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Antimicrobial activity of the essential oils from the

leaves of two morphotypes of Croton cajucara Benth
a b c d
Mariana M.B. Azevedo , Aline Q. Pereira , Francisco C.M. Chaves , Humberto R. Bizzo ,
e e
Celuta S. Alviano & Daniela S. Alviano
a
Programa de Ps-Graduao em Cincia de Alimentos , Universidade Federal do Rio de
Janeiro , Bloco A, Ilha do Fundo, Rio de Janeiro , RJ , 21941-909 , Brazil
b
Programa de Ps-Graduao em Biotecnologia Vegetal , Universidade Federal do Rio de
Janeiro , Bloco K, Ilha do Fundo, Rio de Janeiro , RJ , 21941-590 , Brazil
c
Embrapa Amaznia Ocidental , Rodovia AM 10, km 29, Manaus , AM , 69010-970 , Brazil
d
Embrapa Agroindstria de Alimentos , Avenida das Amricas 29501, Rio de Janeiro , RJ ,
23020-470 , Brazil
e
Instituto de Microbiologia Paulo de Ges , Universidade Federal do Rio de Janeiro , CCS,
Bloco I, Ilha do Fundo, Rio de Janeiro , RJ , 21941-590 , Brazil
Published online: 09 Jul 2012.

To cite this article: Mariana M.B. Azevedo , Aline Q. Pereira , Francisco C.M. Chaves , Humberto R. Bizzo , Celuta S. Alviano
& Daniela S. Alviano (2012) Antimicrobial activity of the essential oils from the leaves of two morphotypes of Croton cajucara
Benth, Journal of Essential Oil Research, 24:4, 351-357, DOI: 10.1080/10412905.2012.692902

To link to this article: http://dx.doi.org/10.1080/10412905.2012.692902

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 The Journal of Essential Oil Research
Vol. 24, No. 4, August 2012, 351357

Antimicrobial activity of the essential oils from the leaves of two morphotypes of
Croton cajucara Benth.
Mariana M.B. Azevedoa, Aline Q. Pereirab, Francisco C.M. Chavesc, Humberto R. Bizzod*, Celuta S. Alvianoe and
Daniela S. Alvianoe
a
Programa de Ps-Graduao em Cincia de Alimentos, Universidade Federal do Rio de Janeiro, Bloco A, Ilha do Fundo, Rio
de Janeiro, RJ, 21941-909 Brazil; bPrograma de Ps-Graduao em Biotecnologia Vegetal, Universidade Federal do Rio de
Janeiro, Bloco K, Ilha do Fundo, Rio de Janeiro, RJ, 21941-590 Brazil; cEmbrapa Amaznia Ocidental, Rodovia AM 10, km 29
Manaus, AM, 69010-970 Brazil; dEmbrapa Agroindstria de Alimentos, Avenida das Amricas 29501, Rio de Janeiro, RJ, 23020-
470 Brazil; eInstituto de Microbiologia Paulo de Ges, Universidade Federal do Rio de Janeiro, CCS, Bloco I, Ilha do Fundo,
Rio de Janeiro, RJ, 21941-590 Brazil
Downloaded by [University of Toronto Libraries] at 17:50 31 October 2014

(Received 2 September 2011; nal form 30 January 2012)

Croton cajucara Benth. (Euphorbiaceae) is a shrub native to Amazon region and locally known as sacaca. Two mor-
photypes are known, namely white sacaca and red sacaca. The essential oils from the leaves of these morphotypes
are rich in linalool and 7-hydroxycalamenene, respectively. The effectiveness of the oils from forty individuals from
a germplasm bank regarding the antimicrobial activity against Staphylococcus aureus and Candida albicans was
investigated by drop diffusion (or agar diffusion) method. Expressive inhibition zones for both microorganisms tested
were observed. Essential oils rich in 7-hydroxycalamenene were more effective against S. aureus while linalool-rich
essential oils acted inhibiting C. albicans. The minimal inhibitory (MIC) and microbicidal concentrations of the main
constituents of the oils were also determined, using broth dilution assay. It was observed that isolated 7-hydroxyca-
lamenene presented high bactericidal activity, with a MIC of 0.3 g/mL, while for fungicidal activity a MIC of 45 g/
mL was recorded. For S. aureus, the isolated 7-hydroxycalamenene was more active than the whole oil, whereas the
linalool-rich essential oil presented higher bactericide activity than pure linalool. For C. albicans, however, pure linal-
ool was more active than the whole essential oil.
Keywords: antimicrobial activity; essential oil composition; Croton cajucara; Candida albicans; Staphylococcus
aureus; linalool; 7-hydroxycalamenene

