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Antibacterial Evaluation of Croton
grewioides Baill Essential Oil on Beta-
Lactam and Aminoglycoside Antibiotics
José Jonas Ferreira Viturino , Cicera Janaine Camilo , Natália Kelly Gomes De Carvalho ,
Joice Barbosa Do Nascimento , Maria Inácio Silva , Mariana Pereira da Silva ,
José Walber José Walber Castro , Geane Gabriele de Oliveira Souza , Fabíola Fernandes Galvão Rodrigues ,
JOSE GALBERTO MARTINS DA COSTA *
Posted Date: 21 August 2024
doi: 10.20944/preprints202408.1514.v1
Keywords: antibacterial activity; Croton grewioides Baill; essential oil; synergistic effect
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Article
Antibacterial Evaluation of Croton grewioides Baill
Essential Oil on Beta-Lactam and Aminoglycoside
Antibiotics
José Jonas Ferreira Viturino 1,2, Cicera Janaine Camilo 1, Natália Kelly Gomes de Carvalho 2, Joice
Barbosa do Nascimento 1,2, Maria Inacio da Silva 1,2, Mariana Pereira da Silva 2, José Walber
Gonçalves Castro 1,2, Geane Gabriele de Oliveira Souza 1,2, Fabíola Fernandes Galvão Rodrigues
1,2 and José Galberto Martins da Costa 1,2*
1 Programa de Pós-graduação em Química Biológica, Departamento de Química Biológica, Universidade
Regional do Cariri, Crato, CE, Brasil.
2 Laboratório de Pesquisa de Produtos Naturais, Departamento de Química Biológica, Universidade
Regional do Cariri, Rua Coronel Antônio Luíz – Pimenta, 63105-010, Crato, CE, Brasil.
* Correspondence: galberto.martins@gmail.com;
Abstract: Croton grewioides Baill., a species native to the Caatinga, popularly known as canela de cunhã, is used
in traditional medicine to treat gastrointestinal diseases such as diarrhea, gastritis and stomach ulcers. This
study aimed to identify the chemical composition of the essential oil of C. grewioides (OECG) and evaluate its
antibacterial and antibiotic-modulating activities against both standard and multidrug-resistant bacteria. To
analyze the compounds, present in the oil, gas chromatography coupled with mass spectrometry (GC/MS) and
gas chromatography with flame ionization detector (GC/FID) techniques were used. In the bacteriological tests,
the Minimum Inhibitory Concentration (MIC) was obtained using the broth microdilution technique. The
modulating effect of the essential oil was determined by calculating the subinhibitory concentration followed
by serial microdilution of the antibiotics. The MIC reduction factor (FRC) was calculated and its data expressed
as a percentage. Analysis of the chemical composition identified the presence of the major compound estragole
with a relative abundance of 50.34%. The MIC values obtained demonstrated efficacy in K. pneumoniae isolated
from urine with MIC values of 512 µg/mL. OECG potentiated the effects of all antibiotics tested on S. aureus
ATCC 29213, K. pneumoniae Carbapnemase, E. coli ATCC 25922 and S. aureus ATCC 29213 strains with their
FRC of 97.65 %, 99.80 %, 99.85 % and 99.88 %, respectively. Thus, the essential oil of C. growioides presents
synergistic effects when combined with the tested antibiotics, in addition to acting in the fight against bacterial
resistance to antibiotics.
Keywords: antibacterial activity; Croton grewioides Baill; essential oil; synergistic effect;
1. Introduction
The discovery of antibiotics led to a reduction in mortality rates caused by bacterial infections in
hospital settings. However, the widespread and uncontrolled use of these drugs has promoted the
emergence of resistant microorganisms [1], making research into natural products a promising
alternative for combating microbial resistance [2].
