Molecular structures and antiproliferative activity of side-chain saturated and homologated analogs of 2-chloro-3-(n-alkylamino)-1,4-napthoquinone

Article abstract of DOI:10.1016/j.molstruc.2013.06.062

Side chain homologated derivatives of 2-chloro-3-(n-alkylamino)-1,4- naphthoquinone {n-alkyl: pentyl; L-5, hexyl; L-6, heptyl; L-7 and octyl; L-8} have been synthesized and characterized by elemental analysis, FT-IR, ¹H NMR, UV-visible spectroscopy and LC-MS. Compounds, L-4, {n-alkyl: butyl; L-4}, L-6 and L-8 have been characterized by single crystal X-ray diffraction studies. The single crystal X-ray structures reveal that L-4 and L-8 crystallizes in P21 space group, while L-6 in P2₁/c space group. Molecules of L-4 and L-8 from polymeric chains through C AH- ? O and NAH- ? O close contacts. L-6 is a dimer formed by NAH- ? O interaction. Slipped π-π stacking interactions are observed between quinonoid and benzenoid rings of L-4 and L-8. Orientations of alkyl group in L-4 and L-8 is on same side of the chain and polymeric chains run opposite to one another to form zip like structure to the alkyl groups. Antiproliferative activities of L-1 to L-8{n-alkyl: methyl; L-1, ethyl; L-2, propyl; L-3 and butyl; L-4} were studied in cancer cells of colon (COLO205), brain (U87MG) and pancreas (MIAPaCa2) where L-1, L-2 and L-3 were active in MIAPaCa2 (L-1 = L-2 > L-3) and COLO205 (L-2 = L-3 > L-1) and inactive in U87MG. From antiproliferative studies with compounds L-1 to L-8 it can be concluded that homologation of 2-chloro-3-(n-alkylamino)-1,4-napthoquinone with saturated methyl groups yielded tissue specific compounds such as L-2 (for MIAPaCa2) and L-3 (for COLO205) with optimal activity.

Full text of DOI:10.1016/j.molstruc.2013.06.062

Journal of Molecular Structure 1049 (2013) 355–361
Contents lists available at SciVerse ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstruc
Molecular structures and antiproliferative activity of side-chain
saturated and homologated analogs of
2-chloro-3-(n-alkylamino)-1,4-napthoquinone
Sanjima Pal a, Mahesh Jadhav b, Thomas Weyhermüller c, Yogesh Patil d, M. Nethaji d, Umesh Kasabe b,
Laxmi Kathawate b, V. Badireenath Konkimalla a, , Sunita Salunke-Gawali b,
⇑
⇑
a School of Biological Sciences, National Institute of Science Education and Research (NISER), Bhubaneswar 751 005, India
b Department of Chemistry, University of Pune, Pune 411 007, India
c MPI für Chemische Energiekonversion, Stiftstr. 34-36, 45470 Müllheim an der Ruhr, Germany
d Inorganic and Physical Chemistry Department, Indian Institute of Science, Bangalore, India
h i g h l i g h t s
ꢀ
ꢀ
ꢀ
ꢀ
ꢀ
Derivatives of 2-chloro-3-(n-alkylamino)-1,4-naphthoquinone are discussed.
n-butyl, n-octyl derivative from polymeric chain through CAHꢁ ꢁ ꢁO and NAHꢁ ꢁ ꢁO close contacts.
Molecules of n-hexyl derivative show dimer through NAHꢁ ꢁ ꢁO interaction.
Antiproliferative activities were studied against in cancer cells of colon brain and pancreas.
Tissue specific antiproliferative activity is observed for ethyl and for propyl derivative.
