Journal of Molecular Structure 993 (2011) 43–51

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Journal of Molecular Structure

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Theoretical and experimental investigation on the spectroscopic properties

of indigo dye
Anna Amat a,⇑, Francesca Rosi a, Costanza Miliani b, Antonio Sgamellotti a,b, Simona Fantacci b,c
a
Dipartimento di Chimica sdi Perugia, Università degli Studi di Perugia, via Elce di Sotto 8, 06123 Perugia, Italy
b
Istituto di Scienze e Tecnologie Molecolari del CNR (CNR-ISTM), C/o Dipartimento di Chimica, Università di Perugia, Via Elce di Sotto 8, I-06123, Perugia, Italy
c
Italian Institute of Technology (IIT), Center for Biomolecular Nanotechnologies, Via Barsanti 73010, Arnesano, Lecce, Italy

a r t i c l e i n f o a b s t r a c t

Article history: The spectroscopic properties of indigo, one of the most important natural dyes present in nature, have
Available online 30 November 2010 been investigated by means of DFT and TD-DFT calculations and Raman and IR spectroscopies. The
absorption spectra of this dye, in vacuo and in different solvents, have been computed. The formation
Keywords: of aggregates in solvent have been investigated by computing the electronic absorption spectra of the
Indigo dimer and the trimer, thus evaluating the effects of the aggregation on the optical properties of indigo.
DFT The IR and Raman spectra have been measured and computed. The comparison between the experi-
TD-DFT
mental and theoretical spectra and the potential energy distribution (PED) of the computed normal
Raman
IR
modes have been used to perform the assignment of the experimental features in terms of functional
Hydrogen bond group displacements. Finally, the effects of the intermolecular hydrogen bond present in the solid state
have been evaluated by computing the vibrational spectra of the dimer.
The intention of the present work is to give an insight into both the vibrational and optical properties of
indigo as well as to evaluate DFT and TD-DFT potentialities in the study of organic dyes’ spectroscopic
properties of interest in the cultural heritage field.
Ó 2010 Elsevier B.V. All rights reserved.

1. Introduction The absorption spectrum of indigo has been shown to be depen-

The historical use of indigo as a blue dyestuff dates to the pre-
Vedic period in India where Indigofera tinctoria L. was its more
common source. India was a primary supplier of indigo to Europe
in the Greco-Roman era. India’s association with indigo is reflected
in the word for the dyestuff, which was indicum (by Pliny and
Vitruvius) or indicon (by Dioscorides) which passed into Italian
dialect and eventually into English as the word indigo. In Europe
it was also obtained from dyer’s woad but prior to the import of
larger amounts of indigo from India in the 16th century they were
not recognized as containing the same colouring matters [1–3].
Indigo has always attracted great interest and many studies can
be found on this dye and its related derivatives [4]. Indigo is also
known as the chromophore of Maya Blue, a pre-colombian pig-
ment prepared by precipitating indigo on a white clay, usually
palygorskite. Maya Blue was used in wall paintings and ceramics
and it has been widely studied due to its unusual stability and
hue of colour. To gain an insight into the great stability of Blue
Maya, interactions between indigo and palygorskite have been
studied, by means of quantum mechanical methods and molecular
dynamics [5–7].
⇑ Corresponding author. Tel.: +39 075 585 5638; fax: +39 075 585 5606.
E-mail address: anna@thch.unipg.it (A. Amat).
dent on the environment ranging from red (540 nm) in the gas
phase to violet (588 nm) in tetrachloromethane or to blue
(606 nm) in polar solvents such as ethanol [8,9] (see Fig. 1).
Several theoretical studies can be found on the electronic
absorption spectra of indigoids. For indigo a good agreement is
found in polar solvents with CASPT2 although gas-phase calcula-
tions underestimates the first transition by ca. 0.30 eV [8]. The first
work employing TD-DFT uses two different xc-functionals (B3LYP
and BPW91) reporting non negligible differences between the
two functionals and with the experiment [10]. Quite large errors
are retrieved by Matsuura et al. [11] using semi-empirical and
TD-DFT calculations in a set of organic dyes as the solvation effects
where neglected. The best agreements with the experiment are
found in a series of works on indigoids by Jaquemin et al. Hybrid
functionals and large basis sets together with a polarizable contin-
uum model (PCM) are used by these authors for indigo [12], thio-
indigo derivatives [13] and for indirubin, isoindigo and related
derivatives [14] obtaining very good results.
Experimental evidence of indigo aggregation in solution has
been detected with the appearance of a new band around
700 nm at large indigo concentrations that has been assigned to
the formation of a dimer [15]. On the other hand, dimer and trimer
attached to the textile have been proposed as feasible aggregates
when dyeing wool with solutions of high indigo concentration