In Brazil, the native fauna and flora have been used since before colonization by native peoples
to treat illnesses, accumulating enormous knowledge of medicinal flora [3]. Despite having one of
the greatest biodiversities in the world, research on native products of Brazilian flora is relatively
underdeveloped [4]. Among the Brazilian biomes, the Caatinga encompasses a large number of plant
species with therapeutic potential, but with few studies focused on the area of chemistry. Therefore,
research on species from the Caatinga is still in its infancy and their use is reserved, in most cases, for
use by traditional and local communities [5].
The species Croton grewioides Baill. (Euphorbiaceae), commonly known as canela de cunhã, is
widely distributed throughout the Nordeste region of Brazil and is characteristic of the Caatinga
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© 2024 by the author(s). Distributed under a Creative Commons CC BY license.
2
biome [6]. Its use in traditional medicine is through teas and infusions of fresh leaves, being
commonly used in the treatment of gastrointestinal disorders such as diarrhea, gastritis and in the
treatment of stomach ulcers, in addition to being used as a sedative agent, relieving pain symptoms
and having a calming effect [7, 8, 9, 10, 11]. Its anti-inflammatory, hepatoprotective and insecticidal
effect is also reported, being used by small and medium-sized farmers for pest control [12]. These
reported activities are likely associated with the chemical constituents anethole and estragole, which
are present in the essential oil of the leaves of C. grewioides.
Given the high prevalence of antibiotic-resistant microorganisms and the limited
microbiological studies on C. grewioides, this study aimed to identify the chemical composition of the
essential oil of C. grewioides and evaluate its antibacterial and antibiotic-modulating activities against
both standard and multidrug-resistant bacteria.
2. Materials and Methods
2.1. Collection and obtaining of essential oil
Aerial parts of C. grewioides were collected in the municipality of Crato, Ceará - Brazil, between
August and September 2022 in the morning. A specimen was deposited in the Herbarium Caririense
Dárdano de Andrade Lima of the Regional University of Cariri, where it was assigned registration
number 1619. The essential oil was obtained by hydrodistillation (Clevenger type apparatus), where
the sample of the dry plant material (2 kg) was crushed and subjected to distillation for 4h. The
obtained oil was dried over Na₂SO₄, yielding 0.20% (w/w).
2.2. GC-MS analysis
The essential oil was analyzed using a Shimadzu GC–MS QP2010 series, provided by Shimadzu
Scientific Instruments Inc. (Columbia, MD, USA), with fused silica capillary column SH-Rtx-5 (30 m
× 0.25 mm I.D.; 0.25 m film thickness) and the following temperature program: 80–180 ºC at 4 ºC/min,
then to 246 ºC at 6.6 ºC/min, closing with 10 min at 280 ºC, at 3.4 ºC/min, totaling analysis time of 30
min. He was used as the carrier gas, flow rate of 1.5 mL/min, split mode (1:100), and injection port
was set at 220 ºC. The quadrupole MS operating parameters: interface temperature (280 ºC) and ion
source (200 ºC); electron impact ionization at 70 eV; scan mass range of 40–350 m/z with sampling
rate of 1.0 scan/s. Injection volume: 1 µL of 500 ppm solution prepared with dichloromethane. The
constituents were identified by computational search using digital libraries of mass spectral data
(NIST 08) and by comparing their authentic mass spectra and reported in the literature [13].
2.3. GC-FID analysis
The essential oil was analyzed using a Shimadzu GC-2010 Plus equipped with flame ionization
detector (FID), provided by Shimadzu Scientific Instruments Inc. (Columbia, MD, USA), with fused
silica capillary column SH-Rtx-5 (30 m × 0.25 mm I.D.; 0.25 m film thickness) and the following
temperature program: 80–180 ºC at 4 ºC/min, then to 246 ºC at 6.6 ºC/min, closing with 10 min at 280
ºC, at 3.4 ºC/min, totaling analysis time of 30 min. He was used as the carrier gas, flow rate of 1.5
mL/min, split mode (1:15), and injection port was set at 220 ºC. Injection volume: 1 µL of 500 ppm
solution prepared with dichloromethane. The detector temperature was set to 300 °C. The linear
retention index was obtained by injecting a mixture of C8-C40 linear hydrocarbons under the same
conditions as the samples, as described by Van Den Dool and Kratz (1963). The identity of the
compounds was confirmed by comparing their retention indices and mass spectra with those taken
from the literature [13].
