**3. Antimicrobial activity**

*Escherichia coli* (ATCC 25922), *Staphylococcus aureus* (ATCC 12600) and *Pseudomonas aeruginosa* (ATCC 27853) strains were cultured in brain heart infusion

**153**

*Comparative Analysis of the Chemical Composition, Antimicrobial and Antioxidant Activity…*

The inoculum (100 μL) of each bacterium was seeded in Mueller-Hinton agar, with filter paper impregnated with 50 μL of essential oil placed on the surface. The plates were incubated at 35°C and after 24 h, the inhibition halo was measured with a millimeter ruler [51]. The minimum inhibitory concentration (MIC) was determined using the broth dilution methodology [52] and performed in triplicates with the same bacterium used in solid media diffusion techniques. Initially, an aliquot of the essential oil prepared in DMSO was transferred to a test tube containing BHI broth. Serial dilutions were then performed resulting in concentra-

bacteria were added at each concentration and incubated at 35°C for 24 h. Tubes without bacteria were reserved for control of broth sterility and bacterial growth. After the incubation period, the minimum essential oil inhibitory concentration was defined as the lowest concentration which visibly inhibited bacterial growth observed by the absence of visible turbidity. To confirm growth inhibition, the BHI broth was subjected to the inoculum microbial seeding test on the surface of

The disc diffusion method evaluated the antibacterial activity of *C. zeylanicum*, *O. vulgare*, *Z. officinale*, *R. officinalis*, *C. longa* and *C. latifolia* essential oils to form inhibition halos against the growth of Gram-positive (*S. aureus*) and Gram-negative (*E. coli* and *P. aeruginosa*) bacteria strains. The diameters of the inhibition halos developed by the essential oils are shown in **Table 5**. The halos ranged from 7.67 to 15.33 mm. The largest inhibition halo against Gram-negative *E. coli* bacteria was quantified at 21 mm by *C. latifolia* essential oil. The best bactericidal activity for *S. aureus* was quantified at 15.66 mm by *C. longa*, while the Gram-positive *P. aeruginosa* was strongly inhibited by *C. longa* essential oil of, quantifying a halo of 12 mm.

concentration that prevents visible microbial growth in the culture medium by the

The bactericidal activity of *C. zeylanicum* essential oil was demonstrated by larger halos against the Gram-positive bacteria. Similar results were found in *C. zeylanicum* essential oil [34, 38, 53] and the authors reported cinnamaldehyde as responsible for the antimicrobial action. However, lower results were also described when evaluating the antimicrobial activity of this oil against *Salmonella* 

The MIC's of *C. zeylanicum* essential oil were quantified using *S. aureus* strain and obtained a similar inhibitory concentration to that in this research, however *E. coli* and *Salmonella typhi* concentrations were far superior to that described in our study [55]. In another research conducted by Trajano et al. [4], concentrations are

To the antimicrobial potential of *R. officinalis* essential oil, Cordeiro [56] also obtained inhibition halos for *S. aureus*, as well as Ribeiro [57] for *E. coli*, both using this same essential oil as antimicrobial. The antimicrobial activity of this oil for the strains tested in broth dilution showed MIC's similar to the experiment performed by Silva et al. [58]. However, values for such concentrations have also been reported

On the other hand, *O. vulgare* essential oil has demonstrated efficiencies for both Gram-positive and Gram-negative strains which can be observed by Stefanakis et al. [60] and Sankar et al. [61] who reported antimicrobial activity of this essential oil

Soković et al. [62] obtained MIC's for *O. vulgare* essential oil smaller than those

described in the results of this study for *S. aureus*, *E. coli*, *Salmonella enteritidis*

*typhimurium* and *E. coli*, being its better inhibition against *S. aureus* [54].

scale, recommended by the Clinical and Laboratory Standards Institute [50].

tions of 5–2000 μg/mL. Microbial suspensions containing 1.5 × 108

The minimum inhibitory concentration (MIC) in μg mL<sup>−</sup><sup>1</sup>

action of the natural product, is being reported in **Table 5**.

relatively lower than those observed in this study.

similar to this research against the same bacteria tested.

in smaller units by Thanh et al. [59].

