In situ Bronsted-Lowry acid catalyzed syntheses, characterization, single crystal XRD, electronic spectral-, DPPH radical scavenging-, and DNA protection studies of aryl-3,3′-bis(indolyl)methanes

Article abstract of DOI:10.1016/j.saa.2013.12.017

A series of novel aryl-3,3′-bis(indolyl)methanes (BIMs) were synthesized using indole and formylphenoxyaliphatic acid(s) in water in the absence of any catalyst. The formylphenoxyaliphatic acid behaves as an in situ Bronsted-Lowry acid catalyst in water. UV-Visible and fluorescence spectra of the compounds were recorded in selected solvents. The gas phase geometry optimization of the compounds were achieved using DFT calculations at B3LYP/3-21G(*) level of theory. The electronic properties, such as HOMO-LUMO energies were calculated using the above method based on the optimized structure. Compounds have better DPPH radical scavenging activity and reduction of oxidative damage of DNA.

Full text of DOI:10.1016/j.saa.2013.12.017

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
Contents lists available at ScienceDirect
Spectrochimica Acta Part A: Molecular and
Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa
In situ Bronsted–Lowry acid catalyzed syntheses, characterization, single
crystal XRD, electronic spectral-, DPPH radical scavenging-, and DNA
protection studies of aryl-3,30-bis(indolyl)methanes
G.S. Suresh Kumar a, A. Antony Muthu Prabhu b, P.G. Seethalaksmi c, N. Bhuvanesh d, S. Kumaresan e,
⇑
a Faculty of Chemistry, Department of Science and Humanities, Dr. Mahalingam College of Engineering and Technology (Dr. MCET), Udumalai Road, Pollachi 642 003, Tamilnadu, India
b Department of Chemistry, Manonmaniam Sundaranar University, Tirunelveli 627 012, Tamilnadu, India
c Department of Chemistry, APC Mahalakshmi College for Women, Toothukkudi 628 002, Tamilnadu, India
d Department of Chemistry, Texas A&M University, College Station, TX 77842, USA
e Department of Chemistry, Noorul Islam University, Kumaracoil, Thuckalay 629 180, Tamilnadu, India
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
ꢀ
Formylphenoxyacetic acids act as
in situ Bronsted–Lowry acid catalysts
for the syntheses of BIMs.
ꢀ
SCXRD and DFT, the two indole rings
and the phenyl ring are found to be
noncoplanar.
ꢀ
ꢀ
ꢀ
All the synthesized compounds
display p–
pꢁ transition in solvents.
BIMs have better DPPH radical
scavenging activity.
BIMs have better reduction of
oxidative damage of DNA.
a r t i c l e i n f o
a b s t r a c t
A series of novel aryl-3,30-bis(indolyl)methanes (BIMs) were synthesized using indole and formylphen-
oxyaliphatic acid(s) in water in the absence of any catalyst. The formylphenoxyaliphatic acid behaves
as an in situ Bronsted–Lowry acid catalyst in water. UV–Visible and fluorescence spectra of the
compounds were recorded in selected solvents. The gas phase geometry optimization of the compounds
were achieved using DFT calculations at B3LYP/3-21G(ꢁ) level of theory. The electronic properties, such as
HOMO–LUMO energies were calculated using the above method based on the optimized structure.
Compounds have better DPPH radical scavenging activity and reduction of oxidative damage of DNA.
Ó 2013 Elsevier B.V. All rights reserved.
Article history:
Received 5 December 2012
Received in revised form 24 November 2013
Accepted 5 December 2013
Available online 18 December 2013
Keywords:
3,30-Bis(indolyl)methanes
Crystal structure
Density functional theory
DPPH radical scavenging
DNA protection
Introduction
antimicrobial-, antifungal-, antibiotic-, antibacterial-, antiangio-
genic-, cytotoxic-, antimetastatic-, analgesic-, anti-inflammatory-,
Bis(indolyl)methanes (BIMs) have received more attention due
to their bioactivities against human pathogens [1]. BIMs reveal
etc. activity. Furthermore, BIMs are used in dyes, laser technolo-
gies, and fluorescent materials for visualization of biomolecules
[2]. There is a continuous search for the formulation of newer
synthetic entities of BIMs via efficient methods due to their versa-
tile application possibilities.
⇑
Corresponding author. Tel.: +91 9443182502.
E-mail address: skumarmsu@yahoo.com (S. Kumaresan).