0022-2860/$ - see front matter Ó 2010 Elsevier B.V. All rights reserved.
doi:10.1016/j.molstruc.2010.11.046
44 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51
Fig. 1. Indigo molecular structure.
[16,17]. Semiempirical simulations of the absorption spectrum
using 2 and 3 molecule clusters based on the crystalline structure
have been performed [16,17] to simulate the aggregation effects on
the optical properties. Finally, a limited analysis has been reported
by us in a general paper on the role of computational chemistry in
the study of art materials [18].
Investigations of the indigoids vibrational spectra by means of
ab initio and DFT methods have been reported [19–22]. First
assignment of indigo vibrational bands with ab initio calculations
was performed by Tatsch and Schraeder in 1995 [19] using HF/3-
21G level of theory as implemented in Gaussian 92. In this work
the experimental bands are compared to the theoretical spectra
for which wavenumbers but no intensities are reported and the
normal mode assignment is performed. A few works have been
found revising this information. Sánchez del Río et al. [20] using
the same level of theory (HF/3-21G) assign the Raman spectrum
and report the computed Raman activities. Tomkinson et al. [21]
use DFT calculations with B3LYP/6-31G** and Inelastic Neutron
Scattering (INS) spectroscopy to study the vibrational spectra of in-
digo. The results of the IR and Raman computed spectra are re-
ported with no intensities and the band assignment is performed
on the basis of a previous assignment on isatin and thioindigo
spectra. Finally, in a very recent paper, Baran et al. [22] assign
the main experimental features of the IR and Raman spectra below
1700 cm 1 using B3LYP/6-311++g** calculations and a potential
energy distribution (PED) analysis.
Due to the great experimental and theoretical interest in indigo
we report a comprehensive investigation rationalizing some
incompletely addressed aspects such as the aggregation in solution
or the effects of crystal packing in the vibrational spectra. Regard-
ing the vibrational properties, the Raman and IR spectra have been
measured and computed.
The comparison between the measured and the computed
vibrational spectra, together with the PED analysis, has allowed
us to assign the experimental modes in terms of functional groups
displacements. Finally, the dimer optimized structure in vacuo has
been used as a simple model to evaluate the hydrogen bond effects,
present in the solid state. The vibrational spectra of the dimer have
been computed and compared to those of the monomer and to the
experiment.
A computational methodology to reproduce the electronic
absorption spectra of indigo has been calibrated and used to com-
pute the spectra in vacuo and in different solvents. Results have
been compared to experimental data, evaluating the accuracy of
DFT methods in reproducing the solvation effects and rationalizing
the observed bathocromic effects.
Successively, the aggregation in solution has been studied by
hypothesizing the presence of dimer and trimer aggregates and
analyzing the changes of the indigo geometry, electronic distribu-
tion and absorption spectrum.
We present therefore a complete theoretical and experimental
study on the absorption and vibrational spectroscopic properties
of indigo. In a more general framework we intend to use indigo
as a prototype molecule to setup and evaluate a computational
strategy suitable for the study of cultural heritage materials.
2. Experimental and computational methodology
Micro-Raman experimental spectra have been obtained using a
JASCO Ventuno double-grating spectrophotometer equipped with
a charge-coupled device (CCD) detector cooled to 50 °C and cou-
pled to an optical microscope. Raman spectra have been excited
using green radiation from an Nd:YAG laser (532 nm). The laser
power at the sample was kept between 0.8 and 4 mW, the expo-
sure time varied between 10 and 20 s with 5–10 accumulations
and the resolution was about 2 cm 1. Calibration of the spectrom-
eter was accomplished using the Raman lines of a polystyrene
standard. Transmission IR spectra were collected with a Jasco FTIR
470-plus spectrophotometer equipped with a ceramic light source,
a Michelson interferometer, a DLATGS detector. The spectra were
carried out with 100 scans at a resolution of 2 cm 1 and an effec-
tive range of 4000–400 cm 1 in KBr pressed disks.
DFT and TD-DFT calculations have been carried using the
Gaussian03 code [23]. The Becke’s hybrid exchange functional B3
[24] with the Lee–Yang–Parr correlation functional LYP [25,26]
(B3LYP) have been used in our calculations. The solvation effects
have been included by means of the conductor-like polarizable
continuum model CPCM [27].
Geometry optimizations, with no symmetry constrains, have
been performed using 6-31G** and 6-311++G** basis set and fre-
quencies have been computed, at the same level of theory, to check
the nature of stationary points.
Vibrational spectra have been computed using 6-31G** and a
0.9614 scaling factor has been applied to the computed frequencies
according to the values suggested by Scott et al. [28] and a Lorentz-
ian convolution has been performed on the scaled frequencies to
obtain the computed spectra. Normal coordinate analysis calcula-
tions in terms of potential energy distributions (PEDs) have been
performed using VEDA4 program [29] allowing us to assign unam-
biguously the computed frequencies to the functional group
displacements.
The absorption spectra of indigo in chloroform and in vacuo
have been simulated by computing the lowest singlet-singlet exci-
tations following three different procedures:
(a) 6-31G** // 6-31G** for the vertical excitations // geometry
optimizations.
(b) 6-311++G** // 6-311++G** for the vertical excitations //
geometry optimizations.
(ab) 6-311++G**// 6-31G for the vertical excitations // geometry
optimizations.
The comparison between the theoretical results and the exper-
imental data has allowed the best methodology to be individual-
ized. This methodology has then been used in the analysis of the
absorption spectra in different solvents and in the study of indigo
aggregation in solution.
3. Results and discussion
3.1. Absorption spectra
Indigo optimizations in vacuo and in chloroform with both
6-31G** and 6-311++G** basis sets have been performed. No signif-
icant differences have been retrieved between the geometry opti-
mized in vacuo and in chloroform. Neither the basis set seem to
affect the results obtaining differences that are kept within
0.03 Å and 0.5° for the bond distances and angles, respectively.
Results are reported in Supporting Info (SI).
To rationalize the effect of the basis set and the solvent on the
optical properties, the absorption spectra of indigo has been
 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51 45

