Ag(I)-N-heterocyclic carbene complexes of N-allyl substituted imidazol-2-ylidenes with ortho-, meta- and para-xylyl spacers: Synthesis, crystal structures and in vitro anticancer studies

Article abstract of DOI:10.1016/j.inoche.2012.05.037

A series of N-allyl substituted xylyl-linked imidazolium salts (7-10) and their respective Ag(I)-N-heterocyclic carbene (NHC) complexes (11-14) have been synthesized and characterized by a number of spectral and analytical techniques. Molecular structure of complexes 13 and 14 were established by single-crystal X-ray diffraction method. The in vitro anticancer activity of all imidazolium salts and their Ag(I)-carbene complexes were investigated using human colorectal (HCT 116) cancer cell lines. Imidazolium salts displayed no activity for HCT 116 cell lines, except for 9; yielding IC₅₀ value of 15.9 μM. Ag(I)-carbene complexes 12-14 showed exceptionally good activity (0.9-1.3 μM) against tested cancer cell lines. Furthermore, complex 11 displayed relatively good anticancer activity with IC₅₀ value of 5.2 μM, which is almost equal to standard used.

Full text of DOI:10.1016/j.inoche.2012.05.037

Inorganic Chemistry Communications 22 (2012) 113–119
Contents lists available at SciVerse ScienceDirect
Inorganic Chemistry Communications
journal homepage: www.elsevier.com/locate/inoche
Ag(I)-N-heterocyclic carbene complexes of N-allyl substituted imidazol-2-ylidenes
with ortho-, meta- and para-xylyl spacers: Synthesis, crystal structures and in vitro
anticancer studies
a
a
a
Rosenani A. Haque a, , Mohammed Z. Ghdhayeb , Abbas Washeel Salman , Srinivasa Budagumpi ,
⁎
Mohamed B. Khadeer Ahamed b, Amin M.S. Abdul Majid b
School of Chemical Sciences, Universiti Sains Malaysia, 11800 USM Penang, Malaysia
a
b
EMAN Research and Testing Laboratory, School of Pharmaceutical Sciences, Universiti Sains Malaysia, 11800 USM Penang, Malaysia
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of N-allyl substituted xylyl-linked imidazolium salts (7–10) and their respective Ag(I)-N-heterocyclic
carbene (NHC) complexes (11–14) have been synthesized and characterized by a number of spectral and
analytical techniques. Molecular structure of complexes 13 and 14 were established by single‐crystal X-ray
diffraction method. The in vitro anticancer activity of all imidazolium salts and their Ag(I)-carbene complexes
were investigated using human colorectal (HCT 116) cancer cell lines. Imidazolium salts displayed no activity
for HCT 116 cell lines, except for 9; yielding IC50 value of 15.9 μM. Ag(I)-carbene complexes 12–14 showed
exceptionally good activity (0.9–1.3 μM) against tested cancer cell lines. Furthermore, complex 11 displayed
relatively good anticancer activity with IC50 value of 5.2 μM, which is almost equal to standard used.
© 2012 Elsevier B.V. All rights reserved.
Received 2 May 2012
Accepted 17 May 2012
Available online 24 May 2012
Keywords:
Anticancer activity
Ag(I)-N-heterocyclic carbene complex
Human colorectal cell line
Xylyl-spacer
X-ray diffraction
1. Introduction
vomiting and ear damage. Consideration in this particular field has
focused in the search of new metal-based diagnostic agents on the
Ag(I) complexes of N-heterocyclic carbenes (NHCs) have played an
important role in the development of other transition/inner transition
metal-carbene systems [1]. A large number of Ag(I)-NHC complexes
have been synthesized, characterized, and their potential uses as
antimicrobial, anticancer and carbene transfer agents have also been
studied [2,3]. Three general procedures for the preparation of Ag(I)-NHC
complexes have been established. The first is the reaction of free NHC
with appropriate silver source normally at the liquid nitrogen tempera-
ture [4]. The second method involves the reaction of azolium salts with
silver bases such as Ag2O, Ag2CO3 and AgOAc at ambient temperature.
Finally, the third method utilizes the reaction of azolium salts with
silver salts under basic phase transfer conditions [5–7]. Among these
approaches, second route is more convenient and most frequently used.
In the past few decades since the discovery of cisplatin for the
treatment of cancer, a substantial amount of research has been directed
towards understanding the basic mode of action of this pharmacologi-
cally important compound [8]. However, chemotherapetic treatment
by cisplatin comes with a price of major side effects including nausea,
basis of structure-activity relationships. In this perspective, Ag(I)-
carbene complexes have shown exceptional anticancer and anti-
biogram activities, in particular for the treatment of chronic lung infec-
tions and cystic fibrosis [9]. Therefore, the organometallic silver
derivatives historically used to maintain human health, and also many
silver salts are being used in healing the wounds to prevent infection
[10]. However, literature survey revealed that the Ag(I)-carbene com-
plexes derived from benzimidazole derivatives showed remarkably high
activity against the human cancer cell line OVCAR-3 (ovarian), human
renal cancer cell line (Caki-1), MB157 (breast) and HeLa (cervical) cell
line [11,12].
