ArticlePDF Available

The Potential of Use Basil and Rosemary Essential Oils as Effective Antibacterial Agents

Authors:

Abstract and Figures

The considerable therapeutical problems of persistent infections caused by multidrug-resistant bacterial strains constitute a continuing need to find effective antimicrobial agents. The aim of this study was to demonstrate the activities of basil (Ocimum basilicum L.) and rosemary (Rosmarinus officinalis L.) essential oils against multidrug- resistant clinical strains of Escherichia coli. A detailed analysis was performed of the resistance of the drug to the strains and their sensitivity to the tested oils. The antibacterial activity of the oils was tested against standard strain Escherichia coli ATCC 25922 as well as 60 other clinical strains of Escherichia coli. The clinical strains were obtained from patients with infections of the respiratory tract, abdominal cavity, urinary tract, skin and from hospital equipment. The inhibition of microbial growth by both essential oils, presented as MIC values, were determined by agar dilution. Susceptibility testing to antibiotics was carried out using disc diffusion. The results showed that both tested essential oils are active against all of the clinical strains from Escherichia coli including extended-spectrum β-lactamase positive bacteria, but basil oil possesses a higher ability to inhibit growth. These studies may hasten the application of essential oils in the treatment and prevention of emergent resistant strains in nosocomial infections.
Content may be subject to copyright.
Molecules 2013, 18, 9334-9351; doi:10.3390/molecules18089334
molecules
ISSN 1420-3049
www.mdpi.com/journal/molecules
Article
The Potential of Use Basil and Rosemary Essential Oils as
Effective Antibacterial Agents
Monika Sienkiewicz
1,
*, Monika Łysakowska
1
, Marta Pastuszka
2
, Wojciech Bienias
2
and Edward Kowalczyk
3
1
Medical and Sanitary Microbiology Department, Medical University of Lodz, pl. Hallera 1,
Lodz 90-647, Poland; E-Mail: monika.lysakowska@umed.lodz.pl
2
Department of Dermatology, Pediatric Dermatology and Oncology, Medical University of Lodz,
Kniaziewicza 1/5, Lodz 91-347, Poland; E-Mail: dermatologia@umed.lodz.pl (M.P. & W.B.)
3
Pharmacology and Toxicology Department, Medical University of Lodz, Pl. Hallera 1,
Lodz 90-647, Poland; E-Mail: edward.kowalczyk@umed.lodz.pl
* Author to whom correspondence should be addressed; E-Mail: monika.sienkiewicz@umed.lodz.pl;
Tel./Fax: +48-42-639-3342.
Received: 19 June 2013; in revised form: 30 July 2013 / Accepted: 1 August 2013 /
Published: 5 August 2013
Abstract: The considerable therapeutical problems of persistent infections caused by
multidrug-resistant bacterial strains constitute a continuing need to find effective
antimicrobial agents. The aim of this study was to demonstrate the activities of basil (Ocimum
basilicum L.) and rosemary (Rosmarinus officinalis L.) essential oils against multidrug-
resistant clinical strains of Escherichia coli. A detailed analysis was performed of the
resistance of the drug to the strains and their sensitivity to the tested oils. The antibacterial
activity of the oils was tested against standard strain Escherichia coli ATCC 25922 as well
as 60 other clinical strains of Escherichia coli. The clinical strains were obtained from
patients with infections of the respiratory tract, abdominal cavity, urinary tract, skin and
from hospital equipment. The inhibition of microbial growth by both essential oils, presented
as MIC values, were determined by agar dilution. Susceptibility testing to antibiotics was
carried out using disc diffusion. The results showed that both tested essential oils are active
against all of the clinical strains from Escherichia coli including extended-spectrum
β-lactamase positive bacteria, but basil oil possesses a higher ability to inhibit growth.
These studies may hasten the application of essential oils in the treatment and prevention
of emergent resistant strains in nosocomial infections.
OPEN ACCESS
Molecules 2013, 18 9335
Keywords: basil oil; ESBL-positive strains; Escherichia coli; minimal inhibitory
concentration; rosemary oil
1. Introduction
The multidrug-resistant pathogenic strains of Escherichia coli are responsible for opportunistic
infections, including nosocomial ones, which are difficult to treat, especially in immunocompromised
patients. E. coli is responsible for severe cases of urinary tract infection, meningitis in newborns,
digestive system illnesses, and even pneumonia. In recent years, strains of Enterobacteriaceae
producing an extended spectrum β-lactamase have become a concern in the antimicrobial treatment of
persistent infections and control of infection in hospitals [1–5]. The most severe clinical cases are
isolated resistant strains of Escherichia coli, Klebsiella pneumonia, Pseudomonas aeruginosa and
Acinetobacter baumanii. Extended-spectrum β-lactamases are enzymes produced by Gram-negative
bacilli that mediate resistance to penicillin, cephalosporins, and monobactams [6–9]. The widespread
use of antimicrobial drugs, primarily antibiotics, and the transmissibility of resistance determinants
mediated by plasmids, transposons, and gene cassettes in integrons contribute to the spread of resistance.
A worrying development is the fast spread of resistant clones of these bacteria on a global
scale [10–12]. Thanks to growing resistance to drugs commonly used in clinical practice, effective
treatment availability is greatly reduced. This problem of increasing resistance has necessitated the
search for safe and effective factors that may be used to treat persistent bacterial infections.
Experimental research confirms the varied pharmaceutical activities of not only chemical compounds,
but also many plant metabolites such as polysaccharides, flavonoids, coumarins, glycosides, phenolic
acids, saponins and also essential oils. Plant metabolites are a very interesting alternative for synthetic
preparations: many of them have strong antimicrobial activity [13–17]. Their synergy of action with
each other and in combination with antibiotic and chemotherapeutic therapy make them a valued
complement to anti-infective therapy [18–23].
The Ocimum L. (basil) and Rosmarinus L. (rosemary) genera belong to the family Lamiaceae.
Among the plants known for medicinal value, basil and rosemary plants are highly regarded for their
therapeutic potentials. Ocimum basilicum L. and Rosmarinus officinalis L. essential oils offer promise
as biologically active constituents, in that they confer antibacterial, antifungal and antioxidant
properties. Basil and rosemary oils have long been used for treating, among other things, colds, fever,
cough, asthma, sinusitis and rheumatism, as well as accelerating the process of wound healing [24–28].
The aim of this work was to determine the antibacterial activity of basil oil from Ocimum basilicum L.
and rosemary oil from Rosmarinus officinalis L. against standard and clinical strains of Escherichia coli
isolated from patients and from hospital equipment.
Molecules 2013, 18 9336
2. Results
2.1. Chemical Composition of the Basil and Rosemary Essential Oils
Chemical analysis showed the presence of forty-eight constituents within basil oil from
Ocimum basilicum L., the main ones being estragole (86.4%), 1,8-cineole (4.9%) and trans-α-bergamotene
(3.0%). The chemical composition of the basil oil is presented in Table 1. The essential oil from
Rosmarinus officinalis L. contains thirty-seven components: the main ones being 1,8 cineole (46.4%),
camphor (11.4%), α-pinene (11.0%), β-pinene (9.2%) and camphene (5.2%). The chemical composition
of the rosemary oil is presented in Table 2.
Table 1. Constituents of basil oil.
Number of
compounds
Compound % RI
1 α-Pinene 0.4 929
2 Camphene 0.1 942
3 Sabinene 0.2 965
4 β-Pinene 0.6 969
5 2,3-Dehydro-1.8-cineole Tr 979
6 Myrcene 0.2 983
7 p-Cymene 0.1 1013
8 1,8-Cineole 4.9 1020
9 Limonene 0.4 1021
10 (E)-β-Ocimene 0.6 1038
11 trans-Linalool oxide (f) Tr 1058
12 Fenchone 0.2 1067
13 cis-Linalool oxide (f) Tr 1073
14 Linalool 1.2 1085
15 endo-Fenchol 0.2 1098
16 Camphor 0.7 1119
17 Menthone 0.1 1134
18 Isomenthone Tr 1143
19 Borneol 0.2 1149
20 Menthol 0.3 1159
21 Terpinen-4-ol 0.1 1163
22 Estragole 86.4 1188
23 Fenchyl acetate 0.3 1209
24 Bornyl acetate 0.3 1269
25 2-Hydroxycineol acetate Tr 1321
26 Eugenol methyl ether 0.5 1373
27 β-Bourbonene Tr 1385
28 β-Elemene 0.3 1389
29 cis-α-Bergamotene Tr 1412
30 β-Caryophyllene 0.1 1419
31 trans-α-Bergamotene 3.0 1435
Molecules 2013, 18 9337
Table 1. Cont.
