Synthesis, Antibacterial Evaluation, and QSAR of Caffeic Acid Derivatives

Article abstract of DOI:10.1155/2019/3408315

The present study evaluates the antibacterial effects of a set of 16 synthesized caffeic acid ester derivatives against strains of Staphylococcus aureus and Escherichia coli, as well as discusses their structure-activity relationship (SAR). The antibacterial assays were performed using microdilution techniques in 96-well microplates to determine minimal inhibitory concentration (MIC). The results revealed that five of the compounds present strong to optimum antibacterial effect. Of the sixteen ester derivatives evaluated, the products with alkyl side chains, as propyl caffeate (3), butyl caffeate (6), and pentyl caffeate (7), presented the best antibacterial activity with MIC values of around 0.20 μM against Escherichia coli and only butyl caffeate (6) showing the same MIC against Staphylococcus aureus. For products with aryl substituents, the best MIC results against the tested strain of Escherichia coli were 0.23 μM for (di-(4-chlorobenzyl)) caffeate (13) and 0.29 μM for diphenylmethyl caffeate (10) and all were less active against the Staphylococcus aureus strain. Preliminary quantitative structure-activity relationship (QSAR) analyses confirmed that certain structural characteristics, such as a median linear carbon chain and the presence of electron withdrawal substituents at the para position of the aromatic ring, help potentiate antibacterial activity.

Full text of DOI:10.1155/2019/3408315

Hindawi
Journal of Chemistry
Volume 2019, Article ID 3408315, 9 pages
https://doi.org/10.1155/2019/3408315
Research Article
Synthesis, Antibacterial Evaluation, and QSAR of Caffeic
Acid Derivatives
1
2
1
3
Marianna O. Ara u´ jo, Hilzeth L. Freire Pessoa, Andressa B. Lira , Yunierkis P. Castillo,
1,4
and Damião P. de Sousa
1
Postgraduate Program in Natural and Synthetic Bioactive Products, Federal University of Para ´ı ba, João Pessoa, PB, Brazil
Department of Molecular Biology, Federal University of Para ´ı ba, João Pessoa, PB, Brazil
Escuela de Ciencias F ´ı sicas y Matem a´ ticas, Universidad de Las Am e´ ricas, Quito, Ecuador
Department of Pharmaceutical Sciences, Federal University of Para ´ı ba, João Pessoa, PB, Brazil
2
3
4
Correspondence should be addressed to Damião P. de Sousa; damiao_desousa@yahoo.com.br
Received 28 November 2018; Revised 10 January 2019; Accepted 27 January 2019; Published 12 February 2019
Academic Editor: Sevgi Kolaylı
Copyright © 2019 Marianna O. Ara u´ jo et al. ꢀis is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
ꢀ
e present study evaluates the antibacterial effects of a set of 16 synthesized caffeic acid ester derivatives against strains of
Staphylococcus aureus and Escherichia coli, as well as discusses their structure-activity relationship (SAR). ꢀe antibacterial assays
were performed using microdilution techniques in 96-well microplates to determine minimal inhibitory concentration (MIC).
ꢀe results revealed that five of the compounds present strong to optimum antibacterial effect. Of the sixteen ester derivatives
evaluated, the products with alkyl side chains, as propyl caffeate (3), butyl caffeate (6), and pentyl caffeate (7), presented the best
antibacterial activity with MIC values of around 0.20 μM against Escherichia coli and only butyl caffeate (6) showing the same MIC
against Staphylococcus aureus. For products with aryl substituents, the best MIC results against the tested strain of Escherichia coli
were 0.23 µM for (di-(4-chlorobenzyl)) caffeate (13) and 0.29 µM for diphenylmethyl caffeate (10) and all were less active against
the Staphylococcus aureus strain. Preliminary quantitative structure-activity relationship (QSAR) analyses confirmed that certain
structural characteristics, such as a median linear carbon chain and the presence of electron withdrawal substituents at the para
position of the aromatic ring, help potentiate antibacterial activity.
alternative; however, there are already case reports of in-
termediate [11] and full resistance to the drug [12].
Escherichia coli is active in the human intestinal tract [13];
1
. Introduction
Bacterial diseases are a global health problem and are prev-
alent in developing and low-income countries [1, 2], causing
it participates in vitamin K synthesis and in defense against
2
13 to 17 million deaths annually, equivalent to 25% of global
pathogenic bacteria [14]. Escherichia coli can cause severe
food poisoning and more severe conditions such as peritonitis
[15], meningitis, and urinary infections [16]. In E. coli strains,
production of extended-spectrum β-lactamases (ESBL) [17]
hydrolyzes most penicillins, cephalosporins, and mono-
bactams (aztreonam), the cephamycins and carbapenems
being among the few exceptions [18]. E. coli strains are re-
sistant to aminoglycosides, fluoroquinolones, tetracyclines,
chloramphenicol, and sulfamethoxazole-trimethoprim [19].
In the most serious infections involving ESBL, carbapenems
are indicated [20], but there are already reports in the lit-
erature of resistance to carbapenems [21, 22].
deaths and 45% for developing countries [3]. Due to the
bacterial resistance of Staphylococcus aureus to methicillin
and vancomycin [4] and Escherichia coli to β-lactams [5],
there is an urgent need for discovery and development of new
classes of antimicrobial compounds to treat bacterial in-
fections. Methicillin-resistant Staphylococcus aureus (MRSA)
is responsible for high hospital infection rates [6, 7]. ꢀese
infections range from simple acnes, boils, and abscesses to
more severe forms such as pneumonia, meningitis, endo-
carditis, and septicemia [8, 9]. β-Lactams are not always fully
effective against strains of MRSA [10]. Vancomycin is an

2
Journal of Chemistry
Many studies have investigated the bioactivity of caffeic
the presence of triethylamine (0.6 ml) and halide
(1.14 mmol) until complete reaction (44–120 hours), which
was verified by a single spot in TLC. ꢀe solvent was then
removed under reduced pressure, and the solution was
diluted with 20 ml of water. ꢀe product was extracted with
ethyl acetate (20 ml). ꢀe organic phase was treated with
water, dried over anhydrous sodium sulfate, filtered, and
concentrated under reduced pressure. ꢀe residue was
purified on a silica gel column (eluent: hexane-ethyl acetate
and an increasing polarity gradient) [45, 46].
acid derivatives finding that some of these analogues possess
remarkable activities, with potential as anti-inflammatory
[
[
23–27], antiviral [28], antiatherosclerotic [29], anti-HIV
30], antioxidant [31, 32], immunomodulatory [33], anti-
tumor [34–37], neuroprotective [38], antifungal [39], and
antibacterial agents [40]. Caffeic acid phenethyl ester
(
CAPE) or phenethyl caffeate is considered one of the most
important derivatives of caffeic acid and has several various
biological activities, such as antibacterial action [41–43]. It is
found in propolis extract and occurs in many plants [41].
Due to the problems associated with treatment of bacterial
infections, such as drug-resistant strains, various studies
have been conducted to discover new and safer antibacterial
agents. In the present study, synthesis and antibacterial
activity evaluations of 16 caffeic acid derivatives against two
representative strains Staphylococcus aureus (ATCC-25619)
and Escherichia coli (ATCC-2536) were performed.