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 1 June 2013
Received in revised form 24 June 2013
Accepted 24 June 2013
Available online 2 July 2013
Side chain homologated derivatives of 2-chloro-3-(n-alkylamino)-1,4-naphthoquinone {n-alkyl: pentyl;
L-5, hexyl; L-6, heptyl; L-7 and octyl; L-8} have been synthesized and characterized by elemental analy-
sis, FT-IR, 1H NMR, UV–visible spectroscopy and LC–MS. Compounds, L-4, {n-alkyl: butyl; L-4}, L-6 and
L-8 have been characterized by single crystal X-ray diffraction studies. The single crystal X-ray structures
reveal that L-4 and L-8 crystallizes in P21 space group, while L-6 in P21/c space group. Molecules of L-4
and L-8 from polymeric chains through CAHꢁ ꢁ ꢁO and NAHꢁ ꢁ ꢁO close contacts. L-6 is a dimer formed by
Keywords:
p–p stacking
NAHꢁ ꢁ ꢁO interaction. Slipped
p–p stacking interactions are observed between quinonoid and benzenoid
rings of L-4 and L-8. Orientations of alkyl group in L-4 and L-8 is on same side of the chain and polymeric
chains run opposite to one another to form zip like structure to the alkyl groups. Antiproliferative activ-
ities of L-1 to L-8{n-alkyl: methyl; L-1, ethyl; L-2, propyl; L-3 and butyl; L-4} were studied in cancer cells
of colon (COLO205), brain (U87MG) and pancreas (MIAPaCa2) where L-1, L-2 and L-3 were active in
MIAPaCa2 (L-1 = L-2 > L-3) and COLO205 (L-2 = L-3 > L-1) and inactive in U87MG. From antiproliferative
studies with compounds L-1 to L-8 it can be concluded that homologation of 2-chloro-3-(n-alkylamino)-
1,4-napthoquinone with saturated methyl groups yielded tissue specific compounds such as L-2 (for
MIAPaCa2) and L-3 (for COLO205) with optimal activity.
Aminonaphthoquinone
Hydrogen bonding
Antiproliferative activity
Ó 2013 Elsevier B.V. All rights reserved.
1. Introduction
cancers. All of these drug molecules contain quinone moieties in
their framework. Quinonoid systems are related to their capacity
Anthracyclines are a class of drugs used in cancer chemother-
apy. They include donorubicin, doxorubicin, epirubicin and idaru-
bicin. These compounds are used in treatment of many cancers,
such as leukemias, lymphomas, breast, uterine, ovarian, and lung
to generate free radicals or semiquinones in redox reactions hence
quinone derivatives have various pharmacological applications [1].
The clinical importance of 1,4-naphthoquinone has stimulated
enormous interest in this class of compounds [2]. The synthesis
of alkyl amino derivatives of 1,4-naphthoquinones is of prime
interest, since these compound exhibit strong antibacterial [3],
antimalarial and antitumor activity [4]. Furthermore, the amino-
1,4-naphthoquinone moiety is the component of molecular frame-
work of many natural products and has been used as synthetic key
⇑
Corresponding authors. Tel.: +91 2025601397x53; fax: +91 2025693981
(S. Salunke-Gawali), tel.: +91 9439864399; fax: +91 6742304121 (V.B. Konkimalla).
E-mail addresses: badireenath@niser.ac.in (V.B. Konkimalla), sunitas@chem.
unipune.ac.in (S. Salunke-Gawali).
0022-2860/$ - see front matter Ó 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.molstruc.2013.06.062

356
S. Pal et al. / Journal of Molecular Structure 1049 (2013) 355–361
intermediate for synthesis of several compounds with important
biological applications [5]. The amino and thioether substituted
derivatives of 1,4-naphthoquinones possess extremely rich biolog-
ical activities because of their redox potentials [5]. Single crystal X-
ray structures of 1,4-naphthoquinone derivatives show molecular
associations and the molecules forms three dimensional polymeric
network through CAHꢁ ꢁ ꢁO and OAHꢁ ꢁ ꢁO hydrogen bonding and
using melting point apparatus (Make- METTLER) and are corrected
using DSC (Differential Scanning Calorimetry) technique (Make-
METTLER, STAR SW 8.10).
2.2. X-ray crystallographic data collection and refinement of the
structures
p–p stacking interactions of benzenoid and quinonoid rings [6].
Single crystals of complexes L-4 and L-8 were coated with per-
fluoropolyether, picked up with nylon loops and were mounted in
the nitrogen cold stream of the diffractometers. Monochromated
This molecular associations through hydrogen bonding may lead
to stabilization of radical species and the redox cycling will decide
the biological activity of 1,4-naphthoquinones.
Mo K
a radiation (Bruker-AXS Kappa Mach3 with Incoatec Helios
Aminonaphthoquinone derivatives, 2-chloro-3-(n-alkylamino)-
1,4-naphthoquinone (n-alkyl: pentyl; L-5, hexyl; L-6, heptyl; L-7
and octyl; L-8) (Scheme 1) are synthesized and characterized by
various analytical tools in present investigation. Molecular associ-
ations of L-4, L-6 and L-8 are revealed by single crystal X-ray dif-
fraction studies.