computed in vacuo and in chloroform with the three computa- with respect to the vacuo case, in which differences are within
tional strategies described in Section 2. In all the cases, the main 0.05 eV and can be considered negligible, with differences of
absorption band, responsible for the colour, is originated by the 0.15 eV that are, however, in the limits of the method accuracy. Ba-
lowest singlet–singlet transition (S0–S1). The computed transitions sis set effects retrieved in solvent are ascribable to the response
are reported in Table 1 compared to the maxima of the main exper- part of the calculations given that no differences have been de-
imental absorption band. The charge distribution of the highest tected in the ground state. Previous article reported the need of
occupied molecular orbital, HOMO (hereafter H), and the lowest using large basis set to quantitatively reproduce the absorption
unoccupied molecular orbital, LUMO (hereafter L), computed using experimental spectra of indigo [12] and 6-311G(d,p) and 6-
6-311++G**, reported in Fig. 2, are equivalent to the results ob- 311+G(2d,p) for the geometry optimization and the vertical excita-
tained using 6-31G**, reported in SI. tions, respectively, was identified as the best computational strat-
The H and L involved in the first transitions are completely delo- egy. Our calculations point out that the use of larger basis set on
calized over the whole molecule and have a p and p* character, the geometry optimizations has no influence on the structural
respectively. The H presents a high degree of conjugation that and electronic properties and does not improve the absorption
can be visualized with the charge delocalization on all the mole- spectra calculation accuracy.
cule through the C@C ring. Having established that (ab) procedure give results in excellent
In vacuo, the increase of the basis set does not affect the molec- agreement with the experiment both in vacuo and in solution at a
ular geometry, the charge distribution, the H–L gap or the first sin- lower computational cost than (b) procedure, this strategy has
glet-singlet excitation energy. On the contrary, TD-DFT been used to explore the spectral changes in different solvents
calculations in solution seem to be more sensitive to the basis sets and the aggregation in solution.
A bathochromic shift of indigo absorption spectra with the sol-
vent polarity has been experimentally observed [32–49]. To get in-
Table 1 sight into these effects the absorption spectra, and electron density
Energy [nm(eV)] and composition of the indigo computed lowest singlet–singlet
distribution change between S0 and S1 in vacuo and in different sol-
excitations together with the experimental absorption maxima.
vents have been computed. Results of the computed transitions, re-
a b ab Exp. ported in Table 2 (vacuo, chloroform, and DMSO) and in SI (CCl4,
Vac. 536 (2.31) 548 (2.26) 553 (2.24) 539 (2.30)a benzene, and ethanol) are in very good agreement with the exper-
H ? L (70%) H ? L (72%) H ? L (72%) iment and consistent with previous data [12] and are able to repro-
H-5 ? L (3%) H-5 ? L (2%) H-5 ? L (3%)
duce even the small bathochromic shifts observed in the
Chlor. 565 (2.20) 600 (2.07) 605 (2.05) 604 (2.05)b experimental absorption spectra. This shift has been interpreted
H ? L (70%) H ? L (74%) H ? L (74%)
as resulting from the increased stability of the charge-separated
H-5 ? L (2%)
structure in the excited state rather than in the ground state,
(a)Computed using 6-31G** // 6-31G** for the excitations // geometry optimizations. resulting in a decrease of the excitation energy [12,32].
(b) Computed using 6-311++G** // 6-311++G** for the excitations // geometry
Going from vacuo to chloroform and DMSO, a estabilization/sta-
optimizations (ab) Computed using 6-31++G**// 6-31G** for the excitations //
geometry optimizations.
bilization on the H/L is retrieved, resulting in a decrease of the H–L
a
From Ref. [30]. gap. The computed absorption maxima have similar composition,
b
From Ref. [31]. except for a minimal contribution of the H-5 ? L in vacuo, and
are in good agreement with the experimental values.
The composition of the H and L in terms of atomic orbitals show
a major contribution of the 2pz and 3pz of the N and the O, respec-
L L tively. For the first transition, that has manly a H to L character, this
is translated in a charge transfer upon excitation from the electron
2.40 eV donor ortho NH to the electron acceptor C@O group, as it can be
2.49 eV
observed from the electron density change computed in vacuo be-
H H tween S0 and S1 reported in Fig. 3. Upon excitation, other than the
decrease of the charge in the nitrogen atoms and the increase in
the oxygen atoms, it can be observed a loss of electron density in
Fig. 2. H and L isodensity plots and H–L gap energy of indigo computed in vacuo the center of inversion of the molecule, reflecting a decrease of
(left) and in chloroform (right). the conjugation in the excited state. The electron density change