3
.2.2. Synthesis of Compounds 11–14. A mixture of caffeic
acid (0.2 g, 1.11 mmol) and potassium carbonate (0.17 g,
.22 mmol) was dissolved in 5.5 mL of dimethylformamide
1
in the presence of halide (1.11 mmol). ꢀe mixture was
stirred at room temperature, until complete reaction (27–50
hours), which was verified by a single spot in TLC. ꢀe
solvent was then removed under reduced pressure, and the
solution was diluted with 15 ml of water. ꢀe product was
extracted with ethyl acetate (10 ml). ꢀe organic phase was
neutralized successively with 5% sodium bicarbonate
(10 ml), washed with brine (10 ml), dried over anhydrous
sodium sulfate, filtered, and concentrated under reduced
pressure. ꢀe residue was purified by using a chromato-
graphic column with hexane and ethyl acetate eluents in an
increasing polarity to isolate the ester derivatives [47].
2
. Chemicals
All of the chemical products used during synthesis were from
1
13
Sigma-Aldrich. H-NMR (200 and 50 MHz) and C-NMR
500 and 125 MHz) spectra were, respectively, recorded on
(
Varian Mercury and Varian-RMN-System spectrometers.
Chemical shifts (δ) are expressed in parts per million (ppm)
using TMS as an internal standard. Spin multiplicities are
given as s (singlet), brs (broad singlet), d (doublet), dd
(
doublet of doublets), t (triplet), q (quartet), quint (quintet),
3
.2.3. Synthesis of Compounds 15 and 16 for the Mitsunobu
sext (sextet), sept (septet), and m (multiplet). High-resolution
mass spectrometry was carried out using an Ultraflex II TOF/
TOF spectrometer with a high performance solid state laser
Reaction. A mixture of caffeic acid (0.1 g, 0.55 mmol) and
alcohol (0.55 mmol) was dissolved in 1.85 ml tetrahydro-
furan. ꢀe reaction mixture was stirred under magnetic
(
λ � 355 nm) and a reflector using the MALDI (matrix-
°
stirring at 0 C for about 30 minutes. Afterwards, di-
assisted laser desorption ionization) technique. Column ad-
sorption chromatography (CC) was performed on silica gel
isopropyl azodicarboxylate (0.11 ml, 0.55 mmol) and tri-
phenylphosphine (0.15 g, 0.55 mmol) were added as esteri-
fication agents, with continuous stirring at room
temperature for about 46–50 hours, which was verified by a
single spot in TLC. ꢀe solvent was then removed under
reduced pressure, and the solution was diluted with 10 ml of
water. ꢀe product was then extracted with ethyl acetate
(10 ml). ꢀe organic phase was neutralized successively with
saturated sodium bicarbonate, washed with brine, dried over
anhydrous sodium sulfate, filtered, and evaporated under
reduced pressure. ꢀe residue was then purified on a silica
gel column (eluent: hexane-ethyl acetate, 7 : 3) [48].
(
Merck 60, 230–400 mesh); analytical TLC was performed on
precoated silica gel plates (Merck 60 F ). FTIR spectra were
254
recorded on a Bruker FTIR spectrometer, Vertex 70 model,
using KBr pellets.
3
. Materials and Methods
3
.1. Synthesis of Compounds 1–8 for Fischer’s Esterification.
A mixture of caffeic acid (0.25 g, 1.39 mmol) and alcohol
50 ml) was heated under reflux in the presence of sulfuric
(
acid (0.4 ml) until completion of the reaction (5–21 hours)
and verified by a single spot in TLC. ꢀe alcohol was then
removed under reduced pressure, and the solution was
diluted with 20 ml of water. ꢀe product was extracted with
ethyl acetate (15 ml). ꢀe organic phase was neutralized
successively with 5% sodium bicarbonate and water, dried
over anhydrous sodium sulfate, and filtered. After evapo-
ration under reduced pressure, this phase yielded the ester
derivatives [44].
(
1) Methoxyethyl Caffeate (5). Brown liquid, 61.64% yield; IR
−1
υmax (KBr, cm ): 3319, 3071, 2945, 1689, 1600 and 1442,
1
1
272 and 1176; H-NMR (DMSO-d , 200 MHz): δH 3.27
6
(
3H; s), 3.56 (2H; t; J � 4.8 Hz), 4.22 (2H; t; J � 4.8 Hz), 6.28
(
1H; d; J � 16.0 Hz), 6.76 (1H; d; J � 8.0 Hz), 7.01 (1H; dd;
J � 8.0 Hz, 2.0 Hz), 7.06 (1H; d; J � 2.0 Hz), 7.47 (1H; d;
13
J � 16.0 Hz); C-NMR (DMSO-d , 50 MHz): δC 58.2, 63.0,
6
6
1
[
9.9, 113.8, 114.9, 115.8, 121.6, 125.5, 145.5, 145.6, 148.5, and
66.7 [49]; HRMS (MALDI) calculated for C H O
14 5
1
2
+
M+H] : 239.0919, found 239.1599.
3.2. ReactionwithAlkylandArylHalidesforCompounds9–14
3
.2.1. Synthesis of Compounds 9 and 10. Caffeic acid (0.2 g,
(2) Diphenylmethyl Caffeate (10). Brown liquid, 27.29%
−
1
1
.11 mmol) in acetone (13.5 ml) was heated under reflux in
yield; IR υmax (KBr, cm ): 3370, 3059, 2970, 1702, 1606 and

Journal of Chemistry
3
1
1
6
7
7
448, 1271 and 1149; H-NMR (DMSO-d , 200 MHz): δH
(ATCC-2536). ꢀe strains belonged to the Bacterioteca of
the Laboratory of Toxicological Tests, Institute of Research
in Drugs and Medications, Para ´ı ba Federal University. ꢀe
strains were maintained in an appropriate culture medium,
6
.43 (1H; d; J � 16.0 Hz), 6.77 (1H; d; J � 8.0 Hz), 6.90 (1H; s),
.06 (1H; dd; J � 8.0 Hz, 2.0 Hz), 7.10 (1H; d; J � 2.0 Hz),
13
.47–7.28 (10H; m), 7.59 (1H; d; J � 16.0 Hz), [44]; C-NMR
°
(
DMSO-d , 50 MHz): δC 76.1, 113.7, 115.0 115.8, 121.7,
BHI (Difco Laboratories/France/USA), and stored at 4 C
6
°
1
1
[
25.5, 126.6, 127.8, 128.6, 140.8, 145.6, 146.0, 148.6, and
65.6 [50]; HRMS (MALDI) calculated for C H O
18 4
and 35 C. ꢀe microorganism suspension was prepared
according to McFarland tube 0.5, corresponding to ap-
22
+
8
−1
M+Na] : 369.1103, found 369.1168.
proximately 10 CFU·ml [54, 55].
(
3) 4-Chlorobenzyl Caffeate (11). Yellow amorphous solid,
Culture Medium. ꢀe antifungal activity assays were per-
formed in brain heart infusion (BHI) broth liquid medium
(Difco Laboratories/France/USA), which was prepared and
used according to the manufacturer’s instructions.