mirror, k = 0.71073 Å) from a Mo-target rotating-anode X-ray
source was used for L-4. Intensity data of L-8 were collected on a
Bruker AXS X8 Proteum diffractometer with focusing mirror optics
(Cu Ka, k = 1.54178 Å). Final cell constants were obtained from
least squares fits of several thousand strong reflections. Intensity
data were corrected for absorption using intensities of redundant
reflections with the program SADABS [8]. The structures were
readily solved by direct methods and subsequent difference Fou-
rier techniques. The Siemens ShelXTL [9] software package was
used for solution and artwork of the structures, ShelXL97 [10]
was used for the refinement. All non-hydrogen atoms were aniso-
tropically refined and hydrogen atoms were placed at calculated
positions and refined as riding atoms with isotropic displacement
parameters. Crystallographic data of the compounds are listed in
Table 1. Crystals of the L-6 suitable for diffraction studies were
chosen carefully under a microscope. The unit cell parameters
and the intensity data were collected on a Bruker SMART APEX
Till date several drugs are available in the market for chemo-
therapy but most of them have limited use due to unwanted and
side/toxic effects. These side effects are mainly attributed to bind-
ing of the drug to unintended targets or tissues; urging a need for
more tissue specific and potent therapeutics. Therefore, selectivity
and specificity are the important parameters to be considered at a
pre-screening level for identification potential anticancer lead
compounds. Here in this study, antiproliferative activities of series
of derivatives (n-alkyl: methyl; L-1, ethyl; L-2, propyl; L-3 and bu-
tyl; L-4) along with L-5 to L-8 derivatives were carried out against
three cell lines COLO205 (human colorectal adenocarcinoma),
U87MG (human primary glioblastoma) and MIAPaCa-2 (human
pancreatic carcinoma) are studied. Further, the effect of homologa-
tion on the aminonaphthoquinone moiety (compounds L-1 to L-8)
with respect to its tissue specific antiproliferative activity is re-
ported in terms of degree of selectivity in the above-mentioned
three cell lines.
CCD diffractometer, both equipped with a fine-focus Mo K
a
(k = 0.71073 Å) X-ray source. The SMART software was used for
data acquisition and the SAINT software for data reduction.
Absorption corrections were made using SADABS [8] program.
The structure was solved and refined using the SHELXL-97 [10]
programs. The crystal data was collected at room temperature
and solved by direct methods. All the non-hydrogen atoms were
refined anisotropically. Hydrogen atoms were located by difference
Fourier synthesis and refined isotropically. Crystallographic data of
the compounds are listed in Table 1.
2. Experimental
2.1. General materials and methods
2.3. Synthesis
The materials used viz. 2,3-dichloro-1,4-naphthoquinone, pen-
tyl amine, hexyl amine, heptyl amine and octyl amine were pur-
chased from Sigma–Aldrich. The reactant 2,3-dichloro-1,
4-naphthoquinone have been recrystallized from methanol and
used for further reactions. The solvents used such as dichlorometh-
ane, toluene, methanol were of analytical grade and purchased from
Merck chemicals, India. Solvents were distilled in nitrogen atmo-
sphere by standard methods [7] and dried where ever necessary.
FT-IR spectra of the compounds (Fig. S1) have been recorded be-
tween 4000–400 cmꢂ1 as KBr pellets on SHIMADZU FT 8400 spec-
trometer and 1H NMR of ligands (Fig. S2) are recorded in CDCl3 and
DMSO-d6, on Varian Mercury 300 MHz with TMS (tetramethylsil-
ane) used as a reference. Elemental analysis has been performed
on Thermo Finnigan EA 1112 Flash series elemental Analyzer. Mass
of L-5 to L-8 has been determined using LC–MS 2010-eV (Make
SHIMADZU). Melting points of L-5 to L-8 (Fig. S3) were determined
All four L-5 to L-8, 2-chloro-3-(n-alkylamino)-1,4-naphthoqui-
nones have been synthesized from 2,3-dichloro-1,4-naphthoqui-
none and using respective n-alkylamines. 1 g of 2,3-dichloro-1,
4-naphthoquinone (4.4 mM) was dissolved in 25 ml of dichloro-
Table 1
Crystallographic data for complexes L-4, L-6 and L-8.
