, Volume 70, Issue 8, pp 1003–1010 | Cite as

Factors influencing synergistic antimicrobial activity of thymol and nisin against Shigella spp. in sugarcane juice

  • Sukrita Punyauppa-Path
  • Parichat Phumkhachorn
  • Pongsak RattanachaikunsoponEmail author


Two food-grade antimicrobial substances, thymol and nisin were tested for their antimicrobial activity against 4 species of Shigella including S. boydii, S. dysenteriae, S. flexneri and S. sonnei inoculated in sugarcane juice at 4°C. When used separately, only thymol, but not nisin, exhibited a dose-dependent and species-dependent inhibitory effect on the bacteria. When thymol and nisin were used together, the concentrations of thymol required for complete inhibition were decreased in all cases of the bacteria as the concentrations of nisin were raised, indicating the synergistic antimicrobial activity between both compounds. The effects of bacterial cell number and temperature on the antimicrobial activity of thymol and nisin against S. sonnei in sugarcane juice were investigated. The doses of both compounds required for complete inhibition were directly proportional to the number of bacterial cells in sugarcane juice. Furthermore, they inhibited the bacterium at 30 °C more efficiently than at 4°C. The sensory evaluation showed that thymol-and nisin-treated sugarcane juice was acceptable to consumers. This study suggests that thymol and nisin can be used as food preservatives with the consideration of their dependence on bacteria species, number of contaminated bacterial cells and temperature.