−
1
2
1
2
6.43% yield; IR υmax (KBr, cm ): 3338, 3053, 2964, 1677,
1
600 and 1435, 1258 and 1175, 1011; H-NMR (DMSO-d ,
6
00 MHz): δH 5.18 (2H; s), 6.33 (1H; d; J � 16.0 Hz), 6.76
(
1H; d; J � 8.0 Hz), 7.02 (1H; dd; J � 8.0 Hz, 2.0 Hz), 7.07 (1H;
Determination of the Minimum Inhibitory Concentration
(MIC). ꢀe MIC values were determined by the micro-
dilution method [54] using 96-well “U”-shaped microtiter
plates in duplicate. In each plate well, 100 μl of twice con-
centrated BHI liquid medium was added, except the first line
which received 160 μl. ꢀen, 40 μl of product solution (also
doubly concentrated) was placed in the first row of the plate
wells. By serial dilution (ratio of two), concentrations from
200–12.5 μg/ml were obtained, such that in the first line of
the plate was the highest concentration and in the latter
lower concentration. Finally, another 100 μl of the BHI
medium was added to all wells, and 10 μl of inoculum was
added to the wells in each plate column that referred spe-
cifically to a strain. ꢀe same was also done in the culture
medium with the bacterial drug chloramphenicol (100 mg/
13
d; J � 2.0 Hz), 7.44 (4H; brs), 7.53 (1H; d; J � 16.0 Hz); C-
NMR (DMSO-d , 50 MHz): δC 64.5, 113.6, 114.9, 115.8,
6
1
21.6, 125.5, 128.5, 129.9, 132.6, 135.5, 145.6, 145.8, 148.5,
and 166.3 [51]; HRMS (MALDI) calculated for C H ClO
16
13
4
+
[
M] : 304.0502, found 304.3855.
(
4) (Di-(4-chlorobenzyl)) Caffeate (13). White amorphous
−
1
solid, 11.82% yield; IR υmax (KBr, cm ): 3332, 3059, 2976,
1
1
695, 1606 and 1442, 1277 and 1169, 1011; H-NMR (DMSO-
d , 200 MHz): δH 5.16 (2H; s), 5.19 (2H; s), 6.41 (1H; d;
6
J � 16.0 Hz), 7.00 (1H; d; J � 8.4 Hz); 7.10 (1H; d; J � 2.0 Hz);
13
7
.14 (1H; s), 7.49–7.43 (8H; m), 7.56 (1H; d; J � 16.0 Hz); C-
NMR (DMSO-d , 50 MHz): δC 64.6, 68.9, 113.7, 114.8, 115.0,
6
1
1
21.2, 127.2, 128.4, 128.5, 129.5, 129.9, 132.4, 132.7, 135.4,
°
36.0, 145.2, 147.0, 148.8, and 166.3 [51, 52]; HRMS (MALDI)
ml). ꢀe plates were incubated at 37 C for 24 h for further
+
calculated for C H ClO [M+H] : 429.0660, found 429.0667.
reading; this was performed with the addition of 10 μl of a
16
13
4
0
.01% (w/v) solution of resazurin (Sigma), a colorimetric
(
5) (Di-(4-methoxybenzyl)) Caffeate (14). Orange amorphous
metabolic activity indicator. For each strain, the MIC was
defined as the lowest concentration capable of inhibiting
bacterial growth in the wells, as visually observed compared
with the control. All of the tests were performed in duplicate,
and the results were expressed as a geometric mean of the
MIC values obtained from both tests [56].
−
1
solid, 36.98% yield; IR υmax (KBr, cm ): 3344, 3059, 2831,
1
1
2
702, 1620 and 1435, 1188 and 1112; H-NMR (DMSO-d ,
6
00 MHz): δH 3.74 (3H; s), 3.74 (3H; s), 5.07 (2H; s); 5.11 (2H;
s), 6.36 (1H; d; J � 15.8 Hz), 6.94–6.86 (5H; m), 7.02 (1H; dd,
J � 8.0 Hz, 2.0 Hz), 7.10 (1H; s), 7.22 (2H; d; J � 8.6 Hz), 7.35
13
(
2H; d; J � 8.6 Hz), 7.51 (1H; d; J � 16.0 Hz); C-NMR (DMSO-
d , 50 MHz): δC 55.1, 55.2, 65.4, 69.6, 113.6, 113.8, 113.9, 114.6,
6
5. 3D-QSAR Modeling
1
1
15.1, 121.3, 127.1, 128.4, 128.9, 129.8, 130.3, 144.9, 147.2, 149.2,
59.1, 159.3, and 166.5 [52, 53]; HRMS (MALDI) calculated for
Otherwise noted, default parameters were employed for all
software. One initial 3D conformer per compound was
generated with OpenEye Omega [57]. Each compound’s
initial conformation was subjected to 200 cycles (100 ps
each) of quenched molecular dynamics simulations with
Open3DAlign [58] using an implicit solvent model. Mole-
cules alignment was also performed with Open3DAlign by
means of its mixed algorithm considering multiple con-
formers for the template and candidate compounds.
+
C H O [M] : 420.1573, found 420.0234.
25 24 6
(
6) 4-Methylbenzyl Caffeate (16). Yellow amorphous solid,
−
1
5
1
2
0.95% yield; IR υmax (KBr, cm ): 3332, 3027, 2976, 1677,
1
600 and 1435, 1271 and 1163; H-NMR (DMSO-d ,
6
00 MHz): δH 2.29 (3H; s), 5.13 (2H; s), 6.30 (1H; d;
J � 16.0 Hz), 6.75 (1H; d; J � 8.0 Hz), 7.00 (1H; dd; J � 8.2 Hz,
2
.0 Hz), 7.06 (1H; d; J � 2.0 Hz), 7.18 (2H; d; J � 8.0 Hz), 7.29
13
(
2H; d; J � 8.0 Hz), 7.50 (1H; d; J � 16.0 Hz); C-NMR
Bioactivities were converted to pIC50 values (pIC50
�
(
DMSO-d , 50 MHz): δC 21.9, 65.3, 113.8, 114.9, 115.8,
−log IC ), and QSAR models were separately trained for
6
10
50
1
21.6, 125.5, 128.3, 129.1, 133.5, 137.4, 145.5, 145.6, 148.6,
Escherichia coli and Staphylococcus aureus using Open3-
˚
and 166.5 [50]; HRMS (MALDI) calculated for C H O
16 4
DQSAR [59]. ꢀe box step for the grid was set to 0.5 A, and
17
+
[
M] : 284.1049, found 284.4357 (Scheme 1).
van der Waals (VDW) and electrostatic molecular in-
teraction fields (MIFs) were computed. ꢀe dielectric con-
stant was set to 4. Grid points providing VDW values larger
4
. Antibacterial Assay
4
than 10 kcal/mol were removed from both MIFs. Values of
ꢀ
e
microbiological testing used single strains of
Staphylococcus aureus (ATCC-25619) and Escherichia coli
the grid exceeding 30 kcal/mol or below −30 kcal/mol were
set to 30 kcal/mol and −30 kcal/mol, respectively, for both

4
Journal of Chemistry
O
O
R
O
O
[9]
HO
O
HO
b
OH
OH
1] R = CH3
2] R = CH CH
a
O
b
[
[
[
[
[
[
[
[
[10]
O
2
3
HO
3] R = CH CH CH
2
2
3
OH
OH
O
4] R = CH[CH3]2
5] R = CH CH OCH
3
2
2
c
6] R = CH CH CH CH
HO
O
O
2
2
2
3
7] R = CH CH CH CH CH
2
2
2
2
3
OH
[
[
11] R = 4-Cl
12] R = 4-OCH
8] R = CH CH CH[CH
2
2
3]2
HO
3
R
d
c
OH
O
O
O
[
[
15] R = 4-H
16] R = 4-CH3
O
HO
[13] R = R = 4-Cl
1 2
R1
R R2
[
14] R = R = 4-OCH
1 2 3
OH
OH
Scheme 1: Synthesis of the caffeic acid derivatives: (a) ROH, H SO , reflux; (b) Et N, RX, acetone, reflux; (c) DMF, K CO , RX, room
2
4
3
2
3
°
temperature; (d) ROH, THF, TPP, DIAD, 0 C to room temperature.