Table 2
Computed H and L stabilities and composition in terms of atomic orbitals, H–L gap, and computed and experimental absorption spectra maxima [nm(eV)] in vacuo, chloroform
and DMSO.

H (eV) H comp. L (eV) L comp. H–L (eV) Comp. S1 Exp.

Vac. 5.67 41% N (2pz, 3pz) 3.18 28 %O (2pz, 3pz) 2.49 553 (2.24) 539 (2.39)a
15% C(C@C) (2pz, 3pz) 21% C(C@C) (2pz, 3pz) H ? L (74%)
15% C(CO) (2pz, 3pz) 18% C(CO) (2pz, 3pz) H-5 ? L (3%)
Chlor. 5.64 41% N (2pz, 3pz) 3.25 28 %O (2pz, 3pz) 2.38 605 (2.05) 604 (2.05)b
15% C(C@C) (2pz, 3pz) 21% C(C@C) (2pz, 3pz) H ? L (74%)
14% C(CO) (2pz, 3pz) 20% C(CO) (2pz, 3pz)
DMSO 5.63 41% N (2pz, 3pz) 3.27 28 %O (2pz, 3pz) 2.36 614 (2.02) 619 (2.00)c.
14% C(C@C) (2pz, 3pz) 21% C(C@C) (2pz, 3pz) H ? L (74%)
14% C(CO) (2pz, 3pz) 20% C(CO) (2pz, 3pz)
a
From Ref. [30].
b
From Ref. [31].
c
From Ref. [33]
46 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51