1386-1425/$ - see front matter Ó 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.saa.2013.12.017

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G.S. Suresh Kumar et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
Aryloxyaliphatic acid derivatives possess an extensive range of
materials (monitored by TLC, chloroform:methanol), the reaction
mixture was cooled to room temperature, the solid was filtered,
washed with water, dried well and recrystallized from hot ethanol.
diverse bioactivities such as antimicrobacterial-, anti-inflamma-
tory-, antioxidant-, antibacterial-, analgesic-, antisickling-, antip-
aemic-, antiplatelet-, non-prostanoid prostacyclin mimetic-,
diuretic-, and growth regulators [3].
2-{2-[Bis(1H-indol-3-yl)methyl]phenoxy}acetic acid (2a)
In the ideal case, a target molecule is prepared from readily
available starting materials in one simple, safe, environment-
friendly operation that proceeds quickly and in reasonable yield
[4]. Water finds an important place in organic syntheses as solvent
because of its abundance and less toxicity [5]. Water-facilitated
reactions are categorized as ‘in-water process’ and ‘on-water pro-
cess’ based on the starting materials and reaction conditions [6].
Fundamental understanding on structural and energetic prop-
erties of this kind of materials could lead to beneficial knowledge
for the design of novel bisarylmethanes. Therefore, it is of interest
to discuss the geometrical and the electronic properties based on
density functional theory (DFT) which is shown as a favorite
among several computational chemistry methods because of its
great accuracy in reproducing the experimental values in molecu-
lar geometry, vibrational frequencies, atomic charges, dipole
moment etc. [7].
To the best of our knowledge, we have not encountered any re-
port regarding the use of aromatic aldehydes bearing carboxylic
acid group in the side chain acting as an in situ acid catalyst for
the formation of BIMs in water. This deliberated us to carry out
the efficient syntheses of BIMs from indole and formylphenoxyace-
tic acid(s) at elevated temperature in water in the absence of any
external acid catalyst. In continuation of our work in organic syn-
theses [2b,3b,3c], herein we report the syntheses, the geometrical
parameters studied by both single crystal XRD and theoretical cal-
culations, the fluorescence activity in selected solvents, DPPH rad-
ical scavenging activity, and DNA strand breakage assay of BIMs.
Pink solid; 84%; m.p. 130–132 °C; IR (KBr): 3402, 1723 cmꢂ1; 1H
NMR (500 MHz, d ppm): 4.53 (s, 2H), 6.41 (s, 1H), 6.46–9.16 (m,
14H), 9.16 (s, 2H) ; 13C NMR (125 MHz, d ppm): 32.4, 66.1, 101.7,
111.1, 111.3, 112.3, 118.5, 118.7, 119.3, 119.9, 120.4, 121.2,
121.4, 124.0, 124.5, 127.0, 127.2, 130.0, 133.6, 136.8, 155.5,
171.1; ESI-MS: 395.50; Anal. Calcd. for C25H20N2O3: C, 75.74; H,
5.08; N, 7.07%. Found: C, 75.72; H, 5.06; N, 7.10%.
2-{2-Bis[1H-indol-3-yl)methyl]-6-methoxyphenoxy}acetic acid (2b)
Pink solid; 86%; m.p. 138–140 °C; IR (KBr): 3408, 1731 cmꢂ1; 1H
NMR (500 MHz, d ppm): 3.89 (s, 3H), 4.19 (s, 2H), 6.22 (s, 1H),
6.55–7.40 (m, 13H), 7.64 (s, 2H); 13C NMR (125 MHz, d ppm):
33.6, 55.8, 71.2, 102.6, 110.1, 111.0, 111.2, 118.6, 119.4, 119.6,
119.8, 120.7, 122.0, 122.1, 123.5, 124.1, 125.0, 126.8, 127.8,
136.7, 138.0, 145.2, 151.2, 170.6; ESI-MS: 426.42; Anal. Calcd. for
C26H22N2O4: C, 73.23; H, 5.20; N, 6.57%. Found: C, 73.25; H, 5.25;
N, 6.60%.
4-{2-[Bis(1H-indol-3-yl)methyl]phenoxy}acetic acid (2c)
Pink solid; 84%; m.p. 145–147 °C; IR (KBr): 3406, 1760 cmꢂ1
;
ESI-MS: 395.45; Anal. Calcd. for C25H20N2O3: C, 75.74; H, 5.08; N,
7.07%. Found: C, 75.78; H, 5.12; N, 7.04%.