Introduction
The leaves and bark of Croton cajucara Benth. (Euphor-
biaceae) have been long used in Amazonian folk medi-
cine for the treatment of diabetes, malaria, cholesterol,
gastrointestinal and liver disorders (1, 2). The essential
oil from the leaves was found to be rich in linalool (3).
During the 1990s, Embrapa Amaznia Ocidental
has established a germplasm bank for agronomic stud-
ies of this species with individuals collected from dif-
ferent areas of the Amazon. Two morphotypes were
described: white sacaca and red sacaca, mainly identi-
ed by young leaf and steam color (4).
Considering chemical studies, the essential oils
from the leaves could be classied in two groups: one
rich in linalool, containing 2045% of this terpenic
alcohol, as well as circa 10% of nerolidol (5), the other
rich in cis-7-hydroxycalamenene, up to 44%, but also
containing 1520% of linalool (6).
Concerning biological studies, a very potent in vitro
activity against Leishmania amazonensis was observed
for the linalool-rich essential oil (7). A high activity was
also veried when the same oil was tested with some
planktonic microorganisms, being more active than pure
linalool (8). The biological activities of essential oils rich
in linalool are well documented (911). However, to the
best of our knowledge, there are few studies on biologi-
cal activity of 7-hydroxycalamenene, and they were
restricted to phytopathogenic fungi (12).
Staphylococcus aureus, a Gram-positive species, is
a ubiquitous colonizer of the skin and mucous mem-
branes of humans and animals. It is the most prominent
etiological agent for skin and soft tissue infection
(SSTI) and is a common target for natural product drug
screenings. Many strains of S. aureus carry resistance
genes for penicillin antibiotics, tetracyclines, methicillin
and now, vancomycin. Methicillin-resistant Staphylo-
coccus aureus (MRSA) presents a signicant threat to
public health in the USA. From a surveillance study on
MRSA, over 90,000 patients in the USA had invasive
MRSA infections in 2005, resulting in an estimation of

