Ultrasonication-assisted formation and characterization of geraniol and carvacrol-loaded emulsions for enhanced antimicrobial activity against food-borne pathogens

Original Paper


Gum arabic stabilised oil-in-water emulsions were prepared through ultrasonication approach for the incorporation of natural, plant-based antimicrobial compounds, geraniol and carvacrol. The oil phase of formulated emulsions was constituted with geraniol and carvacrol, incorporated at various ratios of 1:0, 2:1, 1:1, 1:2, and 0:1 (v/v). The ultrasonication procedure was followed using a frequency of 20 kHz at 40% amplitude for 5 min. These emulsion systems were characterized for mean particle diameter, polydispersity index, ζ-potential, storage stability and creaming index. In addition, emulsion microstructure was studied using confocal laser scanning microscopy (CLSM) and transmission electron microscopy (TEM). Evaluation of antimicrobial activity of the functional emulsions was carried out against Gram-positive bacteria Bacillus cereus MTCC 430 and Gram-negative bacteria Escherichia coli MTCC 443. The results demonstrated that geraniol: carvacrol (1:1) emulsion formulation displayed good stability with particle size (202.7 ± 4.17 nm), polydispersity (0.282 ± 0.001), ζ-potential (− 19.37 ± 0.06 mV) and no visible separation of cream was observed. Furthermore, the CLSM and TEM observations confirmed the presence of stable emulsion. In addition, the antimicrobial susceptibility tests demonstrated collaborative activity and prolonged antibacterial efficacy for the combined essential oil-based emulsion against both the model bacterial pathogens.


Essential oil emulsion Geraniol Carvacrol Food-borne pathogens Antimicrobials 

Greek letters


Zeta potential



The authors would like to thank the Science and Engineering Research Board, Govt. of India, for providing financial support for conducting research work through a research grant to Dr. Preetam Sarkar (YSS/2015/000546). The authors would also like to acknowledge technical support extended by Sukanta, Susanta Pradhan, and Subhabrata Chakraborty, National Institute of Technology, Rourkela, for assistance with Zetasizer Nano ZS, confocal laser scanning microscopy imaging and TEM imaging, respectively.

Compliance with ethical standards

Conflict of interest

The authors report no conflict of interest.


