The Inhibitory Activity of Typified Propolis against. Enterococcus Species

School of Dental Medicine, University of Pittsburgh, Departments of Oral Biology and Endodontics, 3501 Terrace Street, Pittsburgh, PA 15260, USA.
Zeitschrift fur Naturforschung C (Impact Factor: 0.55). 08/2012; 67(5-6):249-56. DOI: 10.5560/ZNC.2012.67c0249
Source: PubMed
ABSTRACT
Propolis, a natural bee product widely used for its antimicrobial activity, was tested against isolates of Enterococcus from humans, pig-tailed macaques, isolates of refractory endodontic treatment cases, and isolates from Lactobacillus-containing food supplements. Typification of the propolis was performed by high-performance liquid chromatography (HPLC) by which prenylated compounds, cinnamic acid derivatives, and flavonoids were detected as the main constituents. Minimum inhibitory concentrations (MIC) were determined using the agar dilution method. All human and animal Enterococcus isolates demonstrated MIC values of 1600 microg/mL. Enterococcal species of human and animal origin were inhibited by propolis. Particularly, human isolates of E. faecium and E. faecalis of refractory endodontic treatment cases were susceptible to propolis of Brazilian origin.

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Available from: Walter Bretz, Aug 18, 2015
© 2012 Verlag der Zeitschrift für Naturforschung, Tübingen · http://znaturforsch.com
Introduction
Enterococcus faecalis and E. faecium naturally
inhabit the gastrointestinal tract and the female
genital tract, are often present in various foods,
and are found in some natural food supplements
by accident or design (Facklam and Washington,
1991; Facklam et al., 1995, 1999; Hughes and Hill-
ier, 1990). These organisms have been of increas-
ing concern in medicine because of their involve-
ment in nosocomial infections, endocarditis, and
septicemia (Kirschner et al., 2001; Park and Walsh,
1997). Vancomycin resistance among strains of
Enterococcus is of concern because the resistance
is transmissible, and these orga nisms may there-
fore act as a reservoir for resistance (Facklam
et al., 1995; Park and Walsh, 1997; Bystrom and
Sundqvist, 1981; Heaton et al., 1996).
Treatment of infected root canals is a very suc-
cessful procedure. However, about 5% of teeth so
treated experience treatment failure (Sundqvist
et al., 1998; Sjogren et al., 1990). Successful treat-
ment requires the sterilization of the root canal
system and complete apical seal (Sundqvist et al.,
1998; Sjogren et al., 1990). E. faecalis, E. faecium,
and other species have been recovered from
root canals and periapical tissues of previously
endodontically treated teeth and are believed to
be involved in treatment failures (Bystrom and
Sundqvist, 1981, Sundqvist et al., 1998; Peciuliene
et al., 2000; Siren et al., 1997). The possible rea-
sons for treatment failure include: 1) failure to
achieve a complete apical seal; 2) incomplete
sterilization of the canal system; 3) persistent
infection at the time of canal obturation; 4) re-
infection or introduction of bacteria during in-
strumentation; and 5) reinfection through apical
dentinal tubules (Siren et al., 1997). The under-
lying source of the Enterococcus species in such
failures is unknown but could likely be endog-
enous, from the diet or from the operator during
root canal instrumentation.
The Inhibitory Activity of Typifi ed Propolis against
Enterococcus Species
Bernard J. Moncla
a,b
, Peter W. Guevara
b
, James A. Wallace
a
, Maria C. Marcucci
c
,
Jacques E. Nor
d
, and Walter A. Bretz
e,
*
a
School of Dental Medicine, University of Pittsburgh, Departments of Oral Biology and
Endodontics, 3501 Terrace Street, Pittsburgh, PA 15260, USA
b
School of Medicine, University of Pittsburgh and Magee-Women’s Research Institute,
3550 Terrace Street, Pittsburgh, PA 15260, USA
c
School of Pharmacy, Universidade Anhanguera-Uniban, Rua Maria Cândida, 1813,
São Paulo, SP, Brazil
d
University of Michigan, School of Dentistry, Department of Cariology, Endodontics and
Restorative Sciences, 1011 N. University Ave, Ann Arbor, MI 48109, USA
e
New
York University College of Dentistry, Department of Cariology and
Comprehensive Care, 345 E. 24
th
Street, New York, NY 10010, USA.