Recently, Tacke and coworkers [13] reported a series of nitrile-
functionalized Ag(I)-NHC complexes derived from 4,5-dichloroimidazole
and benzimidazole cores and their anticancer activity against Caki-1
cancer cell line. These functionalized Ag(I) mono-carbene complexes
showed medium to high cytotoxicity with comparable IC50 values. In
the nearest past, we have reported the synthesis of a series of mono-,
and bis-carbene Ag(I)-NHC complexes [14]. Most notable among the
similarities of reported complexes with cisplatin are the linear C-Ag-C
fragment (N-Pt-Cl in cisplatin) and the basicity of the Ag ion to bind
cellular DNA of the infected cell to arrest cell multiplicity. In continuation
of our studies on the stable Ag(I)-NHC complexes showing potent anti-
cancer activity, herein we report design, synthesis, crystal structures
and anticancer activity of mono- and binuclear Ag(I)-NHC complexes
with different xylyl spacers.
⁎
Corresponding author. Tel.: +60 19 4118 262; fax: +60 604 653 3578.
E-mail address: rosenani@usm.my (R.A. Haque).
1387-7003/$ – see front matter © 2012 Elsevier B.V. All rights reserved.
doi:10.1016/j.inoche.2012.05.037

114
R.A. Haque et al. / Inorganic Chemistry Communications 22 (2012) 113–119
2. Experimental
2.2.3. Synthesis of 1,3–bis(3-allylimidazolium-1-ylmethyl)mesitylene
bis(hexafluorophosphate) (9) [18]
2.1. Materials, methods, and instruments
Salt 9 was prepared in an analogous fashion to 7 from 2,4-bis(3-
imidazol-1-yl methyl)mesitylene (0.59 g, 2.1 mmol) and allyl bromide
(0.7 g, 4.2 mmol) in acetonitrile (20 mL), recrystallized from acetoni-
trile to give colorless solid. Yield: 0.4 g (87%). 1H NMR (d6-DMSO,
500 MHz, 298 K): δ 2.21 (3 H, s, 1×Ar-CH3), 2.30 (6 H, s, 2×Ar-CH3),
4.83 (4 H, d, 2×N-CH2), 5.27 (4 H, d, 2×CH2), 5.48 (4 H, s, 2×benzylic
CH2), 6.03 (2 H, m, 2×CH), 7.20 (1H, s, 1×Ar-H), 7.63 (2 H, t, 2×
imidazolium H5′), 7.75 (2 H, t, 2×imidazolium H4′), and 8.94 (2 H, d,
2×imidazolium H2′). 13C{1H}NMR (125 MHz, d6-DMSO, 298 K): δ
16.5 (Ar-CH3), 20.2 (Ar-CH3), 48.2 (N-CH2), 51.7 (CH2), 120.6, 123.3
(imidazolium C5′ and C4′), 128.9, 129.2 (Ar-C), 136.5 (imidazolium
C2′), and 123.5, 136.3, 140.6 (Ar-C). FTIR (KBr disc) cm−1: 2856, 2932
ν(C\H), 1588, 1190 ν(imidazole ring C_N, C\N vibrations). Anal.
Calc. for C23H30N4F12P2: C 42.3, H 4.6, N 8.6%. Found: C 41.6, H 5.0, N
8.2%.
All chemicals and solvents were obtained from commercial
sources and all reagents and solvents were of analytical grade and
used without further purifications. The imidazole, Ag2O, mesitylene,
1,2-dibromomethyl benzene, 1,3- dibromomethyl benzene, 1,4-
dibromomethyl benzene, allyl bromide, 5-fluorouracil (standard), and
3-[4,5-yl]-2,5-dephenyltetrazolium bromide for the MTT assay were
purchased from Sigma–Aldrich. 1,2- , 1,3- and 1,4-methyl-bis(1′-
imidazole) benzene and 2,4-methyl-bis(1′-imidazole) mesitylene
were prepared according to the literature [15] method with slight mod-
ifications. The carbene ligand precursors were prepared according to an
established procedure [16–18]. The FT-IR spectra of the compounds
were recorded in potassium bromide disks using a Perkin Elmer 2000
system spectrometer in the range 4000 to 400 cm−1. The NMR spectra
were recorded in d6-DMSO using Bruker 500 MHz Ascend spectrometer
at ambient temperature with TMS as an internal reference. The instru-
ments are available in the School of Chemical Sciences, Universiti
Sains Malaysia. All the compounds were analyzed for C, H, and N
using a PerkinElmer 2400 Series II microanalyzer. The X-ray diffraction
data were collected using a Bruker SMART APEX2 CCD area-detector
diffractometer.