4-{2-Bis[1H-indol-3-yl)methyl]-6-methoxyphenoxy}acetic acid (2d)
Experimental
Pink solid; 86%; m.p. 159–161 °C; IR (KBr): 3406, 1760 cmꢂ1; 1H
NMR (500 MHz, d ppm): 3.72 (s, 3H), 4.59 (s, 2H), 5.80 (s,1H), 6.48–
7.40 (m, 13H), 9.34 (s, 2H); 13C NMR (125 MHz, d ppm): 4.75, 60.5,
71.0, 106.4, 116.0, 117.6, 118.4, 123.3, 123.8, 123.9, 124.3, 125.1,
125.3, 126.0, 126.1, 128.6, 129.9, 131.7, 132.5, 141.5, 143.5,
150.3, 153.8, 175.8; ESI-MS: 426.42; Anal. Calcd. for C26H22N2O4:
C, 73.23; H, 5.20; N, 6.57%. Found: C, 73.25; H, 5.25; N, 6.60%.
General methods
Melting points were measured in open capillary tubes and are
uncorrected. The crystal structure was determined using a Bruker
APEX 2 X-ray (three-circle) diffractometer. The 1H NMR and 13C
NMR spectra were recorded on a Bruker (Avance) 500 MHz NMR
instrument using TMS as internal standard and CDCl3 as solvent.
Standard Bruker software was used throughout. Chemical shifts
are given in parts per million (d scale) and the coupling constants
in Hertz. Infrared spectra were recorded on a JASCO FT-IR Model
410 spectrophotometer (in KBr pellet). Band positions are reported
in reciprocal centimetres (cmꢂ1). Absorption measurements were
carried out with a Shimadzu UV 1601 PC model UV–Visible spec-
trophotometer and fluorescence measurements were made by
using a Shimadzu spectrofluorimeter model RF-5301. Silica gel-G
plates (Merck) were used for TLC analysis with a mixture of chlo-
roform and methanol as eluent. The electrospray (ESI) mass spectra
were recorded on a THERMO Finnigan LCQ Advantage max ion trap
2-{2-[Bis(2-methyl-1H-indol-3-yl)methyl]phenoxy}acetic acid (2e)
Pink solid; 78%; m.p. 122–126 °C; IR (KBr): 3395,1727 cmꢂ1; 1H
NMR (500 MHz, d ppm): 2.12 (s, 6H), 4.37 (s, 2H), 6.12 (s, 1H),6.78–
7.51 (m, 12H), 7.83 (s, 2H); 13C NMR (125 MHz, d ppm): 12.2, 33.6,
66.3, 100.1, 110.0, 110.3, 110.5, 112.4, 112.6, 118.8, 119.1, 119.5,
120.2, 120.7, 120.9, 121.6, 125.2, 127.3, 129.0, 130.4, 132.0,
133.2, 135.1, 135.2, 136.1, 137.4, 156.1, 170.8; ESI-MS : 424.00;
Anal. Calcd. for C27H24N2O3: C, 76.39; H, 5.70; N, 6.60%. Found: C,
76.42; H, 5.66; N, 6.62%.
mass spectrometer. Samples (10 lL) (dissolved in solvent such as
methanol/acetonitrile/water) were introduced into the ESI source
through Finnigan surveyor autosampler. Elemental analyses were
performed on a Perkin Elmer 2400 Series II Elemental CHNS ana-
lyzer. Formylphenoxyacetic acids were synthesized by literature
method [3b].
2-{2-[Bis(2-methyl-1H-indol-3-yl)methyl]-6-methoxyphenoxy}acetic
acid (2f)
Pink solid; 76%; m.p. 148–150 °C; IR (KBr): 3389, 1725 cmꢂ1; 1H
NMR (500 MHz, d ppm): 2.09 (s, 6H), 3.87 (s, 3H), 3.91 (s, 2H), 6.17
(s, 1H), 6.81–7.29 (m, 11H), 7.50 (s, 2H); 13C NMR (125 MHz, d
ppm): 12.7, 13.7, 33.7, 55.7, 69.1, 99.7, 110.2, 110.4, 112.6, 118.5,
119.1, 119.2, 119.3, 119.9, 120.4, 122.5, 123.6, 129.0, 132.3,
135.2, 136.2, 138.4, 152.3, 171.1; ESI-MS: 453.50; Anal. Calcd. for
Syntheses of BIMs acids
Formylphenoxyaliphatic acid (2.5 mmol) was dissolved in
water (10 mL) at 80 °C, indole (5 mmol) was added with vigorous
stirring at 80 °C for 5–10 min. After the completion of starting
C28H26N2O4: C, 73.99; H, 5.77; N, 6.16%. Found: C, 74.02; H, 5.76;
N, 6.19%.