L-4
L-6
L-8
Chem. formula
Fw
C
14H14ClNO2
C16H18ClNO2
291.76
P21/c
5.512(1)
13.263(2)
19.213(3)
93.246(3)°
1402.4(13)
4
293(2) K
1.382
12,010/27.7
3310
C18H22ClNO2
319.82
263.71
Space group
a (Å)
b (Å)
c (Å)
b (°)
P21, No. 4
7.3733(13)
4.5915(8)
18.024(3)
90.853(3)
610.1(2)
2
P21, No. 4
7.4007(5)
4.7305(3)
22.8178(15)
91.218(4)
798.65(9)
2
V (Å)3
Z
O
where,
T (K)
100(2)
1.435
100(2)
1.330
q
calcd (g cmꢂ3
)
L-1: R=CH3 ; L-2: R=C2H5 ; L-3:
R=C3H7; L-4: R=C4H9
L-5: R=C5 H11 ; L-6: R=C6H13 ; L-
7: R=C7H15; L-8: R=C8H17
NHR
Cl
refl. collected/2Hmax
unique refl./I > 2 (I)
No. of params/restr.
k (Å/ (K
), cmꢂ1
R1a/goodness of fitb
wR2c (I > 2
(I))
Residual density (eÅꢂ3
17,161/61.42
3789/3551
168/1
0.71073/3.06
0.0286/1.040
0.0655
16,798/127.38
2544/2388
204/1
1.54178/21.67
0.0513/1.122
0.1170
r
253/0
l
a
)
0.71073/2.73
0.0490/1.073
0.1135
+0.259/ꢂ0.265
O
r
)
+0.33/ꢂ0.20
+0.63/ꢂ0.42
Scheme 1. Molecular formula of L-1 to L-8.

S. Pal et al. / Journal of Molecular Structure 1049 (2013) 355–361
357
methane and stirred for 15 min, which was then followed by drop
wise addition of n-alkylamine solution (5.2 mM) L-5; pentyl amine,
L-6; hexyl amine, L-7; heptyl amine, L-8; octyl amine). Reaction
mixture has been stirred for 24 h. at room temperature (26 °C).
Progress of reaction and purity of the ligands have been monitored
by thin layer chromatography. Red color solids obtained by evapo-
ration of the solvent. The crude product was purified by column
chromatography in toluene: methanol (9:1) solvent system. The
crystals for X-ray analysis are obtained by recrystalization in
appropriate solvents.
J = 7.9 Hz), 1.378 (m, 2H, J = 3.4 Hz), 0.931 (d, 1H, J = 7.2 Hz). 13C
NMR; (500 MHz, DMSO-d6) d: 13.87, 22.02, 25.98, 28.53, 28.59,
30.70, 31.14, 39.34, 39.51, 40.05, 43.95, 125.77, 126.46, 128.91,
132.08, 132.56, 134.89, 180.17. UV-; (kmax, nm, DMSO): 476, 338,
300. LC-MS (EI); m/z: 319.95 (M++H).
2.4. Cell lines
COLO205 (human colorectal adenocarcinoma), U87MG (human
primary glioblastoma) and MIAPaCa2 (human pancreatic carci-
noma) cell lines were obtained from National Centre for Cell Sci-
ence (Pune, India). COLO205 cell line was cultured routinely in
RPMI-1640 medium (HiMedia, India) while U87MG and MIAPaCa2
in DMEM media. Both media were supplemented with 2 mM glu-
2.3.1. Characterization of 2-chloro-3-pentylamino-1,4-
naphthoquinone
L-5. Red crystals, 0.465 g (36.4%). m.p.: 97.1 °C. Anal. data calcd.
for C15H16Cl NO2: C, 64.87; H, 5.81; N, 5.04. Found C, 64.36; H, 5.78;
N, 4.73. FT-IR (KBr, vmax/cmꢂ1): 3311, 2933, 2862, 1674, 1641,
1599, 1570, 1516, 1446, 1332, 1300, 1261, 1230, 1132, 1072,
1024, 895, 821, 723, 684, 650, 605, 563, 522, 447, 412. 1H NMR;
(300 MHz, DMSO-d6) d: 8.155 (d, 1H, J = 6 Hz), 8.033 (d, 1H,
J = 7.8 Hz), 7.721 (t, 1H, J = 7.3 Hz), 7.620 (t, 1H, J = 7.9 Hz), 6.082
(br. s, 1H), 3.845(q, 2H, J = 6.8 Hz), 1.675 (m, 4H, J = 7.5 Hz), 1.388
(m, 4H, J = 6.3 Hz), 0.904 (d, 1H, J = 6.6 Hz). 13C NMR; (500 MHz,
DMSO-d6) d: 13.84, 21.79, 28.23, 30.45, 39.00, 43.85, 125.76,
126.45, 132.08,132.55, 134.88, 145.11, 175.31,180.15. UV–Vis;
(kmax, nm, DMSO): 475, 338, 302. LC–MS (EI); m/z: 277.95 (M++H).
tamine (Himedia, India), antibiotics (100
l/mL penicillin A and
100 /mL streptomycin; Himedia, India) and 10% heat-inactivated
l
fetal bovine serum (HiMedia, India). All cell lines were cultured in
25 cm2 flasks with loosened caps and incubated in humidified air
containing 5% CO2 at 37 °C. Origin pro software and MS-Excel were
used for data analysis.