2.2.4. Synthesis of 1,4–bis(3-allylimidazolium-1-ylmethyl)benzene
bis-(hexafluorophosphate) (10)
Salt 10 was prepared in an analogous fashion to 7 from 1,4-
bis(N-imidazole-1-yl methyl)benzene (0.5 g, 2.1 mmol) and allyl bro-
mide (0.7 g, 4.2 mmol) in acetonitrile (20 mL), recrystallized from
acetonitrile/dichloromethane (3:1 v/v) to give colorless solid. Yield:
0.48 g (71.8%). 1H NMR (d6-DMSO, 500 MHz, 298 K): δ 4.86 (4 H, d,
2×N-CH2), 5.33 (4 H, d, 2×CH2), 5.47 (4 H, s, 2×benzylic CH2), 6.06
(2 H, m, 2×CH2), 7.50 (4 H, s, 4×Ar-H), 7.74 (2 H, t, 2×imidazolium
H5′), 7.85 (2 H, t, 2×imidazolium H4′), and 9.40 (2 H, d, 2×
imidazolium H2′). 13C{1H}NMR (125 MHz, d6-DMSO, 298 K): δ 49.9
(N-CH2), 50.4 (CH2), 119.6, 121.4, 121.7 (imidazolium C5′ and C4′),
127.9, 130.1, 134.0 (Ar-C) and 134.9 (imidazolium C2′). FTIR (KBr disc)
cm−1: ~2860, ~2925 ν(C\H), 1592, 1176 ν(imidazole ring C_N,
C\N vibrations). Anal. Calc. for C20H24N4F12P2: C 39.4, H 4.0, N 9.2%.
Found: C 39.9, H 3.6, N 9.6%.
2.2. Syntheses
2.2.1. Synthesis of 1,2–bis(3-allylimidazolium-1-ylmethyl)benzene
bis-(hexafluorophosphate) (7) [16]
A mixture of 1,2-bis(imidazole-1-ylmethyl)benzene (0.5 g, 2.1 mmol)
and allyl bromide (0.7 g, 4.2 mmol) in acetonitrile (20 mL) was refluxed
for 24 h. The solvent was removed under reduced pressure to result
pale-brown oil, which was directly converted to its corresponding
hexafluorophosphate counterpart by metathesis reaction using KPF6
(0.76 g, 4.0 mmol) in methanol (20 mL). The precipitate formed was
collected and washed with distilled water (2×5 ml) to remove unreacted
KPF6, and recrystallized from acetonitrile to give colorless solid. Yield
1.1 g, 80.3%. 1H NMR (d6-DMSO, 500 MHz, 298 K): δ 4.85 (4 H, d, 2×N-
CH2), 5.28–5.43 (4 H, d, 2×CH2), 5.58 (4 H, s, 2×benzylic CH2), 6.09
(2 H, m, 2×CH), 7.28–7.55 (4 H, m, 4×Ar-H), 7.74 (2 H, t, 2×
imidazolium H5′), 7.82 (2 H, t, 2×imidazolium H4′), and 9.15 (2 H, d,
2×imidazolium H2′). 13C{1H}NMR (125 MHz, d6-DMSO, 298 K): δ 49.9
(N-CH2), 51.9 (CH2), 121.3, 123.7, 123.8 (imidazolium C5′ and C4′),
130.5, 130.6, 132.4, 133.6 (Ar-C) and 137.3 (imidazolium C2′). FTIR
(KBr disc) cm−1: 2862, 2925 ν(C\H), 1576, 1192 ν(imidazole ring
C_N, C\N vibrations). Anal. Calc. for C20H24N4F12P2: C 39.4, H 4.0, N
9.2%. Found: C 39.2, H 3.6, N 8.9%.
2.2.5. Synthesis of Ag(I)-carbene complex 11
To a suspension of 7 (0.61 g, 1 mmol) in acetonitrile (50 mL) was
added Ag2O (0.458 g, 2 mmol). The mixture was refluxed at 60 °C for
24 h in glassware wrapped with aluminum foil to exclude light. The
resulted colorless solution was filtered through a pad of celite and
filtrate was slowly evaporated to precipitate a white solid. So obtained
solid was redissolved in acetonitrile (3 mL) and was added diethyl
ether (50 mL) to reprecipitate the complex as white solid. Yield:
96.4%. 1H NMR (d6-DMSO, 500 MHz, 298 K): δ 4.67 (4 H, d, 2×N-
CH2), 5.19 (4 H, d, 2×CH2), 5.32 (4 H, s, 2×benzylic CH2), 5.98 (2 H, m,
2×CH), 7.07 (4 H, m, 4×Ar-H), 7.25 (2 H, t, 2×imidazolium H5′), and
7.44 (2 H, t, 2×imidazolium H4′). 13C{1H}NMR (125 MHz, d6-DMSO,
298 K): δ 49.9 (N-CH2), 51.9 (CH2), 121.3, 123.8 (imidazolium C5′ and
C4′), 123.7, 130.5, 130.6, 132.4, 133.6 (Ar-C) and 177.3 (imidazolium
C2′-Ag). FTIR (KBr disc) cm−1: 2869, 2937 ν(C\H), 1496, 1088
ν(imidazole ring C\N vibrations). Anal. Calc. for C40H44N8F12P2Ag2:
C 42.1, H 3.9, N 9.8%. Found: C 41.5, H 3.8, N 9.2%.