G.S. Suresh Kumar et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
251
2-{2-[Bis(2-phenyl-1H-indol-3-yl)methyl]phenoxy}acetic acid (2g)
Green solid; 72%; m.p. 202–204 °C; IR (KBr): 3622, 3424,
Results and discussion
Syntheses of phenoxyaliphatic acids of 3,30-BIMs
1725 cmꢂ1 1H NMR (500 MHz, d ppm): 3.72 (s, 2H), 6.22 (s, 1H),
;
6.75–7.68 (m, 22H) , 8.12 (s, 2H), 10.26 (s, 1H); 13C NMR
(125 MHz, d ppm): 34.8, 66.3, 70.2, 110.9, 113.1, 114.8, 118.8,
120.9, 121.5, 125.3, 126.9, 127.5, 128.0, 128.4, 128.8, 129.1,
130.9, 133.4, 136.1, 156.3, 170.9; ESI-MS: 547.63; Anal. Calcd. for
Synthesis of 2-[2-{bis(1H-indol-3-yl)methyl}phenoxy]acetic
acid (2a) using indole and formylphenoxyacetic acid (1a) in water
was taken as the model reaction. Synthesis of 2-[2-{bis(1H-indol-
3-yl)methyl}phenoxy]acetic acid (2a) using indole and formyl-
phenoxyacetic acid (1a) in water was achieved in 84% yield at
80 °C in the absence of any acid catalyst. Formylphenoxyacetic acid
and indole were completely miscible in hot water and a pink solid
was precipitated out within 5 min. This reaction was categorized
as ‘in-water process’ [3f,6] (Scheme 1).
C37H28N2O3: C, 81.00; H, 5.14; N, 5.11%. Found: C, 81.02; H, 5.16;
N, 5.10%.
Ethyl 2-{2-[bis(1H-indol-3-yl)methyl]phenoxy}acetate (4b)
The above in situ Bronsted–Lowry acid catalyzed reaction
[3f,13] was checked using ethyl 2-formylphenoxyacetate as well
as the sodium salt of 2-formylphenoxyacetic acid. Formation of
4a, 4b, and 4c from the condensation of indole with the corre-
sponding aldehyde(s) in water at 80 °C was not possible in these
cases even after 12 h. But the same reaction proceeded smoothly
in the presence of acetic acid (pKa 4.76) in water to afford 4a, 4b,
and 2a (vide supporting information, Scheme II). This manifests
the formation of hydronium ion from 2-formylphenoxyacetic acid
(pKa 3.04) for the synthesis of 2a in water.
Pale pink solid; 77%; mp = 176–178 °C; IR (KBr): 3401,
1749 cmꢂ1 1H NMR (300 MHz, d ppm): 1.22 (t, J = 14.1 Hz, 3H),
;
4.18 (q, J = 21.3 Hz, 3H), 4.58 (s, 2H), 6.43 (s, 1H), 6.55 (s, 2H),
6.78–6.87 (m, 2H), 6.98 (t, J = 7.8 Hz, 2H), 7.11–7.17 (m, 4H), 7.34
(d, J = 9.0 Hz, 2H), 7.44 (d, J = 7.8 Hz, 2H), 7.84 (s, 2H); 13C NMR
(75 MHz, CDCl3): 14.0, 32.5, 61.0, 66.2, 110.8, 112.0, 119.0,119.4,
120.1, 121.5, 121.7, 123.5, 127.0, 127.2, 130.0, 133.2, 136.8,
155.8, 169.1; ESI-MS: m/z 424.3; Anal. Calcd. for C27H24N2O3: C,
76.39; H, 5.70; N, 6.60%. Found: C, 76.45; H, 5.7; N, 6.55%.
Thus it could be confirmed that the syntheses of BIMs using
formylphenoxyacetic acid(s) did not require any external acid cat-
alyst. Formylphenoxyacetic acid(s) itself acted as a Bronsted–
Lowry acid catalyst in water. To demonstrate the generality of this
methodology, the replacement of indole with 2-methylindole and
2-phenylindole was studied (Scheme 1). Under the same reaction
conditions, 2-methylindole and 2-phenylindole afforded the BIMs
via ‘on-water process’. Even the hydrophobicity of these indoles
did not forbid the formation of the corresponding BIMs in moder-
ate yields. This is due to the strong Bronsted–Lowry acid strength
of the formylphenoxyacetic acid. Synthesis of 2-phenylindole-
substituted BIM 2g using this aldehyde substantiates that the cor-
responding formylphenoxyacetic acid is a better Bronsted–Lowry
acid than acetic acid in water. Hence the mechanism is proposed
for the formation of 2a from the condensation of 2-formylphenoxy-
acetic acid and indole (vide supporting information, Fig. s1).