  1. Abdollahzadeh E, Rezaei M, Hosseini H (2014) Antibacterial activity of plant essential oils and extracts: the role of thyme essential oil, nisin, and their combination to control Listeria monocytogenes inoculated in minced fish meat. Food Control 35:177–183.  https://doi.org/10.1016/j.foodcont.2013.07.004 CrossRefGoogle Scholar
  2. Anandan S, Keerthiga M, Vijaya S, Asiri AM, Bogush V, Krasulyaa O (2017) Physicochemical characterization of black seed oil-milk emulsions through ultrasonication. Ultrason Sonochem 38:766–771.  https://doi.org/10.1016/j.ultsonch.2016.11.005 CrossRefGoogle Scholar
  3. Bennett SD, Walsh KA, Gould LH (2013) Foodborne disease outbreaks caused by Bacillus cereus, Clostridium perfringens, and Staphylococcus aureus—United States, 1998–2008. Clin Infect Dis 57:425–433.  https://doi.org/10.1093/cid/cit244 CrossRefGoogle Scholar
  4. Bi L, Yang L, Narsimhan G, Bhunia AK, Yao Y (2011) Designing carbohydrate nanoparticles for prolonged efficacy of antimicrobial peptide. J Control Release 150:150–156.  https://doi.org/10.1016/j.jconrel.2010.11.024 CrossRefGoogle Scholar
  5. Burt S (2004) Essential oils: their antibacterial properties and potential applications in foods—a review. Int J Food Microbiol 94:223–253.  https://doi.org/10.1016/j.ijfoodmicro.2004.03.022 CrossRefGoogle Scholar
  6. Chang Y, McLandsborough L, McClements DJ (2015) Fabrication, stability and efficacy of dual-component antimicrobial nanoemulsions: essential oil (thyme oil) and cationic surfactant (lauric arginate). Food Chem 172:298–304.  https://doi.org/10.1016/j.foodchem.2014.09.081 CrossRefGoogle Scholar
  7. Chen W, Viljoen AM (2010) Geraniol—a review of a commercially important fragrance material. S Afr J Bot 76:643–651.  https://doi.org/10.1016/j.sajb.2010.05.008 CrossRefGoogle Scholar
  8. Devi KP, Nisha SA, Sakthivel R, Pandian SK (2010) Eugenol (an essential oil of clove) acts as an antibacterial agent against Salmonella typhi by disrupting the cellular membrane. J Ethnopharmacol 130:107–115.  https://doi.org/10.1016/j.jep.2010.04.025 CrossRefGoogle Scholar
  9. Dickinson E (2009) Hydrocolloids as emulsifiers and emulsion stabilizers. Food Hydrocolloid 23:1473–1482.  https://doi.org/10.1016/j.foodhyd.2008.08.005 CrossRefGoogle Scholar
  10. Dickinson E (2010) Food emulsions and foams: stabilization by particles. Curr Opin Colloid Interface Sci 15:40–49.  https://doi.org/10.1016/j.cocis.2009.11.001 CrossRefGoogle Scholar
  11. Ehling-Schulz M, Fricker M, Scherer S (2004) Bacillus cereus, the causative agent of an emetic type of food-borne illness. Mol Nutr Food Res 48:479–487.  https://doi.org/10.1002/mnfr.200400055 CrossRefGoogle Scholar
  12. Epand RM, Epand RF (2009) Lipid domains in bacterial membranes and the action of antimicrobial agents. Biochim Biophys Acta 1788:289–294.  https://doi.org/10.1016/j.bbamem.2008.08.023 CrossRefGoogle Scholar
  13. Food and Drug Administration (2017) Food additives permitted for direct addition to food for human consumption: synthetic flavoring substances and adjuvants. In: Code of federal regulations, Title 21, vol 3, Part 182.20. https://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfcfr/CFRSearch.cfm?fr=182.20
  14. Ghosh V, Mukherjee A, Chandrasekaran N (2013) Ultrasonic emulsification of food-grade nanoemulsion formulation and evaluation of its bactericidal activity. Ultrason Sonochem 20:338–344.  https://doi.org/10.1016/j.ultsonch.2012.08.010 CrossRefGoogle Scholar
  15. Goyal A, Sharma V, Upadhyay N, Singh AK, Arora S, Lal D, Sabikhi L (2015) Development of stable flaxseed oil emulsions as a potential delivery system of omega-3 fatty acids. J Food Sci Technol 52:4256–4265.  https://doi.org/10.1007/s13197-014-1370-2 CrossRefGoogle Scholar
  16. Guarda A, Rubilar JF, Miltz J, Galotto MJ (2011) The antimicrobial activity of microencapsulated thymol and carvacrol. Int J Food Microbiol 146:144–150.  https://doi.org/10.1016/j.ijfoodmicro.2011.02.011 CrossRefGoogle Scholar
  17. Hernández-González M et al (2017) Polysuccinimide functionalized with oregano’s essential oil extracts, an antimicrobial extended release bio-material. Mater Lett 191:73–76.  https://doi.org/10.1016/j.matlet.2017.01.035 CrossRefGoogle Scholar