UFC/mL following the MacFarland

CFU/mL of the

, the lowest visible

*DOI: http://dx.doi.org/10.5772/intechopen.86576*

the plate-count agar.

broth for 24 h at 37°C and then diluted to 108

#### *Comparative Analysis of the Chemical Composition, Antimicrobial and Antioxidant Activity… DOI: http://dx.doi.org/10.5772/intechopen.86576*

broth for 24 h at 37°C and then diluted to 108 UFC/mL following the MacFarland scale, recommended by the Clinical and Laboratory Standards Institute [50].

The inoculum (100 μL) of each bacterium was seeded in Mueller-Hinton agar, with filter paper impregnated with 50 μL of essential oil placed on the surface. The plates were incubated at 35°C and after 24 h, the inhibition halo was measured with a millimeter ruler [51]. The minimum inhibitory concentration (MIC) was determined using the broth dilution methodology [52] and performed in triplicates with the same bacterium used in solid media diffusion techniques. Initially, an aliquot of the essential oil prepared in DMSO was transferred to a test tube containing BHI broth. Serial dilutions were then performed resulting in concentrations of 5–2000 μg/mL. Microbial suspensions containing 1.5 × 108 CFU/mL of the bacteria were added at each concentration and incubated at 35°C for 24 h. Tubes without bacteria were reserved for control of broth sterility and bacterial growth. After the incubation period, the minimum essential oil inhibitory concentration was defined as the lowest concentration which visibly inhibited bacterial growth observed by the absence of visible turbidity. To confirm growth inhibition, the BHI broth was subjected to the inoculum microbial seeding test on the surface of the plate-count agar.

The disc diffusion method evaluated the antibacterial activity of *C. zeylanicum*, *O. vulgare*, *Z. officinale*, *R. officinalis*, *C. longa* and *C. latifolia* essential oils to form inhibition halos against the growth of Gram-positive (*S. aureus*) and Gram-negative (*E. coli* and *P. aeruginosa*) bacteria strains. The diameters of the inhibition halos developed by the essential oils are shown in **Table 5**. The halos ranged from 7.67 to 15.33 mm. The largest inhibition halo against Gram-negative *E. coli* bacteria was quantified at 21 mm by *C. latifolia* essential oil. The best bactericidal activity for *S. aureus* was quantified at 15.66 mm by *C. longa*, while the Gram-positive *P. aeruginosa* was strongly inhibited by *C. longa* essential oil of, quantifying a halo of 12 mm.

The minimum inhibitory concentration (MIC) in μg mL<sup>−</sup><sup>1</sup> , the lowest visible concentration that prevents visible microbial growth in the culture medium by the action of the natural product, is being reported in **Table 5**.

The bactericidal activity of *C. zeylanicum* essential oil was demonstrated by larger halos against the Gram-positive bacteria. Similar results were found in *C. zeylanicum* essential oil [34, 38, 53] and the authors reported cinnamaldehyde as responsible for the antimicrobial action. However, lower results were also described when evaluating the antimicrobial activity of this oil against *Salmonella typhimurium* and *E. coli*, being its better inhibition against *S. aureus* [54].

The MIC's of *C. zeylanicum* essential oil were quantified using *S. aureus* strain and obtained a similar inhibitory concentration to that in this research, however *E. coli* and *Salmonella typhi* concentrations were far superior to that described in our study [55]. In another research conducted by Trajano et al. [4], concentrations are relatively lower than those observed in this study.

To the antimicrobial potential of *R. officinalis* essential oil, Cordeiro [56] also obtained inhibition halos for *S. aureus*, as well as Ribeiro [57] for *E. coli*, both using this same essential oil as antimicrobial. The antimicrobial activity of this oil for the strains tested in broth dilution showed MIC's similar to the experiment performed by Silva et al. [58]. However, values for such concentrations have also been reported in smaller units by Thanh et al. [59].