MIFs. In addition, all grid points with an absolute value
lower than 0.05 were set to zero, and variables with standard
deviation lower than 0.1 were excluded from the calcula-
tions. Variables spanning up to four levels were also ex-
cluded. After these variables filtering steps, those remaining
in the dataset were scaled according to the BUW algorithm
as implemented in Open3DQSAR.
(di-(4-chlorobenzyl)) caffeate (13), (di-(4-methoxybenzyl))
caffeate (14), benzyl caffeate (15), and 4-methylbenzyl caffeate
(16) each with differing substitutions on the ester side chain
(R ) and the hydroxyl of the para position of the caffeic ring
1
(R ) were tested. ꢀe results were expressed as minimal in-
2
hibitory concentration (MIC) in μM. ꢀe structure-activity
relationship (SAR) study verified which chemical structures
influenced the antibacterial activity. In addition, a quantita-
tive structure-activity relationship (QSAR) was performed to
verify contributions of steric and electrostatic factors towards
the Escherichia coli and Staphylococcus aureus activity.
ꢀe results obtained in this study revealed that five
compounds presented strong to optimum antibacterial ac-
tivity; the compounds were found to be more active against
the Escherichia coli strain. Of the sixteen ester derivatives
evaluated, propyl caffeate (3), butyl caffeate (6), and pentyl
caffeate (7) were found to exhibit the best antibacterial activity
of the products with alkyl side chains, with an MIC value of
0.2 μM against the Escherichia coli strain and only butyl
caffeate (6) showed the same MIC for the Staphylococcus
aureus strain, whereas of the products with aryl side chains,
the best MIC results against the Escherichia coli strain were
We considered up to 10 principal components (PCs) for
models training. ꢀe optimal number of PCs considered in
2
the final model was selected according to the r value of the
leave-one-out (LOO) cross validation of the model (q2 ).
LOO
With this optimal number of PCs, variables were further
grouped using the Smart Region Definition procedure
implemented in Open3DQSAR. ꢀe grouped variables were
subjected to a selection procedure according to the fractional
factorial design using leave-many-out (LMO) cross valida-
tion with 20 runs and 5 groups. Only selected variables in the
previous step remained on the dataset. Finally, given the
limited number of available samples, the PLS model was
recomputed and its quality was evaluated in 100 cycles of
LMO cross validation.
0
.23 µM for (di-(4-chlorobenzyl)) caffeate (13) and 0.29 µM
6
. Results and Discussion
for diphenylmethyl caffeate (10), both being less active against
Staphylococcus aureus. Table 1 summarizes the in vitro sus-
ceptibilities of the two Escherichia coli and Staphylococcus
aureus strains against all of the test compounds.
6
.1. Antibacterial Activity and Structure-Activity Relationship
(
SAR). In the current work, a set of sixteen caffeic acid ester
derivatives were evaluated for their in vitro antibacterial
activity against strains of Staphylococcus aureus and
Escherichia coli. Methyl caffeate (1), ethyl caffeate (2), propyl
caffeate (3), isopropyl caffeate (4), methoxy-ethyl caffeate
Of the screened ester derivatives, certain structural
features were observed that may increase antibacterial ac-
tivity, giving information on functional groups that might be
important to antibacterial effect, such as alkyl ester side
chain, especially a median linear carbon chain, benzyl
radicals with two aromatic rings without substituents, and
(
(
5), butyl caffeate (6), pentyl caffeate (7), isopentyl caffeate
8), decyl caffeate (9), diphenylmethyl caffeate (10), 4-
chlorobenzyl caffeate (11), 4-methoxybenzyl caffeate (12),

Journal of Chemistry
5
Table 1: Minimal inhibitory concentration (MIC) values (expressed
in µM) of the compounds 1–16.
diphenylmethyl caffeate (10; diphenylmethyl group); this
was compared to benzyl caffeate (15; benzyl group), which
presented a better bond to its target.
MIC (µM)
ꢀ
e antimicrobial activity of caffeic acid derivatives has
Compounds
Staphylococcus aureus
Escherichia coli
(ATCC-2536)
been reported in other studies [43, 60–65]. For example,
Merkl et al. [66] investigated the antibacterial activity of a
series of caffeic acid esters (methyl, ethyl, propyl, and butyl
caffeate) against E. coli DMF 7503. All of the esters screened
showed potent antibacterial activity against the test strain,
principally propyl and butyl caffeate, with MIC values of
5.00 mM. ꢀe MIC values of the other caffeic acid esters
tested were roughly 10.00 mM [67]. In the study of Meyuhas
et al. [42], the phenethyl caffeate exhibited antibacterial
activity against S. aureus (48 mM) and E. coli (254 mM). ꢀis
compound has structural similarity to ester 15, which
showed strong antibacterial action (Table 1). ꢀe ester de-
rivatives of caffeic acid were prepared using short linear
carbon chain radicals, such as methyl and ethyl caffeate, and
from them, compounds synthesized with (4-chloro-benzyl)
radicals on the two hydroxyls of the meta and para positions
of the caffeic ring. ꢀe antibacterial activities of the com-
pounds were evaluated against E. coli and S. aureus; the
esters, methyl and ethyl caffeates, showed activity for both
strains, while their by-products were only active for the E.
coli strain. Derivatives thus prepared are promising candi-
dates for treating bacterial pathologies, and their test results
corroborate those already published in the literature.
(
ATCC-25619)
1
2
3
4
5
6
7
8
9
1
1
1
1
1
1
1
0.52
0.48
0.45
0.45
0.84
0.21
0.40
0.80
0.62
0.58
1.31
0.67
0.93
0.95
0.74
1.41
0.31
0.52
0.48
0.22
0.45
0.84
0.21
0.20
0.40
0.62
0.29
1.31
0.33
0.23
0.48
0.37
0.70
0.31
0
1
2
3
4
5
6
Chloramphenicol
electron-withdrawal substituents in the para position such
as chloro.
Data analyses revealed that introduction of alkyl sub-
stituents as short chains and the presence of a heteroatom
and long (10 carbon atoms) or branching chains resulted in
decreased antibacterial effect. For example, butyl caffeate (6:
butyl group) was found to be the most bioactive compound
for both strains (MIC � 0.20 μM), whereas methyl caffeate (1;
methyl group), ethyl caffeate (2; ethyl group), isopropyl
caffeate (4; isopropyl group), methoxyethyl caffeate (5;
methoxyethyl group), isopentyl caffeate (8; isopentyl group),
and decyl caffeate (9; decyl group) presented increased MICs
6
.2. Quantitative Structure-Activity Relationship (QSAR)
Studies. ꢀe statistical parameters of the 3D-QSAR models
obtained for the E. coli and S. aureus inhibitory activities are
summarized in Table 2. As can be seen, both models have
good statistical parameters. Furthermore, for both E. coli and
S. aureus, bioactivity is mainly sterically driven. Also, given
the lower number of PCs included in the S. aureus model
and its statistical parameters, it can be considered statisti-
cally more robust.
