Ferrocenyl chalcones with O-alkylated vanillins: synthesis, spectral characterization, microbiological evaluation, and single-crystal X-ray analysis

Article abstract of DOI:10.1007/s00044-016-1609-8

O-alkylated vanillin derivatives 2a–f and acetylferrocene react under Claisen–Schmidt conditions, resulting in good-to-high yields of the corresponding ferrocene chalcones 3a–f. None of the resultant compounds 3b–f has been previously described in the literature. All synthesized compounds were characterized by spectral and physical data, whereas two of them, 1-ferrocenyl-3-(4-ethoxy-3-methoxyphenyl)-prop-2-en-1-one (3b) and 1-ferrocenyl-3-(4-buthoxy-3-methoxy-phenyl)-prop-2-en-1-one (3e), were crystalline substances, suitable for single-crystal X-ray analysis, which confirmed undoubtedly their structures. Chalcones 3a–f were tested for their biological activity and demonstrated relatively good in vitro antimicrobial activity towards different strains of bacteria and fungi. The best antibacterial activity is expressed by compounds 3b and 3c, while compound 3d shows the best antifungal activity.

Full text of DOI:10.1007/s00044-016-1609-8

MEDICINAL
CHEMISTRY
RESEARCH
Med Chem Res
DOI 10.1007/s00044-016-1609-8
ORIGINAL RESEARCH
Ferrocenyl chalcones with O-alkylated vanillins: synthesis, spectral
characterization, microbiological evaluation, and single-crystal X-ray
analysis
Jovana Muškinja1 Adrijana Burmudžija1 Zoran Ratković1 Branislav Ranković2
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Marijana Kosanić2 Goran A. Bogdanović3 Slađana B. Novaković3
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Received: 16 October 2015 / Accepted: 3 June 2016
© Springer Science+Business Media New York 2016
Abstract O-alkylated vanillin derivatives 2a–f and acetyl
ferrocene react under Claisen–Schmidt conditions, result-
ing in good-to-high yields of the corresponding ferrocene
chalcones 3a–f. None of the resultant compounds 3b–f has
been previously described in the literature. All synthesized
compounds were characterized by spectral and physical
data, whereas two of them, 1-ferrocenyl-3-(4-ethoxy-3-
methoxyphenyl)-prop-2-en-1-one (3b) and 1-ferrocenyl-3-
Introduction
Chalcones are an important class of organic compounds
since they often represent the core structure of many natural
products and pharmaceuticals. Chalcones (1,3-diaryl-2-
propen-1-ones) and their derivatives exhibit different phar-
macological and biological activities, the major ones being
antifungal (Bag et al., 2009; Lahtchev et al., 2008; Mostahar
et al., 2007), antimicrobial (Yayli et al., 2006; Trivedi et al.,
2008; Sivakumar et al., 2010; Opletalova, 2000; Katade
et al., 2008), anticonvulsant (Kaushik et al., 2010), anti-
oxidant (Vasilev et al., 2010, Sivakumar et al., 2010; Vogel
et al., 2008, Cheng et al., 2008), antiprotozoal (Lunardi
et al., 2003), antitrichomonal (Oyedapo et al., 2004), anti-
malarial (Motta et al., 2006; Awasthi et al., 2009; Lim et al.,
2007; Wu et al., 2002; Liu et al., 2001), anti-inflammatory
(Yadav et al., 2010; Zhang et al., 2010; Herencia et al.,
1998; Nowakowska 2007), trypsin inhibition (Maliar et al.,
2004) and anti-cancer activity (Achanta et al., 2006;
Romagnoli et al., 2008; Echeverria et al., 2009; Szliszka
et al., 2009; Ilango et al., 2010; Wattenberg et al., 1994;
Edwards et al., 1988; Kumar et al., 2011; Seo et al., 2010).
Different research groups are involved with the aim to
identify the fragment in chalcones molecules, synthesized
or modified natural ones, responsible for previously
described activities. Tsukiyama et al. (2002) reported the
potent antibacterial activity of licochalcone A and lico-
chalcone B. Following these results, it was estimated that
free phenolic group in the ring at position 4 was the key
factor responsible for their antibacterial activity (Chen et al.,
1997). Starting from the fact that natural products, such as
curcumines, show various bioactivities (Chan, 1995; Mishra
et al., 2005; Dubey et al., 2008), and vanillin fragment is
present in these molecules, we supposed that vanillin is a
suitable aldehyde for chalcone formation due to its easy
(4-buthoxy-3-methoxy-phenyl)-prop-2-en-1-one
(3e),
were crystalline substances, suitable for single-crystal X-
ray analysis, which confirmed undoubtedly their struc-
tures. Chalcones 3a–f were tested for their biological
activity and demonstrated relatively good in vitro anti-
microbial activity towards different strains of bacteria and
fungi. The best antibacterial activity is expressed by
compounds 3b and 3c, while compound 3d shows the best
antifungal activity.