Within each indigo unit no relevant differences are retrieved

Fig. 3. Isodensity plot of the electron density difference between S0 and S1 in vacuo.
A blue (white) colour indicates a decrease (increase) of the electron density upon
excitation.
2.027
(1.996)
2.035
28.9°
29.2°
(25.8°)
(a) (b)
Fig. 4. Indigo dimer(a) and trimer(b) optimized structures. Average values of the
hydrogen bonds (Å) and dihedral angles (°) between the molecules are reported for
the chloroform(vacuo) calculations.
computed in chloroform and DMSO are very similar to those com-
puted in vacuo and are reported in SI.
3.2. Aggregation in solution
Experimental evidence of indigo aggregation in solution with
the appearance of a broad shoulder at 700 nm (1.77 eV) has been
reported. The spectral position of the aggregate indicates the for-
mation of more polar species than the monomer and excludes
the formation of an H-type aggregate [15]. More likely an hydro-
gen-bonded structure is formed retrieving similar effects on the
absorption spectra as the ones detected for the solid state where
the formation of an hydrogen-bonded structure has been proved
[50]. We have therefore investigated the geometry and absorption
spectra of two different aggregates (dimer and trimer) in chloro-
form following the strategy defined for the monomer.
The indigo dimer optimizations in vacuo and in solvent leaded,
in both cases, to stable hydrogen-bonded structures with all posi-
tive frequencies. Structures are reported in Fig. 4(a) together with
the main distances and angles.
between the dimer and the monomer optimized structures and
are also equivalent between vacuo and solvent calculations. Con-
trarily, some structural differences are retrieved in the unit
arrangements, being the angle between the two indigo units
29.2° (25.8°) in chloroform(vacuo) and the average values of the
hydrogen bonds between the two molecules 2.027(1.999) Å.
The geometry optimization in solution has been also performed
for the indigo trimer providing the structure reported in Fig. 4(b)
which shows inappreciable differences with respect to the dimer,
with average intermolecular hydrogen bond distances and dihedral
angles of 2.035 Å and 28.9°, respectively. Energetically, the dimer
and the trimer are computed 16.67 and 26.74 kcal/mol more sta-
ble, respectively, than the corresponding isolated monomer,
though these results are probably overestimated due to the basis
set superposition error (BSSE) that has not been corrected.
On the optimized structures in solution, the lowest singlet–sin-
glet transitions have been computed. The results, reported in Table
3, point out the appearance of new transitions at lower energies
that, even though they are characterized by small oscillator
strengths for both aggregates, they qualitatively reproduce the
new band detected experimentally upon increase of the indigo
concentration. The same results were obtained with calculations
performed in dichloroethane. Experimental spectra from Ref. [15]
at different concentrations together with the positions of the com-
puted vertical excitations for the monomer, the dimer and the tri-
mer are reported in Fig. 5.
The new experimental band at ca. 700 nm (1.77 eV) is com-
puted sligthly blue-shifted by 0.10 eV, a deviation that is within
the accuracy of the methodology. The molecular orbitals are
grouped in almost isoenergetic pairs(trios) for the dimer(trimer).
Only the molecular orbitals belonging to the first pair(trio) are in-
volved in the lowest transitions responsible for the colour band.
Their electronic distribution has been analyzed for the dimer. H,
H-1, L and L+1 molecular orbitals energies and electronic distribu-
tions are reported in Fig. 6.
The electronic delocalization in each indigo unit is very similar
to that computed for the isolated molecule, except for a slight de-
crease of the charge delocalization in the HOMOs and an increase
of the charge in the N in the LUMOs. In the H-1 and the L, the indigo
unit orbitals have the same symmetry while the contrary is true for
the H and L+1. Transitions are computed with a very low oscillator
strength that might be affected by an incomplete treatment of the
weak intermolecular hydrogen bond forces and to a not accurate
description of the long-range charge-transfer excitations [51–53].
3.3. Raman and IR spectra
Vibrational spectroscopy is one of the most powerful tools in
modern materials research. Quantum chemical calculations pre-
dicting harmonic wavenumbers and spectral intensities are essen-
tial when interpreting experimental spectra, particularly for large

Table 3
Computed TD-DFT transitions of indigo dimer and trimer in chloroform.