(
decreased activities). ꢀis reduction in antibacterial activity
probably occurs due to steric hindrance caused by the
presence of bulky groups and also poorly lipophilic short-
chain groups. Activity achieved a maximum for the esters
with a median linear carbon chain conferring lipophilicity to
the molecules.
6
.2.1. Scaffold. Bioactivity is mostly steric-guided. Models
show 77% and 73% contributions of the steric factor to the
Escherichia coli and Staphylococcus aureus activities, re-
spectively (Figure 1).
ꢀ
e compounds presenting aryl radicals resulted in
decreased antibacterial effect, such as aromatic ring sub-
stituents with electron donor groups in the para position
such as methyl and methoxy or with electron withdrawal
groups such as chloro (except compound (13) which was
active against E. coli). ꢀose without substituents also
resulted in decreased antibacterial effect, except compound
6.2.2. Electrostatic Molecular Interaction Field. ꢀe elec-
trostatic contribution to bioactivity shows that certain
substitutions at R can improve bioactivity. In general, the
2
(10) which was active against E. coli. ꢀe presence of an
electronegative molecular interaction field next to the O-R
2
electron-withdrawing group such as di-(4-chlorobenzyl)
caffeate (13; di-(4-chlorobenzyl) group) provided more
activity against the Escherichia coli strain, which probably
occurred due to the para chloro substituent in the caffeic
ring, which was the only difference compared to 4-chlor-
obenzyl caffeate (11; chlorobenzyl group), and presented
similarity to the reference drug having two chloro sub-
stituents. Compound (10) presented good results against
Escherichia coli due to the presence of bulky groups without
substituents with two aromatic rings, such as with
bond indicates a positive contribution to bioactivity of
substitutions at this position. ꢀis positive contribution
cannot be exploited when R � H. ꢀis later observation
2
holds for both Escherichia coli and Staphylococcus aureus.
Figures 2 and 3 summarize the electrostatic contribution to
bioactivity in both strains.
ꢀe comparisons of compounds 11 and 13 and of
compounds 12 and 14 reveal some interesting details. In the
case of Escherichia coli, there is an electronegative molecular
interaction field close to the 4-chlorobenzyl substitution at

6
Journal of Chemistry
Table 2: Statistical parameters of the 3D-QSAR models for the
antibacterial activity against E. coli and S. aureus.
E. coli
4
S. aureus
a
No. PCs
2
2b
R
0.95
0.70
0.68
77%
23%
0.86
0.76
0.74
73%
27%
c
2
c
q
q
LOO
2
d
LMO
(
a)
e
VDW contrib.
Ele. contrib.
f
a
Number of PCs in the final model. bCoefficient of determination. LOO
d
cross-validation performance.
LMO cross-validation performance.
e, f
Contribution of VDW and electrostatic factors to bioactivity, respectively.
O
(
b)
R1
O
Figure 3: Electrostatic molecular interaction field for compounds
1 and 13 in Staphylococcus aureus.
1
R2
O
favorable region around R in both E. coli and S. aureus
1
OH
models. Compounds with small aliphatic substituents at R
1
are favored, while too bulky or too long substituents such as
long aliphatic chains and aromatic substitutions are dis-
favored at this position. ꢀis effect is more pronounced in
the Staphylococcus aureus model where there is a large
unfavorable steric region close to R . For example, the R
Figure 1
1
1
substituent of compound 6 completely falls into the steric
favorable region defined by both strains models. On the
other hand, the 4-methylbenzyl substituent of compound 16
falls beyond the steric favorable region in Escherichia coli
and in the steric unfavorable region defined by the Stha-
phylococcus aureus model. In addition, bulky substitutions at
(
a)
R (such as those present in compound 14) decrease anti-E.
2
coli bioactivity. ꢀe influence of the steric factor in bio-
activity is summarized in Figures 4 and 5.
Considering all these observations, for dual Escherichia
coli and Staphylococcus aureus bioactivity, small aliphatic
substitutions are essential at R . Furthermore, basing on the
1
(
b)
available data, substitutions at R could be beneficial for dual
2
bioactivity. ꢀis last hypothesis should be confirmed by
testing more compounds substituted at R2 with small
groups.
Figure 2: Electrostatic molecular interaction field for compounds
1 and 13 in Escherichia coli.
1
Our QSAR studies confirmed the biological activity of
the tested compounds, connecting with chemical structures
for (di-(4-chlorobenzyl)) caffeate (13), through the elec-
R , indicating that benzyl groups with electronegative para-
2
substitutions can have an additional positive contribution to
bioactivity against this strain. In contrast, substitutions at R2
such as 4-methoxybenzyl present in compound 14 are
tronegative electrostatic molecular interaction field at R as
2
against Escherichia coli and for butyl caffeate (6), due to the
small aliphatic radical completely falling into the sterically
favorable region as defined by both strain models. In con-
trast to these, 4-methylbenzyl caffeate (16) exceeded the
sterically favorable regions of these models and presented
the worst bacterial activity against the tested strains.
unable to increase the bioactivity of R -substituted com-
2
pounds. ꢀis can be clearly exemplified for compounds 12
and 14.
ꢀ
e lack of any additional electrostatic molecular in-
teraction field surrounding R in the Staphylococcus aureus
2
model indicates that no increase in the anti-S. aureus bio-
activity could be obtained with specific substitutions at R .
2
7. Conclusion
ꢀ
e present study investigated the antibacterial activity
6
.2.3. Steric Molecular Interaction Field. ꢀe analysis of the
of sixteen caffeic acid ester derivatives against a Gram-
steric molecular interaction field shows that there is a steric
positive Staphylococcus aureus strain and a Gram-negative

Journal of Chemistry
7
should be performed to optimize antimicrobial activity in
this class of compounds for eventual synthesis of new an-
tibacterial agents.
Data Availability
(
a)
ꢀ
e data used to support the findings of this study have been
deposited in the Federal University of Para ´ı ba repository at
https://repositorio.ufpb.br/jspui/handle/tede/9074.
Conflicts of Interest
ꢀ
e authors declare that they have no conflicts of interest.
(
b)
Figure 4: Steric contribution to Escherichia coli bioactivity for
compounds 6 and 16.
Acknowledgments
ꢀ
is work was supported by the Brazilian agencies: Conselho
Nacional de Desenvolvimento Cient ´ı fico e Tecnol o´ gico
(CNPq) and Coordenação de Aperfeiçoamento de Pessoal de
N ´ı vel Superior (CAPES).
Supplementary Materials
Spectroscopy data of known compounds. 13C-NMR and H-
NMR spectra of the unpublished compounds: methoxyethyl
caffeate, diphenylmethyl caffeate, 4-chlorobenzyl caffeate, (di-
1
(
4-chlorobenzyl)) caffeate, (di-(4-methoxybenzyl)) caffeate,
and 4-methylbenzyl caffeate. (Supplementary Materials)
References
(
a)
[1] I. C. Bygbjerg, “Double burden of noncommunicable and
infectious diseases in developing countries,” Science, vol. 337,
no. 6101, pp. 1499–1501, 2012.