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Keywords Vanillin Ferrocene Ferrocenyl chalcones
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Microbial activity Crystal structure
Electronic supplementary material The online version of this article
(doi: 10.1007/s00044-016-1609-8) contains supplementary material,
which is available to authorized users.
* Zoran Ratković
wor@kg.ac.rs
1
Department of Chemistry, Faculty of Science, University of
Kragujevac, Kragujevac, Serbia
2
Department of Biology, Faculty of Science, University of
Kragujevac, Kragujevac, Serbia
3
Vinča Institute of Nuclear Sciences, Laboratory of Theoretical
Physics and Condensed Matter Physics, University of Belgrade,
Belgrade, Serbia

Med Chem Res
Scheme 1 Synthesis of ferrocenyl chalcones
modification by O-alkylation (Loev et al., 1956, Tatsuzaki
et al., 2006, Katritzky et al., 2006), by coupling reactions
and forming of divanillin (Nishimura et al., 2010), formyla
tions in position 5 (Blažević et al., 1979), and halogenation
in position 5 (Pepper and MacDonald, 1953). On the other
hand, enone system is the key part of substrates and could
be easily converted into various heterocyclic derivatives
(Abdel-Rahman et al., 2007, Kalirajan et al., 2009).
Ferrocenyl derivatives are among the most promising
compounds, which can be used in microbiological research.
The water-soluble ferrocenyl derivatives are more potent as
drugs than the water-insoluble ones. In our previous work
we had reported on the synthesis of different ferrocene
derivatives with expressed biological activities (Damljanović
et al., 2009; Damljanović et al., 2009; Ratković et al., 2010)
and we expected that incorporation of vanillin pharmaco-
phore and the ferrocene scaffold into the same molecule
might have an attractive structural result for the development
of novel antimicrobial agents, expecting the interesting
features due to the coexistence of two kinds of promising
pharmacophores.
Elemental microanalyses for C, H, and N were
performed by standard methods on a Vario III CHNS
Elemental Analyzer, Elemental Analysensysteme GmbH.
Synthesis
Keeping in mind the reactivity of aldehyde functional
group, the synthesis of ferrocene containing chalcones was
carried out as a simple Claisen–Schmidt condensation of
O-alkylated vanilline derivatives and acetylferrocene. A set
of O-alkyl derivatives 2a–f (Scheme 1) was prepared by
alkylation free phenolic group of vanillin 1 with corre-
sponding alkyl halides, according to the described literature
procedures (Loev and Dawson, 1956; Tatsuzaki et al., 2006;
Katritzky et al., 2006). Their spectral data were listed in
another paper (Ratković et al., 2016), describing synthesis
and biological activity of vanillin-based 1-acetyl-5-aryl-4,5-
dihydro-1H-pyrazoles.
In the next step, aldehydes 2a-f were subjected to the
reaction with acetylferrocene following the literature procedure
(Cheng et al., 2008), and the desired ferrocene containing
chalcone products were isolated in 59–85 % yields (Table 1).
The detailed analysis of 1H and 13C NMR spectra (see below)
revealed the structures of compounds 3a–f (Scheme 1),
whereas the structures of 3b and 3e were unambiguously
confirmed by the single-crystal X-ray analysis.
In the present paper we wish to report on synthesis,
spectral characterization and evaluation of antimicrobial
activity of a series of novel chalcone derivatives containing
ferrocene unit, prepared from O-alkylated vanillines and
acetylferrocene, as well as single-crystal X-ray analysis for
two of them.
General procedure for synthesis of chalcones
Experimental
The corresponding vanillin aldehyde 2a–f (10 mmol) and
acetylferrocene (10 mmol) were dissolved in 20 mL of
warm ethanol, and the mixture was stirred for 10 min and
1 mL of 40 % NaOH was added slowly. The reaction
mixture was stirred overnight at 50°C. Crushed ice, 100 g,
was placed in a beaker and the reaction mixture was poured
onto it with stirring. The product in some cases could be
isolated by filtration, if not it was necessary to extract with
toluene or dichloromethane (3 ×50 mL). The organic layer
was washed with water (2 ×50 mL), brine (2 ×50 mL), and
dried over anhydrous Na2SO4. The main part of the solvent
was evaporated at reduced pressure and crude concentrated
solution was filtered through a short column of silica gel.