To further investigate the synthetic potential of this process, the
in situ Bronsted–Lowry acid catalyzed reaction was extended to
phenolic aldehydes in water. We examined the synthesis of
[bis(1H-indol-3-yl)methyl]phenols (5) from indole and hydroxy-
benzaldehyde(s) in the absence of acid catalyst in water. After
1 h, the corresponding BIMs were observed in moderate yields
(5a, 65% and 5b, 68%). Hydroxybenzaldehydes were not completely
miscible in hot water. This reaction was regarded as ‘on-water pro-
cess’ (vide supporting information, Scheme III). The increase in
reaction time and lower yield are due to the less acidic nature of
hydroxybenzaldehydes (pKa for 5a, 6.79 and 5b, 7.66) in water.
Further, we have subjected the four acid derivatives 2a–d of
BIMs for characterizing their structural-, photophysical-, radical
scavenging activity, and DNA strand breakage studies. The opti-
mized structures of the selected derivatives with the substitutions
–OCH2COOH at o-/p-positions and –OCH3 group at m-position in
the phenyl ring are given in supporting information, Fig. s2.
Computational methods
The ground state geometries of BIMs 2a–d were optimized by
using the Gaussian-09 series of programs [8]. For this purpose
the B3LYP/DFT approach, which includes the interchange hybrid
functional from Becke (B3) [9a] in combination with the three-
parameter correlation functional by Lee–Yang–Parr (LYP) [9b]
was employed in combination with the basis set 3-21G(ꢁ).
Evaluation of radical scavenging activity using DPPH model system
The radical scavenging activity of the compounds was evaluated
as per the method of Blois [10] with slight modification [11]. The
compounds (2a–d) and BHA at different concentrations (25, 50,
75 and 100 ppm in 1 mL) were taken in different test tubes. Four
milliliters of 0.1 mM methanolic solution of DPPH was added to
these tubes and shaken vigorously. The tubes were allowed to
stand at 27 °C for 20 min. The control was prepared as above
without any compound and methanol was used for the baseline
correction. Optical density (OD) of the samples was measured at
517 nm. Radical scavenging activity was expressed as the inhibi-
tion percentage and was calculated using the following formula:
%Radical scavenging activity ¼ ðControl OD ꢂ Sample OD=Control ODÞ ꢃ 100:
DNA strand breakage assay
The DNA strand breakage assay was carried out by an estab-
lished method [12]. Briefly, each reaction mixture contained
10
FeSO4, 10
and b); the volume was brought to 100
were incubated for 60 min in a vial at 37 °C. 5
mixtures were mixed with 2 L of gel loading solution (0.1 M
l
L of DNA (calf thymus, pBR 322 or pUC 18), 10
L of 10 mM H2O2, and 10 L of compound (2a–d, 4a
L with PBS. The mixtures
L of the reaction
lL of 1 mM
l
l
l
l
l
Single crystal XRD studies
EDTA, 0.5% sodium dodecyl sulfate, 40% sucrose, and 0.5% bromo-
phenol blue) and loaded into individual wells in a 0.8% agarose
gel for electrophoresis at 5 V/cm in TBE buffer (0.1 M tris; 0.09 M
boric acid; 0.001 M EDTA). The gel was visualized by photograph-
ing the fluorescence of intercalated ethidium bromide under a UV
illuminator.
The molecular structure of the derivative 2d has been deter-
mined by single crystal X-ray diffraction (CCDC No. 911716) and
the ORTEP diagram is shown in Fig. 1. The crystal data and struc-
tural refinement of 2d are summarized in Table 1. The derivative
2d was crystallized as a triclinic system in methanol under dark

252
G.S. Suresh Kumar et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
Scheme 1. Synthesis of acid BIMs (2).
Fig. 1. ORTEP view of 2d.
conditions. Selected bond lengths, bond angles, and torsion angles
for 2d are listed in Table 2.
In the crystal structure of compound 2d, two molecules of 2d
are connected together through H-bonding (O4–H4Aꢄ ꢄ ꢄO3,
1.794 Å) like a normal acid dimer synthon with the Etter’s graph
the exploited wavelength region (200–800 nm), all compounds
showed an absorption peak at 290 nm. The molecules showed a
high extinction coefficient of the order of 104 Mꢂ1 cmꢂ1 at the low-
est energy transition in the solvents. The substituent –OCH2COOH
at o- and p-position; and –OCH3 group at m-position have little
influence on the absorption wavelength and absorbance. However,
there is not much change in the energy of transition in the different
solvents, which implies that solvent stabilization of the ground
state species is not significant in all compounds [17].