2.2.2. Synthesis of 1,3–bis(3-allylimidazolium-1-ylmethyl)benzene
bis-(hexafluorophosphate) (8) [17]
Salt 8 was prepared in an analogous fashion to 7 from 1,3-bis(N-
imidazole-1-yl methyl)benzene (0.5 g, 2.1 mmol) and allyl bromide
(0.7 g, 4.2 mmol) in acetonitrile (20 mL), recrystallized from acetoni-
trile to give colorless solid. Yield: 0.41 g (61.3%). 1H NMR (d6-DMSO,
500 MHz, 298 K): δ 4.86 (4 H, d, 2×N-CH2), 5.36 (4 H, d, 2×CH2),
5.46 (4 H, s, 2×benzylic CH2), 6.00–6.13(2 H, m, 2×CH), 7.39–7.54
(4 H, m, 4×Ar-H), 7.76 (2 H, t, 2×imidazolium H5′), 7.98 (2 H, t,
2×imidazolium H4′), and 9.27 (2 H, d, 2×imidazolium H2′). 13C
{1H}NMR (125 MHz, d6-DMSO, 298 K): δ 51.9 (N-CH2), 52.5 (CH2),
121.3, 121.4, 123.5, 123.7 (imidazolium C5′ and C4′), 129.2, 129.5,
130.6, 132.3, 132.4 (Ar-C) and 137.1 (imidazolium C2′). FTIR (KBr
disc) cm−1: ~2860, 2920 ν(C\H), 1582, 1196 ν(imidazole ring C_N,
C\N vibrations). Anal. Calc. for C20H24N4F12P2: C 39.4, H 4.0, N 9.2%.
Found: C 39.1, H 3.6, N 9.1%.
2.2.6. Synthesis of Ag(I)-carbene complex 12
Complex 12 was prepared in an analogous fashion to 11 from
8 (0.61 g, 1 mmol) and Ag2O (0.458 g, 2 mmol) in acetonitrile (20 mL).
Complex was recrystallized from acetonitrile/diethyl ether to give
white solid. Yield: 96.1%. 1H NMR (d6-DMSO, 500 MHz, 298 K): δ 4.72
(4 H, d, 2×N-CH2), 5.16 (4 H, d, 2×CH2), 5.15 (4 H, s, 2×benzylic
CH2), 6.02 (2 H, m, 2×CH), 7.18 (4 H, m, 4×Ar-H), 7.49 (2 H, t,
2×imidazolium H5′), and 7.53 (2 H, t, 2×imidazolium H4′). 13C{1H}
NMR (125 MHz, d6-DMSO, 298 K): δ 49.0 (N-CH2), 55.0 (CH2), 118.3,
122.0, 124.5 (imidazolium C5′ and C4′), 132.0, 132.5, 135.0, 132.3,
137.4,139.0 (Ar-C) and 180.3 (imidazolium C2′-Ag). FTIR (KBr disc)
cm−1: ~2860, 2933 ν(C\H), 1488, 1093 ν(imidazole ring C\N

R.A. Haque et al. / Inorganic Chemistry Communications 22 (2012) 113–119
115
vibrations). Anal. Calc. for C20H22N4F6P1Ag1: C 42.1, H 3.9, N 9.8%. Found:
C 41.3, H 3.8, N 9.9%.
value of the reported compounds was described as that concentration
reducing cell proliferation of the control cells by 50%.
2.2.7. Synthesis of Ag(I)-carbene complex 13
3. Results and discussion
Complex 13 was prepared in an analogous fashion to 11 from 9
(0.653 g, 1 mmol) and Ag2O (0.458 g, 2 mmol) in acetonitrile (20 mL).