Number of
compounds
Compound % RI
32 β-Sesquifenchene 0.2 1437
33 (Z)-β-Farnesene tr 1447
34 α-Humulene tr 1452
35 Cadina-1(6),4-diene tr 1459
36 trans-β-Bergamotene 0.2 1479
37 α-Bulnesene 0.1 1499
38 γ-Cdinene 0.5 1506
39 Calamenene tr 1510
40 β-Sesquiphellandrene 0.1 1514
41 p-Methoxycinnamaldehyde 0.5 1525
42 p-Methoxycinnamyl alcohol 0.4 1532
43 Spathulenol 0.1 1565
44 Caryophyllene oxide 0.1 1571
45 Humulene epoxide II 0.1 1595
46 1-epi-Cubenol 0.1 1603
47 T-Cadinol 0.7 1627
48 α-Cadinol tr 1639
RI-Retence Index; tr < 0.05%.
Table 2. Constituents of rosemary oil.
Number of
compounds
Compound % RI
1 Tricyclene 0.2 919
2 α-Thujene 0.1 923
3 α-Pinene 11.0 932
4 Camphene 5.2 944
5 Sabinene 0.1 966
6 β-Pinene 9.2 971
7 Myrcene 1.2 983
8 α-Phellandrene 0.2 997
9 Car-3-ene 0.1 1005
10 α-Terpinolene 0.1 1010
11 p-Cymene 1.3 1017
12 1,8-Cineole 46.4 1027
13 Limonene 1.0 1027
14 γ-Terpinene 1.0 1050
15 trans-Sabinene tr 1054
16 Terpinolene 0.2 1079
17 Linalool 0.5 1087
18 α-Campholenol tr 1096
19 endo-Fenchol tr 1102
20 Camphor 11.4 1124
Molecules 2013, 18 9338
Table 2. Cont.
Number of
compounds
Compound % RI
21 Borneol 3.1 1152
22 Terpinen-4-ol 0.4 1163
23 α-Terpineol 1.8 1175
24 Bornyl acetate 1.0 1269
25 α-Cubebene tr 1349
26 α-Ylangene tr 1372
27 α-Copaene 0.1 1377
28 Longifolene 0.1 1407
29 β-Caryophyllene 3.5 1421
30 α-Humulene 0.4 1452
31 γ-Muurolene 0.1 1471
32 α-Selinene tr 1492
33 α-Muurolene tr 1494
34 γ-Cadinene tr 1506
35 trans-Calamenene tr 1511
36 δ-Cadinene 0.1 1514
37 β-Caryophyllene oxide 0.1 1571
RI-Retention Index; tr < 0.05%.
2.2. Susceptibility Testing of Clinical Escherichia coli Strains
2.2.1. Susceptibility Testing of Clinical Escherichia coli (ESBL+) Strains
Extended spectrum β-lactamase production for the tested Escherichia coli clinical strains was
detected for strains from the abdominal cavity (n = 4), bronchia (n = 4), wounds (n = 4), urine (n = 4)
and for strains isolated from blood (n = 3) and catheters (n = 3). The results are shown in Figure 1.
The tested strains of Escherichia coli (ESBL+) were generally resistant to β-lactams, aminoglycosides
and quinolones recommended for susceptibility testing. Most of them were resistant to cephalosporins
and β-lactam antibiotics with such inhibitors as clavulanic acid, sulbactam and tazobactam.
2.2.2. Susceptibility Testing of Clinical Escherichia coli (ESBL) Strains
Escherichia coli ESBL negative strains, characterized by a much lower resistance to β-lactam
antibiotics, were resistant mainly to ampicillin, piperacillin, tikarcillin and also to ticarcillin/clavulanic
acid. Most of them were resistant to aminoglycosides (gentamicin, amikacin) and quinolones
(ciprofloksacin) and tetracycline. The results are shown in Figure 2.
Molecules 2013, 18 9339
Figure 1. The susceptibility testing of clinical Escherichia coli (ESBL+) strains.
The (ESBL+) and (ESBL) Escherichia coli were found to demonstrate significant resistance to the
reference antibiotics in the susceptibility tests. In our tests, all twenty-two of the tested clinical isolates
of ESBL positive E. coli were resistant to AMC (n = 21, 95%), CZ (n = 10, 45%), CXM (n = 12,
54%), GM (n = 10, 45%), AM (n = 21, 95%), PIP (n = 21, 95%), TIC (n = 21, 95%), TIM (n = 18,
81%), FEP (n = 11, 50%), CIP (n = 14, 64%), AN (n = 12, 54%), NET (n = 13, 59%), C (n = 17, 77%),
TE (n = 21, 95%) and SXT (n = 20, 91%). The thirty-eight tested ESBL negative E. coli strains were
generally resistant to AMC (n = 21, 55%), GM (n = 15, 39%), PIP (n = 23, 60%), TIC (n = 26, 68%),
TIM (n = 13, 34%), CIP (n = 16, 42%), AN (n = 14, 37%), C (n = 15, 39%), TE (n = 34, 89%) and
SXT (n = 26, 68%).
Molecules 2013, 18 9340
Figure 2. The susceptibility testing of clinical Escherichia coli (ESBL) strains.
2.3. The Susceptibility Escherichia coli Bacterial Strains to Basil Oil
The MIC values for sixty tested E. coli strains were between 8.0 µL/mL to 11.5 µL/mL. The basil
oil showed inhibitory activity against E. coli ATCC 25922 standard strain at 8.0 µL/mL.
2.3.1. The Susceptibility Escherichia coli (ESBL+) Strains to Basil Oil
Most E. coli ESBL+ strains isolated from the abdominal cavity (n = 4) and from the bronchia
(n = 4) were sensitive to basil oil at a concentration range from 8.25 µL/mL to 9.0 µL/mL. For
(ESBL+) clinical strains from wounds (n = 4), MIC values were between 8.5 µL/mL to 9.25 µL/mL.
The growth inhibition concentrations for bacteria isolated from blood were 8.75 µL/mL (n = 1) and
Molecules 2013, 18 9341
9.25 µL/mL (n = 2). Basil oil at a concentration of 8.75 µL/mL inhibited the growth of one of the
tested clinical strains isolated from urine, while 9.0 µL/mL inhibited the growth of three. The MIC
values for strains from catheters were 8.75 µL/mL (n = 2) and 9.25 µL/mL (n = 1). The susceptibility
of Escherichia coli (ESBL+) strains to basil oil is shown in Figure 3.
Figure 3. The susceptibility of Escherichia coli (ESBL+) and (ESBL) strains to basil oil.
2.3.2. The Susceptibility Escherichia coli (ESBL) Strains to Basil Oil
The basil oil was active against Escherichia coli (ESBL) strains at a concentration range from
8.25 µL/mL to 11.5 µL/mL. The most tested strains isolated from the abdominal cavity (n = 4) were
inhibited by concentrations of 11.0–11.5 µL/mL. Clinical strains from the bronchia (n = 5) were
sensitive to basil oil at concentrations of 10.25–11.5 µL/mL. The MIC values for most strains from
wounds (n = 4) were from 10.0 µL/mL to 11.5 µL/mL. Most Escherichia coli (ESBL) strains isolated
from blood and urine were inhibited by basil oil at concentrations ranging from 9.5 µL/mL to
11.5 µL/mL. The MIC values for bacteria from the catheters (n = 6) were 10.0–11.5 µL/mL.
The susceptibility of Escherichia coli (ESBL) strains to basil oil is demonstrated in Figure 3.
2.4. The Susceptibility Escherichia coli Bacterial Strains to Rosemary Oil
The rosemary oil was less active against the sixty tested Escherichia coli clinical strains obtained
from the diverse clinical materials and the hospital equipment. The MIC values were between
18.0 and 20.0 µL/mL. The standard strain E. coli ATCC 25922 was sensitive to rosemary oil at a
concentration of 18.5 µL/mL.
2.4.1. The Susceptibility of Escherichia coli (ESBL+) Strains to Rosemary Oil
Most Escherichia coli (ESBL+) strains isolated from the abdominal cavity (n = 3) were sensitive to
rosemary oil at concentrations from 18.0 µL/mL to 18.5 µL/mL, and (n = 1) at 19.25 µL/mL
concentration. The MIC values for bacterial strains isolated from the bronchia (n = 4) and wounds
(n = 4) were 18.25–19.0 µL/mL and 18.5–19.25 µL/mL, respectively. E. coli (ESBL+) strains isolated
from blood were sensitive to rosemary oil at concentrations between 18.75 and 19.75 µL/mL. All strains
Molecules 2013, 18 9342
from urine (n = 4) were inhibited in the concentration ranges 18.0 µL/mL to 18.75 µL/mL, while those
from catheters (n = 3) were inhibited from 18.25 µL/mL to 18.75 µL/mL. The results of the tests
detailing the susceptibility of Escherichia coli (ESBL+) strains to rosemary oil are presented in Figure 4.
Figure 4. The susceptibility of Escherichia coli (ESBL+) and (ESBL) strains to rosemary oil.