Fax: +1-212-9989914. E-mail: wb36@nyu.edu
* Author for correspondence and reprint requests
Z. Naturforsch. 67 c, # # (2012); received June 17, 2011/February 29, 2012
Propolis, a natural bee product widely used for its antimicrobial activity, was tested against
isolates of Enterococcus from humans, pig-tailed macaques, isolates of refractory endodontic
treatment cases, and isolates from Lactobacillus-containing food supplements. Typifi cation of
the propolis was performed by high-performance liquid chromatography (HPLC) by which
prenylated compounds, cinnamic acid derivatives, and fl avonoids were detected as the main
constituents. Minimum inhibitory concentrations (MIC) were determined using the agar
dilution method. All human and animal Enterococcus isolates demonstrated MIC values of
1600 μg/mL. Enterococcal species of human and animal origin were inhibited by propolis.
Particularly, human isolates of E. faecium and E. faecalis of refractory endodontic treatment
cases were susceptible to propolis of Brazilian origin.
Key words: Propolis, Enterococcus, MIC
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Running title: B. J. Moncla et al., · Propolis and Enterococci
Propolis is a resin-like product extracted from
plants by honey-bees that mix the resin with sali-
vary secretions and use the resulting mixture to
seal and to repair their hives. Propolis use dates
back to about 300 BC when it was fi rst used for
cosmetics and as a medicine (Dobrowolski et al.,
1991; Kujumgiev et al., 1999). The typical chemical
composition of propolis is 50% resin and vegetable
balsam, 30% wax, 10% essential and aromatic oils,
5% pollen, 5% minerals and fl avonoids. The chemi-
cal composition is dependent on the vegetation
from which the material was collected (Kujumgiev
et al., 1999; Marcucci et al., 2000; Moreno et al., 1999;
Sforcin et al., 2000). There are several methods that
can be used to extract propolis in order to devel-
op preparations and solutions. The most common
method that has been used is by ethanol extraction.
A number of studies have reported that propolis
has antibacterial, anti-infl ammatory, antifungal, an-
tiviral, anesthetic, antiulcer, immunostimulant, and
wound-healing properties (Kujumgiev et al., 1999;
Banskota et al., 2001; Cetinkaya et al., 2000; Koo
et al., 2000, 2002; Santos et al., 1999; Yatsuda et al.,
2000). The antimicrobial properties of propolis may
relate to or be a function of the fl avonoids (Banskota
et al., 2001; Cetinkaya et al., 2000; Mirzoeva et al.,
1997) and of other propolis components such as hy-
droxyacids, sesquiterpenes or phenolics (Banskota
et al., 2001). Accordingly, typifi cation of propolis is
an essential requirement in order to characterize its
moieties, quantify its main active compounds and
their respective biological applications.
Bretz and collaborators (1998) compared the
effects of propolis and calcium hydroxide on di-
rect dental pulp exposures in animals. Propolis
was at least comparable to calcium hydroxide in
exhibiting normal reorganization of the pulp and
no increased vascularity, and in maintaining a low
infl ammatory and microbial cell population.
Because of the suspected importance of Ente-
rococcus species in endodontic treatment failure
and their increasing importance in nosocomial
infections, we have studied a number of human
and animal isolates, reference strains, and strains
from food supplement sources as to determine
their susceptibility to propolis.
Methods and Materials
Propolis and vancomycin solutions
Crude propolis [source: alecrim (Baccharis dra-
cunculifolia)] was obtained from Piracicaba, São
Paulo, Brazil. Propolis was extracted in a Sox-
hlet extractor with 95% ethanol at 50 °C for 24 h
(three 8-h periods). The resulting syrup was dried
under vacuum and stored at –80 °C until used.
Subsequently, the propolis syrup was washed with
100 mL of cold ethanol. The solution was then fi l-
tered and stored at –20 °C until used. Working
stock solutions were prepared at a concentration
of 160 mg/mL in either 100% ethanol or dime-
thyl sulfoxide (DMSO). Serial twofold dilutions
of the stock solutions were used to give a fi nal
concentration of propolis ranging from 50 μg/mL
to 1600 μg/mL. Vancomycin/HCl •?• (Sigma, St.