*Corresponding author. Email: bizzo@ctaa.embrapa.br

ISSN 1041-2905 print/ISSN 2163-8152 online

2012 Taylor & Francis
http://dx.doi.org/10.1080/10412905.2012.692902
http://www.tandfonline.com
 352 M.M.B. Azevedo et al.
18,650 deaths, surpassing the mortality estimates for
AIDS in this country (13).
Yeasts from the genus Candida are usual opportu-
nistic pathogens in humans. They can be found in 50
60% of oral cavities of healthy adults, but also colonize
surfaces such as intestinal and vagina epithelia.
Amongst Candida species, Candida albicans is the
most commonly isolated from oral cavity and is
responsible for most supercial and systemic fungal
infections (14).
The objective of this work was to study the biologi-
cal activity of the essential oils from the leaves of C.
cajucara, against S. aureus and C. albicans.
Experimental
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Plant material
All individuals were kept in a germplasm bank under
the same cultivation practices. Leaves of C. cajucara
were collected between 08:00 and 09:00 hours. Vou-
cher specimens were deposited at Embrapa Amaznia
Ocidental Herbarium (registry IAN 165013). SV coded
samples corresponded to the red morphotype, while SB
to the white one.
Chemicals
All solvents used were spectroscopic grade (Tedia, Fair-
eld, USA). Reagents were from Aldrich (Milwaukee,
USA). Column and planar chromatographic products
were furnished by Merck (Darmstadt, Germany).
Essential oils
The oils were obtained by hydrodistillation on a modi-
ed Clevenger apparatus for 4 hours.
Analysis of the essential oils
The oils were analyzed in an Agilent (Palo Alto, USA)
6890N gas chromatograph tted with a 5% phenyl95%
methylsilicone (HP5, 30 m  0.32 mm  0.25 m) fused
silica capillary column. The oven temperature was
programmed from 60C to 240C at 3C/minute, and
hydrogen was used as carrier gas (1.4 mL/minute);
1.0 L of a 1% solution of the oil in dichloromethane
was injected, in split mode (1:100). The injector was kept
at 250C and the ame ionization detector (FID) at 280
C. All analyses were performed in triplicate.
Mass spectra were obtained in an Agilent 5973N sys-
tem operating in electronic ionization mode (EI) at 70
eV, with scan mass range of 40500 m/z. Sampling rate
was 3.15 scan/second. The ion source was kept at 230C,
mass analyzer at 150C and transfer line at 260C. The
mass detector was coupled to an Agilent 6890 gas chro-
matograph tted with a low bleeding 5% phenyl95%
methylsilicone (HP-5 MS, 30 m  0.25 mm  0.25 m)
fused silica capillary column. Injection procedure and
oven temperature program were the same as above.
Helium was the carrier gas, at 1.0 mL/minute.
Linear retention indices (LRI) were calculated (15)
by injection of a series of n-alkanes (C7C26) in the
same column and conditions as above for gas chroma-
tography analyses.
Identication of the oil components was based on
computer search using the Wiley 6th edition library of
mass spectral data and by comparison of their calcu-
lated LRIs with literature data (16). Standard solutions
of linalool and cis-7-hydroxycalamenene were also
injected for conrmation. The later standard was pre-
pared after isolation of the hydroxycalamenene by pre-
parative column chromatography followed by 1H and
13
C NMR characterization, as described previously (6).
Antimicrobial analysis of the essential oils and
7-hydroxycalamenene
The antimicrobial assay was carried out by the drop
agar diffusion method (1719). The microorganisms
tested were C. albicans Serotype B (ATCC 36802) and
S. aureus MRSA (BMB9393) (Hospital Clementino
Fraga Filho, UFRJ). Microorganisms were spread over
Petri plates containing BHI (brain heart infusion) solid
medium and, after 10 minutes, 10 L of the oil were
added to the center of the plate. 7-hydroxycalamenene
was dissolved in 50 L of dimethyl sulfoxide (DMSO)
and diluted in water (1:3), providing as nal concentra-
tion 48 mg/mL. Then 10 L were added to the center of
the plate. After incubation at 37C for 48 hours, the
diameter of the inhibition halos (in cm) were measured.
Thin-layer chromatography (TLC) and bioautography
The oils were applied on silica-gel 60F254 chromato-
plates and eluted with hexane-ethyl acetate (9:1). Spots
were revealed by observation under UV light (254 nm),
then with sulfuric anisaldehyde and heating. For the
bioautography, the experiment above was repeated, but
after elution and drying of solvents the plates were
exposed to UV for 20 minutes, then added to sterile
Petri dishes and immediately covered with BHI previ-
ously inoculated with the microorganisms to be tested
(c. 106108 cells per dish). After incubation period, the
plates were observed and the inhibition zones were
marked with 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-
2H-tetrazolyum bromide (20).
Minimal inhibitory concentration (MIC) assays
The MIC of 7-hydroxycalamenene against C. albicans
and S. aureus was determined by broth microdilution
technique, in a sterile 96-well microtiter plate, as
described by the Clinical and Laboratory Standards
Institute (21). The isolated 7-hydroxycalamenene was
tested at concentrations ranging from 0.1 to 10 mg/
mL, diluted in 1% DMSO. Each well contained
 The Journal of Essential Oil Research
100 L of two-fold serially diluted 7-hydroxycalamene
in Mueller Hinton growth medium (for bacteria) or
RPMI 1640 (for yeasts), and either 10 L of an