The solvent was evaporated under reduced pressure and the
Material and methods
All the chemicals used were commercially available and
used as received, except that the solvents were purified by
distillation. Chromatographic separations were carried out
using silica gel 60 (Merck, 230-400 mesh ASTM), whereas
silica gel on Al plates, layer thickness 0.2 mm (Merck), was
used for TLC. IR spectra were recorded on a Perkin-Elmer
Spectrum One FT-IR spectrometer with a KBr disc. NMR
spectra were recorded on a Varian Gemini (200 MHz)
spectrometer using CDCl3 as the solvent and TMS as the
internal standard.

Med Chem Res
Table 1 Synthesis of ferrocene
derivatives 3a–f
Entry Vanillines
1
Products
Yield
(%)a
78
85
79
59
68
67
2a
3a
3b
3c
3d
3e
3f
2
2b
3
2c
4
2d
5
2e
6
2f
a
Yields of isolated products based on acetylferrocene

Med Chem Res
residue was chromatographed on silica gel column using
toluene-ethyl acetate (8:2) mixture as eluent for separation
of reaction products 3a–f.
(s, 5H), 4.57 (t, J = 1.6Hz, 2H), 4.92 (t, J = 2Hz, 2H), 6.92
(d, J = 8.4Hz 1H), 6.99 (d, J = 15.6Hz, 1H), 7.15–7.26
(m, 2H), 7.75 (d, J = 15.6Hz, 1H); 13C NMR (50 MHz,
CDCl3): δ 22, 56.2, 69.7, 70, 71.4, 72.5, 80.8, 111.6, 120.9,
122.2, 128.2, 129, 141, 149.6, 150.4, 192.9 (CO). Anal.:
calcd. for C23H24FeO3 (404.280): C, 68.33; H, 5.98; Fe,
13.81; O, 11.87. Found: C, 67.31; H, 5.95.
1-Ferrocenyl-3-(3,4-dimethoxyphenyl)prop-2-en-1-one (3a)
Cinnabar red crystals; m.p. 148–149 °C; yield 78 %;
IR (cm−1): 1648, 1586, 1517, 1263, 1081; 1H NMR
(200 MHz, CDCl3): δ 3.95 (d, J = 6.8 Hz, 6H), 4.22 (s, 5H),
4.58 (t, J = 2Hz, 2H), 4.92 (t, J = 2Hz, 2H), 6.91 (d, J =
8.4Hz 1H), 6.99 (d, J = 15.6Hz, 1H), 7.14–7.27 (m, 2H),
7.76 (d, J = 15.6Hz, 1H); 13C NMR (50 MHz, CDCl3):
δ 55.9, 69.7, 70, 72.5, 80.7, 110.6, 111.3, 121, 122.4,
128.2, 140.9, 149.2, 151.1, 192.8 (CO). Anal.: calcd. for
C21H20FeO3 (376.227): C, 67.04; H, 5.36; Fe, 14.84; O,
12.76. Found: C, 66.10; H, 5.30.
1-Ferrocenyl-3-(4-butoxy-3-methoxyphenyl)prop-2-en-1-
one (3e)
Cinnabar red crystals; m.p. 128 °C; yield 68 %; IR (cm−1):
1651, 1592, 1511, 1264, 1077; 1H NMR (200 MHz,
CDCl3): δ 0.99 (t, J = 7.4Hz, 3H), 1.45–1.57 (m, 2H),
1.76–1.89 (m, 2H), 3.95 (s, 3H), 4.08 (t, J = 6.8Hz, 2H),
4.21 (s, 5H), 4.57 (t, J = 2Hz, 2H), 4.92 (t, J = 2Hz, 2H),
6.91 (d, J = 8.2Hz 1H), 6.99 (d, J = 15.6Hz, 1H), 7.14–7.26
(m, 2H), 7.75 (d, J = 15.6Hz, 1H); 13C NMR (50 MHz,
CDCl3): δ 13.8, 31.1, 56.2, 68.7, 69.7, 70, 72.5, 80.8, 111.2,
112.7, 120.8, 122.4, 127.9, 141, 149.6, 150.8, 192.9 (CO).