2.5. XTT assay for anti-proliferative activity
The effect of isolated L-1 to L-8 on the viability of the cell lines
were measured using XTT assay. The cytotoxicity were evaluated
based on the amount of 2,3-bis[2-methoxy-4-nitro-5-sulfophe-
nyl]-2H-tetrazolium-5-carboxanilide inner salt formed by the via-
ble cells in the treated wells. Fresh stock solutions of 3 and 4 were
prepared in DMSO at a concentration of 100 mM. Serial dilution in
50:50; media: DMSO mixtures produced stock solutions of com-
2.3.2. Characterization of 2-chloro-3-hexylamino-1,4-naphthoquinone
L-6. Red crystals, 0.603 g (46.9%). m.p.: 84.95 °C. Anal. data
calcd. for C16H18ClNO2: C, 65.86; H, 6.22; N, 4.8; Found C, 65.20;
H, 6.23; N, 5.12. FT-IR (KBr, vmax/cmꢂ1): 3309, 3261, 2964, 2947,
2916, 2850, 1672, 1633, 1597, 1568, 1508, 1444, 1377, 1334,
1294, 1246, 1224, 1143, 1124, 1076, 1057, 999, 900, 821, 792,
725, 682, 644, 621, 547, 462, 432. 1H NMR; (300 MHz, DMSO-d6)
d: 8.155 (d, 1H, J = 7.2 Hz), 8.035 (d, 1H, J = 6.9 Hz), 7.720 (t, 1H,
J = 7.2 Hz), 7.622 (t, 1H, J = 7.0 Hz), 6.078 (br. s, 1H), 3.845(q, 2H,
J = 6.8 Hz), 1.664 (m, 5H, J = 7.5 Hz), 1.331 (m, 5H, J = 8.5 Hz),
0.880 (d, 1H, J = 6.9 Hz). 13C NMR; (500 MHz, DMSO-d6) d: 13.82,
21.98, 25.70, 30.73, 30.89, 39.17, 39.93, 40.01, 43.90, 125.78,
pounds ranging from 10ꢂ8 M to 10ꢂ4 M. About 50
lL of the cell
suspension diluted to a final density of 1 ꢃ 105 cells/mL were
sowed into each well of a 96-well culture plate (Axygen, USA)
and treated with the varying concentrations of compounds in
duplicates. The compound treated 96-well plates were left for
incubation at 37 °C and 5% CO2 in humidified atmosphere for
72 h. XTT reagent mixture was freshly prepared with XTT-labeling
reagent and electron-coupling reagent in a ratio of 50:1. Post 72 h
126.48, 132.09, 132.575, 134.90, 180.18 UV–Vis; (kmax
, nm,
of incubation, 50 lL of this mixture was added to each of the 96
DMSO): 476, 338, 301. LC–MS (EI); m/z: 291.95 (M++H).
wells. The plates were incubated at 37 °C, 5% CO2 in humidified
atmosphere and read out after optimal color development in each
of the wells. Quantification of cell viability was performed in an
ELISA plate reader (Bio-Rad, München, Germany) at 490 nm with
a reference wavelength of 655 nm. The above-mentioned cell lines
treated with doxorubicin served as a positive control for cytotoxic-
ity and to evaluate the same [11,12].
2.3.3. Characterization of 2-chloro-3-heptylamino-1,4-
naphthoquinone
L-7. Red crystals, 0.577 g (42.8%). m.p.: 86.69 °C. Anal. data
calcd. for C17H20Cl NO2: C, 66.77; H, 6.59; N, 4.58; Found C,
68.81; H, 6.67; N, 4.73; FT-IR (KBr, vmax/cmꢂ1): 3313, 2926, 2856,
1674, 1641, 1599, 1568, 1514, 1442, 1371, 1330, 1298, 1259,
1163, 1132, 1072, 1022, 871, 821, 721, 682, 650, 603, 563, 526,
441. 1H NMR; (300 MHz, DMSO-d6) d: 8.154 (d, 1H, J = 7.5 Hz),
8.036 (d, 1H, J = 7.8 Hz), 7.728 (t, 1H, J = 7.5 Hz), 7.621 (t, 1H,
J = 7.6 Hz), 6.085 (br. s, 1H), 3.844(q, 2H, J = 7.0 Hz), 1.688 (m, 4H,
J = 7.1 Hz), 1.578 (d, 4H, J = 8.2 Hz), 1.333 (m, 4H, J = 8.2 Hz),
0.884 (d, 1H, J = 6.7 Hz). 13C NMR; (500 MHz, DMSO-d6) d: 13.88,
21.98, 25.98, 28.32, 30.74, 31.14, 43.88, 125.78, 126.47, 129.83,
132.09, 132.57, 134.90, 145.14, 145.34, 175.33, 180.18. UV–Vis;
(kmax, nm, DMSO): 476, 338, 300. LC–MS (EI); m/z: 305.95 (M++H).