Dimer Trimer
Th. nm/eV f MO Th. nm/eV f MO
660/1.88(S1) 0.0071 (85%)H ? L 670/1.85(S1) 0.0113 (83%)H ? L
(7%) H-1 ? L+1 (4%)H ? L+2
647/1.92(S2) 0.0217 (63%)H ? L+1 652/1.90 (S3) 0.0392 (67%)H ? L+2
(37%)H-1 ? L (30%)H-2 ? L
612/2.02(S3) 0.0220 (80%)H-1 ? L+1 618/2.01(S5) 0.0298 (47%)H-2 ? L+2
(2%)H ? L (17%)H-1 ? L+1
603/2.06(S4) 0.5863 (50%)H-1 ? L 602/2.06(S7) 0.8436 (33%)H-2? L
(23%)H ? L+1 (26%)H-1 ? L+1
(26%)H? L+2
 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51
Fig. 5. Experimental absorption spectra in solvent at different indigo concentra-
tions from Ref. [15] and positions of the computed vertical excitations for the
monomer (red), the dimer (blue) and the trimer (green). (For interpretation of the
references in colour in this figure legend, the reader is referred to the web version of
this article.)
Fig. 6. MO electronic distribution of indigo dimer in solvent.
molecules in which the high density of states results in spectral
complexity. Accurate vibrational assignments for such systems
are necessary for the material characterization and they can be
proposed on the basis of wavenumber agreement between the
computed harmonics and the observed fundamentals [54].
The IR and Raman spectra have been experimentally measured
and computed and their comparison has allowed an assignment of
the experimental features in terms of functional groups displace-
ments. Our results have been also compared with previous data
available. Among the theoretical works [19–22] present in litera-
ture we report, alongside our results and assignment, Tatsch’s
assignment [19] which is, the reference paper most recalled by
experimental works on indigo. Moreover is the only work that cov-
ers the whole spectral range and reports IR and Raman spectra
with the experimental intensities and activities.
In Tatsch’s work, normal modes assignment for indigo were
performed on the basis of visual inspection. Usually, many atoms
are involved in a vibrational mode and the assignment to a func-
tional group in terms of stretching (m), in-plane bending (d), and
out-of-plane deformation (c) is not straightforward. We have
therefore performed a normal coordinate analysis in terms of po-
tential energy distributions (PEDs) which provides a strong sup-
port to the frequency assignment.
47
Our simulated and experimental vibrational spectra of indigo
are compared and reported in Fig. 7. Experimentally, the Raman
spectra was partially covered by its own fluorescence complicating
the analysis of the Raman spectra above 3000 nm, therefore exper-
imental data from Ref. [19] has been used for the assignment in
this region.
Even though it has been observed that a better agreement with
experiment is found using a different scaling factor at long wave-
numbers (>1800 cm 1) [55], we have chosen to perform a uniform
scaling in all the spectrum and therefore bands computed in the
high wavenumber region are quite overestimated.
Indigo observes a planar molecule that belongs to the C2h group
and has 84 fundamental modes divided into C = 29Ag + 13B-
g + 14Au + 28Bu. The rule of mutual exclusion holds and gerade
vibrations are Raman active only while ungerade are IR active only.
Ag and Bu types are planar while Au and Bg ones are non-planar.
Although our calculations have been performed without symmetry
constrains, the optimized structure neglectably deviates from the
C2h symmetry, therefore the computed modes are in perfect agree-
ment with the expected mode character distribution of the C2h
group and its 84 modes numbers are Ag(1–29), Au(30–43), Bg(44–
56) and Bu(57–84). The most intense modes of our experimental
and computed spectra are reported in Table 4 alongside with Tats-
ch’s assignment while the complete table with all the experimental
and computed bands is reported in SI. The experimental signals
that have not been related to the computed normal modes are
highlighted in yellow.
Overall, our experimental spectra is very similar to those mea-
sured in precedence with only slight differences mainly in the sig-
nals intensities, probably due to the use of different incident laser
wavelengths. However, a substantial difference is found on the
strong band that we measure at 1075 cm 1 in the IR spectrum that
is reported as a shoulder at 1074 cm 1 by Tatsch et al. No strong
bands are reported in the 1000–1100 cm 1 region by these authors
while a strong band is reported at 1065 cm 1 by Baran et al. [22]
We compute this band at 1052 cm 1 with an intensity of 54% with
respect to the most intense one, in line with the experimental find-
ings and we assign it to in-plane bending modes on the pyrrole
rings.
The computed frequencies show that most of the IR normal
modes have its correspondent mode in the Raman spectrum, with
shifted wavenumbers, differing only from their symmetry.
In the region above 2000 cm 1 10 normal modes are retrieved,
divided in ungerade/gerade pairs with almost the same frequen-
cies, that can be ascribed to NH and CH stretching. While medium
intensity mCN and mCH bands are retrieved in the Raman spectrum,
no intense mCH band is computed in the IR spectra on this region.
C@C and C@O stretching modes are computed in the 1700–
1500 cm 1 range, where we compute the most intense bands.
These modes are followed by a series of mCC (inter-ring stretching)
and in-plane dCH bendings. All the 27 out of plane modes, Au and
Bg are computed below the 1000 cm 1.
Computationally, the main differences with previous article
[19] are retrieved in the assignment of the normal modes to the
specific functional groups probably due to the different methodol-
ogies used to perform the assignments and not to the different le-
vel of theory. Similar wavenumbers are computed and all the
optical character coincide except for the low intensity bands of
the Raman spectrum at 1097 cm 1 that we compute Ag, see SI, in
contrast with the Bg reported by Tatsch, in line with the discrepan-
cies retrieved by Tomkinson [21].
In general a good agreement is found with the experiment. The
bigger discrepancies with the measured wavenumbers are re-
trieved in the high wavenumber region, as discussed above, and
in the IR band at 1627 cm 1 that we compute at 1660 cm 1. Low
intensity bands are sometimes difficult to assign but the good
48 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51