Anal.: calcd. for C24H26FeO3 (418.306): C, 68.91; H, 6.26;
Fe, 13.35; O, 11.47. Found: C, 68.16; H, 6.23.
1-Ferrocenyl-3-(4-ethoxy-3-methoxyphenyl)prop-2-en-1-
one (3b)
Cinnabar red crystals; m.p. 155–156 °C; yield 85 %; IR
(cm−1): 1650, 1588, 1513, 1267, 1076; 1H NMR (200 MHz,
CDCl3): δ 1.49 (t, J = 7Hz, 3H), 3.96 (s, 3H), 4.16 (q, J =
7.2Hz, 2H), 4.21 (s, 5H), 4.57 (t, J = 1.8Hz, 2H), 4.92
(t, J = 1.8Hz, 2H), 6.90 (d, J = 8.4Hz 1H), 6.99 (d, J =
15.6Hz, 1H), 7.14–7.27 (m, 2H), 7.75 (d, J = 15.6Hz, 1H);
13C NMR (50 MHz, CDCl3): δ 14.7, 56.1, 64.4, 69.6, 70,
72.5, 80.8, 110.9, 112.5, 120.9, 122.4, 128, 141, 149.5,
150.5, 192.9 (CO). Anal.: calcd. for C22H22FeO3 (390.253):
C, 67.71; H, 5.68; Fe, 14.31; O, 12.30. Found: C, 66.61;
H, 5.61.
1-Ferrocenyl-3-(4-benzyloxy-3-methoxyphenyl)prop-2-en-
1-one (3f)
Cinnabar red crystals; m.p. 135–136 °C; yield 67 %;
IR (cm−1): 1650, 1591, 1515, 1262, 1084; 1H NMR
(200 MHz, CDCl3): 3.97 (s, 3H), 4.20 (s, 5H), 4.57 (t, J =
2Hz, 2H), 4.91 (t, J = 2Hz, 2H), 5.21 (s, 2H), 6.91 (d, J =
8.2Hz, 1H), 6.98 (d, J = 15.6Hz, 1H), 7.15–7.43 (m, 7H),
7.74 (d, J = 15.6Hz, 1H); 13C NMR (50 MHz, CDCl3):
δ 56.1, 69.6, 70, 70.9, 72.5, 80.7, 111.3, 113.7, 121.1,
122.1, 127.2, 127.9, 128.5, 128.6, 136.6, 140.9, 149.8,
150.2, 192.8 (CO). Anal.: calcd. for C27H24FeO3 (452.323):
C, 71.69; H, 5.35; Fe, 12.35; O, 10. 61. Found: C, 70.61;
H, 5.32.
1-Ferrocenyl-3-(3-methoxy-4-propoxyphenyl)prop-2-en-1-
one (3c)
Cinnabar red crystals; m.p. 104–105 °C; yield 79 %;
IR (cm−1): 1647, 1586, 1515, 1258, 1079; 1H NMR
(200 MHz, CDCl3): δ 1.06 (t, J = 7.6Hz, 3H), 1.80–1.98
(m, 2H), 3.95 (s, 3H), 4.03 (t, J = 6.8Hz, 2H), 4.21 (s, 5H),
4.57 (t, J = 1.6Hz, 2H), 4.92 (t, J = 2Hz, 2H), 6.90 (d, J =
8.4Hz 1H), 6.99 (d, J = 15.6Hz, 1H), 7.14–7.27 (m, 2H),
7.75 (d, J = 15.6Hz, 1H); 13C NMR (50 MHz, CDCl3):
δ 10.3, 22.3, 56.2, 69.6, 70, 70.5, 72.5, 80.8, 111.2, 112.7,
120.8, 122.4, 127.9, 141, 149.6, 150.8, 192.8 (CO). Anal.:
calcd. for C23H24FeO3 (404.280): C, 68.33; H, 5.98; Fe,
13.81; O, 11.87. Found: C, 67.51; H, 5.93.
X-ray crystallography and structural comparisons of 3b
and 3e
Crystal data and experimental details for 3b and 3e com-
pounds are summarized in Table 2. Single-crystal diffrac-
tion data were collected at room temperature on an Agilent
Gemini S diffractometer equipped with Mo Kα radiation
(λ = 0.71073 Å). Data integration and scaling of the
reflections were performed using the CRYSALIS software
(Agilent Technologies, 2013) with multi-scan absorption
corrections applied using SCALE3 ABSPACK (Agilent
Technologies, 2013). Crystal structure was solved by direct
methods, using SIR2002 (Burla et al., 2003) and refined
using SHELXL97 (Sheldrick, 2008) program both incor-
porated in WinGX (Farrugia, 1999) program package.