  18. Hu Q, Gerhard H, Upadhyaya I, Venkitanarayanan K, Luo Y (2016) Antimicrobial eugenol nanoemulsion prepared by gum arabic and lecithin and evaluation of drying technologies. Int J Biol Macromol 87:130–140.  https://doi.org/10.1016/j.ijbiomac.2016.02.051 CrossRefGoogle Scholar
  19. Hussein J, El-Bana M, Refaat E, El-Naggar ME (2017) Synthesis of carvacrol-based nanoemulsion for treating neurodegenerative disorders in experimental diabetes. J Funct Foods 37:441–448.  https://doi.org/10.1016/j.jff.2017.08.011 CrossRefGoogle Scholar
  20. Jain A, Thakur D, Ghoshal G, Katare O, Shivhare U (2015) Microencapsulation by complex coacervation using whey protein isolates and gum acacia: an approach to preserve the functionality and controlled release of β-carotene. Food Bioprocess Technol 8:1635–1644.  https://doi.org/10.1007/s11947-015-1521-0 CrossRefGoogle Scholar
  21. Jayme ML, Dunstan DE, Gee ML (1999) Zeta potentials of gum arabic stabilised oil in water emulsions. Food Hydrocoll 13:459–465.  https://doi.org/10.1016/S0268-005x(99)00029-6 CrossRefGoogle Scholar
  22. Jiménez M, Domínguez JA, Pascual-Pineda LA, Azuara E, Beristain C (2017) Elaboration and characterization of O/W cinnamon (Cinnamomum zeylanicum) and black pepper (Piper nigrum) emulsions. Food Hydrocoll.  https://doi.org/10.1016/j.foodhyd.2017.11.037 Google Scholar
  23. Ju J, Xu X, Xie Y, Guo Y, Cheng Y, Qian H, Yao W (2018) Inhibitory effects of cinnamon and clove essential oils on mold growth on baked foods. Food Chem 240:850–855.  https://doi.org/10.1016/j.foodchem.2017.07.120 CrossRefGoogle Scholar
  24. Khem S, Bansal V, Small DM, May BK (2016) Comparative influence of pH and heat on whey protein isolate in protecting Lactobacillus plantarum A17 during spray drying. Food Hydrocoll 54:162–169.  https://doi.org/10.1016/j.foodhyd.2015.09.029 CrossRefGoogle Scholar
  25. Kowalska M, Zbikowska A, Wozniak M, Kucharczyk K (2017) Long-term stability of emulsion based on rose oil. J Dispers Sci Technol 38:1563–1569.  https://doi.org/10.1080/01932691.2016.1262777 CrossRefGoogle Scholar
  26. Kumar DL, Sarkar P (2017) Encapsulation of bioactive compounds using nanoemulsions. Environ Chem Lett.  https://doi.org/10.1007/s10311-017-0663-x Google Scholar
  27. Kwon SJ, Chang Y, Han J (2017) Oregano essential oil-based natural antimicrobial packaging film to inactivate Salmonella enterica and yeasts/molds in the atmosphere surrounding cherry tomatoes. Food Microbiol 65:114–121.  https://doi.org/10.1016/j.fm.2017.02.004 CrossRefGoogle Scholar
  28. Liang R, Xu S, Shoemaker CF, Li Y, Zhong F, Huang Q (2012) Physical and antimicrobial properties of peppermint oil nanoemulsions. J Agric Food Chem 60:7548–7555.  https://doi.org/10.1021/jf301129k CrossRefGoogle Scholar
  29. Lu WC, Huang DW, Wang CR, Yeh CH, Tsai JC, Huang YT, Li PH (2018) Preparation, characterization, and antimicrobial activity of nanoemulsions incorporating citral essential oil. J Food Drug Anal 26:82–89.  https://doi.org/10.1016/j.jfda.2016.12.018 CrossRefGoogle Scholar
  30. Ma Q, Davidson PM, Zhong Q (2016) Nanoemulsions of thymol and eugenol co-emulsified by lauric arginate and lecithin. Food Chem 206:167–173.  https://doi.org/10.1016/j.foodchem.2016.03.065 CrossRefGoogle Scholar
  31. Martinez-Hernandez GB, Amodio ML, Colelli G (2017) Carvacrol-loaded chitosan nanoparticles maintain quality of fresh-cut carrots. Innov Food Sci Emerg 41:56–63.  https://doi.org/10.1016/j.ifset.2017.02.005 CrossRefGoogle Scholar
  32. McClements DJ, Decker EA, Weiss J (2007) Emulsion-based delivery systems for lipophilic bioactive components. J Food Sci 72:R109–R124.  https://doi.org/10.1111/j.1750-3841.2007.00507.x CrossRefGoogle Scholar
  33. Mirhosseini H, Amid BT (2012) A review study on chemical composition and molecular structure of newly plant gum exudates and seed gums. Food Res Int 46:387–398.  https://doi.org/10.1016/j.foodres.2011.11.017 CrossRefGoogle Scholar
  34. Mirhosseini H, Tan CP, Hamid NSA, Yusof S (2008) Optimization of the contents of Arabic gum, xanthan gum and orange oil affecting turbidity, average particle size, polydispersity index and density in orange beverage emulsion. Food Hydrocoll 22:1212–1223.  https://doi.org/10.1016/j.foodhyd.2007.06.011 CrossRefGoogle Scholar