Fig. 7. Comparison between the computed (red) and the experimental (black) Raman (a) and IR (b) spectra. (For interpretation of the references in colour in this figure legend,
the reader is referred to the web version of this article.)

agreement with the experiment and an accurate analysis taking
into account the computed mode character and relative intensities
has allowed us to determine almost all the experimental signals.
3.4. Inter-molecular hydrogen bond effects on the vibrational spectra
In general, three main sources of divergences can be individu-
ated when comparing experimental spectra and those computed
with harmonic DFT calculations. Firstly, anharmonic effects are
not considered in this computational methodology although they
are partially corrected by the scaling factor used. Secondly, exper-
imental spectra can present resonance effects that are not taken
into account in our calculations and that may enhance particular
frequencies. Finally, calculations regard the isolated molecule,
therefore the possible effects of the intermolecular interactions
are ignored.
 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51 49

Table 4
Vibrational spectra of Indigo. Experimental and computed wavenumbers, computed relative intensities and raman activities, normal modes numbers, computed modes, this work
assignment, and previous work assignment.
1 1
Exp. IR (cm ) Exp. Ra. (cm ) Th. W. IR Int. Ra. Act. m Sym. Mode (prev. work) Modea
a
3270(vw) 3480 2% 23% 1 Ag 100% mNH mNH
3268(w)a 3479 14% 3% 57 Bu 80% mNH mNH
a
3060(w) 3091 <1% 32% 2 Ag 85% mCH mCH
3056(vw)a 3091 7% 3% 58 Bu 78% mCH mCH
1692(s) 1710 0% 8% 6 Ag 26% mC@C mCO
mC@C
1627(vs) 1660 54% 0% 62 Bu 40% mOC mCO
1624(m) 1631 0% 47% 7 Ag 24% mOC mCC
624% mC@C dCH
13% mCC
1614(s) 1602 100% 0% 63 Bu 26% mCC mCC
dCH
1581(s) 1596 0% 100% 8 Ag 13% mOC mC@C
16% mC@C mCO
18% mCC
1585(m) 1575 3% <1% 64 Bu 25% mCC mCC, dCH
1483(s) 1469 48% 0% 65 Bu 20% mCC mCC
29% dHCC
1462(s) 1441 30% 0% 66 Bu 35% dHCC mCC
dCH
1395(m) 1400 58% 0% 67 Bu 9% mNC dNH
16% dHNC dCH
10% dHCC
1362(s) 1361 0% 2% 12 Ag 18% mNC dNH
7% dHCC dCH
1347 0% 13% 13 Ag 32% mNC dNH
1317(m) 1319 14% 0% 68 Bu 33% mCC mCC
12% dHCC
1308(w) 1303 0% 15% 14 Ag 19% mCC mCC
26% dHCC
1299(m) 1274 20% 0% 69 Bu 18%dHCC dCH
1247(m) 1227 0% 15% 15 Ag 16% mNC dCH
22% dHCC dCO
1199(m) 1169 26% 0% 71 Bu 11% mCC mCC
5% dCCC mCN
10% dHCC
1173(m) 1152 63% 0% 72 Bu 13% mNC dCC
7% dHNC
5% dHCC
1127(s) 1122 53% 0% 73 Bu 9% mNC dCC
9% dHNC dC-NH-C
20% dHCC
1075(s) 1052 54% 0% 75 Bu 12% dCCC mC-NH-C
10% dOCC
10% dCNC
754(m) 736 12% 0% 34 Au 32% cHCCC dCH
8% cNCCC dNCC
713(m) 690 4% 0% 79 Bu 27% dCCC c5-Ring
566(m) 550 10% 0% 36 Au 19% cCCCC c5-Ring
6% cNCCN
548 11% 0% 81 Bu 24% dCCC
547(m) 533 0% 3% 26 Ag 12% mCC dC@C–CO–C
39% dCCC
508(w) 475 18% 0% 37 Au 58% cHNCC dC@C–C@O
5% cNCCN
a
Form Ref. [19].