Complex was recrystallized from acetonitrile/diethyl ether to give
white solid. Yield: 94.0%. 1H NMR (d6-DMSO, 500 MHz, 298 K): δ 2.08
(3 H, s, 1×C2-CH3), 2.59 (6 H, s, 1×C1-CH3/1×C5-CH3), 4.63 (4 H, d,
2×N-CH2), 5.06 (4 H, d, 2×CH2), 5.40 (4 H, s, 2×benzylic CH2), 5.93
(2 H, m, 2×CH), 6.83 (1H, s, 1×C6-CH3), 7.13 (2 H, t, 2×imidazolium
H5′), and 7.20 (2 H, t, 2×imidazolium H4′). 13C{1H}NMR (125 MHz,
d6-DMSO, 298 K): δ 16.5 (C2-CH3),18.9 (C1-CH3/C5-CH3) 44.8 (N-CH2),
48.4 (CH2), 121.7, 122.6 (imidazolium C5′ and C4′), 127.3, 128.6(Ar-
C6), 122.3136.6, 138.9 (Ar-C) and 179.5 (imidazolium C2′-Ag). FTIR
(KBr disc) cm−1: ~2870, 2945 ν(C\H), 1482, 1064 ν(imidazole ring
C\N vibrations). Anal. Calc. for C23H28N4F6P1Ag1: C 45.0, H 4.6, N
9.1%. Found: C 45.8, H 4.6, N 9.2%.
3.1. Syntheses
Reactions involved in the syntheses of NHC precursors (7–10) and
their respective Ag(I) bis-carbene complexes (11–14) are summarized
in Schemes 1 and 2, respectively. Bis-imidazoles with ortho-, meta-
and para-xylene cores [15] reacted with allyl bromide (2 equivalents)
in acetonitrile to afford respective bis-imidazolium bromide salts as
thick brown oils in good yield. It is noteworthy that the N-allylation of
bis-imidazoles proceeded predominantly and the mono allyl substitut-
ed salts were not observed. So obtained bromide salts were directly
converted into their corresponding hexafluorophosphate counterparts
by salt metathesis reaction using KPF6 (2 equivalents) in methanol to af-
ford respective bis(hexafluorophosphate) salts 7–10 in good yield. Salts
7–10 were non-hygroscopic, stable to air and moisture, and soluble in
common organic solvents such as DMF, DMSO, acetone and acetonitrile.
All imidazolium salts were fully characterized by 1H, 13C NMR, and ele-
mental analysis. Additionally, we have reported the solid-state struc-
ture of salts 7, 8 and 9 were determined by single-crystal X-ray
diffraction method [16–18].
Ag(I) bis-carbene complexes 11, 12 and 14 were prepared in
exceptionally good yield (>90%) by in situ deprotonation reactions
of the corresponding bis(hexafluorophosphate) imidazolium salts
(7, 8 and 10) with Ag2O in acetonitrile at reflux conditions for 24 h.
The mixture was filtered through a pad of celite, and the filtrate was
slowly evaporated to precipitate a white solid. So obtained solid
was redissolved in acetonitrile and 150 mL of diethyl ether was
added to reprecipitate the solid in pure form. Basic Ag2O played two
2.2.8. Synthesis of Ag(I)-carbene complex 14
Complex 14 was prepared in an analogous fashion to 11 from 10
(0.61 g, 1 mmol) and Ag2O (0.458 g, 2 mmol) in acetonitrile (20 mL).
Complex was recrystallized from acetonitrile/diethyl ether to give
white solid. Yield: 96.3%. 1H NMR (d6-DMSO, 500 MHz, 298 K): δ
4.79 (4 H, d, 2×N-CH2), 5.15 (4 H, d, 2×CH2), 5.28 (4 H, s, 2×benzylic
CH2), 6.04 (2 H, m, 2×CH), 7.10 (4 H, s, 4×Ar-H), 7.44 (2 H, t,
2×imidazolium H5′), and 7.53 (2 H, t, 2×imidazolium H4′). 13C{1H}
NMR (125 MHz, d6-DMSO, 298 K): δ 51.9 (N-CH2), 52.5 (CH2), 121.3,
121.4, 123.5, 123.7 (imidazolium C5′ and C4′), 129.2, 129.5, 130.6,
132.3, 132.4 (Ar-C) and 176.7 (imidazolium C2′-Ag). FTIR (KBr disc)
cm−1: 2832, 2956 ν(C\H), 1479, ~1075 ν(imidazole ring C\N vibra-
tions). Anal. Calc. for C20H22N8F6P1Ag1: C 42.1, H 3.9, N 9.8%. Found: C
40.8, H 4.8, N 10.8%.
important roles in this reaction. First, it serves as
a base to
deprotonate the imidazolium proton and generates the free NHC and
secondly, it is a source of silver for metallation reaction. Additionally,
Ag(I)-carbene complex 13, with a substituted xylyl spacer, containing
imidazolium and allyl arms to facilitate the synthesis, and structural
comparison with its non-substituted counterpart was also prepared.
Complex 13 was also prepared in a analogues manner that of rest
Ag(I)-carbene complexes starting from bis-imidazolium salt 9. All
Ag(I)-carbene complexes are stable to air and moisture, and however,
sensitive to light, hence it is noteworthy to mention that it was not
2.3. Anticancer activity
The antiproliferative effects using MTT assay in HCT-116 cells after
72 h exposure to the imidazolium salts (7–10) and silver complexes
(11–14) were evaluated according to an established procedure [19].