2] R. J. Coker, B. M. Hunter, J. W. Rudge, M. Liverani, and
P. Hanvoravongchai, “Emerging infectious diseases in
southeast Asia: regional challenges to control,” @e Lancet,
vol. 377, no. 9765, pp. 599–609, 2011.
[
[
[
3] L. Wannmacher, “Uso indiscriminado de antibi o´ ticos e
resist eˆ ncia microbiana: uma guerra perdida?,” Uso racional de
medicamentos:temas selecionados, vol. 1, no. 4, 2004.
4] A. Breves, C. A. C. Miranda, C. Flores, I. d. Filippis, and
M. M. Clementino, “Methicillin- and vancomycin-
resistantStaphylococcus aureusin health care workers and
medical devices,” Jornal Brasileiro de Patologia e Medicina
Laboratorial, vol. 51, no. 3, pp. 143–152, 2015.
(
b)
Figure 5: Steric contribution to Staphylococcus aureus bioactivity
for compounds 6 and 16.
[
5] S. Shaikh, J. Fatima, S. Shakil, S. M. D. Rizvi, and M. A. Kamal,
“Antibiotic resistance and extended spectrum beta-
lactamases: types, epidemiology and treatment,” Saudi Jour-
nal of Biological Sciences, vol. 22, no. 1, pp. 90–101, 2015.
Escherichia coli strain. Based on the findings of the present
[6] A. I. Fattom, G. Horwith, S. Fuller, M. Propst, and R. Naso,
“Development of StaphVAX, a polysaccharide conjugate
vaccine against S. aureus infection: from the lab bench to
phase III clinical trials,” Vaccine, vol. 22, no. 7, pp. 880–887,
investigation, we conclude that the propyl, butyl, pentyl (di-
(
4-chlorobenzyl)), and diphenylmethyl caffeate derivatives
present the highest antibacterial activity against the
Escherichia coli strain; only butyl caffeate was active against
the Staphylococcus aureus strain. Further, the QSAR study
2004.
[
7] E. A. Palombo and S. J. Semple, “Antibacterial activity of
Australian plant extracts against methicillin-resistant Staph-
ylococcus aureus (MRSA) and vancomycin-resistant entero-
cocci (VRE),” Journal of Basic Microbiology, vol. 42, no. 6,
pp. 444–448, 2002.
demonstrated that small aliphatic substituents in R (butyl
1
caffeate) and substituents in the aromatic ring, with electron
withdrawal groups in the para position in R ((di-(4-
2
chlorobenzyl)) caffeate), result in bioactive compounds. ꢀis
preliminary study revealed that certain structural features
are important for antibacterial action. Additional studies
[
8] S. Das, C. Lindemann, B. C. Young et al., “Natural mutations
in aStaphylococcus aureusvirulence regulator attenuate cy-
totoxicity but permit bacteremia and abscess formation,” in

8
Journal of Chemistry
Proceedings of the National Academy of Sciences, vol. 113,
no. 22, pp. E3101–E3110, 2016.
9] J. A. Lindsay and M. T. G. Holden, “Staphylococcus aureus:
superbug, super genome?,” Trends in Microbiology, vol. 12,
no. 8, pp. 378–385, 2004.
10] L. C. Gelatti, R. R. Bonamigo, A. P. Becker, and
P. A. d’Azevedo, “Staphylococcus aureus resistentes a` metic-
ilina: disseminação emergente na comunidade,” Anais Bra-
sileiros de Dermatologia, vol. 84, no. 5, pp. 501–506, 2009.
11] G. A. Oliveira, A. M. Dell’Aquila, R. L. Masiero et al., “Iso-
lation in Brazil of nosocomial Staphylococcus aureus with
reduced susceptibility to vancomycin,” Infection Control &
Hospital Epidemiology, vol. 22, no. 7, pp. 443–448, 2015.
[25] Y.-J. Lee, P.-H. Liao, W.-K. Chen, and C.-C. Yang, “Prefer-
ential cytotoxicity of caffeic acid phenethyl ester analogues on
oral cancer cells,” Cancer Letters, vol. 153, no. 1-2, pp. 51–56,
2000.
[26] Z. Orban, N. Mitsiades, T. R. Burke, M. Tsokos, and
G. P. Chrousos, “Caffeic acid phenethyl ester induces leu-
kocyte apoptosis, modulates nuclear factor-kappa B and
suppresses acute inflammation,” Neuroimmunomodulation,
vol. 7, no. 2, pp. 99–105, 2000.
[
[
[
[27] T. Toyoda, T. Tsukamoto, S. Takasu et al., “Anti-inflammatory
effects of caffeic acid phenethyl ester (CAPE), a nuclear factor-
κB inhibitor, onHelicobacter pylori-induced gastritis in
Mongolian gerbils,” International Journal of Cancer, vol. 125,
no. 8, pp. 1786–1795, 2009.
[
12] S. Chang, D. M. Sievert, J. C. Hageman et al., “Infection with
vancomycin-ResistantStaphylococcus aureusContaining the-
vanAResistance Gene,” New England Journal of Medicine,
vol. 348, no. 14, pp. 1342–1347, 2003.
[28] M. R. Fesen, K. W. Kohn, F. Leteurtre, and Y. Pommier,
“Inhibitors of human immunodeficiency virus integrase,” in
Proceedings of the National Academy of Sciences, vol. 90, no. 6,
pp. 2399–2403, 1993.
[
13] M. Rajilic-Stojanovic, H. Smidt, and W. M. De Vos, “Diversity
of the human gastrointestinal tract microbiota revisited,”
Environmental Microbiology, vol. 9, no. 9, pp. 2125–2136,
[29] K. Natarajan, S. Singh, T. R. Burke, D. Grunberger, and
B. B. Aggarwal, “Caffeic acid phenethyl ester is a potent and
specific inhibitor of activation of nuclear transcription factor
NF-kappa B,” in Proceedings of the National Academy of
Sciences, vol. 93, no. 17, pp. 9090–9095, 1996.
[30] C.-n. Xia, H.-b. Li, F. liu, and W.-x. Hu, “Synthesis of trans-
caffeate analogues and their bioactivities against HIV-1
integrase and cancer cell lines,” Bioorganic & Medicinal
Chemistry Letters, vol. 18, no. 24, pp. 6553–6557, 2008.
[31] A. Russo, R. Longo, and A. Vanella, “Antioxidant activity of
propolis: role of caffeic acid phenethyl ester and galangin,”
Fitoterapia, vol. 73, no. 1, pp. S21–S29, 2002.
[32] H. Yang, Y. Dong, H. Du, H. Shi, Y. Peng, and X. Li, “An-
tioxidant compounds from propolis collected in Anhui,
China,” Molecules, vol. 16, no. 4, pp. 3444–3455, 2011.
[33] J. H. Park, J. K. Lee, H. S. Kim et al., “Immunomodulatory
effect of caffeic acid phenethyl ester in Balb/c mice,” In-
ternational Immunopharmacology, vol. 4, no. 3, pp. 429–436,
2004.
[34] Y.-J. Chen, M.-S. Shiao, M.-L. Hsu, T.-H. Tsai, and
S.-Y. Wang, “Effect of caffeic acid phenethyl ester, an anti-
oxidant from propolis, on inducing apoptosis in human
leukemic HL-60 cells,” Journal of Agricultural and Food
Chemistry, vol. 49, no. 11, pp. 5615–5619, 2001.