1-Ferrocenyl-3-(4-isopropoxy-3-methoxyphenyl)prop-2-en-
1-one (3d)
Cinnabar red crystals; m.p. 114 °C; yield 59 %; IR (cm−1):
1653, 1590, 1508, 1264, 1074; 1H NMR (200 MHz,
CDCl3): δ 1.41 (d, J = 6.2Hz, 6H), 3.94 (s, 3H), 4.21

Med Chem Res
Table 2 Crystallographic data for 3b and 3e
(ATCC 12453). All of the bacteria used were obtained from
the American Type Culture Collection (ATCC). The bac-
terial cultures were maintained on Mueller-Hinton agar
substrates (Torlak, Belgrade). The fungi used as test
organisms were: Aspergillus niger (ATCC 16888), Candida
albicans (ATCC 10259), Penicillium italicum (ATCC
10454), Mucor mucedo (ATCC 20094), Trichoderma viride
(ATCC 13233). All of the fungi were from ATCC. The
fungal cultures were maintained on potato dextrose (PD)
agar, except for Candida albicans that was maintained on
Sabourad dextrose (SD) agar (Torlak, Belgrade). All of the
cultures were stored at 4 °C and subcultured every 15 days.
Bacterial inoculum were obtained from bacterial cultures
incubated for 24 h at 37 °C on Mueller-Hinton agar sub-
strates and brought up to approximately 108 CFU/ml by
dilution according to the 0.5 McFarland standard. Suspen-
sions of fungal spores were prepared from freshly mature
(3- to 7-day-old) cultures that grew at 30 °C on a PD agar
substrate. The spores were rinsed with sterile distilled water,
used to determine turbidity spectrophotometrically at
530 nm, and were then further diluted to approximately 106
CFU/ml according to the procedure recommended by
NCCLS (1998).
Compound
3b
3e
Empirical formula
Formula weight
Temperature (K)
Wavelength (Å)
Crystal system
Space group
Unit cell dimensions
a (Å)
C
22H22FeO3
C24H26FeO3
418.30
390.25
293(2)
0.71073
Monoclinic
P21/c
293(2)
0.71073
Monoclinic
P21/c
14.6500(6)
7.9537(2)
16.7216(5)
106.086(3)
1872.14(11)
4
16.1310(6)
7.9881(2)
17.3283(6)
113.533(4)
2047.15(12)
4
b (Å)
c (Å)
β (°)
V (Å3)
Z
Dcalc (Mg/m3)
1.385
1.357
μ (mm−1
)
0.823
0.758
Crystal size (mm3)
0.50 × 0.37 × 0.12 0.48 × 0.24 × 0.17
θ range for data collection (°) 3.04–28.93
2.94–29.02
10841
Reflections collected
11443
Independent reflections, Rint
4333, 0.0244
4731, 0.0280
99.8 %
Completeness to θ = 26° (%) 99.8 %
Data/restraints/parameters
Goodness-of-fit on F2
4333/0/237
4731/0/255
1.041
1.076
Final R1/wR2 indices (I
42σI)
0.0437, 0.0905
0.0397, 0.0827
The MIC was determined by the broth microdilution
method using 96-well micro-titer plates (Sarker et al.,
2007). A series of dilutions with concentrations ranging
from 20 to 0.0195 mg/ml of the tested compounds were
used in the experiment against every microorganism tested.
The starting solutions with tested compounds were obtained
by measuring a certain quantity of the compounds and
dissolving it in 5 % DMSO. Two-fold dilutions of the
compounds were prepared in a Mueller-Hinton broth for
bacterial cultures and a SD broth for fungal cultures. The
MIC was determined with resazurin. Resazurin is an
oxidation-reduction indicator used for the evaluation of
microbial growth. It is a blue non-fluorescent dye that
becomes pink and fluorescent when reduced to resorufin by
oxidoreductases within viable cells. The boundary dilution
without any change in the color of resazurin was defined as
the MIC for the tested microorganism at a given con-
centration. As a positive control of growth inhibition,
streptomycin was used in the case of bacteria and ketoco-
nazole in the case of fungi. A 5 % DMSO solution was used
as a negative control for the influence of the solvents.