For the experiments, the compounds (100 μL) were prepared freshly
as stock solutions in DMSO and diluted with the HCT 116 cell culture
medium to the final assay concentrations (0.1% V/V DMF). The IC50
Scheme 1. Synthesis of N-allyl substituted imidazol-2-ylidenes (7–10).

116
R.A. Haque et al. / Inorganic Chemistry Communications 22 (2012) 113–119
Scheme 2. Synthetic pathway to Ag(I)-bis-carbene complexes (11–14) from NHC precursors (7–10).
necessary to use dehydrated or degassed solvents or carry out the
reaction under inert atmosphere for the synthesis of these bench-
stable compounds. All complexes are insoluble in THF, benzene and
toluene, whereas, completely soluble in the aforementioned organic
solvents.
3.4. Crystal structures
The molecular structures of Ag(I)-carbene complexes 13 and 14
are depicted in Figs. 1 and 2, respectively. The crystal data and refine-
ment details for both the compounds are tabulated in Table 1, where-
as selected bond lengths and bond angles of Ag(I)-carbene complexes
13 and 14 are given along with their respective figure legend. Single
crystals of 13 and 14 suitable for X-ray diffraction analysis were
grown from slow diffusion of diethyl ether into a solution of complex
in acetonitrile at ambient temperature.
Complex 13 crystallizes in monoclinic space group C2/c and pos-
sesses a 3-dimentional framework. Both the counterions were shared
between two metal centers of two independent complex units. The
asymmetric unit of the complex consists of a Ag(I) complex cation
and two hexafluorophosphate anions. Three fluorides of each general-
position hexafluorophosphate anion are distorted over two sets of sites,
and one of the hexafluorophosphate anions is located at a crystallographic
center of symmetry. In the complex cation, the Ag(I) is coordinated by
3.2. FTIR spectral studies
All synthesized compounds 7–14 were characterized by FTIR spectral
technique. FTIR spectra of all the compounds showed a band of medium
intensity in the range 2840–2670 cm−1, which is assigned to aliphatic
and aromatic C\H vibrations. In the spectra of NHC precursors 7–10,
two sharp and medium intensity bands appeared in the range
1588–1565 and 1175–1190 cm−1, were ascribed to the imidazole ring
C_N and C\N stretching vibrations, this observation is in accordance
with the similar reported compounds [20,21]. However, in the complex
spectra these bands showed a shift of ~120 and ~25 cm−1, respectively,
indicating the formation of free C\N module in the imidazole core,
suggesting dequarternization followed by coordination of the carbene
carbon to Ag(I). Finally, the spectra of all compounds contain prominent
bands at around 1400, 1220 and 990 cm−1, and are assigned for the
other modes of vibrations of C\H, C_N and C\N modules.
3.3. NMR spectral studies
Structures of all reported compounds 7–14 in solution were
deduced from the 1H and 13C NMR spectra. The imidazolium C(2)-H
showed a typical absorption in the 1H NMR spectra of 7–10 in the
range δ 9–9.4, which is consistent with those of other reported imid-
azole based NHC precursors [22]. However, this peak is completely
disappeared in the complex spectra indicating coordination of the
carbene carbon after deprotonation. Three typical signals due to allyl
protons were resonated at around δ 4.8, 5.1 and 5.3 in the spectra.
Sharp singlets and multiplets at round δ 6.0 and 7.3–7.7 in all spectra
were attributed to the resonance of benzylic and aromatic protons,
respectively.
Inspection of 13C NMR spectra of all the complexes revealed a signif-
icant change of the carbene carbon chemical shift between imidazolium
NHC precursors 7–10 and bis-carbene complexes 11–14. The NMR
resonance of the carbene carbon nuclei of all bis-carbene complexes,
11–14, drastically shifted downfield (in the range δ 177–179) from
that of NHC precursors observed in the range δ 132–135. These reso-
nances were observed as singlet peaks without showing any carbene
carbon-Ag couplings, though similar complexes have been reported to
show such couplings [23–25]. Further, there were no significant
changes in the peak position of the other resonances were observed.
Fig. 1. X-ray structure of 13 with displacement ellipsoids drawn at 50% probability.
Counterion and solvent molecule are omitted due to clarity. Selected bond lengths
(Å): Ag1-C6 2.1215(17); Ag1-C18 2.1229(17); C3-N1 1.455(3); N1-C6 1.351(2); C6-N2
1.355(2); N2-C7 1.477(3); C7-C8 1.509(3); C10-C17 1.502(3); C17-N3 1.480(3); N3-C18
1.352(2); C18-N4 1.353(2); N4-C21 1.467(3). Selected bond angles (°): C6-Ag1-C18
176.81(7); C3-N1-C6 124.42(16); N1-C6-N2 104.02(15); N2-C7-C8 111.10(15);
C10-C17-N3 110.86(16); N3-C18-N4 103.63(15); C18-N4-C21 125.16(17).