2.4.2. The Susceptibility of Escherichia coli (ESBL) Strains to Rosemary Oil
Rosemary oil was found to have MIC values from 18.5 µL/mL to 19.75 µL/mL for isolates from the
abdominal cavity (n = 6). Growth inhibition was found to occur at concentrations of 19.0–20.0 µL/mL
for clinical bacterial strains obtained from the bronchia (n = 6). Rosemary oil at concentrations ranging
from 18.25 µL/mL to 20.0 µL/mL inhibited the growth of all tested Escherichia coli (ESBL) strains
isolated from wounds. For the blood (n = 7) and urine (n = 6) isolates, the MIC values were in the
range 18.25-19.75 µL/mL. Finally, similar MIC values were found (18.25–19.75 µL/mL) against
E. coli (ESBL) strains isolated from catheters (n = 7). The susceptibility of the Escherichia coli
(ESBL) strains to rosemary oil is shown in Figure 4.
3. Discussion
In our investigation, all clinical strains of Escherichia coli were found to be sensitive to basil
(Ocimum basilicum L.) and rosemary (Rosmarinus officinalis L.) essential oils, irrespective of the
clinical conditions they were obtained under or the pattern of antibiotic resistance they demonstrated,
but basil oil was more active against the tested bacteria. Out of the sixty clinical strains of E. coli,
twenty-two strains were extended-spectrum beta-lactamase positive (ESBL+). Concentrations of basil
oil ranging from 8.25 µL/mL to 9.25 µL/mL were seen to inhibit the growth of eighteen Escherichia coli
(ESBL+) strains. For the extended-spectrum beta-lactamase negative strains, seventeen out of the
thirty-eight were sensitive to basil oil at concentrations between 11.25–11.50 µL/mL. The MIC values
for the other twenty-one Escherichia coli (ESBL) strains ranged from 8.0 µL/mL to 11.0 µL/mL.
The rosemary oil demonstrated significantly lower activity. No apparent differences in activity of
the essential oil were found against extended-spectrum β-lactamase-positive and negative strains. Out
of the twenty-two clinical strains of E. coli (ESBL+), rosemary oil concentrations ranging from
18.0 µL/mL to 19.0 µL/mL were effective for eighteen of them. Eleven of the Escherichia coli (ESBL)
Molecules 2013, 18 9343
strains out of the thirty-eight obtained under various conditions were inhibited by 19.75 µL/mL of
rosemary oil. MIC values ranging from 18.25 µL/mL to 19.5 µL/mL for comparable numbers of
strains was obtained.
The results of our tests clearly demonstrate that basil and rosemary essential oils can be widely used
to eliminate clinical strains of Escherichia coli found in different clinical conditions. It is also
significant that extended-spectrum β-lactamase (ESBL) producing clinical strains of E. coli are
sensitive to these oils. Studies Orhan at al [29], confirm that, essential oils from Foeniculum sp.,
Mentha sp., Ocimum sp., Origanum sp. and Satureja sp. (Lamiaceae family) possess strong antibacterial
activity against extended-spectrum beta-lactamase (ESBL) positive strains of Klebsiella pneumoniae
isolated from food. The obtained MIC values ranged from 32 to 64 μg/ml for a number of strains
resistant to trimetoprime-sulfametoxazole, sulbactam-ampicilin, clavulonate-amoxicilin, ceftriaxon,
cefepime, imipenem, ceftazidime, tobramicine, gentamisine, ofloxacin, and ciprofloxacin. According
our investigations, the Escherichia coli (ESBL+) responsible for human infectious diseases were
significantly more resistant to the basil and rosemary essential oils.
The antimicrobial properties of essential oils are strictly connected with their chemical composition.
The usefulness of essential oils as effective antimicrobial agents can be evaluated only by analysing
their individual components. Therefore, a thorough GC-FID-MS analysis of the tested basil
and rosemary essential oils was conducted. The composition of the essential oil obtained from
Ocimum basilicum L., is as given in the ISO-11043 standard. The composition of the tested basil oil
corresponded to required standards, according to which the content of estragole must be higher than
75.0%. The composition of the essential oil derived from Rosmarinus officinalis L. was found to meet
the requirements of the European Pharmacopoeia 6 [30] and of the Polish Pharmacopoeia VIII [31] for
the nine main components. The content of β-pinene amounted to 9.2% (required 4.0%–9.0%) and
limonene to 1.0% (required 1.5%–4.0%). Verbenone was not found among the components of the
tested rosemary oil, although EP 6 and the Polish Pharmacopoeia VIII specify its content to be a
maximum of 0.4%. For the tested rosemary essential oil, nine of the thirteen main constituents of the
oil met the requirements given in the ISO-1342 standard: α-pinene, camphene, myrcene, 1,8-cineole,
p-cymene, camphor, bornyl acetate, α-terpineol and borneol.
The highest antibacterial activity is demonstrated by phenolic compounds such as carvacrol, thymol
and eugenol. Another effective group of active compounds are alcohols: terpinen-4-ol, γ-terpineol,
geraniol, cytronellol, menthol and linalol. Many of them are synthesized by plants from the Lamiaceae
family [32]. For instance, the essential oil of Satureja hortensis L. demonstrates high levels of activity.
Mihajilov-Kristev et al. [33], showed that essential oil containing mainly carvacrol (67.0%) and
γ-terpinene (15.3%) is effective against Gram-negative strains, including Escherichia coli, with MIC
values from 0.025 µL/mL to 0.78 µL/mL according to the broth microdilution method. In our study for
basil and rosemary essential oils, we obtained significantly higher MIC values. This high activity
demonstrated by Satureja hortensis L. essential oil is certainly related to the high content of carvacrol,
which is one of the most potent antimicrobial compounds.
The literature reports that basil oil, which contains mainly estragole and linalool, also possesses
antibacterial agents which are effective against a variety of Gram-positive and Gram-negative
bacteria [34]. According to Saković et al. [35], Ocimum basilicum essential oil possesses antibacterial
activity against the human pathogenic bacteria Bacillus subtilis, Enterobacter cloacae, Escherichia coli
Molecules 2013, 18 9344
O157:H7, Micrococcus flavus, Proteus mirabilis, Pseudomonas aeruginosa, Salmonella enteritidis,
S. typhimurium, Staphylococcus epidermidis and S. aureus. According to the authors, basil essential
oil containing 69.3% linalool as a main component possesses antibacterial properties against
Escherichia coli O157:H7 with MIC and MBC values of 6.0 µg/mL when assessed by microdilution.
The multidrug clinical strains of E. coli tested in this study were more resistant to basil essential oil
containing mainly estragole (86.4%).
Results obtained by Sartoratto et al. [36], show that basil oil from Ocimum basilicum containing
mainly linallol 32.6% and eugenol 28.1%, and oil from Ocimum gratissimum containing 93.9%
eugenol, have a broad spectrum of antibacterial activity against reference strains of Gram-positive,
Gram-negative bacteria and Candida albicans. According to the authors, essential oils obtained from
these two genera of Ocimum were active against Escherichia coli CCT0547 standard strain with a MIC
value of >2 mg/mL according to the microplate method. These results also confirm that oils with
active constituents such as eugenol tend to have high antibacterial properties. Our results are slightly
higher than those obtained by Sartoratto et al. The MIC values were in the range from 7.92 mg/mL to
11.04 mg/mL for clinical strains of Escherichia coli which were both positive and negative for
extended-spectrum beta-lactamase activity.
Nakamura et al. [37], demonstrated that the essential oil from Ocimum gratissimum, with eugenol
as its main constituent, possesses antibacterial activity against clinical strains of Escherichia coli
ATCC 25922 with MIC—6 mg/mL. The MICs of this essential oil against a group of other
Gram-negative bacteria comprising Shigella flexneri, Salmonella enteritidis, Klebsiella sp. and
Proteus mirabilis were found to be from 0.3 to 12 mg/mL. The MIC values given in our study were
slightly higher than those given by Nakamura et al. Pereira et al. [10], in their investigations, showed
that oil from Ocimum gratissimum possesses antibacterial activity against clinical Escherichia coli
strains isolated from urinary tract infections. The essential oil was found to be active against
about 70% of tested E. coli clinical isolates. The results of the present study demonstrate that a
chemotype containing mainly 1,8-cineole, eugenol, methyleugenol, thymol, p-cimene, cis-ocimene and
cis-caryophyllene has lower activity.
In our tests, basil oil obtained from Ocimum basilicum, containing mainly estragole (86.4%),
inhibited the growth all strains isolated from various clinical materials. Among them were bacteria
isolated from urine, which were also extended-spectrum beta-lactamase positive. Our studies confirm
that antibacterial activity is possessed by not only basil oil chemotypes with linalool or eugenol as their
main components, but also that of Ocimum basilicum, containing mainly estragole. Our research
showed that basil essential oil was significantly more effective against all clinical isolates than
rosemary essential oil. Similar results were obtained by Hammer et al [38], who studied the antimicrobial
activity of basil and rosemary essential oils against Acinetobacter baumanii, Aeromonas veronii
biogroup sobria, Candida albicans, Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae,
Pseudomonas aeruginosa, Salmonella enterica subsp. enterica serotype typhimurium, Serratia marcescens
and Staphylococcus aureus, using an agar dilution method. The authors confirmed that basil (Ocimum
basilicum) oil is more active against Escherichia coli than rosemary (Rosmarinus officinalis). The
MIC values were 0.5 and 1.0% (v/v), respectively.
