Preeti Sirohi*, Alok Kumar Yadav1 and Nand K. Singh2
Affiliation: Department of Biotechnology, Motilal Nehru National Institute of Technology Allahabad, Prayagraj-211004 (U.P) India
1,2Department of Biotechnology, Motilal Nehru National Institute of Technology Allahabad, Prayagraj-211004(U.P)India
Email:rbt1503@mnnit.ac.in
Received-03.09.2019, Revised-22.09.2019
Abstract: Bacillus related microbes are mainly found in the severe cases of foodborne infection and many other lethal diseases. Few herbal medicines are effective against Bacillus licheniformis, but their mechanism of action is not reported. The antimicrobial potential of herbal extract from Clerodendrum phlomidis (n-hexane and dichloromethane extract) was assessed by well diffusion assay which showed a zone of inhibition of 19 and 15.5 mm respectively. Underlying mechanism of action behind the n-hexane and dichloromethane extract of C. phlomidis is reported by an in vitro study over Bacillus licheniformis through FACS, SEM, and DNA fragmentation analysis. Fluorescent activated cell sorting was done to determine membrane potential disruption caused by extracts that revealed dichloromethane has more activity for membrane potential disruption. The SEM analysis of bacteria showed irregular structure in cell wall, blebbing and leakage of cellular contents and disruption of membrane. DNA fragmentation analysis showed a precise smear formation of bacterial cellular DNA providing evidence of its apoptosis. The FTIR spectroscopic analysis of herbal extract was done to determine the dominance of functional groups present within purified antimicrobial extracts and which had shown the dominance of Alcoholic (OH-) group, halogen groups(C-X), Aldehydic (CHO-) groups within them. The antibacterial action is mainly contributed by disruption of membrane potential, degradation of bacterial genomic DNA, damage of bacterial cell membrane and leakage of cellular content. The result provides a significant contribution towards understanding the antibacterial mechanism of n-hexane and dichloromethane extract of C. phlomidis.
Keywords: Clerodendrum phlomidis, Bacillus licheniformis, herbal extract, FACS, SEM, DNA fragmentation
References
Ahsan, N., Paul, N., Islam, N. and Akhand, A.A. (2012). Leaf extract of Syzygiumcumini shows anti-vibrio activity involving DNA damage. Dhaka Univ. J. Pharm. Sci., 11(1):25-28.
Al-Bayati, F.A. (2009). Isolation and identification of antimicrobial compound from Mentha longifolia L. leaves grown wild in Iraq. Annals of Clinical Microbiology and Antimicrobials, 8:20. https://doi.org/10.1186/1476-0711-8-20
Anandhi, D., Srinivasan, P.T., Kumar, G.P. and Jagatheesh, S. (2014). DNA fragmentation induced by the glycosides and flavonoids from C. coriaria. Int. J.Curr. Microbiol. App. Sci., 3(12): 666-673.
Bharvad, N.J. (2005). Control of worms in cotton, brinjal, sesame and chilli. Honey Bee, 16(4):15.
De, A., Boyadjieva, N.I., Pastorcic, M., Reddy, B.V. and Sarkar, D.K. (1994). Cyclic AMP and ethanol interact to control apoptosis and differentiation in hypothalamic beta-endorphin neurons. J. Biol. Chem., 269: 26697-26705.
Dhanabal, S.P., Murgaraja, M.K.M. and Suresh, B. (2008). Antidiabetic activity of Clerodendrumphlomidis leaf extract in alloxan induced diabetic rats. Indian J. Pharm. Sci., 70(6):841-844.
Frank, J.F. (1997). Milk and dairy products. In: Doyle M P, Beuchat L R, Montville T J, editors. Food microbiology: fundamentals and frontiers. Washington, D.C., American Society for Microbiology.
Gu, L.Y., Chen, Z., Zhao, J.,Ruan, X.J., Zhao, S.Y. and Gao, H. (2015). Antioxidant, anticancer and apoptotic effects of the Bupleurum Chinese root extract in HO-8910 ovarian cancer cells. JBUON, 20(5):1341-1349.
Gupta, A., Mahajan, S. and Sharma, R. (2015). Evaluation of antimicrobial activity of Curcuma longa rhizome extract against Staphylococcus aureus. Biotechnology Reports, 6: 51–55.
Haydushka, I.A., Markova, N.,Kirina, V. and Atanassova, M. (2012). Recurrent sepsis due to Bacillus licheniformis. J. Glob. Infect. Dis., 4(1): 82–83.
Huang, D.F., Xu, J.G., Liu, J.X., Zhang, H. and Hu, Q.P. (2014). Chemical constituents, antibacterial activity and mechanism of action of essential oils from Cinnamomum cassia bark against four food related bacteria. Microbiology, 83(4):357-365.
Ignacimuthu, S., Pavunraj, M., Duraipandiyan, V., Raja, N. and Muthu, C. (2009). Antibacterial activity of a novel quinone from the leaves of Pergulariadaemia (Forsk.), a traditional medicinal plant. Asian Journal of Traditional Medicines, 4(1): 36–40.
Johnson, C.T., Lupson, G.R. and Lawrence, K.E. (1994). The bovine placentome in bacterial and mycotic abortions. Vet. Rec., 134: 263–266.
Kaur, C.J. and Renu, S. (2012). Anti-microbial efficacy of medicinally important plant (C. phlomidis) used in folkaric medicines in arid zones. Int. Res. J. Pharmacy, 3(5): 415-17.
Keshabhai, J.R. (1992). Treatment of skin disease: leaf extract of ‘Arani’(Clerodendrumphlomidis). Honey Bee, 3(2): 21.