R.A. Haque et al. / Inorganic Chemistry Communications 22 (2012) 113–119
117
13, the hexafluorophosphate anions link the complex cations into three-
dimensional networks through two different types of intermolecular
C\H—F hydrogen bonds with bond distances being 2.632 and 2.648 Å,
which are almost parallel to ab plane.
Complex 14 crystallizes in triclinic space group P-1 and has been
defined as a well ordered ionic Ag(I)-carbene complex, possesses a
3-dimentional framework. Molecular structure of 14 is consistent
with one binuclear Ag(I) complex cation and two hexafluorophosphate
anions. In the binuclear Ag(I) complex cation, each of the Ag(I) center is
coordinated by two carbene carbon atoms of the two ligands to form a
bilayer complex network with 2:2 ligand to metal ratio. The X-ray anal-
ysis revealed that both the sets of imidazole rings at Ag1 and Ag1A are
orientated at a dihedral angle of 7.50 (3)° and all imidazole rings are
almost perpendicular to the plane of central para-xylyl spacer. Both
the Ag atoms are in a two-coordinate distorted linear environment,
with two carbene carbon coordination (Ag1-C4=2.081(2) Å, Ag1-
C17=2.082(2) Å) with the same bond angle of about 172.50 (7)°. The
internal ring angle of all imidazole rings at the carbene center are in
the range 104.40(18)–104.60(18) °, which are in well agreement with
the reported complexes having similar ligand architecture [26–28].
Thus the complex adopted a two layer structure with two metal centers.
In the extended structure of 14, the hexafluorophosphate anions link
the binuclear complex cations into three-dimensional networks
through two different types of intermolecular C\H—F hydrogen
bonds with bond distances 2.478 and 2.578 Å, which are almost parallel
to bc and ac planes, respectively.
Fig. 2. X-ray structure of 14 with displacement ellipsoids drawn at 50% probability.
Counterions and solvent molecule are omitted due to clarity. Selected bond lengths
(Å): Ag1-C4 2.081(2); Ag1-C17 2.082(2); C3-N2 1.458(3); N2-C4 1.351(3); C4-N1
1.356(3); N1-C7 1.470(3); C7-C8 1.520(3); C11-C14 1.508(3); C14-N3 1.468(3);
N3-C17 1.350(3); C17-N4 1.355(3); N4-C18 1.457(3). Selected bond angles (°): C4-
Ag1-C17A 172.50(7); N2-C4-N1 104.40(18); C3-N2-C4 122.82(19); N1-C7-C8 110.48(18);
C11-C14-N3 111.39(17); N3-C17-N4 104.60(18); N4-C18-C19 113.60(19).
3.5. Anticancer activity
Percentage inhibition of cell proliferation was determined at
different concentrations of the test compounds ranging from 1.5625
to 100 μM. Figs. 3 and 4 show the effect of concentration of test com-
pounds on HCT 116 cell line after 72 h of incubation time.
two carbene carbon atoms of the ligand, and both the imidazole rings are
orientated at a dihedral angle of 3.19 (3)° and are almost perpendicular to
the plane of central mesitylene spacer. The Ag atom is in a two-coordinate
distorted linear environment, with two carbene carbon coordination
(Ag1-C6=2.1215(17) Å, Ag1-C18=2.1229(17) Å) the C6-Ag1-C18
bond angle about 176.81 (7)°; while, the molecule possesses ‘chair-like’
shape. The internal ring angle of imidazole (N–C–N) at the carbene center
are 104.02(15) and 103.63(15) Å for N1-C6-N2 and N3-C18-N4, respec-
tively, which are in well agreement with the reported complexes con-
taining similar ligand architecture [26–28]. In the extended structure of
All reported metal and non-metal imidazole derivatives have been
tested for anticancer activity against human derived cancer cell was
investigated using HCT 116 cell line and a standard bioassay, MTT
method after incubating for 72 h. Cytotoxic potency of the imidazolium
salts and their respective Ag(I)-carbene complexes was measured in
terms of IC50 (in μM) values. Imidazolium salts 7, 8 and 10 displayed
no effect on either cell growth or viability for the cancer cell line even
after prolonged salt exposure. However, the analogues salt 9 displayed
significant antiproliferative effect against tested cell line, yielding IC50
value 15.9 μM. This higher activity of the salt 9 among others is attributed
to the structural difference of the salt especially with the central core. On
the other hand, the experiments performed with Ag(I)-carbene com-
plexes 11–14 on aforementioned cell line in turn showed that the com-
plexes are significantly active as very efficient anticancer drugs, with
IC50 values in the range 1.1–5.3 μM. The most active complexes 14, 13
Table 1
Crystal data and structure refinement details for carbene complexes 13 and 14.