According to Lopez et al. [39], the oils from Ocimum basilicum and Rosmarinus officinalis have an
antibacterial potential against the Gram-positive bacteria Staphylococcus aureus, Enterococcus faecalis
Molecules 2013, 18 9345
and Listeria monocytogenes and against Gram-negative bacteria Escherichia coli, Yersinia enterocolitica,
Salmonella choleraesuis and Pseudomonas aeruginosa as foodborne bacterial strains. The authors
present a detailed analysis of the tested oils and their ability to inhibit the growth of bacteria. Their
basil and rosemary essential oils were of a similar composition to the essential oils in our
investigations. The main component of the basil essential oil was estragole—82% ± 1.2%, while the
rosemary essential oil contains 1.8-cineole—48% ± 9.1%, camphor—17% ± 4.0, β-pinene—4.8% ± 0.9%
and β-caryophyllene—6.8% ± 3.7%. The authors confirm that the basil oil is more effective at
inhibiting the growth of Escherichia coli strains.
Probuseenivasan et al [27], confirmed that rosemary essential oil strongly inhibits Escherichia coli
ATCC 25922. Although the basil oil was also seen to demonstrate low activity against the tested
bacteria, no data was given about the constituents of the essential oils. The minimal inhibitory
concentration for rosemary oil against E. coli was >6.4 mg/mL. The MIC values obtained by the
present study were higher and ranged from 16.02 mg/mL to 17.35 mg/mL against E. coli clinical
strains. Fabio et al [40], report that rosemary oil has an antibacterial effect on a number of
microorganisms responsible for respiratory infections, isolated from clinical specimens, among which
were antibiotic-sensitive and antibiotic-resistant strains such as Streptococcus pyogenes, S. agalactiae,
S. pneumoniae and Klebsiella pneumoniae, Staphylococcus aureus and Stenotrophomonas maltophilia.
In our tests, rosemary oil was also found to demonstrate antibacterial activity against Escherichia coli
strains with different patterns of resistance, including extended-spectrum β-lactamase positive strains
isolated from various clinical materials. The rosemary essential oil used in the present study obtained
from Rosmarinus officinalis contains mainly 1,8-cineole (46.4%), camphor (11.4%) and α-pinene
(11.0%). The composition of the rosemary essential oil used by Jiang et al. [41], was similar to that
used by us: mainly 1,8-cineole (26.54%) and α-pinene (20.14%). The authors show that the tested oil
possesses antibacterial activity against Gram-positive bacteria (Staphylococcus epidermidis,
Staphylococcus aureus and Bacillus subtilis), Gram-negative bacteria (Proteus vulgaris, Pseudomonas
aeruginosa and Escherichia coli) and fungi (Candida albicans and Aspergillus niger). Bendeddouche
et al [42], showed that of essential oil from Rosmarinus tournefortii De Noé growing wild in the
occidental region of Algeria possesses antimicrobial activity also against Gram-negative (Escherichia
coli and Pseudomonas aeruginosa) and Gram-positive (Staphylococcus aureus) pathogenic bacteria.
The main constituents of the tested essential oil were camphor (37.6%), 1,8-cineole (10.0%),
p-cymene-7-ol (7.8%) and borneol (5.4%).
A number of studies show that essential oils and their constituents possess useful properties
concerning human health. Many of them may be applied in anticancer therapy, cardiovascular and
nervous system disorders to reduce the level of cholesterol, to regulate the glucose level or to stimulate
hormone production [43]. They also might have great value in preventing and treating infectious
diseases. Essential oils not only have bactericidal activity but also can inhibit multidrug bacterial strain
formation. Their multiple antibacterial, antifungal, antiviral and also anti-inflammatory and antioxidant
effects, have made them valuable agents in human treatment and for the prevention of pathological
changes. In addition, essential oils have a number of beneficial properties as natural preservatives in
cosmetics, toiletries, drugs and food products [44–46]. Considering the huge increase in the number of
multidrug resistant bacterial strains in health care facilities, essential oils may prove to be effective
natural antimicrobial agents.
Molecules 2013, 18 9346
4. Experimental
4.1. Bacterial Strains
The standard strain, E. coli ATCC 25922, was obtained from the collection of the Medical and
Sanitary Microbiology Department, Medical University of Lodz. The clinical strains of Escherichia
coli were collected in 2011 and 2012 from a range of clinical materials recovered from patients and
from the hospital equipment in various wards from one of the Medical University hospitals in Lodz:
internal medicine, surgery, urology and the intensive care unit. The tested bacterial strains were
isolated from the abdominal cavity (n = 10), bronchia (n = 10), wounds (n = 10), blood (n = 10), urine
(n = 10) and from catheters (n = 10).
4.2. Bacterial Strain Identification
E. coli strains were cultured on Columbia Agar (bioMerieux, Craponne, France) and on Mac
Conkey Agar (bioMerieux). They were identified to the species by using API 20 E tests (bioMerieux).
The bacteria were incubated at 37 °C for 24 h.
4.3. Essential Oil Analysis
Commercial essential oils from basil—Ocimum basilicum L. and rosemary—Rosmarinus officinalis L.
were purchased from the manufacturer (POLLENA-AROMA, Warsaw, Poland) and analyzed by
GC-FID-MS in the Institute of General Food Chemistry, Lodz University of Technology, using a Trace
GC Ultra apparatus (Thermo Fisher Scientific Inc., Waltham, MA, USA) MS DSQ II detectors and FID-MS
splitter (SGE). Operating conditions: apolar capillary column Rtx-1ms (Restek Corporation, Bellefonte,
PA, USA), 60 m × 0.25 mm i.d., film thickness 0.25 µm; temperature program, 50–300 °C at 4 °C/min;
SSL injector temperature 280 °C; FID temperature 300 °C; split ratio 1:20; carrier gas helium at regular
pressure 200 kPa.; FID temperature 260 °C; carrier gas, helium; 0.5 mL/min; split ratio 1:20. Mass
spectra were acquired over the mass range 30–400 Da, ionization voltage 70 eV; ion source temperature
200 °C. The analysis of the constituents of the oils were performed two times independently.
Identification of components was based on the comparison of their MS spectra with those of the
laboratory-made MS library, commercial libraries (NIST 98.1, Wiley Registry of Mass Spectral Data,
8th Ed. and MassFinder 3.1) and with literature data [47,48] along with the retention indices on the
apolar column (Rtx-1, MassFinder 3.1) associated with a series of alkanes with linear interpolation
(C
8
-C
26
). A quantitative analysis, expressed as percentages of each component, was carried out by
peak area normalization measurements without correction factors.
4.4. Antibacterial Tests
The standard and clinical strains were cultivated in Columbia Agar medium and incubated at 37 °C
for 48 h in aerobic conditions. The microbial suspension was standardized to a cell density of
1–2·× 10
8
cells/mL, equal to an optical density of 0.5 on the Mc Farland scale, by a bioMerieux
densitometer. The agar dilution method was employed for the screening of antimicrobial activities of
the essential oils [32,38,49–51]. The tested essential oils were diluted in 96% ethanol PURE (POCH,
Molecules 2013, 18 9347
Gliwice, Poland) yielding a concentration of 97% v/v of oils. Although the tested essential oils dissolve
well in ethanol, only minimum amounts were used, as it can inhibit the growth of the tested bacteria.
This solution was mixed with a culture medium to obtain concentrations from 7.25 µL/mL to
11.75 µL/mL for basil oil and 17.75 µL/mL to 20.25 µL/mL for rosemary oil and poured into sterile
Petri dishes. An inoculum containing 1–2 × 10
8
cells/mL (0.1 mL) per spot was seeded upon the
surface of the agar with various oil concentrations, as well as on agar with no oil added (acting as a
control for strain growth). The Minimal Inhibitory Concentration, MIC, was determined after 24 h of
incubation at 37 °C under aerobic conditions. The MIC was considered the lowest concentration of the
sample at which no visible growth was observed. The analysis of the antibacterial activity of the oil
was performed three times independently. Control media containing only alcohol at concentrations
used in the dilutions of tested essential oils did not inhibit the growth of bacterial strains.