The fluorescence spectra of compounds were measured at room
temperature in cyclohexane, acetonitrile, methanol, and water as
shown in Fig. 2. Compounds 2a–d excited at 290 nm in solvents
showing the maximum emission peak around 340 nm. Unlike
absorption spectra, this implies that the substituent of the phenyl
group has a significant change on the fluorescence wavelength and
fluorescence intensity of the compounds. The red shift on the fluo-
rescence spectra on changing the solvent from cyclohexane to
water indicates the effect of hydrogen bonding in the excited state
of the compounds [18].
set notation R22 (8). C–Hꢄ ꢄ ꢄ
p
and N–Hꢄ ꢄ ꢄ
p interactions were ob-
served. Similar interactions are found in 3,30-BIMs derivatives
[2b,14], other indole derivatives [15], and globular proteins [16].
These interactions may afford stability and contribute to the fold-
ing process or have a functional role in proteins and DNA. The dihe-
dral angle between two indole rings is 72.79 Å, aryl ring - indole (1)
is 89.41 Å, and aryl ring – indole (2) is 84.36 Å.
Solvent effects on absorption and fluorescence spectrum
Fig. 2 shows a typical example of the solvent effect on the elec-
tronic absorption and emission spectrum of compound 2d. The
absorption and fluorescence spectra of 2a, 2b, and 2c are almost
similar to those of 2d. The absorption and emission wavelengths
and Stokes shift of BIMs (2a–d) in selected solvents (cyclohexane,
acetonitrile, methanol, and water) are presented in Table 3. In
The absorption and emission spectra of the compounds mainly
originate from the indole rings. The presence of a methine group in

G.S. Suresh Kumar et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
253
Table 1
Crystal data and structure refinement of 2d.
0.40
Empirical formula
C27 H24 N2 O3
0.30
0.20
CCDC No.
911716
424.48
110(2) K
0.71073 Å
Monoclinic
P2(1)/c
Formula weight
Temperature
Wavelength
Crystal system
Space group
Unit cell dimensions
a,b,c (Å)
3
4
1
0.10
2
9.6067(11), 13.9907(15), 16.2903(18)
90°, 95.91(10)°, 90°
2177.8(4) Å3
a,b,c (°)
Volume
360
200
320
240
280
Wavelength (nm)
Z
4
Density (calculated)
Absorption coefficient
F(000)
Crystal size
Theta range for data collection
Index ranges
Reflections collected
Independent reflections
Completeness to theta = 27.50°
Absorption correction
Max. and min. transmission
Refinement method
Data/restraints/parameters
Goodness-of-fit on F2
Final R indices [I > 2sigma(I)]
R indices (all data)
Largest diff. peak and hole
1.295 Mg/m3
0.085 mmꢂ1
400
300
896
0.58 ꢃ 0.22 ꢃ 0.16 mm3
2
2.13–27.50°
ꢂ12 6 h 6 12, ꢂ18 6 k 6 18, ꢂ20 6 l 6 20
24448
4955 [R(int) = 0.0317]
99.1%
Semi-empirical from equivalents
0.9865 and 0.9524
Full-matrix least-squares on F2
4955/0/290
1
200
100
4
3
360
440
320
400
280
1.045
Wavelength (nm)
R1 = 0.0407, wR2 = 0.0951
R1 = 0.0508, wR2 = 0.1014
0.276 and ꢂ0.211 eÅꢂ3
Fig. 2. Absorption and fluorescence spectra of 2d in selected solvents. 1. Cyclo-
hexane, 2. Acetonitrile, 3. Methanol, and 4. Water.
in Table 2. The calculated geometric parameters (Table 2) represent
good correlations with single crystal XRD data and can be used as a
Table 2
Selected bond lengths and bond angles of 2a–d.
source to calculate other parameters such as total energy, EHOMO
,
ELUMO, and dipole moment for the compound. Similarly, the
Atom numbering
Experimental DFT
ground-state geometric parameters (bond lengths and bond an-
gles) of compounds 2a–d are very similar to each other.