[35] C.-P. Chuu, H.-P. Lin, M. F. Ciaccio et al., “Caffeic acid
phenethyl ester suppresses the proliferation of human pros-
tate cancer cells through inhibition of p70S6K and Akt sig-
naling networks,” Cancer Prevention Research, vol. 5, no. 5,
pp. 788–797, 2012.
[36] H. P. Lin, S. S. Jiang, and C. P. Chuu, “Caffeic acid phenethyl
ester causes p21(Cip1) induction, akt signaling reduction, and
growth inhibition in PC-3 human prostate cancer cells,” PLoS
One, vol. 7, no. 2, pp. 1–12, 2012.
[37] T. Nagaoka, A. H. Banskota, Y. Tezuka, I. Saiki, and S. Kadota,
“Selective antiproliferative activity of caffeic acid phenethyl
ester analogues on highly liver-metastatic murine colon 26-L5
carcinoma cell line,” Bioorganic & Medicinal Chemistry,
vol. 10, no. 10, pp. 3351–3359, 2002.
[38] A. Ilhan, M. Iraz, A. Gurel, F. Armutcu, and O. Akyol, “Caffeic
acid phenethyl ester exerts a neuroprotective effect on CNS
against pentylenetetrazol-induced seizures in mice,” Neuro-
chemical Research, vol. 29, no. 12, pp. 2287–2292, 2004.
[39] J. Breger, B. B. Fuchs, G. Aperis, T. I. Moy, F. M. Ausubel, and
E. Mylonakis, “Antifungal chemical compounds identified
using a C. elegans pathogenicity assay,” PLoS Pathogens, vol. 3,
no. 2/e18, pp. 168–178, 2007.
2
007.
[
14] S. Hudault, J. Guignot, and A. L. Servin, “Escherichia coli
strains colonising the gastrointestinal tract protect germfree
mice against Salmonella typhimurium infection,” Gut, vol. 49,
no. 1, pp. 47–55, 2001.
[
15] A. Sabr a´ , “ECEP, ECET, ECEA, ECEH, ECEI, ECAD: a E. coli
revisitada no contexto da diarr e´ ia aguda,” Jornal de Pediatria,
vol. 77, no. 1, pp. 5–7, 2002.
16] J. B. Kaper, J. P. Nataro, and H. L. T. Mobley, “Pathogenic
Escherichia coli.,” Nature Reviews Microbiology, vol. 2, no. 2,
pp. 123–140, 2004.
17] K. Bush and G. A. Jacoby, “Updated functional classification
of -lactamases,” Antimicrobial Agents and Chemotherapy,
vol. 54, no. 3, pp. 969–976, 2009.
18] D. M. Livermore, “Defining an extended-spectrum β-lacta-
mase,” Clinical Microbiology and Infection, vol. 14, no. 1,
pp. 3–10, 2008.
19] D. L. Paterson, “Recommendation for treatment of severe
infections caused by Enterobacteriaceae producing extended-
spectrum β-lactamases (ESBLs),” Clinical Microbiology and
Infection, vol. 6, no. 9, pp. 460–463, 2000.
20] D. L. Paterson, W.-C. Ko, A. Von Gottberg et al., “Antibiotic
therapy for Klebsiella pneumoniae bacteremia: implications of
production of extended-spectrum -lactamases,” Clinical In-
fectious Diseases, vol. 39, no. 1, pp. 31–37, 2004.
[
[
[
[
[
[
21] M. Gonzalez-Padilla, J. Torre-Cisneros, F. Rivera-Espinar
et al., “Gentamicin therapy for sepsis due to carbapenem-
resistant and colistin-resistant Klebsiella pneumoniae,” Jour-
nal of Antimicrobial Chemotherapy, vol. 70, no. 3, pp. 905–
913, 2014.
[
22] R. Moayednia, D. Shokri, S. Mobasherizadeh, A. Baradaran,
S. M. Fatemi, and A. Merrikhi, “Frequency assessment of
β-lactamase enzymes in Escherichia coli and Klebsiella isolates
in patients with urinary tract infection,” Journal of Research in
Medical Sciences, vol. 1, no. 19, pp. S41–S45, 2014.
[
[
23] F. M. da Cunha, D. Duma, J. Assreuy et al., “Caffeic acid
derivatives: in vitro and in vivo anti-inflammatory proper-
ties,” Free Radical Research, vol. 38, no. 11, pp. 1241–1253,
2
009.
24] S. Juman, N. Yasui, K. Ikeda et al., “Caffeic acid phenethyl
ester suppresses the production of pro-inflammatory cyto-
kines in hypertrophic adipocytes through lipopolysaccharide-
stimulated macrophages,” Biological and Pharmaceutical
Bulletin, vol. 35, no. 11, pp. 1941–1946, 2012.

Journal of Chemistry
9
[
40] A. Kujumgiev, V. Bankova, A. Ignatova, and S. Popov,
[56] P. Gerhardt, R. G. E. Murray, W. A. Wood, and N. R. Krieg,
Methods for General and Molecular Bacteriology, American
Society for Microbiology, Washington, USA, 1994.
[57] P. C. D. Hawkins, A. G. Skillman, G. L. Warren,
B. A. Ellingson, and M. T. Stahl, OMEGA, “Santa Fe, NM:
OpenEye Scientific Software”, 1997, http://www.eyesopen.
com.
[58] P. Tosco, T. Balle, and F. Shiri, “Open3DALIGN: an open-
source software aimed at unsupervised ligand alignment,”
Journal of Computer-Aided Molecular Design, vol. 25, no. 777,
2011.
“
Antibacterial activity of propolis, some of its components
and their analogs,” Pharmazie, vol. 48, no. 10, pp. 785–6, 1993.
41] G. Murtaza, S. Karim, M. R. Akram et al., “Caffeic acid
phenethyl ester and therapeutic potentials,” BioMed Research
International, vol. 2014, pp. 1–9, 2014.
42] S. Meyuhas, M. Assali, M. Huleihil, and M. Huleihel, “An-
timicrobial activities of caffeic acid phenethyl ester,” Journal of
Molecular Biochemistry, vol. 4, pp. 21–31, 2015.
[
[
[
43] C. Velazquez, M. Navarro, A. Acosta et al., “Antibacterial and
free-radical scavenging activities of Sonoran propolis,”
Journal of Applied Microbiology, vol.103, no. 5, pp.1747–1756,
[59] P. Tosco and T. Balle, “Open3DQSAR: a new open-source
software aimed at high-throughput chemometric analysis of
molecular interaction fields,” Journal of Molecular Modeling,
vol. 17, no. 1, pp. 201–208, 2010.
2
007.
[
[
[
[
[
[
[
44] M. O. De Farias, T. C. Lima, A. L. A. L. P ´e rez et al., “Anti-
fungal activity of ester derivatives from caffeic acid against
Candida species,” International Journal of Pharmacy &
Pharmaceutical Research, vol. 7, no. 1, pp. 151–159, 2016.
45] P. Boeck, M. M. S a´ , B. S. d. Souza et al., “A simple synthesis of
kaurenoic esters and other derivatives and evaluation of their
antifungal activity,” Journal of the Brazilian Chemical Society,
vol. 16, no. 6B, pp. 1360–1366, 2005.