overnight bacterial culture, representing approximately
5  104 bacterial cells/mL, or 100 L of an overnight
yeast culture, representing circa 2.5  103 yeast cells/
mL. Amphotericin B (MIC 1.6 g/mL) and ciprooxa-
cin (MIC 0.25 g/mL) were used as positive controls.
The negative controls comprised pure growth media
and inoculated growth media without test agent. The
plates were incubated at 37C for 24 hours (bacteria)
or 37C for 48 hours (yeast). The results were based
on visible growth or inhibition. The plates were sha-
ken for 30 seconds before each reading.
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Minimal bactericidal concentration (MBC) and
minimal fungicidal concentration (MFC)
To assess whether the action of the essential oil was
microbiostatic or microbicidal, 20 L of cultures of
each microorganism, from concentrations equal or
higher than MIC, were resuspended in 200 L of sterile
saline solution. This volume was streaked on BHI agar
plates and incubated at 37C for 48 hours. No growth
at concentrations higher than or equal to the MIC indi-
cated bactericidal action. Control tests were run simul-
taneously by adding solvents, without the essential oil.
All experiments were made in triplicate.
Results and discussion
The average essential oil yields were 0.8% and 1.0%
for the red and white morphotypes, respectively. The
chemical composition of the oils is presented in Tables
1 and 2 (red sacaca and white sacaca). The major com-
pounds present in red morphotype samples (Table 1)
were 7-hydroxycalamenene (up to 44.3%), linalool, -
caryophyllene and nerolidol. It is worth of note that
there is chemical variability inside the red morphotype,
with samples presenting no 7-hydroxycalamenene.
Therefore, this substance could not be used as chemical
marker for this morphotype.
For the white morphotype (Table 2), linalool was
usually the major component, followed by nerolidol, -
caryophyllene and germacrene D. In general, 7-hydrox-
ycalamenene was not present, but samples SB022,
SB025 and SB027 contained this compound.
From these data, the occurrence of chemotypes can-
not be associated with the observed morphotypes.
The results obtained for the antimicrobial activity
are presented in Table 3. For C. albicans, inhibition
halos varied from 0.9 to 1.3 cm, and no signicant dif-
ferences were observed between the oils tested. The
growth inhibition activity seemed to be independent of
the presence of 7-hydroxycalamenene for C. albicans,
since no expressive variation in halo diameters was
353
observed with high (SV012), low (SB025) or no con-
tent (SB034) of 7-hydroxycalamenene.
For S. aureus, however, a wide variation between
inhibition halos was recorded. The largest inhibition
halo (1.8 cm) was observed when oil from sample
SV010 was tested. This oil was the one with highest
content of 7-hydroxycalamenene (44.3%). The smallest
inhibition (0.2 cm) occurred in the test of the sample
SB032, with presented the highest concentration of lin-
alool (45.0%) and absence of 7-hydroxycalamenene.
According to these results, it was suggested that the
antibacterial activity against S. aureus is due mainly to
7-hydroxycalamenene and that it is not affected by lin-
alool content. Since some samples exhibited some
activity even in the lack of 7-hydroxycalamenene, the
contribution of other components in the oil to the anti-
bacterial activity should be considered.
From the drop tests carried out previously, a higher
activity of the oils from the red morphotype (coded
SV) was observed against bacteria, while for fungus no
generalization could be made.
To evaluate which compounds were responsible for
the activity observed in the drop test, some samples
were subjected to bioautography. For the sample
SV005 (with 31.9% of 7-hydroxycalamenene and
16.1% of linalool), no growth was observed for S. aur-
eus in the band of elution of 7-hydroxycalamenene.
The effect on the growth of C. albicans, however, was
very small.
The test was run also with sample SB016, which
contained no 7-hydroxycalamenene and 18.6% of linal-
ool. Only a small inhibition was observed for S. aureus
and no inhibition at all when C. albicans was essayed.
Since this last result seemed to be in contradiction with
the drop test data, the drop test was repeated using pure
linalool, in the same concentration of the oil. The C.
albicans growth inhibition due solely to linalool was
conrmed. Comparisons of inhibition halos between
oils and pure compounds are presented in Table 4.
Results for MIC are presented in Table 5. After
establishing the MIC, no growth of the tested organ-
isms was observed when placed in new media, leading
to the conclusion that at the MIC the oils tested were
bactericidal and fungicidal.
It was observed that S. aureus was more sensitive to
7-hydroxycalamenene than C. albicans. The MIC was
also expressed as minimum bactericidal concentration
(MBC) and minimum fungicidal concentration (MFC).
The MBC (0.3 g/mL) was similar to that observed for
the antibiotic ciprooxacin (0.25 g/ml), but result of the
MFC (45 g/mL) was not as efcient when compared
with the MIC/MFC of amphotericin B (1.6 g/mL).
The inhibitory capacity against S. aureus is classi-
ed as high in samples with MIC values between 50
and 500 g/ml (0.05 to 0.50 mg/mL), as moderate with
 Downloaded by [University of Toronto Libraries] at 17:50 31 October 2014