4.5. Susceptibility Testing
The following antibiotics (Becton Dickinson) were used for susceptibility testing of Escherichia coli
strains (R—resistance; I—intermediate susceptibility; S—susceptibility): AM—ampicillin (10 µg)
(R 13, 14 I 16, S 17), AMC—amoxicillin/clavulanic acid (20 µg/10 µg) (R 13, 14 I 17,
S 18), CF—cefalotin (30 µg) (R 14, 15 I 17, S 18), CZ—cefazoline (30 µg) (R 14, 15 I 17,
S 18), CXM—cefuroxime (30µg) (R 14, 15 I 17, S 18), GM—gentamicin (10 µg) (R 12,
13 I 14, S 15), TE—tetracycline (30 µg) (R 14, 15 I 18, S 19), NOR—norfloxacin
(10 µg) (R 12, 13 I 16, S 17) (only for the isolates from urine), FTN—nitrofurantoin (300 µg)
(R 14, 15 I 16, S 17) (as above), FOS—fosfomycin (200 µg) (R 12, 13 I 15, S 16)
(as above), STX—trimethoprim/sulfamethoxazole (1.25 µg/23.75 µg) (R 10, 11 I 15, S 16),
PIP—piperacillin (100 µg) (R 17, 18 I 20, S 21), TIC—tikarcillin (75 µg) (R 14, 15 I 19,
S 20), TZP—piperacyllin/tazobaktam (100/10 µg) (R 17, 18 I 20, S 21), TIM—
ticarcillin/clavulanic acid (75 µg/10 µg) (R < 16, S > 16), FOX—cefoxitin (30 µg) (R 14, 15 I 17,
S 18), CTX—cefotaxim (30 µg) (R 14, 15 I 22, S 23), CAZ—ceftazidime (30 µg) (R 14,
15 I 17, S 18), FEP—cefepim (30 µg) (R 14, 15 I 17, S 18), ATM—aztreonam (30 µg)
(R 15, 16 I 21, S 22), IMP—imipenem (10 µg) (R 13, 14 I 15, S 16), MEM—meropenem
(10 µg) (R 13, 14 I 15, S 16), ETP—ertapenem 10 µg) (R 15, 16 I 18, S 19), DOR—
doripenem (10 µg) (R 19, 20 I 23, S 24), CIP—ciprofloxacin (5 µg) (R 15, 16 I 20, S 21),
AN—amikacin (30 µg) (R 14, 15 I 16, S 17), NET—netilmicin (30 µg) (R 12, 13 I 14,
S 15), TOB—tobramycin (10 µg) (R 12, 13 I 14, S 15), C—chloramphenicol (30 µg)
(R 12, 13 I 17, S 18), TGC—tigecyclin (15 µg) (R 14, 15 I 18, S 19).
Susceptibility testing was carried out using the disc-diffusion method, on Mueller-Hinton II Agar
(bioMerieux). Cultures were incubated at 37 °C for 16–18 h. The results were interpreted according to
EUCAST guidelines [52].
The double-disk synergy test and combination disk method were used to determine ESBL
production. The sensitivity of the DDST can be improved by reducing the distance between the disks
of cephalosporins and clavulanate. The disk approximation method was performed on a Muller-Hinton
agar plate inoculated with the clinical bacterial strain, by placing disks containing CAZ—ceftazidime
(30 µg), CTX—cefotaxime (30 µg) and ATM—aztreonam (30 µg) 20 mm (edge to edge) from a disk
Molecules 2013, 18 9348
of AMC—amoxicillin/clavulanic acid (20/10 µg). Following incubation for 16–18 h at 37 °C, any
enhancement of the zone of inhibition between a cephalosporin and monobactam-aztreonam disk from
the amoxicillin/clavulanic acid disk, was indicative of the presence of an ESBL. E. coli ATCC 25922
was used as a positive control [11,53].
5. Conclusions
(1) The results of these experiments indicate the potential use of basil and rosemary essential
oils against resistant Escherichia coli clinical strains, and also against extended-spectrum
β-lactamase positive bacteria.
(2) The tested basil oil was more active against all Escherichia coli clinical strains.
(3) The action of essential oils against bacteria exhibiting different mechanisms of resistance may
be useful, not only in treating but also preventing the spread of resistant strains.
Acknowledgements
The authors wish to thank Danuta Kalemba from the Institute of General Food Chemistry,
Lodz University of Technology for essential oil analysis. The research reported in this manuscript was
supported by grant no 503/5-020-03/503-01 and has not been submitted elsewhere.
Conflict of Interest
The authors declare no conflict of interest.
References
1. Canton, R.; Novais, A.; Valverde, A.; Machado, E.; Peixe, L.; Baquero, F.; Coque T.M.
Prevalence and spread of extended-spectrum β-lactamase-producing Enterobacteriaceae in Europe.
Clin. Microbiol. Infect. 2008, 14, 144–153.
2. Cornaglia, G.; Akova, M.; Amicosante G.; Cantón R.; Cauda R.; Docquier J.D.; Edelstein, M.;
Frère, J.M.; Fuzi, M.; Galleni, M.; et al. ESCMID Study Group for Antimicrobial Resistance
Surveillance (ESGARS). Metallo-β-lactamases as emerging resistance determinants in Gram-negative
pathogens. Int. J. Antimicrob. Agents 2007, 29, 380–388.
3. Cornaglia, G.; Garau, J.; Livermore, D.M. Living with ESBLs. Clin. Microbiol. Infect. 2008, 14, 1–2.
4. Empel, J.; Baraniak, A.; Literacka, E.; Mrówka, A.; Fiett, J.; Sadowy, E., Hryniewicz, W.;
Gniadkowski, M. Molecular survey of β-lactamases conferring resistance to newer β-lactams in
Enterobacteriaceae isolates from Polish hospitals. Antimicrob. Agents Chemother. 2008, 52,
2449–2454.
5. Pitout, J.D.D.; Nordmann, P.; Laupland, K.B.; Nordmann, P. Emergence of Enterobcteriaceae
producing extended-spectrum β-lactamases (ESBLs) in the community. J. Antimicrob. Chemother.
2005, 56, 52–59.
6. Nicolas-Chanoine, M.H.; Blanco, J.; Leflon-Guibout, V.; Demarty, R.; Alonso, M.P.; Caniça, M.M.;
Park, Y.J.; Lavigne, J.P.; Pitout, J.; Johnson, J.R. Intercontinental emergence of Escherichia coli
clone O25:H4-ST131 producing CTX-M-15. J. Antimicrob. Chemother. 2008, 61, 273–281.
Molecules 2013, 18 9349
7. Woodford, N.; Reddy, S.; Fagan, E.J.; Hill, R. L.; Hopkins, K.L.; Kaufmann M.E.; Kistler, J.;
Palepou, M.F.; Pike, R.; Ward, M.E.; et al. Wide geographic spread of diverse acquired AmpC
beta-lactamases among Escherichia coli and Klebsiella spp. in the UK and Ireland. J. Antimicrob.
Chemother. 2007, 59, 102–105.
8. Kang, H.Y.; Jeong, Y.S.; Oh, J.Y.; Jeong, J.H., Seol, S.Y.; Cho, D.T.; Kim, J.; Lee, Y.C.
Characterization of antimicrobial resistance and class 1 integrons found in Escherichia coli
isolates from humans and animals in Korea. J. Antimicrob. Chemother. 2005, 55, 639–644.
9. Tenover, C.; Raney, P.M.; Williams, P.P.; Rasheed, J.K.; Biddle, J.W.; Oliver A.; Fridkin, S.K.;
Jevitt, L.; McGowan, J.E., Jr. Evaluation of the NCCLS extended-spectrum β-lactamase confirmation
methods for Escherichia coli with isolates collected during Project ICARE. J. Clin Microbiol.
2003, 41, 3142–3146.
10. Pereira, R.S.; Sumita, T.C.; Furlan, M.R.; Jorge, A.O.; Ueno, M. Antibacterial activity of essential
oils on microorganisms isolated from urinary tract infection. Rev. Saude Publica 2004, 38, 326–328.
11. Drieux, L.; Brossier, F.; Sougakoff, W.; Jarlier, V. Phenotypic detection of extended-spectrum
beta-lactamase production in Enterobacteriaceae: Review and bench guide. Clin. Microbiol. Infect.
2008, 14, 90–103.
12. Canton, R.; Coque, T.M. The CTX-M beta-lactamase pandemic. Curr. Opin. Microbiol. 2006, 9,
466–475.
13. Silva, N.C.C.; Fernandes, A., Jr. Biological properties of medicinal plants: A review of their
antimicrobial activity. J. Venom. Anim. Toxins Trop. Dis. 2010, 16, 402–413.
14. Rios, J.L.; Recio, M.C. Medicinal plants and antimicrobial activity. J. Ethnopharm. 2005, 100, 80–84.
15. Sparg, S.G.; Light, M.E.; van Staden, J. Biological activities and distribution of plant saponins.
J. Ethnopharm. 2004, 94, 219–243.
16. Tajkarimi, M.M.; Ibrahim, S.A.; Cliver, D.O. Antimicrobial herb and spice compounds in food.
Food Control 2010, 21, 1199–218.
17. Burt, S. Essential oils: Their antibacterial properties and potential applications in foods—A review.
Int. J. Food Microb. 2004, 94, 223–253.
18. De Rapper, S.; Kamatou, G.; Viljoen, A.; van Vuuren, S. The in vitro antimicrobial activity of
Lavandula angustifolia essential oil in combination with other aroma-therapeutic oils. Evid. Based
Complement. Alternat. Med. 2013, 2013, 852049.