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2.4. Antibacterial activity
2.4.1. Determination of minimum inhibitory concentration (MIC)
To determine the antibacterial activity, the broth microdilution method containing the Brain
Heart Infusion Broth (BHI) culture medium was used based on the CLSI document [14]. The
following strains were used: Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29213, Klebsiella
pneumoniae Clinical Urine Isolate (ICU), Escherichia coli 06, Staphylococcus aureus 10, Klebsiella
pneumoniae Carbapnemase (KPC) and Klebsiella pneumoniae Betalactamase (BLSE).
The essential oil of C. grewioides (OECG) was prepared at a concentration of 1024 µg/mL diluted
in sterile water and DMSO. In the assay, serial microdilutions of 100 µL of OECG were performed
where the control group contained only the inoculum. The tests were conducted in triplicate and
incubated in the microbiological incubator at 37º C for 24 h. The readings were performed by the
colorimetric method using 25 µL of 0.01% sodium resazurin [14].
2.4.2. Antibiotic modulation tests
The antibiotics used to evaluate the modulating activity of OECG were amoxicillin, penicillin,
gentamicin and amikacin. The methodology for performing the test was adapted from Coutinho et
al. [15]. Based on the calculation of the subinhibitory concentrations (MIC/8), serial microdilution of
the respective antibiotics was performed. The control group contained only OECG and the inoculum.
The plates were placed in an oven at 37°C for 24 h and then bacterial growth was evaluated by the
colorimetric method with sodium resazurin (0.01%).
2.4.3. Determination of the MIC reduction factor
To determine the MIC Reduction Factor (FRC) expressed as a percentage, the value of X was
calculated using the formula below. To do this, a simple rule of three was used, which obtained the
following conclusions: X is equal to the values of the test MIC (which contains the essential oil
together with the antibiotic) multiplied by the value of 100 divided by the values of the control MIC
(antibiotics and bacteria). Thus, the values obtained in the CRF calculation represent the percentage
of reduction in the MIC values in the presence of the essential oil.
FRC = 100 - X
X = CIM test x 100 / CIM control
FCR = 100 – (CIM test x 100 / CIM control)
2.4.4. Statistical analysis
The tests were analyzed by two-way ANOVA followed by the Bonferroni test using GranphPad
Prism 10.0 software. Results expressed at p<0.05 were considered statistically significant.
3. Results and discussion
3.1. Composition of C. grewioides essential oil
The evaluation of the chemical composition of the essential oil extracted from the aerial parts of
C. grewioides revealed the presence of 2.92 % of monoterpenoid compounds, 5.45 % of oxygenated
monoterpenes, 4.40 % of sesquiterpenes, 0.42 % of oxygenated sesquiterpenes, 69.73 % of
phenylpropanoids and 10.99 % of ethers. The major compound identified in the chemical analysis
was estragole with 50.34 %, followed by the compounds methyl eugenol with 19.39 % and anisole
with 10.99 %. In total, 18 compounds were identified by chromatography analysis (Figure 1 and
Figure 2), whose percentages are described in Table 1.
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Figure 1. Chromatogram GC/MS.
Figure 2. Chromatogram GC/MS.
Table 1. Compounds identified in the essential oil of the aerial parts of C. grewioides.