Experiment was carried out in triplicate.
Final R1/wR2 indices (all
data)
0.0633, 0.0975
0.0622, 0.0929
Largest differential density
0.285 and −0.218
0.285 and −0.306
peak and hole (e Å−3
)
All non-H atoms were refined anisotropically to con-
vergence. All H atoms were placed at geometrically calculated
positions with the C-H distances fixed to 0.93 from Csp2, and
0.96 and 0.97 Å from methyl and methylene Csp3, respec-
tively. The corresponding isotropic displacement parameters of
the hydrogen atoms were equal to 1.2Ueq and 1.5Ueq of the
parent Csp2 and Csp3, respectively. Figures were produced
using ORTEP-3 (Farrugia, 1997) and MERCURY, Version
2.4 (Macrae et al., 2006). The software used for the prepara-
tion of the materials for publication are WinGX (Farrugia,
1999), PLATON (Spek, 2003) and PARST (Nardelli, 1995).
Antimicrobial activity
The antimicrobial activity was estimated by determining the
minimal inhibitory concentration (MIC) using the broth micro
dilution method against five species of bacteria and five species
of fungi. As a result of the study, the measured MIC values for
tested compounds varied from 0.0352 to 0.8873 mg/ml.
The following bacteria were used as test organisms in
this study: Staphylococcus aureus (ATCC 25923), Bacillus
subtilis (ATCC 6633), Bacillus cereus (ATCC 10987),
Escherichia coli (ATCC 25922), and Proteus mirabilis
Results and Discussion
Spectral characterization
1
All synthesized compounds were characterized by IR, H,
and 13C NMR spectral data. In IR spectra of the synthesized

Med Chem Res
Fig. 2 The molecules of 3b (blue) and 3e (red) are overlapped using
the C1–C5 ring as a common structural part
Table 3 Selected geometrical parameters in the crystal structures of
3b and 3e compounds
Compound
3b
3e
Selected bond lengths (Å)
O1−C11
1.224(2)
1.367(2)
1.426(2)
1.355(2)
1.439(2)
1.466(3)
1.472(3)
1.320(3)
1.451(3)
1.222(2)
1.362(2)
1.432(2)
1.362(2)
1.435(2)
1.467(3)
1.474(3)
1.324(3)
1.459(3)
O2−C16
Fig. 1 Molecular structures of 3b (a) and 3e (b) with atom numbering
scheme
O2−C20
O3−C17
O3−C21
C1−C11
compounds the most recognizable band appears in the
carbonyl group region 1680 cm–1 and alkoxy group region
1261–1269 cm–1.
C11−C12
C12−C13
1HNMR spectra of compounds 2a–f could be recognized
as sharp signals, corresponding to the aldehyde proton, δ =
9.83–9.86 ppm, and strong sharp signals corresponding
to three protons, approximately δ = 3.94 ppm, belongs to
methyl protons of methoxy group [-OCH3] from vanillin.
1HNMR spectra of compounds 3a–f could be recognized
as the most important signals, such as doublet signals,
appear at δ = 7.75 and δ = 6.99 ppm, corresponding to
protons of chalcone double bond with characteristic J =
15.6 Hz, ferrocene unsubstituted ring at 4.22 area, and
strong sharp signal corresponding to three protons,
approximately δ = 3.94–3.98 ppm, belongs to methyl pro-
tons of 3-methoxy group from vanillin ring.
C13−C14
Selected bond angles (°)
C2−C1−C11
C5−C1−C11
O2−C16−C15
O2−C16−C17
O3−C17−C18
O3−C17−C16
Selected torsion angles (°)
C20−O2−C16−C17
C21−O3−C17−C16
128.01(19)
124.8(2)
127.51(19)
125.10(19)
125.51(17)
114.99(18)
125.60(18)
115.24(18)
125.40(17)
114.46(18)
125.58(18)
115.78(17)
−174.48(19)
−170.25(18)
174.15(18)
169.16(17)
C1-substituent regarding the ferrocene unit. Structure 3e is
somewhat larger deformed than 3b and this is well illustrated
in Fig. 2, where two molecules are overlapped by Cp1 rings.
This perspective view illustrates similarity in spatial orien-
tation of the C1-substituent in the two molecules.