Louis, MO, USA) dilutions ranging from 0.5 μg/
mL to 16 μg/mL served as positive controls.
Bacterial samples
Human isolates of Enterococcus species used
in this study represented rectal and vaginal iso-
lates from 87 •cf. Table II• women and animal
isolates that were obtained from 3• pig-tailed ma-
caques (Dr. S. L. Hillier, University of Pittsburgh,
School of Medicine, Department of Obstetric and
Gynecology, Pittsburgh, PA, USA). Two human
isolates were attained from clinical cases of en-
dodontic treatment failures (G. Sundqvist, Umeå
University, Faculty of Dentistry, Department
of Oral Microbiology, Umeå, Sweden). Twenty-
four Enterococcus isolates were recovered from
six Lactobacillus-containing health food supple-
ments. Control organisms included Staphylococ-
cus aureus ATCC 29213, Escherichia coli ATCC
25922, and E. faecalis ATCC 29212.
Health food supplements were purchased lo-
cally and stored as recommended by the manufac-
turers. Capsules were aseptically removed, placed
into 10 mL of Mueller-Hinton broth (•manufac-
turer, city, country•), and incubated at 37 °C as to
dissolve the capsules. Samples were dispersed with
a vortex mixer, and serial ten-fold dilutions were
prepared in the same media, plated on blood agar
plates, and incubated overnight at 37 °C. Typical
Enterococcus colonies were selected, subcultured
to establish purity, and identifi ed at the genus level
as described previously (Facklam and Washington,
1991; Facklam et al., 1995, 1999).
Microbiological procedures and minimum
inhibitory concentration (MIC)
MIC values of propolis and vancomycin were
determined using agar dilution methods in accord-
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Running title: B. J. Moncla et al., · Propolis and Enterococci
3
ance to CLSI standard methods for susceptibility
testing (Watts and Shryock, 2005). Briefl y, Mueller-
Hinton agar (Difco, Dearborn, MI, USA) was
used to carry out experiments. Each test solution
(at the various concentrations tested) was mixed
with the agar to give a fi nal content of the solvent
of 2%. Bacterial strains were cultured on blood
agar, isolated colonies selected and suspended in
saline to a density of 0.5 MacFarland units and
then diluted 1:10 in saline. Bacterial samples were
placed in 3 different wells of a Steer’s replicator
(400 μL) (•manufacturer, city, country•) and in-
oculated onto agar plates. The bacterial samples
were allowed to dry on the agar at room tem-
perature and incubated overnight at 37 °C in air
enriched with 5% CO
2
. Then the bacterial growth
was determined. All experiments were conducted
in duplicate and were repeated on three separate
days.
Composition of propolis assessed by HPLC
The typifi ed propolis sample (named BRP1)
used in this study was analyzed by high-perfor-
mance liquid chromatography (HPLC) (Merck-
Hitachi, Darmstadt, Germany) with L-7100 pumps
and an L-7200 auto-sampler. The chromatograph-
ic column was a reverse phase column Lichro-
chart 100 RP-18 (12.5 x 0.4 cm, particle dia meter
of 5 μm; Merck). The mobile phase was water/
formic acid (95:5, v/v) (solvent A) and methanol
(solvent B) at the fl ow of 1 mL/min using a lin-
ear gradient. The time of analysis was 50 min, and
the detection was performed at 280 and 340 nm
using a diode array (detector L-7450; Merck-
Hitachi). The software used for data analysis was
that provided by the manufacturer (DAD Man-
ager, Darmstadt, Germany). All compounds were
identifi ed by comparison with authentic standards
(same retention time and UV spectra) evaluated
by diode array.
Results
MIC values for typifi ed propolis were deter-
mined using two different solvents, ethanol and
DMSO. Comparable results were observed for
both solvents (Table I). The majority of reference
strains and isolates from patients refractory to en-
dodontic treatment exhibited MIC values equal
to the maximum concentration of propolis tested
(Table I), with the exception of S. aureus MIC val-
ue s for both propolis solvents which differed from
each other and were of lower concentration. Only
one strain of E. faecium demonstrated resistance
to the maximum concentration of propolis tested
when ethanol was used as the carrier for propo-
lis. Similar resistance patterns were found for E.
coli for both solvents. The reference strains of S.
aureus and E. faecalis remained within control
limits of 0.5 – 2 μg/mL and 1 4 μg/mL, respective-
ly, for the vancomycin assays (data not shown).