3. Result and discussion
Biological activity of quinones depends on their ortho- and
para- tautomeric form [13]. Three mechanisms were proposed to
cytotoxic activity of quinones in various biological systems [14].
These mechanisms involve generation of active oxygen species
by redox cycling, intercalation between nucleotides in the DNA
double helix or alkylation of biomolecules. Quinones generally ac-
cepts one and two electrons by redox cycling to form in situ corre-
sponding radical anion and dianions species respectively, this
creates intracellular hypoxic conditions due to excess of superox-
ide radical [14a]. This radical will be converted to hydrogen perox-
ide by superoxide dismutase enzyme or it may form hydroxyl
radical by Fenton reaction[14b,c]. These oxygen intermediates or
reactive oxygen species (ROS) may react directly with DNA or other
biomolecules such as proteins and lipids, which leads to cell dam-
age [14g,h]. Cytotoxicity of quinones to mammalian and cancer
cells can also be explained due inhibition of topoisomerases, a
2.3.4. Characterization of 2-chloro-3-octylamino-1,4-naphthoquinone
L-8. Red crystals, 0.591 g (41.7%). m.p.: 89.95 °C. Anal. data
calcd. for C18H22ClNO2: C, 67.60; H, 6.93; N, 4.38; Found C, 67.05;
H, 6.88; N, 4.39; FT-IR (KBr, vmax/cmꢂ1): 3313, 2924, 2854, 1674,
1641, 1597, 1570, 1514, 1442, 1371, 1330, 1298, 1259, 1132,
1072, 1003, 823, 721, 684, 650, 565, 530. 1H NMR; (300 MHz,
DMSO-d6) d: 8.157 (d, 1H, J = 7.8 Hz), 8.034 (d, 1H, J = 7.5 Hz),
7.727 (t, 1H, J = 7.3 Hz), 7.622 (t, 1H, J = 7.6 Hz), 6.083 (br. s, 1H),
3.848(q, 2H, J = 6.8 Hz), 1.697 (m, 2H, J = 7.1 Hz), 1.475 (m, 2H,

358
S. Pal et al. / Journal of Molecular Structure 1049 (2013) 355–361
group of enzyme that are important for DNA replication in
cells[14f].
assigned to n–pꢅ charge transfer transition [16], this imparts red
color to all the compounds.
Biological activity of the quinones could also be related to their
interactions with biomolecules. Noncovalent interactions and p–p
3.2. X-ray crystal structures of L-4, L-6 and L-8
stacking ability of quinone derivatives will play major role in bind-
ing of quinone molecules to the biomolecules [15]. Single crystal
structures of several previously reported [6] naphthoquinone li-
gands are polymeric in nature where intermolecular CAHꢁ ꢁ ꢁO,
OAHꢁ ꢁ ꢁO and NAHꢁ ꢁ ꢁO interactions were common in these ligands.
L-1 and L-3 crystallizes in triclinic space group P-1, while
L-2 crystallizes in orthorhombic space group Pca2 (CCDC 907096-
907098). L-1 to L-3 shows, intra (NAHꢁ ꢁ ꢁO) as well as intermolec-
ular (NAHꢁ ꢁ ꢁO and CAHꢁ ꢁ ꢁO) hydrogen bonding interactions.
Some of them do possess
p–p stacking interactions, which could
Quinonoid rings of adjacent chains in L-1 and L-3 are
while this interaction is absent in L-2.
p–p stacked,
help them to intercalate with DNA.
To reveal their roles in molecular interactions besides redox cy-
cling, we have synthesized homologated series of aminonaphtho-
ORTEP plots of L-4, L-6 and L-8 are shown in Fig. 1a–c respec-
tively and crystallographic data is represented in Table 1. L-4 and
L-8 crystallizes in monoclinic space group P21 No. 4, while L-6 crys-
tallizes in P21/c (Table 1). Carbonyl bond distances are within range
quinone
viz.
2-chloro-3-(n-alkylamino)-1,4-naphthoquinone.
