the mechanism of action of natural products on antibiotic-induced microbial death remains largely unknown. Programmed cell death (PCD) is a genetically controlled process associated with development and homeostasis of bacteria, fungi, plants and animals [5]. E. coli exhibited apoptotic markers such as exposure of phosphatidylserine, condensation of chromosomes and DNA fragmentation with antibiotic treatment [6]. R. oryzae cells treated with triazoles in combination with hyperthermia (42 °C) exhibited characteristic markers of early apoptosis such as externalization of phosphatidylserine, fragmentation, and condensation of DNA, membrane depolarization and increased metacaspase activity [7]. An apoptosis-like process similar to eukaryotes with activation of a caspase-3-like protease (C3LP) followed by DNA fragmentation was reported in B. subtilis [8]. A more recent study by Anandhi et al., (2014)[11] demonstrated marked DNA fragmentation in E. coli and S. aureus with glycosides and flavonoids [9]. PCD has been proposed as a potential target for new antibacterial therapy [10]. Flavonoids reported having good antibacterial potential with bacteriostatic activities against different Gram-negative bacteria such as E. coli and P. aeruginosa. They also possess an inhibitory effect on S. aureus and on the growth of spore-forming B. subtilis [11]. Flavonoids also reported having synergistic behavior in combination with other flavonoids and antimicrobial agents [12]. Earlier reports claimed that the natural compounds like alkyl gallates and gallic acid with different classes of β-lactams antibiotics showed maximum inhibitory activity in clinical isolates [13]. The antibiotic additive and synergistic action of flavonoids against methicillin-resistant S. aureus. C-glycosyl flavone are a class of glycosylated flavonoids with good health benefits. Tatsuya Hasegawa et al., (2008) [14] reported that C-glycosyl flavone exhibits stronger radical scavenging activity than ascorbic acid [15]. The C- glycosyl flavonoids showed higher antioxidant potential than their corresponding O-glycosyl flavonoids and aglycones [11]. Urginea indica is a rare, endangered and threatened medicinal plant belongs to Liliace family and commonly called as sea onion. Its bulbs are used to cure wound and infections [16]. Recently, antioxidant [17] and antibacterial activities of crude extracts from U. indica bulb were evaluated. However, antimicrobial efficacy and synergistic effect of purified glycosyl flavonoids from U. indica were not reported. In this study, one novel glycosyl flavonoid together with two known compounds was isolated from U. indica bulb. Synergistic antimicrobial efficacy isolated glycosyl flavonoids together with known antibiotics was evaluated against pathogenic microorganisms, E. coli, P. aeruginosa, S. aureus, B. subtilis, R. oyzae and A. niger. Further, the mode of synergistic action of C-glycosyl flavone on ciprofloxacin-induced cell death in S. aureus and B. subtilis or clotrimazole in R. oryzae and A. niger. International Journal of Pharmacy and Pharmaceutical Sciences ISSN- 0975-1491 Vol 8, Issue 5, 2016