2d
2d
2a
2b
2c
Bond length
C17–C15
C17–C18
C17–C7
N1–C8
N2–C16
C22–O1
C20–O4
C21–O2
C19–O1
C26–O3
C25–O2
O2–C25
O3–C25
O1–C24
O4–C26
1.513(3)
1.518(3)
1.508(3)
1.388(3)
1.379(3)
1.378(3)
–
1.374(3)
–
1.218(3)
–
1.515
1.535
1.518
1.397
1.397
1.384
–
1.394
–
1.218
–
1.517
1.534
1.520
1.397
1.397
–
1.515
1.534
1.517
1.397
1.399
–
1.515
1.534
1.518
1.397
1.397
–
Table 3
Spectral properties of compounds 2a–d.
Compounds
2a
Solvent
kmax
log e
kflu
Stokes shift
4211
Cyclohexane
288.2
219.6
290.4
236.1
291.6
234.0
290.2
230.6
3.11
3.42
3.50
4.12
3.59
4.07
3.38
4.15
328
–
–
1.413
–
Acetonitrile
Methanol
Water
345
349
350
5450
5640
5887
1.397
–
1.225
–
1.370
–
–
1.398
–
1.220
–
1.379
–
–
1.412
–
1.224
–
1.376
–
1.485
1.420(3)
–
1.422(3)
1.307(3)
1.384
–
1.458
–
2b
2c
2d
Cyclohexane
Acetonitrile
Methanol
Water
287.2
219.1
289.0
235.8
290.3
233.8
290.8
231.6
2.23
3.22
3.60
4.31
3.65
4.34
3.48
4.24
329
341
345
350
4423
5277
5462
5816
–
Bond angle
C7–C17–C15
C7–C17–C18
C15–C17–C18
112.4(2)
113.11(19)
110.6(2)
111.33
110.14
112.24
113.04
109.01
112.32
110.53
109.01
112.34
111.58
110.33
111.71
Torsion angle
C8–C7–C17–C18
C7–C17–C18–C23
ꢂ15.74
ꢂ81.88
ꢂ14.87 ꢂ73.73 ꢂ17.78 ꢂ10.98
ꢂ83.35 ꢂ92.19 ꢂ78.14 85.99
ꢂ30.47 ꢂ39.74 ꢂ22.62 ꢂ38.74
Cyclohexane
Acetonitrile
Methanol
Water
287.7
219.5
289.2
228.6
290.2
230.5
291.9
235.3
2.57
3.40
3.61
4.26
3.69
4.38
3.45
4.20
327
340
344
350
4177
5167
5389
5687
C15–C17–C18–C19 ꢂ32.60
C16–C15–C17–C18 116.60
114.29
134.90
100.13
113.88
between the two indole rings and substituted phenyl ring prevents
the extension of conjugation. The absorption and emission maxima
are thus very close to that of an individual indole ring.
Cyclohexane
Acetonitrile
Methanol
Water
286.7
220.4
288.8
234.6
289.4
234.0
290.4
235.3
2.66
3.65
3.76
4.32
3.81
4.40
3.62
4.30
328
348
349
351
4392
5891
5901
5945
Ground state geometries at the B3LYP level
The optimized structure of the compounds was calculated at
the B3LYP/3-21G(ꢁ) level of theory by DFT method in the gas phase,
carried out from the experimental structures. The geometric
parameters and corresponding experimental values of 2d are listed

254
G.S. Suresh Kumar et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
Fig. 3. Molecular orbital surfaces for the HOMO and LUMO of 2a–d computed by B3LYP/3-21G(ꢁ) method in gas-phase.
Table 4 presents the total energies, frontier molecular orbital
amount in phenyl ring, while the electron density of HOMO is
located on one of the indole rings. However, 2d has the electron
density of LUMO located on the indole ring, while the electron den-
sity of HOMO is located on the indole and phenyl rings. The energy
gap value of compound 2d is more negative than other three com-
pounds. This result suggests that 2d is more stable than 2a, 2b and
2c. Consequently, these systems present a small HOMO–LUMO
gap, enabling lesser electron-transfer process in all molecules
[20]. Compound 2d has a larger dipole moment than the other
compounds.
energies, and dipole moments of the compounds in gas phase. Ob-
tained energy values are ꢂ811200 kcal/mol for 2a, ꢂ882674 kcal/
mol for 2b, ꢂ811200 kcal/mol for 2c, and ꢂ882667 kcal/mol for
2d. From Table 4, it is understood that the substituent –OCH2COOH
present at ortho/para position of the phenyl ring exhibits higher
energy (2a and 2c) compared to –OCH3 substituent in the phenyl
ring of compounds 2b and 2d.