Compounds (%) RT (min) IR1 IR2
estragole 50.34 6.53 1.246 1.226
methyl eugenol 19.39 8.10 1.339 1.359
anisole 10.99 7.25 1.283 1.264
methyl-trans-isoeugenol 6.09 8.94 1.403 1.422
1,8-cineole 4.2 5.01 1.156 1.164
α-selinene 1.97 11.32 1.628 1.648
β-elemene 1.37 8.31 1.355 1.375
β-ocimene 1.09 5.15 1.165 1.145
camphor 1.08 6.09 1.223 1.243
α-pinene 0.82 3.97 1.070 1.050
trans-caryphyllene 0.81 8.64 1.380 1.400
caryphyllene oxide 0.42 10.35 1.553 1.572
camphene 0.36 4.14 1.090 1.097
β-myrcene 0.29 4.53 1.122 1.142
2-β-pinene 0.25 4.45 1.116 1.119
isoledene 0.25 8.80 1.391 1.373
isoborneol 0.17 6.34 1.236 1.255
sabinense 0.11 4.38 1.111 1.123
Total 100 %
RT: Retention time (min); IR1: Linear retention index obtained; IR2: Linear retention index obtained from the
literature.
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In the study by Santos et al. [16] that evaluated the chemical composition of essential oils from
three chemotypes of C. grewioides extracted from branches, in the first chemotype the major
component was (E)-anethole with 70.5 %. In the second chemotype, eugenol with 84.2 %. Finally, in
the third chemotype, (Z)-methyl with 53.4 % was presented. In another study carried out by Silva et
al. [17], the composition of the essential oil of C. grewioides extracted from branches revealed (E)-
anethole as the major component with 47.8 %. Comparing these studies with the components
identified in this research, the absence of the compound (E)-anethole in the chromatographic analyses
is demonstrated, which can be explained by environmental factors such as collection site, collection
period, part of the plant used, vegetative state of the plant, genetic factors, among others [18, 19].
Studies evaluating the chemical composition of the essential oil extracted from the leaves of C.
grewioides, reported by Costa et al. [20] and Siqueira et al. [21], reveal the presence of estragole with
76.8 % and 45.95 % respectively as the major components present in its oil. As in the studies by Leite
et al. [22] and Andrade et al. [8], its percentage present in the essential oil is 78 % and 84.7 %, whose
estragole compound and its percentages are close to those found in this study.
The variation in the chemical composition of essential oils is likely a result of the plant's adaptive
mechanisms, which utilize secondary metabolites to defend against predators and to adapt to
variations in both biotic and abiotic environmental conditions [23].
3.2. Antibacterial activity
The antibacterial activity of C. grewioides essential oil tested on standard strains demonstrated
significant results against the K. pneumoniae (ICU) strain with an MIC value of 512 µg/mL. In
multidrug-resistant strains, the oil presented better results on the K. pneumoniae (KPC) and K.
pneumoniae (BLSE) strains, with MIC values of 512 and 256 µg/mL, respectively. The other MIC values
of the strains used in this study are described in Table 2.
Table 2. Minimum Inhibitory Concentration (MIC) valuesof OECG against standard and
multiresistant bacteria.
Organisms MIC (µg/mL)
Multidrug-resistant strains
SA 10 ≥1024
EC 06 ≥1024
KPC 512
KP BLSE 256
Standard strains
EC 25922 ≥1024
SA 29213 ≥1024
KP ICU 512
SA 10 - S. aureus 10; EC 06 - E. coli 06; KP BLSE - K. pneumoniae BLSE; KPC - K. pneumoniae Carbapenemáse; KP
ICU – K. pneumoniae Clinical Urine Isolate; SA 29213 - S. aureus ATCC 29213; EC 25922 - E. coli ATCC 25922;.
In the strains tested, there was no inhibition of bacterial growth even at a concentration of 1024
µg/mL of OECG, this was due to the fact that the strains tested present biochemical mechanisms
related to bacterial resistance, which implies a high degree of pathogenicity, since the strains tested
in this study are involved in various infections in hospitalized patients, mainly due to urinary tract
infections, as is the case with strains of E. coli and K. pneumoniae [24].
Studies such as those by Rodrigues and collaborators [25], which demonstrate the antibacterial
potential of C. grewioides essential oil against standard strains of Pseudomonas aeruginosa ATCC 15442
and S. aureus ATCC 12692, obtaining minimum inhibitory dose (MID) results of 1 and 0.5 mg/L of air
in S. aureus. These findings corroborate the results obtained in this research, suggesting that
compounds present in the essential oil, such as estragole and anethole, which are commonly reported
in chemical studies, are directly related to the antibacterial effects observed in the tests.
