Document Type : Full Research Paper


Department of Food Sciences, Islamic Azad University, Damghan Branch, Damghan, University, Damghan Branch, Damghan, Iran.


Introduction: Plant extracts have antimicrobial properties and can be substituted as chemical flavorings in food. Preservatives are compounds that are used to inhibit the growth or to eliminate harmful microorganisms to increase the shelf life of foods. Nowadays, the use of natural preservatives derived from medicinal plants is increasing due to their greater compatibility and the presence of volatile compounds (Celiktas et al., 2007). The antimicrobial properties, antioxidant and anticancer effects of these natural additives have been proven (Tajkarimi et al., 2010). In this study, the antimicrobial activity enhancement effect of the combination of hydroalcoholic extracts of basil, cinnamon and salvia on Escherichia coli, Staphylococcus aureus and Saccharomyces cerevisiae yeast in Iranian yoghurt drink (doogh) was investigated for 4 weeks at ambient temperature.
Materials and Methods: The milk was boiled in hygienic conditions at 95 ° C for 5 min. After cooling to 45 ° C, they were inoculated. The aerial parts of the studied plants were milled and powdered. Cinnamon, basil and salvia extracts were then soaked. The food strains of Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 700728 and Saccharomyces cerevisiae PTCC 5193 were prepared as lyophilized ampoules. The microdilution broth method was used to determine the minimum inhibitory concentration. The minimum bactericidal concentration (MBC) was determined in accordance with the minimum inhibitory concentration (MIC) results. To determine the antimicrobial effect of the extracts on the yoghurt drink, 9 ml of the prepared yoghurt drink was poured into each test tube. Then 1 cc of each extract and 1 cc of microbial suspension (1.5 × 108 CFU / ml) were added. One tube was considered as positive control (bacteria + yoghurt drink), one negative control tube (extract + yoghurt drink) and one control tube (yoghurt drink only) and the tubes were placed at ambient temperature. Then, on day 7, 14, 21 and 30 during storage, 20 µl of each tube, were cultured on Muller Hinton agar medium and the same amount of yeast was cultured on blood agar medium. After incubation for 24 h at 37 ° C, the colonies were counted. The interaction of antimicrobial activity of the combination of basil, cinnamon and sage extracts was evaluated on the basis of differential inhibitory concentration index (FIC).
Results and discussion: The results showed that the minimum inhibitory concentration of the combined basil, cinnamon and sage extracts against Staphylococcus aureus and Saccharomyces cerevisiae and Escherichia coli were 0.4, 0.4 and 3.12 mg / ml, respectively. Also, Staphylococcus aureus was more susceptible to the extracts, which were used alone, and was inhibited by a lower concentration of used extract (Ahmadi et al., 2018). However, due to the maximum minimum inhibitory concentration (MIC), Escherichia coli was less sensitive to the extracts which was used alone. The combined usage of all three extracts significantly reduced the maximum minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC), indicating that their antimicrobial activity was more effective against the studied microorganisms than each of the extracts which was used alone. The results of this study showed that synergistic effect against Staphylococcus aureus and Saccharomyces cerevisiae was more effective with the combination of the aforementioned extracts (p <0.05). The results of the Escherichia coli population counting during storage indicated that most of the extracts had a good effect on preventing the growth of Escherichia coli in the doogh environment compared to the control sample (p <0.05). Addition of the extract significantly prevented pH reduction over time (p <0.05), so that the control sample had the highest and the sample containing all three extracts showed the lowest pH changes over time (p <0.05). Therefore, it can be concluded that acid production by Lactobacillus bacteria in samples containing extract during storage period was higher than control. The results of this study were in agreement with the results of Mortazavian et al., (2007) and Voosogh et al., (2009). The acidity of the doogh samples increased significantly over time (P<0.05), while the addition of the extract significantly prevented the acidity increase over time (P<0.05). Sensory properties are the main factors of acceptance and satisfaction of many products. In this study, the mean viewpoint of doogh samples containing the combination of the three above mentioned extracts in terms of taste, odor, color and general acceptance are acceptable. Additionally, it was found that about 90% of participants were willing to use it. Although studies of the overlapping or synergistic effects of medicinal plant extracts in real food environments are not extensive, their results have shown that, the addition of different medicinal plants compounds can increase antibacterial effects. Therefore, the simultaneous use of these extracts can be used in effective control of food pathogens as a complementary method. The results of this study were in agreement with the results of Karimi (2007).