Although redox cycling is present in all compounds of homolo-
gated series, however their molecular interactions viz. hydrogen
of oxidized form of naphthoquinone ligands [17] (Tables S1–S3).
bonding and
p–p
stacking interactions may vary and this will af-
0
CAN distance is ꢄ1.34 ÅA is also within range while L-6 crystallizes
in P21/c (Table 1). Carbonyl bond distances observed for similar to
aminonaphthoquinone ligands while C(2)AC(3) bond distance is
observed to be longer as compared to similar aminonaphthoqui-
none ligands [18].
Inter molecular hydrogen bonding interaction is observed in
L-4, L-6 and L-8 (Table 2). Molecular structures of L-4 and L-8 are
hydrogen bonded polymer while molecular structure of L-6 is a
dimer. Although L-4 and L-8 crystallizes in the same space group,
their hydrogen bonding interactions are differs.
Bifurcated intermolecular hydrogen bonding is observed to O(1)
in L-4. Molecules of L-4 form polymeric chain through CAHꢁ ꢁ ꢁO
and NAHꢁ ꢁ ꢁO interactions [19] (Fig. 2a). Orientations of alkyl group
when viewed down b-axis, is on same side of the chain while the
polymeric chains runs opposite to one another and zip like struc-
ture is formed to the alkyl groups of the neighboring chains.
fect their tissue specific antiproliferative activity.
3.1. Synthesis and characterization of homologated, 2-chloro-3-(n-
alkylamino)-1,4-naphthoquinone
Ligands L-5 to L-8 are purified by column chromatography
using toluene and methanol as solvents and purity have been
determined by LC–MS experiments and purity of all the com-
pounds is >99.5%. Melting points of L-5 to L-8 were corrected by
Differential Scanning Calorimetry (DSC) (Fig. S3).
mNAH vibrations are observed in FT-IR spectra ꢄ3300 cmꢂ1 and
m
C@O vibration frequency is observed ꢄ1674 cmꢂ1 [6b]. The charac-
teristics of paranaphthoquinone (p-NQ) vibrations are observed
ꢄ1260 cmꢂ1 in L-5, L-7 and L-8, while these vibrations are de-
creased by ꢄ15 cmꢂ1 in L-6. p-NQ vibrations are susceptible to
strength of intermolecular hydrogen bonding. UV–vis spectra of
L-5 to L-8 (Fig. S4) in DMSO show two bands in UV region viz.
Slipped
p–p stacking interaction (Fig. S5) are observed between
benzenoid and quinonoid rings; C(1),C(9) [3.362 Å] and C(1), C(8)
[3.359 Å].
Molecules of L-8 form polymeric chain through CAHꢁ ꢁ ꢁO and
NAHꢁ ꢁ ꢁO interaction (Fig. 2c). An orientation of alkyl groups are
ꢄ301 nm and 338 nm are assigned to
p
–
pꢅ transitions of naphtho-
quinone ring and a broad band between 400 and 600 nm is
Fig. 1. ORTEP plots of: (a) L-4, (b) L-6 and (c) L-8.

S. Pal et al. / Journal of Molecular Structure 1049 (2013) 355–361
359
Table 2
Hydrogen bond geometries for L-4, L-6 and L-8.
0
0
0
Sr. no
DAHꢁ ꢁ ꢁA
DAHꢁ ꢁ ꢁA(°)
DAH(AÅ)
Hꢁ ꢁ ꢁA(AÅ)
2.320 (2)
2.548(1)
2.609(1)
Dꢁ ꢁ ꢁA(AÅ)
2.935(2)
3.242(2)
3.337(2)
L-4
1
2
3
N(11)AH(11)ꢁ ꢁ ꢁO(1)i
0.820(2)
0.951(1)
0.950(1)
132(2)
130(1)
130(1)
C(9)AH(9)ꢁ ꢁ ꢁO(4)ii
C(13)AH(13)Bꢁ ꢁ ꢁO(1)iii
L-6
L-8
4
5
N(11)AH(11)ꢁ ꢁ ꢁO(4)iv
0.840(2)
0.990(3)
2.650(2)
2.600(2)
3.458(2)
3.384(3)
163(2)
136(2)
C(17)AH(17A)ꢁ ꢁ ꢁO(1)v
6
7
N(11)AH(11)ꢁ ꢁ ꢁO(1)ii
C(9)AH(9)ꢁ ꢁ ꢁO(4)ii
C(7)AH(7)ꢁ ꢁ ꢁO(4)vi
0.880(4)
0.950(4)
0.949(5)
2.310(2)
2.562(1)
2.708(1)
2.945(4)
3.257(5)
3.297(4)
129(4)
130(2)
121(1)
(i) ꢂ1+x,y,z; (ii) 1+x,y,z; (i) ꢂx,1/2+y,ꢂz; (iv) ꢂx,ꢂy,ꢂz; (v) 1+x,1/2ꢂy,1/2+z; (vi) 2ꢂx, ꢂ1/2+y,1ꢂz.