354

Table 1. Chemical composition of Croton cajucara essential oils red morphotype.

Componentsa LRI SV001 SV002 SV003 SV004 SV005 SV006 SV007 SV008 SV009 SV010 SV011 SV012 SV013
-Pinene 937 1.2 0.7 0.2 0.4 0.2 0.6 2.1 9.5 3.5 0.7 n.d. n.d. 0.5
Sabinene 976 10.5 0.1 0.1 n.d. 0.8 n.d. n.d. 0.2 n.d. 3.8 n.d. n.d. n.d.
-Pinene 979 0.6 n.d. n.d. n.d. n.d. n.d. 0.8 3.5 1.3 0.0 n.d. n.d. n.d.
(E)--Ocimene 1050 1.0 0.1 0.4 n.d. 0.4 0.7 0.5 n.d. n.d. 0.5 n.d. n.d. 0.2
Linalool 1101 8.3 21.5 28.6 16.3 16.1 17.3 18.6 18.5 14.5 3.0 19.4 15.2 15.6
-Copaene 1376 1.9 0.6 0.5 2.5 1.9 1.3 1.9 1.7 1.0 2.3 1.4 1.3 2.1
-Bourbounene 1380 0.6 2.4 1.9 1.5 1.2 1.8 1.5 1.0 1.5 0.7 1.1 1.1 1.4
-Elemene 1389 n.d. 0.3 0.4 0.4 0.1 0.7 n.d. n.d. 1.0 n.d. n.d. n.d. 0.1
-Caryophyllene 1419 4.8 10.4 9.1 3.6 3.2 8.9 3.2 3.2 10.0 6.2 3.3 2.9 3.8
-Copaene 1428 n.d. 1.4 1.1 0.5 0.6 0.5 0.6 n.d. 0.6 n.d. n.d. n.d. 0.2
-Humulene 1453 1.6 1.9 1.8 1.4 0.8 2.0 1.1 0.4 2.1 1.9 1.2 1.1 1.3
allo-Aromadendrene 1460 2.3 n.d. 0.8 2.5 2.1 1.2 1.7 1.7 1.1 2.8 1.9 1.8 2.4
-Muurolene 1476 0.8 0.3 0.3 1.0 0.8 0.8 0.5 0.7 0.4 1.0 n.d. 0.6 0.8
Germacrene D 1480 6.5 9.8 8.9 8.0 5.9 8.5 7.3 4.5 8.5 8.9 5.7 4.3 5.1
Bicyclogermacrene 1494 0.7 7.5 5.5 1.8 0.7 2.9 2.5 0.7 5.2 0.6 0.9 0.7 0.7
-Muurolene 1498 0.6 1.8 1.4 0.8 1.0 0.8 0.9 0.6 0.9 0.6 n.d. 0.4 0.5
n.i. 1502 0.4 0.7 0.6 0.3 0.5 0.7 0.5 n.d. 1.7 n.d. n.d. n.d. 0.2
-Cadinene 1523 2.6 2.1 1.6 6.6 4.6 2.7 3.9 3.4 0.3 5.0 5.9 2.0 7.6
Germacrene B 1556 1.0 2.8 2.6 0.9 0.7 3.1 1.6 0.7 4.1 1.3 1.2 1.0 0.7
Nerolidol 1567 n.d. 12.6 10.0 0.8 2.0 10.4 4.3 n.d. 0.7 n.d. 3.4 2.6 2.2
M.M.B. Azevedo et al.

Spathulenol 1577 1.2 2.2 2.3 2.3 1.7 1.3 1.7 n.d. 2.3 n.d. n.d. n.d. 1.6
Caryophyllene oxide 1581 0.6 1.1 0.9 0.6 0.6 1.2 n.d. 0.9 1.5 0.5 n.d. n.d. 0.7
Viridiorol 1589 n.d. n.d. n.d. 1.2 2.1 n.d. n.d. n.d. 0.5 n.d. 2.4 2.5 1.8
Rosifoliol + guaiol 1600 0.8 0.6 0.6 0.8 1.0 1.0 n.d. 0.6 0.7 0.5 1.2 1.2 0.6
n.i. 1628 n.d. 1.0 1.0 0.8 1.6 n.d. 1.2 0.4 n.d. n.d. n.d. n.d. 0.7
epi--Cadinol 1641 0.6 0.5 0.7 0.8 1.0 0.7 n.d. n.d. n.d. n.d. n.d. n.d. 0.8
-Muurolol 1646 n.d. 1.6 n.d. 0.5 0.7 0.7 0.5 n.d. n.d. n.d. n.d. n.d. 0.7
-Cadinol 1654 0.5 1.3 1.0 1.1 1.5 1.6 0.9 n.d. n.d. n.d. n.d. n.d. 1.3
n.i. 1673 n.d. n.d. n.d. n.d. n.d. n.d. 3.1 n.d. n.d. n.d. n.d. n.d. n.d.
n.i. 1688 n.d. 0.6 0.6 0.4 0.5 0.6 0.5 n.d. 4.9 n.d. 1.8 1.5 1.2
Zinzanal 1693 n.d. n.d. n.d. 0.8 1.4 0.5 0.2 n.d. n.d. n.d. n.d. n.d. n.d.
Drimenol 1762 1.0 4.2 4.4 1.1 0.8 n.d. 2.7 0.9 0.1 1.7 1.3 1.0 n.d.
7-Hydroxycalamenene 1806 35.9 n.d. n.d. 27.9 31.9 4.9 22.2 27.2 9.1 44.3 30.2 34.5 31.2

Notes: aCompounds present at 1.0% or higher in at least one sample. LRI, linear retention indices, on a HP-5 column. n.d., not detected; n.i., not identied.
 Downloaded by [University of Toronto Libraries] at 17:50 31 October 2014

Table 2. Chemical composition of Croton cajucara essential oils white morphotype.