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3.3. Modulating activity
The beta-lactam antibiotics used in the study were amoxicillin and penicillin, which have a
mechanism of action associated with the inhibition of bacterial cell wall synthesis, leading to
destabilization of the cell wall's macrostructure and leakage of intracellular fluid due to membrane
lysis. The aminoglycoside antibiotics were amikacin and gentamicin, which act by inhibiting the
protein synthesis mechanism specifically in the 30S subunit of bacterial ribosomes [26].
From the results obtained in the modulation tests, OECG enhanced the effects of amoxicillin on
strains of S. aureus 10, K. pneumoniae (ICU) and S. aureus ATCC 29213, with their respective MIC
Reduction Factors (FRC) of 85 %, 84.16 % and 97.65 %. As for penicillin, it presented results on strains
of S. aureus 10 and S. aureus ATCC 29213, with its most potent effects on K. pneumoniae (KPC) with
a FRC of 99.95 %, E. coli 06 with 99.80 % and K. pneumoniae (ICU) with 98.02 %. The remaining results
are detailed in Table 3.
When associated with gentamicin, OECG demonstrated promising results in both standard and
multidrug-resistant strains, except for K. pneumoniae (KPC), which presented negative FRC,
indicating an antagonistic effect. Of these results, the standard strains obtained FRC of 99.85 % in E.
coli ATCC 25922, 99.80 % in S. aureus ATCC 29213 and 99.75 % in K. pneumoniae (ICU). As for
amikacin, OECG was more effective against S. aureus ATCC 29213 with FRC of 99.88 %.
Table 3. Minimum inhibitory concentrations (µg/mL) of antibiotics in the absence and presence of C.
growioides essential oil and their MIC reduction factors (FRC).
Substance Organisms MIC of antibiotics (µg/mL)
Amoxicillin Penicillin Gentamicin Amikacin
Control SA 10 40 8 8 40
EC 06 40 256 8 40
KP BLSE 256 1024 1024 32
KPC 645 1024 812 512
KP ICU 101 406 203 161
SA 29213 256 256 128 256
EC 25922 256 512 203 128
OECG test SA 10 6 0.5 0.5 2
EC 06 161 0.5 0.5 3
KP BLSE 256 1024 256 161
KPC 1024 0.5 1024 512
KP ICU 16 8 0.5 4
SA 29213 6 40 0.25 0.30
EC 25922 203 512 0.30 20
FRC SA 10 85 % 93.75 % 93.75 % 95 %
EC 06 - 302.5 % 99.80 % 93.75 % 92.5 %
KP BLSE 0 % 0 % 75 % - 403.12 %
KPC - 58.76 % 99.95 % - 26.10 % 0 %
KP ICU 84.16 % 98.02 % 99.75 % 97.51 %
SA 29213 97.65 % 84.37 % 99.80 % 99.88 %
EC 25922 20.7 % 0 % 99.85 % 84.37 %
SA 10 - S. aureus 10; EC 06 - E. coli 06; KP BLSE - K. pneumoniae BLSE; KPC - K. pneumoniae Carbapenemáse; KP
ICU – K. pneumoniae Clinical Urine Isolate; SA 29213 - S. aureus ATCC 29213; EC 25922 - E. coli ATCC 25922;
OECG - C. grewioides essential oil; FRC - MIC reduction factor.
In the graph shown in Figure 3, the modulating effect of OECG associated with the antibiotic
amoxicillin against the strains used in this study, a synergistic effect was observed in the
multiresistant strain S. aureus 10, and in all standard strains tested. In the other strains tested, an
antagonistic effect was observed in S. aureus 10, E. coli 06, K. pneumoniae (BLSE), K. pneumoniae (KPC).