19. Fernandes, T.G.; Carneiro de Mesquita, A.R.; Randau, K.P.; Franchitti, A.A.; Ximenes, E.A.
In Vitro Synergistic Effect of Psidium guineense (Swartz) in Combination with Antimicrobial
Agents against Methicillin-Resistant Staphylococcus aureus Strains. Scient. World J. 2012, 2012,
158237.
20. Fankam, A.G.; Kuete, V.; Voukeng, I.K.; Kuiate, J.R.; Pages, J.-M. Antibacterial activities of
selected Cameroonian spices and their synergistic effects with antibiotics against multidrug-resistant
phenotypes. BMC Complement. Altern. Med. 2011, 11, 104.
21. Nweze, E.I.; Eze, E.E. Justification for the use of Ocimum gratissimum L. in herbal medicine and
its interaction with disc antibiotics. BMC Complement. Altern. Med. 2009, 9, 37.
22. Sung, W.S.; Lee, D.G. The combination effect of Korean red ginseng saponins with kanamycin
and cefotaxime against methicillinresistant Staphylococcus aureus. Biol. Pharm. Bull. 2008, 31,
1614–1617.
Molecules 2013, 18 9350
23. Zhao, J.; Lou, J.; Mou, Y.; Li, P.; Wu, J.; Zhou, L. Diterpenoid tanshinones and phenolic acids
from cultured hairy roots of Salvia miltiorrhiza Bunge and their antimicrobial activities.
Molecules 2011, 16, 2259–2267.
24. Lee, S.J.; Umano, K.; Shibamota, T.; Lee, K.G. Identification of volatile components in basil
(Ocimum basilicum L.) and thyme leaves (Thymus vulgaris L.) and their antioxidant properties.
Food Chem. 2005, 91, 131–137.
25. Wannissorn, B.; Jarikasem, S.; Siriwangchai, T.; Thubthimthed, S. Antibacterial properties of
essential oils from Thai medicinal plants. Fitoterapia 2005, 76, 233–236.
26. Opalchenova, G.; Obreshkova, D. Comparative studies on the activity of basilan essential oil from
Ocimum basilicum L. against multidrug resistant clinical isolates of the genera Staphylococcus,
Enterococcus and Pseudomonas by using different test methods. J. Microbiol. Methods 2003, 54,
105–110.
27. Prabuseenivasan, S.; Jayakumar, M.; Ignacimuthu, S. In vitro antibacterial activity of some plant
essential oils. BMC Complement. Altern. Med. 2006, 6, 39.
28. Bozin, B.; Mimica-Dukic, N.; Samojlik, J.; Jovin, E. Antimicrobial and antioxidant properties of
rosemary and sage Rosmarinus officinalis L. and Salvia officinalis L., Lamiaceae essential oils.
J. Agric. Food Chem. 2007, 55, 7879–7885.
29. Orhan, I.E.; Ozcelik, B.; Kan, Y.; Kartal, M. Inhibitory effects of various essential oils
and individual components against extended-spectrum beta-lactamase (ESBL) produced by
Klebsiella pneumoniae and their chemical compositions. J. Food Sci. 2011, 76, 538–546.
30. European Pharmacopoeia, 6th ed.; Council of Europe: Strasbourg, France, 2008.
31. Polish Pharmacopeia VIII, 8th ed.; Polish Pharmaceutical Society: Warsaw, Poland, 2008.
32. Kalemba, D.; Kunicka, A. Antibacterial and antifungal properties of essential oils. Curr. Med. Chem.
2003, 10, 813–829.
33. Mihajilov-Kristev, T.; Radnovic, D.; Kitic, D.; Stajnovic-Radic, Z.; Zlatkovic, B. Antimicrobial
activity of Satureja hortensis L. essential oil against pathogenic microbial strains. Bioterchnol.
Biotechnol. Equip. 2009, 23, 1492–1496.
34. Wan, J.; Wilcock, A.; Coventry, M.J. The effect of essential oil basil on the growth of
Aeromonas hydrophila and Pseudomonas fluorescens. J. Appl. Microbiol. 1998, 84, 152–158.
35. Soković, M.; Glamočlija, J.; Marin, P.D.; Brkić, D.; van Griensven, L.J.L.D. Antibacterial effects
of the essential oils of commonly consumed medicinal herbs using an in vitro model. Molecules
2010, 15, 7532–7546.
36. Sartoratto, A.; Machado, A.L.M.; Delarmelina, C.; Figueira, G.M.; Duarte, M.C.T.;
Rehder, V.L.G. Composition and antimicrobial activity of essential oils from aromatic plants used
in Brazil. Braz. J. Microbiol. 2004, 35, 275–280.
37. Nakamura, C.V.; Ueda-Nakamura, T.; Bando, E.; Melo, A.F.; Cortez, D.A.; Dias Filho, B.P. Antibacterial
activity of Ocimum gratissimum L. essential oil. Mem. Inst. Oswaldo Cruz 1999, 94, 675–678.
38. Hammer, K.A.; Carson, C.F.; Riley, T.V. Antimicrobial activity of essential oils and other plant
extracts. J. Appl. Microbiol. 1999, 86, 985–990.
39. Lopez, P.; Sanchez, C.; Battle, R.; Nerin, C. Soilid- and vapour-phase antimicrobial activities of
six essential oils: Susceptibility of selected foodborne bacterial and fungal strains. Agric. Food Chem.
2005, 53, 6939–6946.
Molecules 2013, 18 9351
40. Fabio, A.; Cermelli, C.; Fabio, G.; Nicoletti, P.; Quaglio, P. Screening of the antibacterial
effects of a variety of essential oils on microorganisms responsible for respiratory infections.
Phytother. Res. 2007, 21, 374–377.
41. Jiang, Y.; Wu, N.; Fu, Y-J.; Wang, W.; Luo, M.; Zhao, C.J.; Zu, Y.G.; Liu, X.L. Chemical
composition and antimicrobial activity of the essential oil of Rosemary. Environ. Toxicol. Pharmacol.
2011, 32, 63–68.
42. Bendeddouche, M.S.; Benhassaini, H.; Hazem, Z.; Romane, A. Essential oil analysis and antibacterial
activity of Rosmarinus tournefortii from Algeria. Nat. Prod. Commun. 2011, 6, 1511–1514.
43. Edris, A.E. Pharmaceutical and therapeutic potentials of essential oils and their individual volatile
constituents: A review. Phytother. Res. 2007, 21, 308–323.
44. Bakkali, F.; Averbeck, S.; Averbeck, D.; Idaomar, M. Biological effect of essential oils-A review.
Food Chem. Toxicol. 2008, 46, 446–475.
45. Reichling, J.; Schnitzler, P.; Suschke, U.; Saller R. Essential oils of aromatic plants with
antibacterial, antifungal, antiviral and cytotoxic properties—An overview. Forsch Komplementmed
2009, 16, 79–90.
46. Sienkiewicz, M.; Kowalczyk, E.; Wasiela, M. Recent patents regarding essential oils and the
significance of their constituents in human health and treatment. Recent Pat. Anti Infect. Drug
Discov. 2012, 7, 133–140.
47. Adams, R.P. Identification of Essential Oil Components by Gas Chromatography/Mass
Spectroscopy, 4th ed.; Allured Publishing Corporation: Carol Stream, IL, USA, 2007.
48. Joulain, D.; Konig, W.A. The Atlas of Spectral Data of Sesquiterpene Hydrocarbons;
E.B.-Verlag: Hamburg, Germany, 1998.
49. Maino, M.; Bersani, C.; Comi, G. Impedance measurements to study the antimicrobial activity of
essential oils from Lamiaceae and Compositae. Int. J. Food Microbiol. 2001, 67, 187–195.
50. Sivropoulou, A.; Nikolaou, C.; Papanikolaou, E.; Kokkini, S.; Lanaras T.; Arsenakis, M.
Antimicrobial, cytotoxic and antiviral activities of Salvia fructicosa essential oil. J. Agric.
Food Chem. 1997, 45, 3197–3201.
51. Vander Berghe, D.A.; Vietinck A.J. Screening Methods for Antibacterial and Antiviral Agents
from High er Plants. In Methods In Plant Biochemistry; Dey, P.M., Harborne J.B., Eds.;
Academic Press: London, UK, 1991.
52. European Committee on Antimicrobial Susceptibility Testing (EUCAST). Breakpoint Tables for
Interpretation of MICs and Zone Diameters, Version 2.0; valid from 1 January 2012. Available
online: http://www.eucast.org/ (accessed on 16 August 2012).
53. Jarlier, V.; Nicolas, M.; Fournier, G.; Philippon, A. Extended broad-spectrum β-lactamases
conferring transferable resistance to newer β-lactam agents in Enterobacteriaceae: Hospital
prevalence and susceptibility patterns. Rev. Infect. Dis. 1988, 10, 867–878.
Sample Availability: Samples of the basil and rosemary essential oils (POLLENA-AROMA Poland)
are available from the authors.