Componentsa LRI SB014 SB015 SB016 SB017 SB018 SB019 SB020 SB021 SB022 SB023 SB024 SB025 SB026 SB027 SB028 SB029 SB030 SB031 SB032 SB033 SB034 SB035 SB036 SB037 SB038 SB039 SB040

-Pinene 937 0.7 1.0 0.3 0.5 0.5 0.2 0.7 0.5 0.4 0.1 0.3 0.5 0.7 0.8 0.1 0.41 n.d. 0.3 0.8 0.5 0.4 0.2 0.5 n.d. 0.6 0.6 0.4
1,8-Cineole 1034 0.8 0.8 0.4 0.3 0.9 0.1 0.3 0.3 0.5 n.d. 0.6 1.6 1.2 1.1 0.3 0.6 0.3 0.3 2.8 1.2 0.6 0.5 0.3 0.6 n.d. 0.4 0.4
Linalool 1101 20.1 27.0 18.6 22.0 29.7 16.3 25.0 23.8 22.6 2.9 22.5 24.1 31.3 34.1 15.6 25.6 13.9 15.9 45.0 31.1 29.1 19.1 18.5 25.1 16.8 23.5 24.8
-Elemene 1338 0.5 0.5 0.8 0.8 0.5 0.6 0.7 0.6 0.5 1.1 0.6 0.4 0.7 0.5 0.9 0.5 0.6 0.5 0.3 0.4 0.5 0.9 0.6 0.5 0.8 0.8 0.8
n.i. 1365 1.0 0.8 1.0 1.0 0.7 0.6 0.9 0.8 0.8 1.6 0.9 0.6 0.6 0.7 1.0 0.7 1.0 0.8 0.4 0.7 0.7 0.9 1.0 0.8 1.0 0.8 0.7
-Copaene 1376 0.7 0.6 0.7 0.5 0.5 0.5 0.6 0.6 0.8 1.1 0.6 0.5 0.5 0.5 0.8 0.5 0.7 0.6 0.3 0.5 0.5 0.6 0.7 0.6 0.7 0.6 0.5
-Bourbonene 1380 2.6 2.6 3.3 2.2 2.0 2.0 2.1 2.8 2.7 5.1 3.0 2.5 1.6 2.2 3.2 2.4 3.2 2.6 1.5 1.9 2.3 2.2 3.3 2.6 3.0 1.7 2.4
-Cubebene 1387 n.d. 0.1 0.1 n.d. n.d. 0.1 n.d. n.d. n.d. 0.1 n.d. n.d. n.d. n.d. T n.d. n.d. 0.1 n.d. n.d. n.d. n.d. n.d. n.d. n.d. 0.1 n.d.
-Elemene 1389 0.6 0.3 0.6 0.5 0.6 0.6 0.6 0.6 0.6 1.1 0.6 0.5 0.6 0.6 0.8 0.5 0.7 0.6 0.4 0.5 0.6 0.6 0.7 0.6 0.7 0.6 0.6
-Gurjunene 1408 3.4 0.1 n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d.
-Caryophyllene 1419 7.3 9.0 11.5 13.4 9.3 11.0 13.0 14.1 7.8 13.8 10.6 6.2 9.0 7.8 9.5 9.4 9.8 8.0 5.5 8.2 6.7 11.9 8.5 10.5 13.6 9.4 11.9
-Copaene 1428 1.0 1.3 1.6 1.4 1.2 1.1 1.3 1.4 1.3 2.6 1.4 1.1 0.9 1.1 1.7 1.2 1.7 1.3 0.7 1.0 1.1 1.3 1.7 1.4 1.5 1.1 1.2
-Humulene 1453 1.9 1.7 2.1 2.3 1.8 2.2 2.3 2.6 1.6 2.5 1.9 1.4 1.7 1.5 1.8 1.8 1.9 1.7 1.1 1.6 1.4 2.1 1.7 2.0 2.5 1.8 2.2
allo-Aromadendrene 1460 1.0 0.8 1.1 0.9 0.8 0.7 0.9 0.8 1.1 1.5 1.0 0.8 0.8 0.7 1.1 0.8 1.1 0.9 0.5 0.8 0.7 1.0 1.0 0.9 0.9 0.8 0.8
Germacrene D 1480 8.4 7.9 9.2 9.8 7.7 8.8 9.8 7.8 4.1 13.8 7.4 5.2 8.7 6.7 10.4 6.7 8.8 6.4 3.5 6.2 6.9 10.7 9.1 7.5 9.2 10.8 9.1
Bicyclogermacrene 1494 5.5 5.4 6.1 5.0 5.0 5.1 4.4 4.2 2.1 9.1 5.6 3.5 5.9 4.3 7.1 5.0 6.5 5.3 2.9 4.4 6.0 8.7 7.5 5.4 4.7 5.1 4.8
-Muurolene 1498 1.8 1.4 2.0 1.8 1.5 1.5 1.7 1.7 2.1 3.0 1.7 1.4 1.1 1.4 1.9 1.4 2.1 1.6 0.9 1.3 1.3 1.6 1.9 1.7 1.9 1.5 1.5
-Cadinene 1523 1.6 1.6 2.2 2.5 1.9 1.8 2.0 2.0 2.9 3.3 1.9 1.9 1.8 1.8 2.3 1.6 2.3 2.2 1.1 1.5 1.6 2.0 2.1 1.7 2.2 1.8 1.8
Germacrene B 1556 2.4 2.2 2.6 2.7 2.2 2.7 2.4 2.2 2.1 3.9 2.3 1.9 2.4 2.0 2.8 2.1 2.6 2.3 1.3 1.9 2.2 2.6 2.7 2.3 2.5 2.8 2.4
Nerolidol 1567 10.1 10.7 10.3 10.8 9.2 10.7 10.4 9.0 8.8 8.4 10.7 12.1 8.6 8.8 10.1 11.9 12.2 13.3 8.5 n.d. 12.2 11.1 12.5 9.5 12.4 11.9 10.6
Spathulenol 1577 2.3 2.2 1.7 1.5 1.8 2.1 1.0 1.5 1.8 1.7 2.5 3.0 1.5 2.3 2.5 3.0 3.4 3.1 2.4 n.d. 3.2 2.1 3.1 2.8 1.6 1.2 1.5
Caryophyllene oxide 1581 1.4 1.3 1.1 1.2 1.3 1.6 1.0 1.6 1.4 0.8 1.6 2.0 0.9 1.5 1.1 1.8 1.8 1.8 1.6 10.7 1.4 1.0 1.3 1.8 1.4 0.9 1.2
n.i. 1586 n.d. T n.d. n.d. n.d. 0.1 0.2 n.d. 0.1 n.d. n.d. n.d. n.d. n.d. n.d. n.d. n.d. 0.1 n.d. 2.4 n.d. n.d. n.d. 0.6 n.d. 0.1 n.d.
The Journal of Essential Oil Research