Fig. 2. Molecular packing in: (a) L-4, down b-axis; (b) L-6 down a-axis and (c) L-8 down b-axis.
on same side of the chain and the two polymeric chain runs oppo-
site to one another. The chains are further intermolecular hydrogen
bonded through CAHꢁ ꢁ ꢁO interactions [C(7)AH(7)ꢁ ꢁ ꢁO(4)]. There is
homologated series. Quinonoid as well as benzenoid rings are in-
volved in stacking interactions. Molecules of L-1 and L-3 show
stacking of the quinonoid rings of the neighboring chains
while, molecules of L-4 and L-8 shows slipped stacking inter-
action between benezenoid and quinonoid rings in a chain and
p–p
p–p
slipped
p–p
stacking interaction is observed between Cl and C(1),
p–p
C(10) and C(1), C(8) [x, 1+y, z] (Fig. S6).
Molecules of L-6 form dimers through NAHꢁ ꢁ ꢁO(1) interaction
(Fig. 2b). Orientations of molecules are ‘anti’ to one another. The di-
mers further interlinked by CAHꢁ ꢁ ꢁO interaction.
molecules of L-2 and L-6 do not possess p–p stacking interactions.
3.3. Antiproliferative activities
Molecular structures of homologated series vary with respect to
p–p stacking and hydrogen bonding interactions. This variation in
molecular interactions could affect the biological properties in
Table 3 illustrates the IC50 values obtained from cell prolifera-
tion studies on three carcinoma cell lines (COLO205, U87MG and
MIAPaCa2). Compounds L-1 to L-8 has no effect on the brain tumor
type (U87MG) but in case of COLO205 and MIAPaCa2, compounds
with carbon chain P3 (C = 4, 5, 6, 7 and 8; L-4 to L-8) were inactive
in all three cell lines.
Table 3
Effect of homologation on antiproliferative for L-1 to L-8 in COLO205, U87MG and
MIAPaCa2 in terms of IC50
.
In COLO205, an increase in activity was observed from L-1, L-2
and L-3 (C = 1, 2 and 3 respectively) and no effect from L-4 to L-8
indicating L-3 to be an optimal structure for antiproliferative activ-
ity in COLO205 cell line (Table 4). Whereas, a vice versa was ob-
served in MIAPaCa2 for L-1 to L-3 where a decrease in the
antiproliferative activity and no effect from L-4 to L-8 indicating
L-2 as an optimal structure of antiproliferative activity in MIA-
PaCa2 (Table 4). Treatment of cell lines with doxorubicin showed
marked increase in antiproliferative activity non-specifically in
all three cases.
Compound
Carbon chain
COLO205
U87MG
MIAPaCa2
11.5 6.2 lM
L-1
L-2
L-3
L-4
L-5
L-6
L-7
L-8
ACH3
223 3.5
91.4 9.5
92.2 12
>100 mM
>100 mM
>100 mM
>100 mM
>100 mM
l
l
l
M
M
M
>100 mM
>100 mM
>100 mM
>100 mM
>100 mM
>100 mM
>100 mM
>100 mM
AC2H5
AC3H7
AC4H9
AC5H11
AC6H13
AC7H15
AC8H15
12.8
8 lM
1.3 2 mM
>100 mM
>100 mM
>100 mM
>100 mM
>100 mM
0.07 0.4 nM
Doxorubicin
67.8 0.25 nM
0.34 lM
Table 4
Correlation of homologation of L-1, L-2 and L-3 compounds to degree of specificity to COLO205 and MIAPaCa2 cell line.
*
#
Carbon chain
Degree of specificityCOLO205
Degree of specificityMIAPaCa2
L-1
L-2
L-3
ACH3
AC2H5
AC3H7
0.05
0.14
14.10
19.39
7.14
0.07
*
Degree of specificityCOLO205 = IC50(MIAPaCa2)/IC50(COLO205)
Degree of specificityMIAPaCa2 = IC50(COLO205)/IC50(MIAPaCa2)
.
#
.

360
S. Pal et al. / Journal of Molecular Structure 1049 (2013) 355–361
3.4. Correlation of homologation to degree of specificity
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