© 2013 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
(http://creativecommons.org/licenses/by/3.0/).
... Hussain and coauthors [41] found that 1,8-cineol (38.5%), camphor (17.1%), αpinene (12.3%), limonene (6.23%), camphene (6.00%), and linalool (5.70%) were the major constituents of R. officinalis. Dıraz Yıldırım [42], Sienkiewicz and coauthors [43] and Jiang and coauthors [44] stated that the major compounds of R. officinalis were found as α-pinene, 1.8-cineole, α-terpineol, α-pinene, camphor, isoborneol. ...
... It was reported that essential oils of R. officinalis had weak or moderate antibacterial activity [55]. Sienkiewicz and coauthors [43], Burt [56] Sirocchi and coauthors [57] and Yildirim and coauthors [58] demonstrated the antibacterial activities of R. officinalis in their studies. Rosmarinic acid, rosmaridiphenol, carnosol, epirosmanol, carnosic acid, rosmanol and isorosmanol in rosemary are inhibitory in cells due to their interaction with the cell membrane, which causes loss of membrane functionality and structure, changes in genetic material and nutrients, changes in electron transport, leakage of cellular components, and changes in fatty acid production [59]. ...
Article
Full-text available
Medicinal and aromatic plants have been widely using in folk medicine as antimicrobial, anti-inflammatory and antinociceptive agents. The aim of this study was to determine essential oil composition and antimicrobial activity of T. spicata, L. X Intermedia, S. macrantha and R. officinalis. Essential oil components of these plants were obtained by water vapor distillation method using Neo-Clevenger apparatus. Essential oil components were determined by gas chromatography-mass spectroscopy (GC-MS). The main components of these plants are carvacrol (74.26 %) and γ-terpinene (10.28%) in T. spicata, 1,8-cineol (32.48%), linalool (24.38%) and camphor (14.73%) in L. X Intermedia, p-cymene (56.70%), carvacrol (10.96 %) in S. macrantha and camphor (18.26 %), α-pinene (15.51%), 1,8-cineole (11.86%) and borneol (10.39%) in R. officinalis were determined. T. spicata and S. macrantha showed strong effects against three microorganisms. L. X Intermedia and R. officinalis showed strong activity against Candida albicans, while they had moderate effects against Staphylococcus aureus, Escherichia coli.
... Regarding the antibacterial properties of R. officinalis extract, other studies based on the minimal inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) also came to the same conclusion that R. officinalis extract has a bactericidal effect. For example, Sienkiewicz et al. [41] showed inhibition of E. coli growth with an R. officinalis essential oil. Various studies also confirmed the efficacy of R. officinalis against S. aureus [26,42], Bacillus cereus [43], Clostridium perfringens [44], and various Salmonella species [45,46]. ...
Article
Full-text available
Objective In the last few decades, there has been a growing worldwide interest in the use of plant extracts for the prevention of oral diseases. The main focus of this interest lies in the identification and isolation of substances that limit the formation of microbial biofilm which plays a major role in the development of caries, periodontitis, and peri-implantitis. In this clinical ex vivo study, we investigated the antimicrobial effects of Rosmarinus officinalis extract against oral microorganisms within in situ initial oral biofilms. Materials and methods Initial in situ biofilm samples (2 h) from six healthy volunteers were treated ex vivo with R. officinalis extract at concentrations of 20 mg/ml and 30 mg/ml. The number of viable bacterial cells was determined by counting the colony-forming units. All surviving bacteria were isolated in pure cultures and identified using MALDI-TOF and biochemical testing procedures. Additionally, live/dead staining in combination with epifluorescence microscopy was used for visualizing the antimicrobial effects in the initial biofilms. Results The number of colony-forming units in the R. officinalis–treated biofilms was significantly lower than in the untreated controls (p < 0.001). The reduction range of log10 was 1.64–2.78 and 2.41–3.23 for aerobic and anaerobic bacteria, respectively. Regarding the bacterial composition, large intra- and interindividual variability were observed. Except for Campylobacter spp., the average amount of all bacterial taxa was lower after treatment with R. officinalis than in the untreated biofilms. A total of 49 different species were detected in the untreated biofilms, while only 11 bacterial species were detected in the R. officinalis–treated biofilms. Live/dead staining confirmed that the R. officinalis–treated biofilms had significantly lower numbers of surviving bacteria than the untreated biofilms. Conclusions The treatment with R. officinalis extract has a significant potential to eliminate microbial oral initial biofilms. Clinical relevance The results of this study encourage the use of R. officinalis extracts in biofilm control and thus in the treatment of caries and periodontitis as a herbal adjuvant to synthetic substances.
... Structure and molecular biological properties of selected compounds in R. officinalis has been the determinant of the biological activity as depicted in Fig. 2a-f. Similarly, Sienkiewicz et al. 26 reported that ROEO contains 1,8-cineole (46.4%), camphor (11.4%) and α-pinene (11.0%) and precursors of other significant bioactive molecules used for therapeutic, cosmetics, and food industries. ROEO used by Jiang et al 18 had 1,8-cineole (26.54%) and α-pinene (20.14%) present significant antimicrobial activity. ...
Article
Full-text available
Plant Based Natural Products (PBNPs) have contributed to the development of drugs for diverse indications. Worldwide interest in use of PBNPs has been growing, and its beneficial effects being rediscovered for the development of drug leads. Literature survey on indigenous traditional knowledge bestows ethnopharmacological potentials of PBNPs, has inspired research in drug design and discovery; PBNPs provide a baseline for the development of novel drug leads against various pharmacological targets. Reports indicate that rosemary essential oil (ROEO) extracts show biological bioactivities such as hepatoprotective, antifungal, insecticide, antioxidant and antibacterial. However, their application is limited because of their odor, color and taste. Phytochemical screening indicates the presence of phenol, flavonoids, tannins, alkaloids, carbohydrates, proteins, glycosides, saponins, coumarins, terpenoids, quinones, steroids. Owing to widespread applications of phyto-compounds in ROEO - GCMS was performed. GCMS analysis detected the presence of 22 compounds (α-Pinene, Camphene, β-Myrcene, α-Terpinine, p-Cymene, trans-3-Caren-2-ol, 1,8-Cineole, γ-Terpinene, α-Terpinolene, Linalool, Isopulegol, Eucalyptol, Terpinen-4-ol, 2-Naphthalenol, (-)-Myrtenol, Verbenone, Terpine, α-Copaene, β-Caryophyllene, γ-Cadinene, Caryophyllene oxide) of which 6 compounds (α-Pinene, p-Cymene, Isopulegol, Eucalyptol, 2-Naphthalenol, Terpine) were in abundant. These compounds have been prospected for their molecular and biological properties in the present study. Keywords: Rosmarinus officinalis; Rosemary Essential Oils (ROEO); GCMS; Bioprospecting; PBNPs
... [37,38,41,42] Vats (2017) has explained the various methods and strategies where phytochemicals can be extracted from forest biowastes. [4] In this study, the author has also mentioned the [43][44][45][46] This study is also the continuation of the same. [16] Vats et al., [9,10] This study is also the continuation of the previous study. ...
Article
Aim: This study aims to extract the phytochemicals from thyme, rosemary, and oregano and evaluate their shelf life and antimicrobial properties. Introduction: The world has witnessed unprecedented time with another virus-based pandemic, severe acute respiratory syndrome coronavirus 2 or novel coronavirus disease 2019. A good immune system is the ultimate shield toward pathogenic diseases. Use of natural and herbal products and extracts to boost immune system has gained momentum. Phytochemicals present in the herbs, spices, and plants have positive impact on fight against pathogens. Materials and Methods: This study has taken singular and synergistic effect of the Thymus vulgaris, Rosmarinus officinalis L., and Origanum vulgare for health-care application. Extraction from the herbs was done with the two methods low temperature for long time and high temperature for short time. Results and Discussion: In the present study, qualitative study of the phytochemicals present in the each sample was determined, antimicrobial activities and shelf life study were carried out. Conclusion: The present study open door for the application of phytochemical extracts against new emerging pandemics related to microbes present in air, water and soil. All the three herbs have been traditionally used in the medicinal systems. A detailed study can reveal the individual and synergistic applications of phytochemicals.
... Oregano oil, rosemary extract, methylparaben and green tea extract have been identified as the most promising antimicrobial substances according to the authors of [27][28][29][30]. The MIC (Minimal Inhibitory Concentration) tests were carried out, as the recommended method by EUCAST (European Committee on Antimicrobial Susceptibility Testing). ...