13
14
Formula
Formula weight
Crystal system
Space group
Unit cell dimensions
a (Å)
b (Å)
c (Å)
α (°)
β (°)
C
23H28Ag1F6N4P1
C40H40Ag4F12N20O4P4
613.33
Monoclinic
C2/c
1790.08
Triclinic
P1
24.0323 (10)
8.7154(3)
25.5864 (10)
90.00
112.5630(10)
90.00
9.07290(10)
12.0869(2)
12.5392(2)
115.4880(10)
90.5560(10)
95.6990(10)
1233.02(3)
1
γ (°)
V (Å3)
4969.3(3)
8
Z
Density (calcd) (gm/cm3) 1.640
2.411
2.029
876
Abs coeff (mm−1
F(000)
)
0.940
2480
Crystal size (mm)
Temperature (K)
Radiation (Å)
θ Min, max (°)
Dataset
0.57×0.49×0.38
293
MoKa 0.71073
1.72, 30.09
−33:33, −10:12, −36:36 −13:13, −18:18, −19:19
25,993
0.42×0.33×0.32
293
MoKa 0.71073
1.80, 32.75
Tot.; uniq. data
R (int)
25,122
0.0173
0.0176
Nref, Npar
R, wR2, S
7246, 383
0.0266, 0.0772, 1.077
9029, 311
0.0373, 0.1532, 1.324
Fig. 3. MTT assay results of NHC precursor 9, and Ag-carbene complexes 11–14 vs the
HCT 116 cell lines.

118
R.A. Haque et al. / Inorganic Chemistry Communications 22 (2012) 113–119
11 compared to other complexes, also displayed antiproliferative and
cytotoxic effects in these cancer cell line, with IC50 value of 5.2 μM,
which is almost equal to the standard used. Further, complex 13 dis-
played 15 times higher activity compared to its NHC precursor 9, and
the activity of all other complexes could not be compared with their
respective ligand counterparts as they are all totally inactive against
HCT 116 cell line.
HCT116 cell images were taken under an inverted phase-contrast
microscope at ×200 magnification with a digital camera at 72h after
treatment with the samples. Cells from the control group showed
fully confluent growth. Treatment with 13 showed marked inhibition
in cell proliferation as the population of the cells reduced drastically
whereas its ligand precursor 9 showed mild antiproliferative effect
as the IC50 was moderately high (15.9 μM). Complex 14 exhibited
strong cytotoxicity as the cells showed abnormal morphology. Photo-
micrograph of complex 12 treated cells showed cytotoxicity as the
IC50 found to be considerably low (1.3 μM). Treatment with complex
11 showed the modest inhibition but all cells from the group showed
unhealthy transformation as the figure depicts that large number of
vacuoles can be seen in the 11 treated cells. Images of HCT116 cells
treated with compounds 9 and 11–14 after 72 h of incubation are
depicted in Fig. 5.
Fig. 4. Effects of increasing amounts of 9, 11–14 on the percentage inhibition of cell
proliferation.
and 12 displayed IC50 value of 0.9, 1.1 and 1.3 μM, respectively, which are
almost four times more active than the standard used, i.e., 5-fluorouracil
(IC50 =5 μM) after 72 h of incubation. Although less effective complex
It is thus concluded that the number of Ag centers within the com-
plex molecule and the substitutions on the ligand determines its
Fig. 5. Images of HCT116 cells treated with compounds 9 and 11–14 after 72 h of incubation.

R.A. Haque et al. / Inorganic Chemistry Communications 22 (2012) 113–119
119
antiproliferative and cytotoxic activity, which was found for HCT 116
cell line, to increase with the number of Ag(I) ions coordinated to the
carbene ligand and the presence of methyl substitutions at the appro-
priate position of the central spacer moiety [29]. In contrast, the
macrocyclic\like Ag(I) complexes 11 and 14 were found to display
clear and distinct cytotoxic activities toward the tested cancer cell
line, could be ascribed to their structural difference with the central
spacer moiety.
free of charge via http://www.ccdc.cam.ac.uk/conts/retrieving.html,
or from the Cambridge Crystallographic Data Centre, 12 Union Road,
Cambridge CB2 1EZ, UK; fax: (+44) 1223-336-033; or e-mail:
deposit@ccdc.cam.ac.uk. Crystal packing diagrams of the complexes
are also given. Supplementary data to this article can be found online
at http://dx.doi.org/10.1016/j.inoche.2012.05.037
References
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Acknowledgments
RAH thanks Universiti Sains Malaysia (USM) for the Short term
grant (203/PKIMIA/6311123) and the Research University (RU) grant
1001/PKIMIA/811157. MZG thanks USM for the RU grant 1001/
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Appendix A. Supplementary material
CCDC 853549 and 853551 contains the supplementary crystallo-
graphic data for 13 and 14, respectively. These data can be obtained