Table II presents the percentage of isolates and
the corresponding MIC values for propolis with
DMSO as a solvent. The 97 • human and ani-
mal isolates showed susceptibility to propolis at
1600 μg/mL. In addition, these isolates were sen-
sitive to vancomycin in the concentration range
of 0.5 – 8 μg/mL (data not shown). Enterococcus
strains isolated from food supplements had MIC
Table I. Propolis MIC values of reference strains and
human isolates.
Organism MIC in
ethanol
[μg/mL]
MIC in
DMSO
[μg/mL]
E. coli ATCC 25922
>1600 >1600
S. aureus ATCC 29213
400 <50
E. faecalis ATCC 29212
1600 1600
E. faecalis 3199
a
1600 1600
E. faecium 3266
a
>1600 1600
a
Isolates were recovered from refractory cases of
endodontic treatment.
Table II. Propolis MIC values for Enterococcus species from various sources.
Source Number of
isolates
% of isolates
MIC <400
a
μg/mL MIC 800 μg/mL MIC 1600 μg/mL MIC >1600 μg/mL
Human root canal 2 100
Human other sites 88 100
Pig-tailed macaque 7 100
Food supplements 24 19 14 39 28
a
Propolis solutions dissolved in DMSO.
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Running title: B. J. Moncla et al., · Propolis and Enterococci
values that ranged from <400 to >1600 μg/mL of
propolis.
Fig. 1 shows the HPLC chromatogram for the
typifi ed sample of propolis employed in this study.
Table III shows the compounds identifi ed by
HPLC. The majority of the compounds were de-
rived from cinnamic acid and p-coumaric acid. Fla-
vonoids were also detected but to a lesser extent.
Discussion
Previous reports have demonstrated differenc-
es in propolis antibacterial action against Gram-
positive and Gram-negative organisms, as well as
variations in the chemical composition of propolis
material dependent upon the location from where
the material was derived and how preparations
or solutions were designed (Banskota et al., 2001;
Marcucci et al., 2000; Marcucci and Bankova,
1999). Most propolis studies have used ethanol as
the solvent and have relied on either agar or disc
diffusion methods, or agar dilution methodology
to determine the MIC values for various bacterial
species.
We investigated the use of DMSO and etha-
nol in a standard protocol recommended for the
measurement of MIC values of Enterococcus
species (Watts and Shryock 2005). Agar dilution
plates prepared with propolis dissolved in etha-
nol were much less homogeneous in appearance,
especially at higher propolis concentrations, when
compared with plates prepared with propolis dis-
solved in DMSO. Their comparisons were gener-
ally in agreement (Table I) with two exceptions
where one strain of E. faecium required more
than 1600 μg/mL of propolis dissolved in ethanol
while complete inhibition was observed on plates
prepared with 1600 μg/mL of propolis dissolved in
DMSO. S. aureus had a MIC value of 400 μg/mL
in the former and <50 μg/mL in the latter. These
observations may be explained as a result from
a more homogeneous suspension of propolis di-
luted in DMSO than propolis diluted in ethanol.
Alternatively, DMSO may facilitate the transport
of propolis biologically active compounds into the
cells. There are other solvents that can be used for
propolis such as Tween 80 and sorbitol that are
as equally effective as DMSO (unpublished data).
These solvents are safe and would be indicated
for clinical use as opposed to DMSO.