Ahmadi, S.M., Moslehishad, M., Rahimi, A. 2018. Evaluation of antimicrobial activity of oleoresin essential oil of Pistacia atlantica on Staphlicoccus aureus, Escherichia coli, Kluyveromyces matxianus, Penicillum natatum and its impact on Iranian doogh shelf life. JFST. 85(15):113-124 [In Persian].
Azizkhani, M., Misaghi, A., Akhondzadeh Basti, A., Gandomi Nasrabadi, H., Hosseini, H. 2012. Effect of Zataria multiflora Boiss. Essential Oil on Growth and Enterotoxin E Production of Staphylocuccus aureus ATCC 29213. J Med plants. 11(44): 185-192 [In Persian].
Abolhosseini, Sh., Golestan, L., Kaboosi, H. 2019. The effect of fresh and ripe Pistacia atlantica on Shelf life and sensory properties of Iranian doogh. JFST. 85(15):417-424 [In Persian].
Baron, E.J., and Finegold, S.M. 1990. Bailey and Scott's Diagnostic Microbiology. 8th Ed., CV Mosby Co., St. Louis, USA. pp: 286-402, 435-438.
Behradmanesh, S., Derees, F., Rafieian-kopaei, M. 2013. Effect of Salvia officinalis on diabetic patients. J Renal Inj Prev. 2(2): 51-54.
Celiktas, O.Y., Kocabas, E.H., Bedir, E., Sukan, F.V., Ozek, T., and Baser, K.H.C. 2007. Antimicrobial activities of methanol extracts and essential oils of Rosmarinus officinalis, depending on location and seasonal variations. Food Chem. 14:323-8.
Dinakar, P., and Mistry, V. 1994. Growth and viability of Bifidobact erium bifidum in cheddar cheese. J. Dairy Sci. 77: 2854-2864.
Dini, A., Razavi, S.H., and Ebrahimzadeh Mousavi, S.A. 2013. Effect of incubation and storage temperatures and final pH on the viability of probiotic bacteria and sensory characteristics in probiotic Doogh. J. Food Sci. 23(3):367-380 [In Persian].
Farajvand, N., Alimohammadi, M. 1393. Prevalence of Staphylococcus aureus in Four Famous Brand of Doogh Produced in Iran. Iran. J. Health & Environ. 7 (1):85-94.
Goni, P., Lopez, P., Sanchez, C., Go.mez-Lus, R., Becerril, R., and Nerin, C. 2009. Antimicrobial activity in the vapour phase of a combination of cinnamon and clove essential oils. Food Chem. 116:982-989.
Heinrich, M., Barne, J., Gibbons, S., Williamson, M. 2012. Fundamentals of pharmacognosy and phytotherapy. Elsevier Ltd, p. 232.
Hassanzadeh, A. 2012. Cinnamon and its properties. Monthly Training and New Technology Research of Food. Haghjoo R., and Tadjalli M. 2016. Effect of Persian Sage (Salvia rhytidia) Extract on Histomorphometric Changes of Cerebral Cortex and Hippocampus Following Ischemia-Reperfusion Injuries in Rat. Zahedan J Res Med Sci. 18(2):59-93.
ISIRI. 2007. Doogh – Specifications and test method. Institute of Standards and Industrial Research of Iran, 2453, 1th ed. Tehran.
ISIRI. 2007. Institute of Standards and Industrial Research of Iran. Milk and milk products–Enumeration of colony forming units of yeasts and/or moulds-colony-Count technique at 25°C, No. 10154. 1st. Ed Tehran.
ISIRI. 1999. Basic principles of sensory evaluation of milk and its products. Institute of Standards and Industrial Research of Iran, No. 4691.
Kermanshah, H., HashemiKamangar, S., Arami, S., Mirsalehian, A., Kamalinejad, M., Karimi, M., JabalAmoli, F. 2009. In vitro evaluation of antibacterial activity of hydroalcoholic extract of Salvia officinalis and Pimpinellaanisum against cariogenic bacteria. JIDA. 22:149-54.
Karimi, A. 1386. Study the antimicrobial effect of essential oils of mint, oregano, rose water and effect of the temperature on the growth of Debaryomyces hansenii in Doogh. Ph.D. thesis in food hygiene. IAU. Oloum tahghighat branch.
Lino, C.M., and Pen, A. 2010. Occurrence of caffeine, saccharin, benzoic acid and sorbic acid in soft drinks and nectars in Portugal and subsequent exposure assessment. Food Chem. 121(2):503-508.
Mortazavian, A. M., Ehsani, M. R., Sohrabvandi, S., and Reihemer, J. A. 2007. MRS-Bile agar: Its suitability for the enumeration of mixed probiotic in cultured dairy products. Milchwissenschaft. 62:270–272.