Khare, C.P. (2007). Indian Medicinal plants. New York: Springer 169.
Lu, L., Huang, Q., Chen, Z., Huang, X., Liang, J., Xia, S., Wang, Y., Yang, X. and Zhang, Y. (2012). Knowledge, attitudes and practices of food-borne diseases and surveillance among physicians in Guangdong, China. Food Control, 28:69–73.
Mori, A., Nishino, C., Enoki, N. and Tawata, S. (1987). Antibacterial activity and mode of action of plant flavonoids against Proteus vulgaris and Staphylococcus aureus. Phytochemistry, 26:2231- 4.
Murugan, M. and Mohan, V.R. (2014). Phytochemical, FT-IR and antibacterial activity of whole plant extract of Aervalanata (L.) Juss. Ex. Schult. Journal of Medicinal Plants Studies, 2(3):51-57.
Pandey, C.N., Raval, B.R., Mali, S. and Salvi, H. (2005). Medicinal plants of Gujarat, Gandhinagar. Ecological education and research (GEER) Foundation.156.
Pepe, O., Blaiotta, G., Moschetti, G. and Greco, T. (2003). Villani F. Rope-producing strains of Bacillus spp. from wheat bread and strategy for their control by lactic acid bacteria. Appl. Environ. Microbiol., 69(4):2321-9.
Pieme, C.A., Kumar, S.G., Dongmo, M.S., Moukette, B.M., Boyoum, F.F., Ngogang, J.Y. and Saxena, A.K. (2014). Antiproliferative activity and induction of apoptosis by Annona muricata (Annonaceae) extract on human cancer cells. BMC Complement Altern. Med., 14:516.
Salonen, S.S., Vuorio, R., Andersson, M.A., Kämpfer, P., Andersson, M.C., Buzalski, T.H. and Scoging, A.C. (1999). Toxigenic strains of Bacillus licheniformis related to food poisoning. Appl. Environ. Microbiol., 65(10):4637–4645.
Saritha, K., Rajesh, A., Manjulatha, K., Setty, O.H. and Yenugu, S. (2015). Mechanism of antibacterial action of the alcoholic extracts of Hemidesmus indicus (L.) R. Br. Ex Schult, Leucas aspera (Wild.), Plumbago zeylanica L., and Tridaxprocumbens (L.) R. Br. ex Schult. Front. Microbiol., 6:577.
Sathish, M., Priyadarsini, R., Sunitha, P.G. and Saraswathy, T. (2013). Antimicrobial activity of the extracts and isolated compounds of Clerodendrumphlomidis. Int. J. Pharm. Pharm. Sci., 5(4): 362-366.
Shi, L., Gunther, S., Hubschmann, T., Wick, L.Y., Harms, H. and Muller, S. (2007). Limits of propidium iodide as a cell viability indicator for environmental bacteria. International Society for Analytical Cytology 71A:592-598.
Teanpaisan, R., Senapong, S. and Puripattanavong, J. (2014). In vitro antimicrobial and antibiofilm activity of Artocarpuslakoocha (Moraceae) extract against some oral pathogens. Tropical Journal of Pharmaceutical Research, 13(7): 1149-1155.
Ulanowska, K., Tkaczyk, A., Konopa, G. and Wegrzyn, G. (2006). Differential antibacterial activity of genistein arising from global inhibition of DNA, RNA and protein synthesis in some bacterial strains. Archi.Microbio., 184:271-278.
Vaidyar, M.S. (1997). Hypothermia or shivering in cattle. Honey Bee, 8(4):9.
Valgas, C., de Souza, S.M., Smânia, E.F.A. and Smânia, Jr. (2007). Screening methods to determine antibacterial activity of natural products. Brazilian Journal of Microbiology, 38:369-380.
Yadav, A.K., Saraswat, S., Sirohi, P., Rani, M., Srivastava, S., Singh, M.P. and Singh, N.K. (2017). Antimicrobial action of methanolic seed extracts of SyzygiumcuminiLinn. on Bacillus subtilis. AMB Expr., 7:196-205. DOI 10.1186/s13568-017-0500-4
Yadav, A.K., Sirohi, P., Saraswat, S., Rani, M., Srivastava, S., Singh, M.P. and Singh, N.K. (2018). Inhibitory mechanism on combination of phytic acid with methanolic seed extract of Syzygiumcuminiand sodium chloride over Bacillus subtilis. Current Microbiology, 75:849-856. https://doi.org/10.1007/s00284-018-1457-5
Yang, J.Y., Park, J.H. and Lee, H.S. (2013). Isolation of 8-hydroxyquinoline from Sebastianiacorniculata and antimicrobial activity against food-borne bacteria. J. Korean Soc. Appl. Biol. Chem., 56:763−766.
Yeh, C.C., Yang, J.I., Lee, J.C., Tseng, C.N., Chan, Y.C., Hseu, Y.C., Tang, J.Y., Chuang, L.Y., Huang, H.W., Chang, F.R. and Chang, H.W. (2012). Anti-proliferative effect of methanolic extract of Gracilariatenuistipitata on oral cancer cells involves apoptosis, DNA damage, and oxidative stress. BMC Complement.Altern. Med., 12:142. doi: 10.1186/1472-6882-12-142.
Yenugu, S., Hamil, K.G., French, F.S. and Hall, S.H. (2006). Antimicrobial actions of human and macaque sperm associated antigen (SPAG)11 isoforms: influence of the N-terminal peptide. Mol. Cell Biochem., 284:25–35. doi: 10.1007/s11010- 005-9009-2.