Santos and colleagues (2002a) reported that
Gram-negative anaerobes and microaerophilic
organisms (Actinobacillus actinomycetemcomi-
tans, Fusobacterium spp., and Bacteroides fragilis)
Fig. 1. Retention time (min) at 2.43: caffeic acid; 3.95: p-coumaric acid; 4.92: ferulic acid; 20.57: 3-prenyl-4-hydroxy-
cinnamic acid; 22.90: 2,2-dimethyl-6-carboxyethenyl-2H-1-benzopyrane; 29.23: 3,5-diprenyl-4-hydroxycinnamic acid;
32.29: compound E; and 32.96: 6-propenoic-2,2-dimethyl-8-prenyl-2H-1-benzopyran acid. •Use “. in fi gure to sepa-
rate decimals. Write “min” not “nin”.•
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Running title: B. J. Moncla et al., · Propolis and Enterococci
5
were susceptible to concentrations of propolis
ethanolic extracts and commercial preparations
in the range of 50 5000 μg/mL. The same group
of investigators reported MIC values of propo-
lis ethanolic extracts and commercial prepara-
tions for Prevotella intermedia, P. nigrescens,
and Porphyromonas gingivalis in the range of
64 – 256 μg/mL (Santos et al., 2002b). It has been
suggested that Gram-positive organisms are more
sensitive to propolis than Gram-negative bacteria
(Banskota et al., 2001). Our results have demon-
strated that Enterococcus species required a mod-
erate to high concentration of propolis before in-
hibition of growth was observed. All human and
animal isolates were inhibited by 1600 μg/mL of
propolis dissolved in DMSO (Tables I and II).
Interestingly, the Enterococcus isolates obtained
from food supplements demonstrated greater di-
versity in the MIC values ranging from less than
400 μg/mL to greater than 1600 μg/mL. Differenc-
es in MIC profi les for the various pathogens de-
scribed above could be explained by plant origin
of the propolis and by different methodologies in
the preparation of propolis solutions.
There is evidence in the literature suggesting
that propolis ethanolic extracts can inhibit the
Table III. Quantitative analysis of propolis compounds identifi ed by HPLC. •check all names•
No.
a
Compound Identifi cation
b
Content (mg/g)
c
1
(E)-3-•?•4-Hydroxy-3-[(E)-4-(2,3)-dihydrocynamoyloxy-3-methyl-2-
butenyl]-5-prenylphenyl-2-propenoic acid
P 3.67 0.10
2
2,2-Dimethyl-6-carboxyethenyl-2H-1-benzopyran
- 5.05 0.02
3
2,2-Dimethyl-8-prenyl-2H-1-benzopyran-6-propenoic acid
- 13.24 0.23
4 3,4-Dihydroxy-5-prenylcinnamic acid P 1.49 0.01
5 3,5-Diprenyl-4-hydroxycinnamic acid
d
(derivative 11) P 0.84 0.04
6 3,5-Diprenyl-4-hydroxycinnamic acid
d
(derivative 12) P 1.15 0.05
7 3,5-Diprenyl-4-hydroxycinnamic acid
d
(derivative 13) P 2.34 0.11
8 3,5-Diprenyl-4-hydroxycinnamic acid
d
(derivative 2) P 1.21 0.03
9 3,5-Diprenyl-4-hydroxycinnamic acid
d
(derivative 6) P 3.30 0.12
10 3,5-Diprenyl-4-hydroxycinnamic acid (ARTEPILLIN C
®
) P 26.39 1.23
11 3-•?•4-Hydroxy-3-(oxobutenyl)-phenylacrylic acid P 0.82 0.01
12
3-Prenyl-3(E)-(4-hydroxy-3-methyl-2-butenyl)-5-prenylcinnamic acid
P 1.57 0,05
13 3-Prenyl-4-(2-methylpropionyloxy)-cinnamic acid P 0.91 0.04
14 3-Prenyl-4-dihydrocinnamoyloxycinnamic acid P 5.09 0.11
15 3-Prenyl-4-hydroxycinnamic acid P 5.43 0.20
16
6-Propenoic-2,2-dimethyl-8-prenyl-2H-1-benzopyran acid
P 4.39 0.07
17 Betuletol P 0.21 0.02
18 Caffeic acid P 1.55 0.03
19 Caffeoylquinic acid 1 P 13.61 0.67
20 Caffeoylquinic acid 2 P 0.69 0.03
21 Caffeoylquinic acid 3 P 2.91 0,04
22 Cinnamic acid
e
(derivative 2) P 2.39 0.02
23 Cinnamic acid
e
(derivative 3) P 65.68 4.57
24 Dihydrokaempferide P 2.41 0.13
25 Ferulic acid P 6.06 0.95
26 Kaempferide P/F 15.89 1.07
27 Kaempferol P/F 5.13 0.08
28
p-Coumaric acid
P 16.95 1.03
29 Pinobanksin P/F 30.29 2.95
Total amount (mg/g) - 240.67 14.01
Total amount (%) (w/w) - 24.07 1.40
a
There is no correlation with the retention time.
b
P, phenol; F, avonoid.
c
In milligrams of each compound per gram of crude resin.
d
Same UV spectra of 3,5-diprenyl-4-hydroxycinnamic acid with different retention times, expressed as 3,5-di-
prenyl-4-hydroxycinnamic acid derivative.
e
Same UV spectra of cinnamic acid with different retention time. Expressed as cinnamic acid.