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Figure 3. Modulating potential of OECG on the antibiotic activity of amoxicillin against strains SA 10
- S. aureus 10; EC 06 - E. coli 06; KP BLSE - K. pneumoniae BLSE; KPC - K. pneumoniae Carbapenemáse;
KP ICU – K. pneumoniae Clinical Urine Isolate; SA 29213 - S. aureus ATCC 29213; EC 25922 - E. coli
ATCC 25922. Source: Author. Two-way ANOVA followed by Bonferroni post-test, using GraphPad
Prism 10.0 software. * p <0.0001; **p <0.0005.
OECG also showed synergistic effects in association with penicillin. Figure 4 demonstrates the
synergistic effects in all strains tested, with the exception of K. pneumoniae (BLSE) and E. coli ATCC
29213, which showed equivalence of means. One of the main mechanisms of bacterial resistance to
beta-lactam antibiotics is the production of the beta-lactamase enzyme that hydrolyzes the beta-
lactam ring of these antibiotics, inhibiting their action [27]. The lack of effects of OECG associated
with penicillin against K. pneumoniae (BLSE) may be related to its resistance mechanism of the type
of producer of the beta-lactamase enzyme that acts to inhibit the action of these drugs [28].
Figure 4. Modulating potential of OECG on the antibiotic activity of penicillin against strains SA 10 -
S. aureus 10; EC 06 - E. coli 06; KP BLSE - K. pneumoniae BLSE; KPC - K. pneumoniae Carbapenemáse;
KP ICU – K. pneumoniae Clinical Urine Isolate; SA 29213 - S. aureus ATCC 29213; EC 25922 - E. coli
ATCC 25922. Source: Author. Two-way ANOVA followed by Bonferroni post-test, using GraphPad
Prism 10.0 software. * p <0.0001; **p <0.0005.
Carbapinemase-producing bacteria exhibit a broad resistance mechanism against several beta-
lactam antibiotics, acting in the hydrolysis of drugs such as penicillins, cephalosporins, monobactams
and carbapenems [29, 30]. In this study, OECG potentiated the effect of penicillin, giving evidence
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that it may be involved in increasing membrane permeability, in which the entry of the drug into the
cell is facilitated, in addition to acting at the genetic level, altering its resistance mechanism, since the
drug is able to enter the cell, overcoming enzymatic barriers and inhibiting peptidoglycan synthesis,
leading to rupture and extravasation of intracellular content [31].
Amoxicillin, which also acts to inhibit cell wall synthesis, showed synergism in standard strains
and multiresistant S. aureus 10. The mechanism is similar to that of penicillin. Furthermore, more
effective inhibition by the drug was expected in standard strains, since these do not have as many
resistance mechanisms as multiresistant strains.
In corroboration with these data, Medeiros et al. [32] evaluated the effects of C. grewioides
essential oil on the modulation of the antibiotics norfloxacin and tetracycline against Staphylococcus
aureus (SA-1199B) with efflux pump resistance mechanism, and demonstrated that in the absence of
the oil, the antibiotic presents an inhibitory concentration of 64 µg/mL for norfloxacin and 32 µg/mL
for tetracycline. In the presence of the oil, these values dropped to 16 µg/mL for norfloxacin and 0.5
µg/mL for tetracycline, enhancing the effect of the antibiotic even in the presence of the overexpressed
gene for efflux pump.
The effect of OECG against aminoglycoside antibiotics showed that amikacin (Figure 5)
presented synergistic effects in all strains tested except K. pneumoniae (BLSE), which presented an
antagonistic effect, and K. pneumoniae (KPC), which demonstrated equivalence to the control. As for
gentamicin (Figure 6), synergistic effects were observed in all strains tested, except for K. pneumoniae
(KPC), which exhibited an antagonistic effect.
Figure 5. Modulating potential of OECG on the antibiotic activity of amikacin against strains SA 10 -
S. aureus 10; EC 06 - E. coli 06; KP BLSE - K. pneumoniae BLSE; KPC - K. pneumoniae Carbapenemáse;
KP ICU – K. pneumoniae Clinical Urine Isolate; SA 29213 - S. aureus ATCC 29213; EC 25922 - E. coli
ATCC 25922. Source: Author. Two-way ANOVA followed by Bonferroni post-test, using GraphPad
Prism 10.0 software. * p <0.0001; **p <0.0005.