Article
Full-text available
Bioactive polypropylene (PP) films with active agents) presence for food packaging application have been prepared and characterized. The novel modified PP films were obtained via PP/additives systems regranulation and cast extrusion. The influence of two types of plasticizers (natural agents as well as commercial synthetic product) and bioactive additives on final features, e.g., mechanical properties, was evaluated. Moreover, the biocidal activity of the films was determined. Due to their functional properties, they are developed as additives to packaging plastic materials such as polyolefins. The study results presented in our work may indirectly contribute to environmental protection by reducing food waste. The aim of the work was to obtain innovative, functional packaging materials with an ability to prolong the shelf life of food products. The best antimicrobial properties were observed for the sample based on 5 wt.% oregano oil (OO) and 5 wt.% cedar oil (OC) in PP matrix. A microbial test revealed that the system causes total reduction in the following microorganisms: B. subtilis, E. coli, S. aureus, P. putida, C. albicans, A. alternata, F. oxysporum.
... Additionally, the compounds identified within EOs and their retention time have also been added to the Supplementary Materials (Table S1). The identified compounds are consistent with previously published studies investigating the compositional concentration of thyme, rosemary, and basil EOs [46][47][48][49]. It can be seen that the major compound identified within the thyme EO-functionalized samples is thymol and p-cymen-7-ol. ...
Article
Full-text available
Recent years have witnessed a tremendous interest in the use of essential oils in biomedical applications due to their intrinsic antimicrobial, antioxidant, and anticancer properties. However, their low aqueous solubility and high volatility compromise their maximum potential, thus requiring the development of efficient supports for their delivery. Hence, this manuscript focuses on developing nanostructured systems based on Fe3O4@SiO2 core–shell nanoparticles and three different types of essential oils, i.e., thyme, rosemary, and basil, to overcome these limitations. Specifically, this work represents a comparative study between co-precipitation and microwave-assisted hydrothermal methods for the synthesis of Fe3O4@SiO2 core–shell nanoparticles. All magnetic samples were characterized by X-ray diffraction (XRD), gas chromatography-mass spectrometry (GC-MS), Fourier-transform infrared spectroscopy (FTIR), dynamic light scattering (DLS), zeta potential, scanning electron microscopy (SEM), transmission electron microscopy (TEM), thermogravimetry and differential scanning calorimetry (TG-DSC), and vibrating sample magnetometry (VSM) to study the impact of the synthesis method on the nanoparticle formation and properties, in terms of crystallinity, purity, size, morphology, stability, and magnetization. Moreover, the antimicrobial properties of the synthesized nanocomposites were assessed through in vitro tests on Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, and Candida albicans. In this manner, this study demonstrated the efficiency of the core–shell nanostructured systems as potential applications in antimicrobial therapies.
... At a high concentration (20 µl/ml), the essential oil exhibited a marked inhibition activity against bacteria, table 3. Many literature studies have provided support for the antimicrobial activities of several of the compounds in the Ocimum basilicum essential oil [16,[27][28][29][30]. For instance, essential oils from Ocimum basilicum and Thymus algeriensis cultivated in the Algerian Saharan Atlas was reported that have a good effect against several pathogenic gram-positive and gram-negative bacteria [16]. ...
Article
Full-text available
Determination of Chemical Composition, Antioxidant activity, and Antimicrobial activity of essential oils of Damask Ocimum Basilicum L.
Book
The Chemistry inside Spices & Herbs: Research and Development brings comprehensive information about the chemistry of spices and herbs with a focus on recent research in this field. The book is an extensive 2-part collection of 20 chapters contributed by experts in phytochemistry with the aim to give the reader deep knowledge about phytochemical constituents in herbal plants and their benefits. The contents include reviews on the biochemistry and biotechnology of spices and herbs, herbal medicines, biologically active compounds and their role in therapeutics among other topics. Chapters which highlight natural drugs and their role in different diseases and special plants of clinical significance are also included. Part II continues from the previous part with chapters on the treatment of skin diseases and oral problems. This part focuses on clinically important herbs such as turmeric, fenugreek, ashwagandha (Indian winter cherry), basil, Terminalia chebula (black myrobalan). In terms of phytochemicals, this part presents chapters that cover resveratrol, piperine and circumin. Audience: This book is an ideal resource for scholars (in life sciences, phytomedicine and natural product chemistry) and general readers who want to understand the importance of herbs, spices and traditional medicine in pharmaceutical and clinical research.
Article
Full-text available
Plant Based Natural Products (PBNPs) have contributed to the development of many drugs for diverse indications. Worldwide interest in use of plants based natural products (PBNPs) has been growing, and its beneficial effects being rediscovered for the development of new drugs. Literature survey on indigenous traditional knowledge bestows ethnopharmacological potentials of PBNPs, which has inspired research in drug discovery; further it provides a baseline for the development of novel drug leads against selected pharmacological targets. Studies report that rosemary essential oil (ROEO) extracts have hepatoprotective, antifungal, insecticide, antioxidant and antibacterial properties. However, their application is limited because of their odor, color and taste. Owing to the widespread applications of phyto-compounds in ROEO - GCMS was performed. GCMS analysis detected 22 compounds of which 6 compounds were in abundant. In the present study, isopulegol - a Prenol Lipid (Monoterpenoid) from Rosmarinus officinalis has been ADMET characterized from biomedical application point of view. Keywords: Rosmarinus officinalis; Rosemary officinalis Essential Oils (ROEO); Pharmacological Activity; ADMET; GCMS;
Article
Full-text available
Adams, R. P. 2007. Identification of essential oil components by gas chromatography/ mass spectrometry, 4th Edition. Allured Publ., Carol Stream, IL Is out of print, but you can obtain a free pdf of it at www.juniperus.org
Article
Full-text available
Background: To evaluate the antibacterial activity of 21 plant essential oils against six bacterial species. Methods: The selected essential oils were screened against four gram-negative bacteria (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus vulgaris) and two gram-positive bacteria Bacillus subtilis and Staphylococcus aureus at four different concentrations (1:1, 1:5, 1:10 and 1:20) using disc diffusion method. The MIC of the active essential oils were tested using two fold agar dilution method at concentrations ranging from 0.2 to 25.6 mg/ml. Results: Out of 21 essential oils tested, 19 oils showed antibacterial activity against one or more strains. Cinnamon, clove, geranium, lemon, lime, orange and rosemary oils exhibited significant inhibitory effect. Cinnamon oil showed promising inhibitory activity even at low concentration, whereas aniseed, eucalyptus and camphor oils were least active against the tested bacteria. In general, B. subtilis was the most susceptible. On the other hand, K. pneumoniae exhibited low degree of sensitivity. Conclusion: Majority of the oils showed antibacterial activity against the tested strains. However Cinnamon, clove and lime oils were found to be inhibiting both gram-positive and gram-negative bacteria. Cinnamon oil can be a good source of antibacterial agents.
Article
Full-text available
A hydro-distilled oil of Satureja hortensis L. was investigated for its antimicrobial activity against a panel of 11 bacterial and three fungal strains. The antimicrobial activity was determined using the disk-diffusion and broth microdilution methods. The essential oil of S. hortensis L. showed significant activity against a wide spectrum of Gram (-) bacteria (MIC/MBC=0.025-0.78/0.05-0.78 ?l/ml) and Gram (+) bacteria (MIC/MBC=0.05-0.39/0.05-0.78 ?l/ml), as well as against fungal strains (MIC/MBC=0.20/0.78 ?l/ml). The results indicate that this oil can be used in food conservation, treatment of different diseases of humans, and also for the treatment of plants infected by phytopathogens. Detected autoplagiarism. Link to the Editorial Decision 10.2298/ABS1004251U
Article
Full-text available
Plants have been used for thousands of years to flavor and conserve food, to treat health disorders and to prevent diseases including epidemics. The knowledge of their healing properties has been transmitted over the centuries within and among human communities. Active compounds produced during secondary vegetal metabolism are usually responsible for the biological properties of some plant species used throughout the globe for various purposes, including treatment of infectious diseases. Currently, data on the antimicrobial activity of numerous plants, so far considered empirical, have been scientifically confirmed, concomitantly with the increasing number of reports on pathogenic microorganisms resistant to antimicrobials. Products derived from plants may potentially control microbial growth in diverse situations and in the specific case of disease treatment, numerous studies have aimed to describe the chemical composition of these plant antimicrobials and the mechanisms involved in microbial growth inhibition, either separately or associated with conventional antimicrobials. Thus, in the present work, medicinal plants with emphasis on their antimicrobial properties are reviewed.
Article
Full-text available
Plants have been used for thousands of years to flavor and conserve food, to treat health disorders and to prevent diseases including epidemics. The knowledge of their healing properties has been transmitted over the centuries within and among human communities. Active compounds produced during secondary vegetal metabolism are usually responsible for the biological properties of some plant species used throughout the globe for various purposes, including treatment of infectious diseases. Currently, data on the antimicrobial activity of numerous plants, so far considered empirical, have been scientifically confirmed, concomitantly with the increasing number of reports on pathogenic microorganisms resistant to antimicrobials. Products derived from plants may potentially control microbial growth in diverse situations and in the specific case of disease treatment, numerous studies have aimed to describe the chemical composition of these plant antimicrobials and the mechanisms involved in microbial growth inhibition, either separately or associated with conventional antimicrobials. Thus, in the present work, medicinal plants with emphasis on their antimicrobial properties are reviewed.