  35. Moon H, Rhee MS (2016) Synergism between carvacrol or thymol increases the antimicrobial efficacy of soy sauce with no sensory impact. Int J Food Microbiol 217:35–41.  https://doi.org/10.1016/j.ijfoodmicro.2015.10.009 CrossRefGoogle Scholar
  36. Niu F, Pan W, Su Y, Yang Y (2016) Physical and antimicrobial properties of thyme oil emulsions stabilized by ovalbumin and gum arabic. Food Chem 212:138–145.  https://doi.org/10.1016/j.foodchem.2016.05.172 CrossRefGoogle Scholar
  37. Noshad M, Mohebbi M, Shahidi F, Koocheki A (2015) Freeze–thaw stability of emulsions with soy protein isolate through interfacial engineering. Int J Refrig 58:253–260.  https://doi.org/10.1016/j.ijrefrig.2015.05.007 CrossRefGoogle Scholar
  38. Oussalah M, Caillet S, Saucier L, Lacroix M (2007) Inhibitory effects of selected plant essential oils on the growth of four pathogenic bacteria: E. coli O157: H7, Salmonella Typhimurium, Staphylococcus aureus and Listeria monocytogenes. Food Control 18:414–420.  https://doi.org/10.1016/j.foodcont.2005.11.009 CrossRefGoogle Scholar
  39. Pol IE, Smid EJ (1999) Combined action of nisin and carvacrol on Bacillus cereus and Listeria monocytogenes. Lett Appl Microbiol 29:166–170.  https://doi.org/10.1046/j.1365-2672.1999.00606.x CrossRefGoogle Scholar
  40. Salvia-Trujillo L, Rojas-Grau MA, Soliva-Fortuny R, Martin-Belloso O (2014) Impact of microfluidization or ultrasound processing on the antimicrobial activity against Escherichia coli of lemongrass oil-loaded nanoemulsions. Food Control 37:292–297.  https://doi.org/10.1016/j.foodcont.2013.09.015 CrossRefGoogle Scholar
  41. Salvia-Trujillo L, Rojas-Grau A, Soliva-Fortuny R, Martin-Belloso O (2015) Physicochemical characterization and antimicrobial activity of food-grade emulsions and nanoemulsions incorporating essential oils. Food Hydrocoll 43:547–556.  https://doi.org/10.1016/j.foodhyd.2014.07.012 CrossRefGoogle Scholar
  42. São José JFBd, Andrade NJd, Ramos AM, Vanetti MCD, Stringheta PC, Chaves JBP (2014) Decontamination by ultrasound application in fresh fruits and vegetables. Food Control 45:36–50.  https://doi.org/10.1016/j.foodcont.2014.04.015 CrossRefGoogle Scholar
  43. Sarkar P, Bhunia AK, Yao Y (2017) Impact of starch-based emulsions on the antibacterial efficacies of nisin and thymol in cantaloupe juice. Food Chem 217:155–162.  https://doi.org/10.1016/j.foodchem.2016.08.071 CrossRefGoogle Scholar
  44. Shah B, Ikeda S, Michael Davidson P, Zhong Q (2012) Nanodispersing thymol in whey protein isolate-maltodextrin conjugate capsules produced using the emulsion–evaporation technique. J Food Eng 113:79–86.  https://doi.org/10.1016/j.jfoodeng.2012.05.019 CrossRefGoogle Scholar
  45. Sharma M, Mann B, Sharma R, Bajaj R, Athira S, Sarkar P, Pothuraju R (2017) Sodium caseinate stabilized clove oil nanoemulsion: physicochemical properties. J Food Eng 212:38–46.  https://doi.org/10.1016/j.jfoodeng.2017.05.006 CrossRefGoogle Scholar
  46. Silva-Angulo AB, Zanini SF, Rosenthal A, Rodrigo D, Klein G, Martinez A (2015) Combined effect of carvacrol and citral on the growth of Listeria monocytogenes and Listeria innocua and on the occurrence of damaged cells. Food Control 53:156–162.  https://doi.org/10.1016/j.foodcont.2015.01.028 CrossRefGoogle Scholar
  47. Sivapratha S, Sarkar P (2018) Multiple layers and conjugate materials for food emulsion stabilization. Crit Rev Food Sci Nutr 58:877–892.  https://doi.org/10.1080/10408398.2016.1227765 CrossRefGoogle Scholar
  48. Sosa N, Schebor C, Pérez OE (2014) Encapsulation of citral in formulations containing sucrose or trehalose: emulsions properties and stability. Food Bioprod Process 92:266–274.  https://doi.org/10.1016/j.fbp.2013.08.001 CrossRefGoogle Scholar
  49. Sugumar S, Ghosh V, Nirmala MJ, Mukherjee A, Chandrasekaran N (2014) Ultrasonic emulsification of eucalyptus oil nanoemulsion: antibacterial activity against Staphylococcus aureus and wound healing activity in Wistar rats. Ultrason Sonochem 21:1044–1049.  https://doi.org/10.1016/j.ultsonch.2013.10.021 CrossRefGoogle Scholar