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Figure 6. Modulating potential of OECG on the antibiotic activity of gentamicin against strains SA 10
- S. aureus 10; EC 06 - E. coli 06; KP BLSE - K. pneumoniae BLSE; KPC - K. pneumoniae Carbapenemáse;
KP ICU – K. pneumoniae Clinical Urine Isolate; SA 29213 - S. aureus ATCC 29213; EC 25922 - E. coli
ATCC 25922. Source: Author. Two-way ANOVA followed by Bonferroni post-test, using GraphPad
Prism 10.0 software. * p <0.0001; **p <0.0005.
Protein synthesis inhibitors that act on bacterial ribosomes in the 30S and 50S subunits are
mainly classified as aminoglycosides, tetracyclines, and aphenicols, among others. These
antibacterial drugs have greater selectivity precisely because they target components of the bacterial
ribosome that are distinct from those of eukaryotic ribosomes [26]. For amikacin, promising results
were observed in both standard and multidrug-resistant strains, with synergistic effects reported in
all standard strains as well as in the multidrug-resistant E. coli 06 and S. aureus 10.
Essential oils, due to their hydrophobic nature, interfere in essential biochemical processes for
the cell when they come into contact with bacterial membranes, such as the respiratory chain and
energy production, which leads to the deregulation of membrane permeability, facilitating the entry
of antibiotic drugs, in addition to interfering in the efflux pump resistance mechanism, since this
requires the binding of the adenosine triphosphate (ATP) molecule for its functioning in the
“expulsion” of toxic substances [25, 33].
Linked to the hydrophobic character of essential oils, their effects on the antibacterial and
modulating activity reported in this research may be related to the various compounds of the terpene
class present in their composition that have an intrinsic relationship to this activity [34].
4. Conclusions
The data obtained in this research allow us to conclude that OECG has synergistic effects when
associated with the antibiotics amoxicillin, penicillin, amikacin and gentamicin against all strains
tested, with the most potent effects observed in E. coli 06, K. pneumoniae (KPC), S. aureus ATCC 29213
and E. coli ATCC 25922. Thus, OECG has the potential to be used in future antimicrobial therapies,
acting as a modulating agent in the fight against bacterial resistance to antibiotics. However, further
research on its action on bacterial membrane permeability, genetic alterations of its resistance
mechanism, in addition to analysis of isolated compounds are necessary to verify whether the efficacy
of the essential oil is linked to the major compound and thus be able to use it in future antibacterial
treatments.
Author Contributions: Author Contribution Statement: Research, writing, formatting, methodology,
investigation, J. J.F,V.; methodology, investigation, C.J.C.; formal analysis, data curation, N.K.G.C.; formal
analysis, data curation, writing, J.B.N.; methodology, investigation, M.I.S.; methodology, investigation, M.P.S.;
formal analysis, data curation, J. W. G. C.; methodology, investigation, G.G.O.S.; methodology, investigation,
validation, writing, F.F.G.R.; conceitualization, methodology, supervision, writing, J.G.M.C.
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Data Availability Statement: The raw data supporting the conclusions of this article will be made available by
the authors on request.
Acknowledgments: This work was carried out at the Laboratório de Pesquisa de Produtos Naturais (LPPN) of
the Departamento de Química Biológica at the Universidade Regional do Cariri (URCA), with support from the
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), Conselho Nacional de
Desenvolvimento Científico e Tecnológico (CNPq), Financiadora de Estudos e Projetos (FINEP), Fundação
Cearense de Apoio ao Desenvolvimento Científico e Tecnológico (FUNCAP), Empresa Brasileira de Pesquisa
Agropecuária (EMBRAPA) and Instituto Nacional de Ciência e Tecnologia – Alimentos (INCT-ALIM).
Conflicts of Interest: The authors declare no conflict of interest.
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