The highest occupied molecular orbital (HOMO) and lowest
unoccupied molecular orbital (LUMO) of the derivatives are shown
in Fig. 3. The LUMO as an electron acceptor represents the ability to
accept an electron and HOMO represents the ability to donate elec-
tron. Moreover, a lower HOMO–LUMO energy gap explained the
Table 4
Calculated energies, frontier orbital energies, and dipole moments of 2a–d in gas
phase.
eventual stability of the compound. Further the (EHOMO–ELUMO
)
2a
2b
2c
2d
gap is an important scale of stability [19] and compounds with
lower (EHOMO–ELUMO) values tend to have higher kinetic stability
and lower chemical reactivity. So we have investigated the
electronic structure of the compounds with these considerations
using DFT method. The electron density of LUMO for 2a, 2b, and
2c is located mainly on the –OCH2COOH group and very little
Energy (kcal/mol)
EHOMO (eV)
ELUMO (eV)
ꢂ811200
ꢂ5.05
ꢂ0.18
ꢂ4.87
1.66
ꢂ882674
ꢂ5.03
ꢂ0.10
ꢂ4.93
3.30
ꢂ811200
ꢂ5.30
ꢂ0.35
ꢂ4.94
2.81
ꢂ882667
ꢂ5.21
ꢂ0.02
ꢂ5.18
4.03
E
HOMO–ELUMO (eV)
Dipole moment (D)

G.S. Suresh Kumar et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 123 (2014) 249–256
255
Table 5
DPPH radical scavenging activity of 2a–d and 4b.
Samples
Radical Scavenging Activity at different concentration (ppm)
25
50
75
100
2a
2b
2c
2d
4b
BHA
92.84 0.83d
52.38 4.00a
70.39 2.26c
56.43 1.32b
55.23 1.43b
93.81 0.01d
90.23 4.41b
74.76 8.38a
84.02 5.03a,b
76.21 3.25a
75.76 8.18a
90.85 0.89b
95.57 1.22b
71.07 17.59a
93.96 3.07b
87.00 1.11b
85.04 5.00a,b
96.58 0.19b
94.04 1.22b
66.91 6.93a
91.95 2.82b
88.27 2.38b
87.30 1.11b
95.04 0.13b
a,b,c,d
Values are mean SD of triplicate values; values not having similar superscripts in the same column are significantly (P 6 0.05) different.
Conclusion
Table 6
DNA protection of compounds 2a–d and 4a–b.
It may be concluded that this study describes the efficient syn-
theses of BIMs from the condensation of indole with formylphen-
oxyacetic acid(s) in water in the absence of any catalyst. The
formylphenoxyacetic acids behave as in situ Bronsted–Lowry acid
catalysts in water. From the observation of single crystal X-ray
analysis and DFT calculation, the two indole rings and the phenyl
ring are found to be noncoplanar. All the synthesized compounds
Lane
Samples
Calf thymus DNA
pBR 322 DNA
pUC 18 DNA
3
4
5
6
7
8
4b
4a
2a
2b
2c
2d
No protection
Smear
Smear
Smear
Smear
No protection
Smear
Smear
Smear
Smear
Partial protection
Smear
Partial protection
Partial protection
Partial protection
Partial protection
Smear
Smear
display
p–
pꢁ transition and can potentially serve as photoactive
materials. Theoretical calculations were used to obtain optimized
structures as well as spatial distributions of the HOMO and LUMO
levels of the compounds. Compounds have a better DPPH radical
scavenging activity and a decrease in oxidative damage of DNA.
Both the HOMOs and the LUMOs are mostly
p
-antibonding type
orbitals and thus, the electronic transitions from the highest
occupied molecular orbital (HOMO) to the lowest unoccupied
molecular orbital (LUMO) are mainly derived from the contribution
of p–
pꢁ bands.
Acknowledgements
One of the authors (G.S. Suresh Kumar) thanks Ms. K. Rama-
lakshmi, Plantation Products Spices and Flavour Technology
Department, Central Food Technological Research Institute, Coun-
cil of Scientific and Industrial Research (CSIR), Mysore, India. CDRI,
Lucknow, India is acknowledged for the spectral studies.
DPPH radical scavenging assay
The free radical scavenging activity of the compounds was
tested using DPPH model system, and the results are presented
in Table 5. It reveals that 25 ppm concentration of 2a has a better
scavenging activity (92.84 0.83) and has nearly the same as BHA,
followed by 2c. Methoxy group in the meta-position reduces the
scavenging activity compared to its unsubstituted counterparts.
Appendix A. Supplementary material
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.saa.2013.12.017.
DNA protection studies
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