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Running title: B. J. Moncla et al., · Propolis and Enterococci
growth of Streptococcus mutans, E. faecalis, and
S. aureus (Koo et al., 2000) indicating that our re-
sults are consistent with these reports. In fact, re-
cent reports corroborate our results where it has
been shown that propolis has signifi cant antimi-
crobial activity against E. faecalis (Kandaswamy
et al., 2010; Kayaoglu et al., 2011; Arslan et al.,
2011).
The assessment of antimicrobials that are not
water-soluble is diffi cult at best. Propolis is not
soluble in water and requires an organic solvent
as a carrier such as ethanol or DMSO. The chem-
ical composition of propolis may be crucial for
its solubility. For example, the typifi ed Brazilian
propolis includes numerous phenolic acid com-
pounds derived from cinnamic acid (Marcucci and
Bankova, 1999), which have different solubilities
in •ethanol• and water resulting in variations that
could affect MIC values. Most studies looking at
MIC values for propolis have used agar diffusion
from fi lter paper discs or wells cut into the agar
to assess the antimicrobial properties of propolis.
In such systems one cannot expect the propolis to
easily diffuse out. In this respect, the agar dilution
method may be more appropriate for determina-
tion of the action of water-insoluble antimicro-
bials. Accordingly, if the use of propolis for the
prevention of refractory endodontic treatment is
to be foreseen, a gel or paste vehicle for propolis
may be more appropriate for intracanal sealing.
In a review of the antimicrobial effects of
propolis, Banskota and collaborators (2001) cited
other studies that demonstrated that a minimum
of 60– 80 μg/ml of propolis is required to inhibit
S. aureus and Bacillus subtilis while a minimum
concentration of 600 800 μg/mL is required to
kill E. coli. Bankova and collaborators (•2000•)
have demonstrated that polar phenolic com-
pounds are responsible for the antimicrobial ef-
fects of propolis. Our results have shown that S.
aureus was inhibited by 400 μg/mL when ethanol
was used as a carrier, suggesting that our particu-
lar lot of propolis had a lower content of polar
phenolics. When DMSO was used as a carrier, the
propolis had an at least eightfold greater activity
against S. aureus (Table I) suggesting some syn-
ergistic effect of propolis with DMSO. The cin-
namic acid and fl avonoid derivatives have been
shown to uncouple energy transduction across
the cytoplasmic membranes of E. coli and B. sub-
tilis (Mirzoeva et al., 1997). Other components
of propolis have been isolated which are active
against other organisms (Koo et al., 2002; Marcuc-
ci et al., 2001; Bankova et al., 2000). Fractionation
of a propolis aqueous ethanol extract revealed
that these fractions exhibited antimicrobial activ-
ity against periodontal pathogens. The propolis
extract, however, was more active than were the
individual fractions suggesting a synergistic effect
of the different propolis compounds (Santos et al.,
2002a). The propolis used in our study was classi-
ed as BRP1 (Brazilian propolis with the highest
content of prenylated compounds) as previously
described by Miorin and colleagues (2003). The
compounds found in our propolis sample con-
rm previous studies that have examined com-
pounds found in Brazilian propolis (Marcucci et
al., 2000, 2001). The main compounds identifi ed
in these studies were derivatives of caffeic acid
and of p-coumaric acid, 3-prenyl-4-hydroxycin-
namic acid (PHCA), 3,5-diprenyl-4-hydroxycin-
namic acid (DHCA), 2,2-dimethyl-8-prenyl-2H-
1-benzopyran -6-propenoic acid (DCBEN), and
2,2-dimethyl-6-carboxyethenyl-2H-1-benzopyran
(DPB). The compounds DHCA and DPB were
inhibitory against E. coli, S. aureus, and S. faecalis.