n.i. 1628 1.0 0.8 0.8 0.9 0.9 1.1 0.7 0.9 1.1 1.0 0.9 1.4 0.8 1.0 0.9 1.1 1.2 1.4 1.0 1.0 1.2 0.6 1.1 1.1 0.9 0.8 0.8
n.i. 1637 0.7 0.6 0.6 0.5 0.6 0.8 0.4 0.6 0.5 0.6 0.8 0.8 0.5 0.7 0.7 0.9 0.9 1.0 0.7 0.7 0.9 0.5 0.8 0.8 0.6 0.5 0.6
epi--Cadinol 1641 0.6 0.7 0.7 0.6 0.7 0.8 0.7 0.8 0.9 0.7 0.7 1.1 0.6 0.8 0.7 0.8 0.9 1.3 0.8 0.7 0.8 0.6 0.8 0.7 0.8 0.6 0.7
-Muurolol 1646 1.5 1.4 1.4 1.5 1.6 2.1 1.5 1.7 1.6 1.5 1.5 1.9 1.5 1.6 1.6 1.7 1.8 2.2 1.4 1.5 1.7 1.4 1.7 1.7 1.7 1.6 1.5
-Cadinol 1654 1.0 1.0 1.3 1.0 1.2 1.3 1.1 1.3 1.5 1.3 1.3 1.9 1.1 1.4 1.0 1.4 1.4 2.3 1.3 1.2 1.4 0.9 1.3 1.3 1.3 1.0 1.2
n.i. 1731 1.0 0.8 0.4 0.6 0.6 1.2 0.6 1.2 0.9 1.0 1.0 1.0 0.6 0.3 1.0 1.9 1.1 1.0 0.7 0.9 0.8 0.6 0.8 1.0 1.0 0.5 0.9
Drimenol 1762 4.3 2.9 4.3 3.3 4.4 5.6 3.5 4.3 3.9 4.5 3.8 4.2 3.9 3.3 4.1 3.9 4.8 4.3 2.7 3.7 3.2 3.3 3.1 3.6 3.8 3.5 4.1
7-Hydroxycalamenene 1806 n.d. n.d. n.d. n.d. n.d. 0.1 n.d. n.d. 8.0 n.d. n.d. 2.7 n.d. n.d. 2.6 n.d. n.d. 0.1 n.d. n.d. n.d. n.d. n.d. n.d. n.d. 0.6 n.d.