Marino, M., Bersani, C., and Comi, G. 2001. Impedance measurements to study the antimicrobial activity of essential oils from Lamiaceae and Compositae. Int. J. Food Microbial. 67(3):187-195.
Mehraban, A., Edalatian Dovom, M. R., Haddad Khodaparast, M.H., Mehraban Sang Atash, M. 2017. Study of antibacterial effect of aqueous, ethanolic and droalcoholic extracts of aerial oranges of Salvia chorassanica against some spoilage and poisoning bacteria. JFST. 66(14):215-225 [In Persian].
NCCLS. 2000. National Committee for Clinical Laboratory Standards, Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. Approved Standard, M7-A5.
Nourizadeh, E., Ghasemi, K., Latifi, S. 2003. Anti-bacterial effects of Licorice on Helicobacter Pylori. The 3th National Congress of Biotechnology. 9-11. Mashad, Iran.
Oussalah, M., Caillet, S., and Lacroix, M. 2006. Mechanism of action of Spanish oregano, Chinese cinnamon, and savory oils against cell membrane and walls of Escherichia coli O157:H7 and Listeria monocytogenes. J. Food Prot. 69(5):1046-1055.
Oroojalian, F., Kasra kermanshahi, R., Azizi, M., Basaami, M.R. 2010. Synergistic antibacterial activity of the essential oils from three medicinal plants against some important food-borne pathogens by microdilution method. IJMAP. 26 (2):133-146 [In Persian].
Shirazi, M.H., Ranjbar, R., Eshraghi, S., Amin, G., Seyed Nouri, M., and Bazzaz, N. 2008. Inhibitory Effects of Sage Extract on the Growth of Enteric Bacteria. PJBS. 11:487-489.
Sandri, I.G., Zacaria, J., Fracaro, F., Delamare, A.P.L., Echeverrigaray, S. 2007. Antimicrobial activity of the essential oils of Brazilian species ofthe genus Cunila against foodborne pathogens and spoiling bacteria. Food Chem. 103:823–828.
Sendra, E., Fayos, F., Lario, Y., Ferna’ndez-Lo’pez, J., Sayas-Barbera’, E. and Pe’rez Alvarez, J.A. 2008. Incorporation of citrus fibers in fermented milk containing probiotic bacteria. Food microbiol. 25:13-21.
Sajjadi, SE. Analysis of the essential oils of two cultivated basil (Ocimum basilicum L) from Iran. 2006. Daru J. Pharm. Sci. 14(3):128 - 130.
Swamy, M.K., Akhtar, M.S. and Sinniah, U.R. 2016. Antimicrobial properties of plant essential oils against human pathogens and their mode of action: an updated review. Evid-Based. Compl. Alt. p: 1-20.
Tajkarimi, M.M., Ibrahim, S.A. and Cliver, D.O. 2010. Antimicrobial herb and spice compounds.
Tamime, A. and Robinson, R. 1999. Yogurt Science and Technology. Woodhead Publ. Cambridge, England, and CRC Press, Boca Raton, FL.
Tabatabaie yazdi, F., Alizadeh Behbahani, B., and Mortazavi, S. A. 2016. Investigation Effects of Lamiaceae plants (Thymus vulgaris L., Mentha spp. and Ziziphora tenuir L.) Inhibitory Staphylococcus aureus and Geotrichum candidium in Razavi Khorasan Province Industrial Doogh Samples with Response Surface Method (RSM). IJSTS. 51(13):15-28 [In Persian].
Vlase, L., Benedec, D., Hanganu, D., Damian, G., Caillag, l., Sevastre, B. 2014. Evaluation of Antioxidant and Antimicrobial Activities and phenolic profile for Hyssopus officinalis, Ocimum basilicum and Teucrium chamaedry S. Molecules. 19:5940-5507.
Voosogh, A.S., Khomeiri, M., Kashani Nijad, M., Jafari, S.M., 2009. Effects of mint extract on the viability of probiotic bacteria in a native Iranian dairy drink (Doogh). ANRES. 16(1):156-164. [In Persian].
Roberfroid, MB. 1996. Functional effects of food components and the gastrointestinal system: chicory fructooligosaccharides. Nutr Rev. 54(11):38-42.
Zareali, M., Hojjati, M., Tahmozi didehban, S., Jouyandeh, H. 2015. Effect of echinophora (Echinophora cinerea Boiss) and Stachys (Stachys lavandulifolia Vahl) extracts on qualitative and sensory charactristics of Doogh. IJBSE. 46(3):327-337 [In Persian].