Key words

nisin Shigella sugarcane juice thymol 



colony-forming unit


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  1. Bassole I.H., Lamien-Meda A., Bayala B., Tirogo S., Franz C., Novak J., Nebie R.C. & Dicko M.H. 2010. Composition and antimicrobial activities of Lippio, multiflora Moldenke, Mentha X piperita L. and Ocimum basilicum, L. essential oils and their major monoterpene alcohols alone and in combination. Molecules 15: 7825–7839.CrossRefGoogle Scholar
  2. Bhattacharya S.K. & Sur D. 2003. An evaluation of current shigellosis treatment. Expert Opin. Pharmacother. 4: 1315–1320.CrossRefGoogle Scholar
  3. Burt S. 2004. Essential oils: their antibacterial properties and potential applications in foods - a review. Int. J. Food Microbiol. 94: 223–253.CrossRefGoogle Scholar
  4. Cosentino S., Tuberoso C.I.G., Pisano B., Satta M., Mascia V., Arzedi E. & Palmas F. 1999. In vitro antimicrobial activity and chemical composition of Sardinian Thymus essential oils. Lett. Appl. Microbiol. 29: 130–135.CrossRefGoogle Scholar
  5. Cotter P.D., Hill C. & Ross R.P. 2005. Bacteriocins: developing innate immunity for food. Nat. Rev. Microbiol. 3: 777–788.CrossRefGoogle Scholar
  6. Cowan M.M. 1999. Plant products as antimicrobial agents. Clin. Microbiol. Rev. 12: 564–582.CrossRefGoogle Scholar
  7. Davidson P.M. & Harrison M.A. 2002. Resistance and adaptation to food antimicrobials and other process controls. Food Technol. 56: 69–78.Google Scholar
  8. Delves-Broughton J. 1990. Nisin and its uses as food preservative. Int. J. Dairy Technol. 43: 73–76.CrossRefGoogle Scholar
  9. Ettayebi K., El Yamani J. & Rossi-Hassani B. 2000. Synergistic effects of nisin and thymol on antimicrobial activities in Listeria monocytogenes and Bacillus subtilis. FEMS Microbiol. Lett. 183: 191–195.Google Scholar
  10. Gould G.W. 1996. Industry perspectives on the use of natural antimicrobials and inhibitors for food applications. J. Food Prot. 59 (Supplement): 82–86.CrossRefGoogle Scholar
  11. Gould I.M. 2000. A review of the role of antibiotic policies in the control of antibiotic resistance. J. Antimicrob. Chemother. 43: 459–465.CrossRefGoogle Scholar
  12. Gross E. & Morell J.L. 1971. The structure of nisin. J. Am. Chem. Soc. 93: 4634–4635.CrossRefGoogle Scholar
  13. Hyldgaard M., Mygind T. & Meyer R.L. 2012. Essential oils in food preservation: mode of action, synergies, and interactions with food matrix components. Front. Microbiol. 3: 12.CrossRefGoogle Scholar
  14. Isenbarger D.W., Hoge C.W., Srijan A., Pitarangsi C., Vithayasai N., Bodhidatta L., Hickey K.W. & Cam P.D. 2002. Comparative antibiotic resistance of diarrheal pathogens from Vietnam and Thailand, 1996-1999. Emerg. Infect. Dis. 8: 175–180.CrossRefGoogle Scholar
  15. Kivanc M. & Akgul A. 2006. Antibacterial activities of essential oils from Turkish spices and citrus. Flavour Frag. J. 1: 175–179.CrossRefGoogle Scholar
  16. Klaenhammer T.R. 1988. Bacteriocins of lactic acid bacteria. Biochimie 70: 337–349.CrossRefGoogle Scholar
  17. Kordel M. & Sahl H.G. 1986. Susceptibility of bacterial, eukary-otic and artificial membranes to the disruptive action of the cationic peptides Pep 5 and nisin. FEMS Microbiol. Lett. 34: 139–144.CrossRefGoogle Scholar
  18. Kotloff K.L., Winickoff J.P., Ivanoff B., Clemens J.D., Swerd-low D.L., Sansonetti P.J., Adak G.K. & Levine M.M. 1999. Global burden of Shigella infections: implications for vaccine development and implementation of control strategies. Bull. World Health Organ. 77: 651–666.PubMedPubMedCentralGoogle Scholar
  19. Kuo C.Y., Su L.H., Perera J., Carlos C., Tan B.H. & Kumaras-inghe G. 2008. Antimicrobial susceptibility of Shigella isolates in eight Asian countries, 2001-2004. J. Microbiol. Immunol. Infect. 41: 107–111.PubMedGoogle Scholar
  20. Lambert R.J.W., Skandamis P.N., Coote P. & Nychas G.J.E. 2001. A study of the minimum inhibitory concentration and mode of action of oregano essential oil, thymol and carvacrol. J. Appl. Microbiol. 91: 453–462.CrossRefGoogle Scholar
  21. Liu W. & Hansen J.N. 1990. Some chemical and physical properties of nisin, a small-protein antibiotic produced by Lactococcus lactis. Appl. Environ. Microbiol. 56: 2551–2558.PubMedPubMedCentralGoogle Scholar
  22. Mahmoud B.S., Yamazaki K., Miyashita K., Kawai Y., Shin I.S. & Suzuki T. 2006. Preservative effect of combined treatment with electrolyzed NaCl solutions and essential oil compounds on carp fillets during convectional air-drying. Int. J. Food Microbiol. 106: 331–337.CrossRefGoogle Scholar
  23. Manou I., Bouillard L., Devleeschouwer M.J. & Barel A.O. 1998. Evaluation of the preservative properties of Thymus vulgaris essential oil in topically applied formulations under a challenge test. J. Appl. Microbiol. 84: 368–376.CrossRefGoogle Scholar
  24. Morris S.L., Walsh R.C. & Hansen J.N. 1984. Identification and characterization of some bacterial membrane sulfhydryl groups which are targets of bacteriostatic and antibiotic action. J. Biol. Chem. 259: 13590–13594.PubMedGoogle Scholar
  25. Niyogi S.K. 2005. Shigellosis. J. Microbiol. 43: 133–143.PubMedGoogle Scholar
  26. Niyogi S.K. 2007. Increasing antimicrobial resistance - an emerging problem in the treatment of shigellosis. Clin. Microbiol. Infect. 13: 1141–1143.CrossRefGoogle Scholar
  27. Periago P.M. & Moezelaar R. 2001. Combined effect of nisin and carvacrol at different pH and temperature levels on the viability of different strains of Bacillus cereus. Int. J. Food Microbiol. 68: 141–148.CrossRefGoogle Scholar
  28. Rattanachaikunsopon P. & Phumkhachorn P. 2010a. Synergistic antimicrobial effect of nisin and p-cymene on Salmonella enterics serovar Typhi in vitro and on ready-to-eat food. Biosci. Biotechnol. Biochem. 74: 520–524.CrossRefGoogle Scholar
  29. Rattanachaikunsopon P. & Phumkhachorn P. 2010b. Assessment of factors influencing antimicrobial activity of carvacrol and cymene against Vibrio cholerae in food. J. Biosci. Bioeng. 110: 614–619.CrossRefGoogle Scholar
  30. Saei-Dehkordi S.S., Fallah A.A., Saei-Dehkordi S.S. & Kousha S. 2012. Chemical composition and antioxidative activity of Echinophora platyloba DC. essential oil, and its interaction with natural antimicrobials against food-borne pathogens and spoilage organisms. J. Food Sci. 77: M631–M637.CrossRefGoogle Scholar
  31. Sjolund Karlsson M., Bowen A., Reporter R., Folster J.P., Grass J.E., Howie R.L., Taylor J. & Whichard J.M. 2013. Outbreak of infections caused by Shigella sonnei with reduced susceptibility to azithromycin in the United States. Antimi-crob. Agents Chemother. 57: 1559–1560.CrossRefGoogle Scholar
  32. Stevens K.A., Klapes N.A., Sheldon B.W. & Klaenhammer T.R. 1992. Antimicrobial action of nisin against Salmonella ty-phimurium lipopolysaccharide mutants. Appl. Environ. Microbiol. 58: 1786–1788.PubMedPubMedCentralGoogle Scholar
  33. Soulsby E.J. 2005. Resistance to antimicrobials in humans and animals. Br. Med. J. 331: 1219–1220.CrossRefGoogle Scholar
  34. Swartz M.N. 1997. Use of antimicrobial agents and drug resistance. N. Engl. J. Med. 337: 491–492.CrossRefGoogle Scholar
  35. Weissinger W.R., McWatters K.H. & Beuchat L.R. 2001. Evaluation of volatile chemical treatments for lethality to Salmonella on alfalfa seeds and sprouts. J. Food Prot. 64: 442–450.CrossRefGoogle Scholar
  36. Xu J., Zhou F., Ji B.P., Pei R.S. & Xu, N. 2008. The antibacterial mechanism of carvacrol and thymol against Escherichia coli.] Lett. Appl. Microbiol. 47: 174–179.CrossRefGoogle Scholar

Copyright information

© Slovak Academy of Sciences 2015

Authors and Affiliations

  • Sukrita Punyauppa-Path
    • 1
  • Parichat Phumkhachorn
    • 1
  • Pongsak Rattanachaikunsopon
    • 1
    Email author
  1. 1.Department of Biological Science, Faculty of ScienceUbon Ratchathani UniversityUbon RatchathaniThailand

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