On the other hand, *O. vulgare* essential oil has demonstrated efficiencies for both Gram-positive and Gram-negative strains which can be observed by Stefanakis et al. [60] and Sankar et al. [61] who reported antimicrobial activity of this essential oil similar to this research against the same bacteria tested.

Soković et al. [62] obtained MIC's for *O. vulgare* essential oil smaller than those described in the results of this study for *S. aureus*, *E. coli*, *Salmonella enteritidis*

*Essential Oils - Oils of Nature*

**Table 3.**

**152**

**Table 4.**

**3. Antimicrobial activity**

*Chemical composition of lemon tahiti essential oil.*

component [46]. Same results were observed by us in the present study. However, there are also descriptions of geranial as its major constituent in this oil [47].

**Peak** *Citrus latifolia* **(tahiti lemon) E.O.**

*Chemical composition of Curcuma longa (saffron) and Zingiber officinale (ginger) essential oils.*

 ρ-Cymene **10.86** D-Limonene 8.85 Cyclooctanone 1.54 ρ-Mentha-E-2,8(9)-dien-1-ol 2.47 trans-Pinocarveol 3.23 14.70 Pinocarvone 2.02 ρ-Cymen-8-ol 3.02 Bicyclo(3.1.1)hept-2-ene-2-carboxaldehyde,6,6-dimethyl- 5.34 Myrtenol 6.31 trans-Carveol 1.58 cis-Carveol **11.59** Carvone 1.68 19.02 Carvone oxide 3.69 Limonene dioxide **25.92** 1,2-Cyclohexanediol 1-methyl-4-(1-methyleth) 8.10 7-Oxabicyclo[4.1.0]heptane,1-methyl-4-1-(1-methylethyl) 1.24 2,7-Octadiene-1,6-diol,2,6-dimethyl 2.56

**Peak** *C. longa* **(saffron) E.O.** *Z. officinale* **(ginger) E.O.**

 ar-Curcumene 1.58 ar-Curcumeno 3.33 **Turmerone 55.43 α-Zingibereno 27.14** β-Sesquiphellandrene 1.10 **Nerolidol 13.51** — - β-Sesquifelandreno 9.45

**Compounds (%) Compounds (%)**

**Compounds (%)**

*Escherichia coli* (ATCC 25922), *Staphylococcus aureus* (ATCC 12600) and *Pseudomonas aeruginosa* (ATCC 27853) strains were cultured in brain heart infusion

The chemical composition obtained from *C. latifolia* leaves is presented in **Table 4**. The essential oil obtained presented from 17 components with the main constituents being limonene dioxide (25.92%), cis-carveol (11.59%) and ρ-cymene (10.86%). Similarly, it was found in the researches carried out in *C. latifolia* tanaka, identifying 17 compounds and limonene as the major compound with 46.3% [48] and 58.43% [49].


#### **Table 5.**

*Diameters of inhibition halos (IH) and minimum inhibitory concentrations (MIC) for the essential oils activity against bacterial strains.*

and *Salmonella typhimurium*. Sarikurkcu et al. [38, 63], also performed an assay to determine the MIC against *S. aureus* and *E. coli*, obtaining results similar to those observed.

For *C. longa* essential oil, the largest halos were quantified for Gram-positive bacteria, similar to results found by Gupta et al. [64], Teles et al. [38] and Mishra et al. [65] who reported the formation of halos against the same bacteria in this study submitted to antimicrobial activity assays. Singh et al. [66] also observed satisfactory MIC values for the control of the microorganisms tested in this study.

In relation to the bactericidal effect of the *Z. officinale* essential oil, the values obtained in our disc diffusion test are superior to those obtained in the study by Singh et al. [67], where the authors did not obtain inhibition halos for *E. coli* and *S. aureus*, and the same were reported by Grégio et al. [68]. However, MIC values were similar to those found by Sasidharan and Menon [69].