  50. Tontul I, Topuz A (2014) Influence of emulsion composition and ultrasonication time on flaxseed oil powder properties. Powder Technol 264:54–60.  https://doi.org/10.1016/j.powtec.2014.05.002 CrossRefGoogle Scholar
  51. Topuz OK, Ozvural EB, Zhao Q, Huang Q, Chikindas M, Golukcu M (2016) Physical and antimicrobial properties of anise oil loaded nanoemulsions on the survival of foodborne pathogens. Food Chem 203:117–123.  https://doi.org/10.1016/j.foodchem.2016.02.051 CrossRefGoogle Scholar
  52. Ultee A, Gorris LG, Smid EJ (1998) Bactericidal activity of carvacrol towards the food-borne pathogen Bacillus cereus. J Appl Microbiol 85:211–218.  https://doi.org/10.1046/j.1365-2672.1998.00467.x CrossRefGoogle Scholar
  53. Ultee A, Kets EP, Smid EJ (1999) Mechanisms of action of carvacrol on the food-borne pathogen Bacillus cereus. Appl Environ Microbiol 65:4606–4610Google Scholar
  54. Ultee A, Bennik M, Moezelaar R (2002) The phenolic hydroxyl group of carvacrol is essential for action against the food-borne pathogen Bacillus cereus. Appl Environ Microbiol 68:1561–1568.  https://doi.org/10.1128/AEM.68.4.1561-1568.2002 CrossRefGoogle Scholar
  55. Visalli MA, Jacobs MR, Appelbaum PC (1996) MIC and time-kill study of activities of DU-6859a, ciprofloxacin, levofloxacin, sparfloxacin, cefotaxime, imipenem, and vancomycin against nine penicillin-susceptible and -resistant pneumococci. Antimicrob Agents Chemother 40:362–366Google Scholar
  56. WHO (2015) WHO estimates of the global burden of foodborne diseases: foodborne disease burden epidemiology reference grou. World Health Organization, Geneva, pp 2007–2015Google Scholar
  57. Wu JE, Lin J, Zhong QX (2014) Physical and antimicrobial characteristics of thyme oil emulsified with soluble soybean polysaccharide. Food Hydrocoll 39:144–150.  https://doi.org/10.1016/j.foodhyd.2013.12.029 CrossRefGoogle Scholar
  58. Xue J, Michael Davidson P, Zhong Q (2015) Antimicrobial activity of thyme oil co-nanoemulsified with sodium caseinate and lecithin. Int J Food Microbiol 210:1–8.  https://doi.org/10.1016/j.ijfoodmicro.2015.06.003 CrossRefGoogle Scholar
  59. Yao X et al (2013) Physical and chemical stability of gum arabic-stabilized conjugated linoleic acid oil-in-water emulsions. J Agric Food Chem 61:4639–4645.  https://doi.org/10.1021/jf400439d CrossRefGoogle Scholar
  60. Ye HQ, Shen SX, Xu JY, Lin SY, Yuan Y, Jones GS (2013) Synergistic interactions of cinnamaldehyde in combination with carvacrol against food-borne bacteria. Food Control 34:619–623.  https://doi.org/10.1016/j.foodcont.2013.05.032 CrossRefGoogle Scholar
  61. Ye F, Miao M, Jiang B, Hamaker BR, Jin Z, Zhang T (2017) Characterizations of oil-in-water emulsion stabilized by different hydrophobic maize starches. Carbohydr Polym 166:195–201.  https://doi.org/10.1016/j.carbpol.2017.02.079 CrossRefGoogle Scholar
  62. Yegin Y, Perez-Lewis KL, Zhang M, Akbulut M, Taylor TM (2016) Development and characterization of geraniol-loaded polymeric nanoparticles with antimicrobial activity against foodborne bacterial pathogens. J Food Eng 170:64–71.  https://doi.org/10.1016/j.jfoodeng.2015.09.017 CrossRefGoogle Scholar
  63. Yildirim ST, Oztop MH, Soyer Y (2017) Cinnamon oil nanoemulsions by spontaneous emulsification: formulation, characterization and antimicrobial activity. Lwt-Food Sci Technol 84:122–128.  https://doi.org/10.1016/j.lwt.2017.05.041 CrossRefGoogle Scholar
  64. Zhang Z, Vriesekoop F, Yuan Q, Liang H (2014) Effects of nisin on the antimicrobial activity of d-limonene and its nanoemulsion. Food Chem 150:307–312.  https://doi.org/10.1016/j.foodchem.2013.10.160 CrossRefGoogle Scholar
  65. Zhang W, Chen L, Fang X (2015) Optimizing the preparation conditions for shea butter nanoemulsions via response surface methodology. J Dispers Sci Technol 36:983–990.  https://doi.org/10.1080/01932691.2014.942317 CrossRefGoogle Scholar

Copyright information

© Institute of Chemistry, Slovak Academy of Sciences 2018

Authors and Affiliations

  1. 1.Department of Food Process EngineeringNational Institute of Technology RourkelaRourkelaIndia

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