In summary, enterococcal species of human
and animal origin were found to be susceptible
to propolis with moderate to high MIC values
(1600 μg/mL). Particularly, human isolates of
E. faecium and E. faecalis of refractory endodon-
tic treatment cases were susceptible to typifi ed
propolis of Brazilian origin at these concentra-
tions. These ndings would warrant future inves-
tigations on the clinical applications of typifi ed
propolis against organisms that are associated
with endodontic treatment failure.
Acknowledgements
This work was supported by NIH grant 6PO1
AI39061-07 and The University of Pittsburgh,
School of Dental Medicine’s Deans’ Fund. We
ac knowledge the excellent technical work of
B. Troy in this study.
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    • "The most evaluated bacterial strains were S. aureus, E.coli, Streptococcus pyogenes, Enterococcus spp., and some anaerobes. The reported data reveals that the Propolis extract has a broad antibacterial activitiy aginst gram positive and negative strains.[3][13] Although Propolis have either gram positive or gram negative activitiy as mentioned above, Stepanović S et al reported that antimicrobial activity of Propolis were frank against gram positive bacteria while gram negatives were less susceptible. "
    [Show abstract] [Hide abstract] ABSTRACT: Propolis, an old and a new natural product collected from beehives by honeybees (Apis mellifera) has been used from ancient times. The aim of this study is to determine the antibacterial activity of Propolis against gram positive and negative clinical isolates besides standard bacterial strains. Two different crude Propolis samples obtained from Konya-Turkey and Moscov-Russian Federation (2012 October) were used in this study. Methicillin sensitive Staphylococcus aureus (MSSA), methicillin resistant Staphylococcus aureus (MRSA), extended spectrum beta lactamases producing (ESBL) Escherichia coli and multi-drug resistant (MDR) Acinetobacter baumannii clinical isolates were tested. Thirty isolates from each bacterium enrolled. S.aureus ATCC 29213, MRSA ATCC 43300, E.coli ATCC 25922, P.aeruginosa ATCC 27853 strains were also tested as standard strains. The MICs of the strains were determined by broth dilution method. MSSA and MRSA clinical isolates’ MIC ranges of Turkish Propolis extract were between 512-2048 μg/mL, while 256-1024 μg/mL for Russian Propolis extract. For ESBL+ E.coli and A.baumannii the MIC ranges of both Turkish Propolis extract and Russian Propolis extract were greater than 16,384 μg/mL. The MIC values of standard strains were also compatible with clinical isolates. The bactericidal concentration of both Propolis samples were greater than 16,384 μg/mL for all strains containing gram positive and gram negative microorganisms used in this study. There was no statistically difference between methicillin resistance and MIC values (p< 0,091). There was statistically significant difference between Gram positive MIC values and Gram negative MIC values for both Turkish and Russian Propolis samples (p<0,001). According to our results crude Propolis is more effective against gram positive clinical isolates than gram negatives. Clinical strains may show different susceptibility patterns from standard strains. Thus to evaluate clinical strains besides standard strains in vitro should demonstrate more accurate results. Before marketing of Propolis containing products standardization, further laboratory assays, laboratory animal researches and clinical trials are to be needed.
    Full-text · Article · Jan 2015
  • Source
    • "The most evaluated bacterial strains were S. aureus, E.coli, Streptococcus pyogenes, Enterococcus spp., and some anaerobes. The reported data reveals that the Propolis extract has a broad antibacterial activitiy aginst gram positive and negative strains.[3][13] Although Propolis have either gram positive or gram negative activitiy as mentioned above, Stepanović S et al reported that antimicrobial activity of Propolis were frank against gram positive bacteria while gram negatives were less susceptible. "
    Full-text · Article · Jan 2015
  • Source
    • "The most evaluated bacterial strains were S. aureus, E.coli, Streptococcus pyogenes, Enterococcus spp., and some anaerobes. The reported data reveals that the Propolis extract has a broad antibacterial activitiy aginst gram positive and negative strains.[3][13] Although Propolis have either gram positive or gram negative activitiy as mentioned above, Stepanović S et al reported that antimicrobial activity of Propolis were frank against gram positive bacteria while gram negatives were less susceptible. "
    Full-text · Article · Jan 2015
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