46] W. Li, N. Li, Y. Tang et al., “Biological activity evaluation and
structure-activity relationships analysis of ferulic acid and
caffeic acid derivatives for anticancer,” Bioorganic & Medic-
inal Chemistry Letters, vol. 22, no. 19, pp. 6085–6088, 2012.
47] H. Shi, D. Xie, R. Yang, and Y. Cheng, “Synthesis of caffeic
acid phenethyl ester derivatives, and their cytoprotective and
neuritogenic activities in PC12 cells,” Journal of Agricultural
and Food Chemistry, vol. 62, no. 22, pp. 5046–5053, 2014.
48] J. G. Handique, D. Mahanta, A. Devi, and M. P. Boruah,
[60] C. Balachandran, V. Duraipandiyan, N. A. Al-Dhabi et al.,
“Antimicrobial and antimycobacterial activities of methyl
caffeate isolated from solanum torvum swartz. Fruit,” Indian
Journal of Microbiology, vol. 52, no. 4, pp. 676–681, 2012.
61] J. Fu, K. Cheng, Z.-m. Zhang, R.-q. Fang, and H.-l. Zhu,
[
“Synthesis, structure and structure-activity relationship
analysis of caffeic acid amides as potential antimicrobials,”
European Journal of Medicinal Chemistry, vol. 45, no. 6,
pp. 2638–2643, 2010.
[
62] V. D. Gangan, L. Jazly, C. T. Chakraborty et al., “Ethyl caffeate
ether derivatives as future potential drug,” Journal of Phar-
macy Research, vol. 8, no. 6, pp. 818–821, 2014.
[
63] M. Kartal, S. Yıldız, S. Kaya, S. Kurucu, and G. Topçu,
“Antimicrobial activity of propolis samples from two different
regions of Anatolia,” Journal of Ethnopharmacology, vol. 86,
no. 1, pp. 69–73, 2003.
“
Synthesis and electrochemical behavior of some dendritic
[
[
[
[
64] I. K. Lee, K. H. Kim, S. U. Choi, J. H. Lee, and K. R. Lee,
polyphenols as antioxidants,” Letters in Organic Chemistry,
vol. 10, no. 1, pp. 53–59, 2013.
“
Phytochemical constitutents of ꢀesium chinense Turcz and
their cytotoxic activities in vitro,” Natural Product Scienses,
49] P. Xiao, S. Zhang, H. Ma, A. Zhang, X. Lv, and L. Zheng,
vol. 15, no. 4, pp. 246–249, 2009.
“
Stereoselective synthesis of caffeic acid amides via enzyme-
¨
¨
¨
65] A. Uzel, K. y. Sorkun, O. Onça g˘ , D. Ço g˘ ulu, O. Gençay, and
B. r. Sali_h, “Chemical compositions and antimicrobial ac-
tivities of four different Anatolian propolis samples,” Mi-
crobiological Research, vol. 160, no. 2, pp. 189–195, 2005.
catalyzed asymmetric aminolysis reaction,” Journal of Bio-
technology, vol. 168, no. 4, pp. 552–559, 2013.
50] J. T. Sanderson, H. Clabault, C. Patton et al., “Anti-
proliferative, antiandrogenic and cytotoxic effects of novel
caffeic acid derivatives in LNCaP human androgen-
dependent prostate cancer cells,” Bioorganic & Medicinal
Chemistry, vol. 21, no. 22, pp. 7182–7193, 2013.
ˇ
66] R. Merkl, I. Hr a´ dkov a´ , V. Filip, and J. Smidrkal, “Antimi-
crobial and antioxidant properties of phenolic acids alkyl
esters,” Czech Journal of Food Sciences, vol. 28, no. 4,
pp. 275–279, 2010.
[
51] J. Zhang, L. X. Xu, X. S. Xu, B. W. Li, R. Wang, and J. J. Fu,
67] L. Jazly, V. D. Gangan, C. T. Chakraborty, A. V. Tamhankar,
J. J. Kadam, and S. Bhalekar, “Synthesis and antibacterial
activity of novel Caffeic acid ester derivatives,” Journal of
Pharmacy Research, vol. 1, no. 10, pp. 942–946, 2013.
“
Synthesis and effects of new caffeic acid derivatives on
nitric oxide production in lipopolysaccharide-induced
RAW 264.7 macrophages,” International Journal of Clini-
cal and Experimental Medicine, vol. 7, no. 4, pp. 1022–1027,
2
014.
[
[
[
[
52] N.-G. Li, R. Wang, Y.-P. Tang et al., “Design, synthesis and
biological study of novel NO-donor-caffeic acid hybrids as
potential anti-atherosclerotic drug candidates,” Letters in
Drug Design & Discovery, vol. 8, no. 6, pp. 550–557, 2011.
53] K. D. Kim, M. H. Song, E. K. Yum, O. S. Jeon, Y. W. Ju, and
M. S. Chang, “2,4-Dihydroxycinnamic esters as skin depig-
menting agents,” Bulletin of the Korean Chemical Society,
vol. 30, no. 7, pp. 1619–1621, 2009.
54] R. Cleeland and E. Squires, “Evaluation of new antimicrobials
“
in vitro” and in experimental animal infections,” in Anti-
biotics in Laboratory Medicine, V. M. D. Lorian, Ed., Lip-
pincott Williams & Wilkins, Baltimore, MD, USA, 1991.
55] F. Hadacek and H. Greger, “Testing of antifungal natural
products: methodologies, comparability of results and assay
choice,” Phytochemical Analysis, vol. 11, no. 3, pp. 137–147,
2
000.

Journal of
Nanomaterials
Journal of
International Journal of
The Scientifc
Analytical Methods
Journal of
Photoenergy
in Chemistry
World Journal
Applied Chemistry
Hindawi
Hindawi
Hindawi Publishing Corporation
Hindawi
www.hindawi.com
Hindawi
www.hindawi.com
Volume 2018
www.hindawi.com
Volume 2018
hw t wt p
w
:/./h
w
i nwd wa w. hi i. nc od ma wi.com
Volume 20183
Volume 2018
www.hindawi.com
Volume 2018
Advances in
International Journal of
Physical Chemistry
Medicinal Chemistry
Hindawi
www.hindawi.com
Hindawi
Volume 2018
www.hindawi.com
Volume 2018
Submit your manuscripts at
www.hindawi.com
Bioinorganic Chemistry
Journal of
and Applications
Materials
Hindawi
Hindawi
www.hindawi.com
Volume 2018
www.hindawi.com
Volume 2018
Advances in
Journal of
BioMed
International Journal of
International Journal of
Tribology
Chemistry
Research International
Spectroscopy
Electrochemistry
Hindawi
Hindawi
Hindawi
www.hindawi.com
Hindawi
Hindawi
www.hindawi.com
www.hindawi.com
Volume 2018
www.hindawi.com
Volume 2018
Volume 2018
www.hindawi.com
Volume 2018
Volume 2018
Journal of
Journal of
Enzyme
Research
Biochemistry
Spectroscopy
Nanotechnology
Research International
Hindawi
Hindawi
www.hindawi.com
Hindawi
www.hindawi.com
Hindawi
www.hindawi.com
www.hindawi.com
Volume 2018
Volume 2018
Volume 2018
Volume 2018