All non-H atoms in the C1-substituent are approximately
coplanar in both the molecules (Fig. 1). However, the
substituent is significantly displaced from the C1–C5 (Cp1)
ring plane. For example, dihedral angle between phenyl and
Cp1 rings is 12.4(2) and 15.4(2)° for 3b and 3e, respec-
tively, while the C21 alkyl group is bent in the direction of
Cp2 ring plane. Such an orientation of the substituent is
attained by rotation around the C1–C11 bond and as a
consequence the C5–C1–C11–C12 torsion angle has values
of −167.3(2) and −162.1(2)° for 3b and 3e, respectively. In
the two compounds, the corresponding bond lengths and
bond angles are very similar (Table 3). The C12–C13 bond
X-ray structure determination
Single-crystal X-ray crystallographic studies of 3b and 3e
show that two compounds have very similar molecular
geometries (Fig. 1; displacement ellipsoids are drawn
at the 30 % probability level; additional projections are
shown in the Supplementary Material file, Fig. S1).
The cyclopentadienyl rings (Cp) of both molecules
have an almost ideal eclipsed conformation. The C1–
Cg1–Cg2–C6 torsion angle is 1.8(2)° in 3b and –0.8(2)° in
3e (Cg1 and Cg2 are centroids of the corresponding Cp
rings, the C1–C5 and C6–C10). The Cp1 and Cp2 rings
show only small mutual tilting as evidenced from the cor-
responding Cp1/Cp2 dihedral angles which are 1.7(2) and
1.0 (2)° in 3b and 3e, respectively. Most importantly, two
molecules are very similar in spatial orientation of the

Med Chem Res
with bond distance of 1.32 Å is the shortest C–C bond in
both the molecules and it could be accepted as the only
localized double C–C bond.
is only the noteworthy intermolecular H-bond. The C21–H…
O1 together with C21–H…π (the C14–C19 phenyl ring)
interaction interconnect the molecules of 3b and 3e into
infinite tapes where all C1-substituents are parallel to each
other as illustrated in Fig. 3. It is interesting to note that in
both intermolecular interactions the C21 methylene group
plays the role of a C–H donor (C21–H…O1 and C21–H…π
interactions are shown with dotted blue lines; H atoms not
involved in intermolecular interactions have been omitted for
clarity; corresponding diagram for 3e is shown in the Sup-
plementary Material file, Fig. S2).
It is interesting to analyze positions of O2 and O3 atoms.
The O2–C16–C15 and O2–C16–C17 angles (which directly
reflect directionality of the O2 atom regarding the phenyl
ring) are extremely different (Table 3). The same could be
found for the O3 atom (Table 3) and in this way two oxygen
atoms are shifted close to each other. As a result, the O2 and
O3 atoms attain a relatively short O…O distance (2.574(2)
and 2.571(3) Å for 3b and 3e, respectively) with lone pair of
electrons directed to each other. The present nearly coplanar
orientation of O-alkyl groups (see the corresponding torsion
angles in Table 3) with a short O2…O3 interatomic contact
is also noticed in majority of the crystal structures (pos-
sessing similar structural fragment) that are found by
searching the Cambridge Structural Database (Allen, 2002).
Obviously this structural feature is not coincidental and
probably it plays specific role in the stabilization of mole-
cular conformation.
Antimicrobial activity
The antimicrobial activity of the investigated compounds
against the test bacteria is shown in Table 4 and fungi in
Table 5. The tested compounds exhibited a similar anti-
microbial activity. They inhibited all the tested bacteria and
antifungal activities. The MIC for bacteria ranged from
0.312 to 5 mg/ml, while MIC for fungi varied from 0.312 to
10 mg/ml. The most sensitive, among the bacteria, were
B. subtilis and B. cereus, whereas the highest resistance
was shown by the Gram-negative bacteria (E. coli and
P. mirabilis). Among the fungi, the most sensitive appeared
to be C. albicans, while A. niger was the most resistant.
The antimicrobial activity was compared with the standard
antibiotics, streptomycin (for bacteria) and ketoconazole
(for fungi). The results showed that standard antibiotics had
stronger activity than the tested samples as shown in
Tables 4 and 5.
