**5. Mechanism of action**

It has been demonstrated that the antimicrobial effects of the Eos acts by causing structural and functional damages to the bacterial cell membrane. It is also indicated that the optimum range of hydrophobicity is involved in the toxicity of the EOs.

Application of antimicrobials by different exposure methods, such as vapor phase compared to direct contact method, of mustard and clove EOs showed noteworthy differences [63]. The stereochemistry, lipophilicity and other factors affected the biological activity of these compounds which might be altered positively or negatively by slight modifications [64]. It has been shown that plant substances affect microbial cells by various antimicrobial mechanisms, including

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*Essential Oil as Antimicrobial Agents: Efficacy, Stability, and Safety Issues for Food Application*

Carvacrol increases the heat shock protein 60 HSP 60 (GroEL) protein and inhibited the synthesis of flagellin highly significantly in *E. coli* O157:H7 [66]. There are concerns regarding the enhanced aroma and taste of oregano at the higher levels of application in food items, especially at 1% [72]. The apparent antimicrobial efficacy of plant origin antimicrobials depends on factors such as the method of extracting EOs from plant material, the volume of inoculum, growth phase, culture medium used, and intrinsic or extrinsic properties of the food such as pH, fat, protein, water content, antioxidants, preservatives, incubation time/temperature, packaging procedure, and physical structure of food [73, 74]. Another important parameter regarding effects of food preservatives is ability to reduce the pH level inside the bacterial cell (pHin). It has been shown that pHin of both *E. coli* and

Generally, Gram-negative bacteria are less sensitive to the antimicrobials because of the lipopolysaccharide outer membrane of this group, which restricts diffusion of hydrophobic compounds. However, this does not mean that Grampositive bacteria are always more susceptible [27]. Gram-negative bacteria are usually more resistant to the plant-origin antimicrobials and even show no effect,

When the combined effect of substances is higher than the sum of the individual effects, this is synergy; antagonism happens when a combination shows less effect compared to the individual applications [27]. Synergistic effects of some compounds, in addition to the major components in the EOs, have been shown in some studies [76–78]. Application of a certain combination of carvacrol-thymol can

Synergism between carvacrol and p-cymene, a very weak antimicrobial, might facilitate carvacrol's transportation into the cell by better swelling the *B. cereus* cell wall [27]. Antimicrobial activity of combination of cinnamon and clove EOs in vapor phase showed better antimicrobial with less active concentration in the vapor phase compare to liquid phase [63]. Thymol and carvacrol showed synergistic and antagonistic effects, in different combinations of cilantro, coriander, dill and eucalyptus EOs (each containing several components) and mixtures of cinnamaldehyde and eugenol, against *Staphylococcus sp*., *Micrococcus sp*., Bacillus sp. and *Enterobacter sp*. [27]. An antagonistic effect on *Bacillus cereus* was seen in rice when carvacrol and p-cymene were used with salt; high-hydrostatic pressure showed a synergistic effect in combination with thymol and carvacrol against *L. monocytogenes*. Vacuum packing in combination with oregano EOs showed a synergistic effect against *L. monocytogenes* with 2–3 log10 reduction. Similar results have been recorded when clove and coriander EOs have been used against *Aeromonas hydrophila* on vacuumpacked pork. Application of oregano EO has a synergistic effect in modified-atmosphere packaging (MAP) including 40% CO2, 30% N2 and 30% O2. The available oxygen is another factor antagonistic on EO activities; by decreasing the oxygen level, the sensitivity of micro-organisms to the EOs has been increased [27].

Salmonella has been reduced by the effect of mustard's EOs [71].

**6. Synergistic and antagonistic effects of components**

improve the efficacy of Eos against pathogenic micro-organisms [79].

compared to Gram-positive bacteria [61, 75].

attacking the phospholipid bilayer of the cell membrane, disrupting enzyme systems, compromising the genetic material of bacteria, and forming fatty acid hydroperoxidase caused by oxygenation of unsaturated fatty acids [65–70]. Allyl isothiocyanate derived from mustard seems to have multi-targeted mechanisms of action in metabolic pathways, membrane integrity, cellular structure and statistically significant higher release of the cell compounds of *Escherichia coli*

*DOI: http://dx.doi.org/10.5772/intechopen.92305*

O157:H7 [71].

#### *Essential Oil as Antimicrobial Agents: Efficacy, Stability, and Safety Issues for Food Application DOI: http://dx.doi.org/10.5772/intechopen.92305*

attacking the phospholipid bilayer of the cell membrane, disrupting enzyme systems, compromising the genetic material of bacteria, and forming fatty acid hydroperoxidase caused by oxygenation of unsaturated fatty acids [65–70]. Allyl isothiocyanate derived from mustard seems to have multi-targeted mechanisms of action in metabolic pathways, membrane integrity, cellular structure and statistically significant higher release of the cell compounds of *Escherichia coli* O157:H7 [71].

Carvacrol increases the heat shock protein 60 HSP 60 (GroEL) protein and inhibited the synthesis of flagellin highly significantly in *E. coli* O157:H7 [66]. There are concerns regarding the enhanced aroma and taste of oregano at the higher levels of application in food items, especially at 1% [72]. The apparent antimicrobial efficacy of plant origin antimicrobials depends on factors such as the method of extracting EOs from plant material, the volume of inoculum, growth phase, culture medium used, and intrinsic or extrinsic properties of the food such as pH, fat, protein, water content, antioxidants, preservatives, incubation time/temperature, packaging procedure, and physical structure of food [73, 74]. Another important parameter regarding effects of food preservatives is ability to reduce the pH level inside the bacterial cell (pHin). It has been shown that pHin of both *E. coli* and Salmonella has been reduced by the effect of mustard's EOs [71].

Generally, Gram-negative bacteria are less sensitive to the antimicrobials because of the lipopolysaccharide outer membrane of this group, which restricts diffusion of hydrophobic compounds. However, this does not mean that Grampositive bacteria are always more susceptible [27]. Gram-negative bacteria are usually more resistant to the plant-origin antimicrobials and even show no effect, compared to Gram-positive bacteria [61, 75].