Notes: aCompounds present at 1.0% or higher in at least one sample. LRI, linear retention indices, on a HP-5 column. n.d., not detected; n.i., not identied.
355
 356 M.M.B. Azevedo et al.

Table 3. Inhibition growth zones (in cm) for microorganisms treated with Croton cajucara oil.
Sample Linalool 7-HC Candida albicans Staphylococcus aureus
SV001 8.3 35.9 1.3 1.6
SV002 21.5 n.d. 1.0 0.5
SV003 28.6 n.d. 1.1 0.5
SV004 16.3 27.9 1.2 1.3
SV005 16.1 31.9 1.2 1.4
SV006 17.3 4.9 0.9 0.8
SV007 18.6 22.2 1.2 1.1
SV008 18.5 27.2 1.2 1.1
SV009 14.5 9.1 1.1 0.9
SV010 3.0 44.3 1.2 1.8
SV011 19.4 30.2 1.2 1.1
SV012 15.2 34.5 1.1 1.5
SV013 15.6 31.2 1.1 1.5
SB014 20.1 n.d. 1.1 0.6
Downloaded by [University of Toronto Libraries] at 17:50 31 October 2014

SB015 27.0 n.d. 1.1 0.4

SB016 18.6 n.d. 1.0 0.8
SB017 22.0 n.d. 1.0 0.5
SB018 29.7 n.d. 1.0 0.9
SB019 16.3 0.1 n.t. n.t.
SB020 25.0 n.d. 0.9 0.9
SB021 23.8 n.d. 1.0 0.9
SB022 22.6 8.0 1.0 1.0
SB023 2.9 n.d. 1.1 1.0
SB024 22.5 n.d. 1.0 0.8
SB025 24.1 2.7 1.1 0.9
SB026 31.3 n.d. 1.0 0.6
SB027 34.1 n.d. 1.0 0.9
SB028 15.6 2.6 1.0 1.0
SB029 25.6 n.d. 1.0 0.6
SB030 13.9 n.d. 1.0 0.7
SB031 15.9 0.1 n.t. n.t.
SB032 45.0 n.d. 1.0 0.2
SB033 31.1 n.d. 1.0 0.7
SB034 29.1 n.d. 1.1 0.6
SB035 19.1 n.d. 1.2 0.7
SB036 18.5 n.d. 1.0 1.0
SB037 25.1 n.d. 1.2 0.6
SB038 16.8 n.d. 1.3 0.9
SB039 23.5 0.1 n.t. n.t.
SB040 24.8 n.d. 1.1 0.3

Notes: 7-HC, cis-7-hydroxycalamenene-rich oil; n.d., not detected; n.t., not tested (oil amount obtained not sufcient for biological testing).

Table 4. Growth inhibition zones (in cm) for the oil rich in Table 5. Minimum inhibitory concentration (MIC) for linalool
7-hydroxycalamenene and the isolated compound.a and 7-hydroxycalamenene against Staphylococcus aureus and
Candida albicans.
Whole essential oil Pure
Microorganism (SV010) 7-HC 7-hydroxy- Reference
Microorganism Linalool calamenenea drug
Candida albicans 1.2 1.0
Staphylococcus aureus 1.8 2.0 S. aureus >2500 0.3 g/mL 0.25 g/mLb
g/mL
Notes: aResults from one of three independent experiments per- C. albicans 2500 45 g/mL 1.6 g/mLc
formed. 7-HC, cis-7-hydroxycalamenene. g/mL

Notes: aResults from one of three independent experiments per-

MIC between 600 and 1500 g/mL (0.6 to 1.50 mg/
mL), and as low with MIC >1500 g/mL (1.50 mg/
mL), using chloramphenicol as control with a MIC of
20 g/mL (0.02 mg/mL) for S. aureus (22). Since the
MIC for 7-hydroxycalamenene for this organism is
formed. bCiprooxacin. cAmphotericin B.
0.3 g/mL (0.0003 mg/mL), this substance shows very
promising potential use against this organism.
 The Journal of Essential Oil Research
Acknowledgements
This work was supported by Coordenao de Aperfeioamento
de Pessoal de Nvel Superior (CAPES), Conselho Nacional de
Desenvolvimento Cientco e Tecnolgico (CNPq), Fundao 11. L. Jirovetz, G. Buchbauer, Z. Denkova, A. Stoyanova,
de Amparo a Pesquisa do Estado do Rio de Janeiro (FAPERJ)
and Universidade Federal do Rio de Janeiro (UFRJ).
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