Both the compounds do not possess any significant
hydrogen bond donor and as a consequence the C21–H…O1
In these experiments, the compounds examined at the
same concentrations showed a slightly stronger antibacterial
than antifungal activity. These results could be expected due
to the fact that numerous tests proved that bacteria are more
sensitive to the antibiotic compared to fungi (Hugo and
Russel, 1983). The reason for different sensitivities between
fungi and bacteria can be found in different permeabilities
of the cell wall. The cell wall of the Gram-positive bacteria
consists of peptidoglycans (murein) and teichoic acids,
while the cell wall of Gram-negative bacteria consists of
Fig. 3 The crystal packing fragment of 3b illustrating one-
dimensional association of the molecules via intermolecular interac-
tions between C1-substituents
Table 4 Antibacterial activity
of tested compounds
Bacteria
Staphylococcus
aureus
Bacillus
subtilis
Bacillus
cereus
Escherichia coli Proteus
mirabilis
Test compounds
MIC (mg/ml)
0.312
3a
1.25
0.625
0.625
2.5
0.625
0.312
0.312
0.625
1.25
1.25
1.25
2.5
2.5
3b
0.625
1.25
1.25
1.25
1.25
2.5
3c
0.312
3d
0.312
1.25
2.5
3e
2.5
0.625
3f
5
1.25
1.25
5
Streptomycin
0.031
0.016
0.016
0.062
0.062

Med Chem Res
Table 5 Antifungal activity of
tested compounds
Fungi
Mucor
mucedo
Trichoderma
viride
Aspergilus
niger
Candida
albicans
Penicillium
italicum
Test compounds
MIC (mg/ml)
3a
1.25
5
2.5
2.5
5
0.625
0.625
1.25
2.5
2.5
1.25
1.25
5
3b
1.25
2.5
3c
2.5
2.5
5
2.5
5
3d
1.25
2.5
0.312
1.25
3e
5
3f
10
5
10
2.5
5
Ketoconazole
0.156
0.078
0.078
0.039
0.156
some halcones and their derived pyrazoles, pyrazolines, iso-
xazolines, and 5,6-dihydropyrimidine-2-(1H)-thiones. Monat-
shefte 138:889–897
lipopolysaccharides and lipopoliproteins (Heijenoort,
2001), whereas the cell wall of fungi consists of poly-
saccharides such as chitin and glucan (Farkaš, 2003).
Achanta G, Modzelewska A, Feng L, Khan SR, Huang P (2006) A
boronic-chalcone derivative exhibits potent anticancer activity
through inhibition of the proteasome. Mol Pharmacol 70:426–433
Agilent Technologies (2013) CrysAlis PRO. Yarnton, Oxfordshire,
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Conclusion
Allen FH (2002) The Cambridge Structural Database: a quarter of a
million crystal structures and rising. Acta Crystallogr B58:
380–388
Awasthi SK, Mishra N, Kumar B, Sharma M, Bhattacharya A, Mishra
LC, Bhasin VK (2009) Potent antimalarial activity of newly
synthesized substituted chalcone analogs in vitro. Med Chem Res
18:407–420
We described herein synthesis, spectral, analytical, single-
crystal X-ray characterization, and in vitro microbiological
activity of a series of ferrocenyl chalcones (3a–f). Chalcones
were prepared in high yields from O-alkylated vanillines
and acetylferrocene.
Single-crystal X-ray analysis of 3b and 3e compounds
show that they have very similar conformation and inter-
molecular association via the C21–H…O1 and C21–H…π
interactions. In both molecules the terminal alkyl groups
attain an approximately coplanar position with phenyl ring
while the methoxy O2 and alkoxy O3 atoms are mutually
located at a surprisingly short interatomic distance of 2.57 Å.
This structural feature could be a significant supplementary
factor for antimicrobial activity of this class of molecules.
We found out that these organometallic derivatives show
noticeable activity toward different strains of bacteria and
fungi. Compared with our compounds, standard antibiotics,
streptomycin (for bacteria) and ketoconazole (for fungi)
showed stronger activity.
Bag S, Ramar S, Degani MS (2009) Synthesis and biological eva-
luation of α,β-unsaturated ketone as potential antifungal agents.
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Acknowledgments This work was supported by the Ministry of
Science and Technological Development of the Republic of Serbia,
Projects No. 172034 and 172014.
Damljanović I, Čolović M, Vukićević M, Manojlović D, Radulović N,
Wurst K, Laus G, Ratković Z, Joksović M, Vukićević RD (2009)
Synthesis, spectral characterization and electrochemical proper-
ties of 1H-3-(o-, m- and p-ferrocenylphenyl)-1-phenylpyrazole-4-
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Joksović M, Vukićević RD (2009) Synthesis and antimicrobial
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Compliance with ethical standards
Conflict of interest The authors declare that they have no conflict of
interests.
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