Anharmonicity and resonance effects can be taken into account

a=9.24 Å ; b=5.77 Å with the more advanced quantum mechanical codes and, even
c=12.22 Å ; ß = 117° though it would be interesting to perform a detailed analysis of
their influence on the computed spectra, such a study is beyond
the aim of this work, where we have focused our attention on
the intermolecular interactions and their effects on the optical
properties of indigo.
In the solid, molecules are disposed in parallel planes with two
different orientations (see Fig. 8).
Each indigo molecule form four hydrogen bonds of ca 2.113 Å
with the four neighboring molecules belonging to the other orien-
tation planes. Frequency calculations on a 5 unit indigo structure
Fig. 8. Crystal structure of indigo from Ref. [50] were too time demanding and attempts to optimize a 3 unit cluster
50 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51
from the crystal structure, with indigo bonding the two neighbor-
ing molecules in parallel planes, were vain. However, it can be rea-
sonably expected that most of the solid effects are due to the
formation of hydrogen bonds rather than to weaker forces as Van
der Waals forces between planes. Therefore, the optimized dimer
structure in vacuo, where intermolecular hydrogen bonds are pres-
ent, has been used to evaluate the effects of the hydrogen bonding
in the solid. The use of the dimer is clearly a simplification of the
real case but we expect the hydrogen bonding effects to be at least
qualitatively reproduced by the model.
Comparison between the computed Raman and IR spectra of the
monomer and the dimer is reported in Fig. 9.
In the Raman spectrum of the dimer, the band conputed in the
3400 cm 1 region is divided into not hydrogen-bonded mNH
(3459 cm 1) and hydrogen-bonded mNH (3423 cm 1). This duplica-
tion is due to the simplified model used, where only one side of the
molecule is bonded to another unit. In the solid, where all the NH
are bonded to the neighboring molecules, band at 3459 cm 1
should disappear. The hydrogen-bonded mNH is computed at short-
er wavenumbers with respect to the monomer improving the
agreement with the experiment. Also mC@C band at 1710 cm 1 is
shifted to 1702 cm 1 in the dimer, being comparable with the
1692 cm 1 measured value. Other differences can be found in the
1600 cm 1 region where a new low intensity band appears at
1653 cm 1 corresponding to mCO. On the other hand, frequencies
at 1596 and 1631 cm 1 related to mCO and mC@C, increment their
relative intensity differences and are shifted to 1585 and
Fig. 9. Comparison between the monomer (black) and the dimer (red) Raman (a) and IR (b) computed spectra. (For interpretation of the references in colour in this figure
legend, the reader is referred to the web version of this article.)
1621 cm 1, respectively, improving the agreement with the exper-
imental bands measured at 1581 and 1624 cm 1. A new band ap-
pears at 1395 cm 1 associated to dCNH that matches the
unassigned experimental shoulder at 1411 cm 1 (see SI). In the
1100–1250 cm 1 region wavenumbers computed for the dimer
are almost unvaried with respect to the corresponding frequencies
in the monomer though the appearance of several new bands
changes the spectrum shape improving the agreement with the
experimental spectrum.
In the IR spectrum the mNH modes are also shifted to smaller
wavenumbers improving the agreement with the experiment. In
this case a very intense band at 3566 cm 1 is computed corre-
sponding to an asymmetric mNH in the hydrogen bonded NH. Con-
trarily to what retrieved for the Raman activities in the 1600 cm 1
region, mCO and mC@C IR relative intensities differences diminish
becoming equivalent manly due to an increase of intensity of the
asymmetric mCO band in line with the experiment where the mCO
band at 1627 cm 1 is retrieved more intense than mCC at
1614 cm 1. This band is also shifted to 1651 cm 1 in the dimer
improving the agreement with the experiment (1627 cm 1), with
respect to the monomer (1660 cm 1) and to wavenumber com-
puted in previous article [19] (1692 cm 1) although it is still quite
overestimated. In the 1100–1250 cm 1 region the IR spectrum of
the dimer is almost unchanged with respect to the monomer, since
bands in this region do not involve NH and CO modes.
As it could be expected, all the perceptible changes in the vibra-
tional spectra of the hydrogen-bonded aggregates with respect to
 A. Amat et al. / Journal of Molecular Structure 993 (2011) 43–51
the monomer can be traced back to modes involving the CO and
NH. Other modes affected involve the C@C bond that is tightened
with the aggregation. Obviously, our simple two-molecule model
can not completely reproduce all the solid state interactions but
can give hints regarding the effects of the hydrogen bonds between
molecules.
4. Conclusions
A combined experimental–theoretical strategy has been used in
this work to further understand the vibrational and electronic
properties of indigo, paying particular attention to the role of
aggregation on the absorption and vibrational spectra.
The absorption spectra in vacuo and in different solvents have
been computed and compared to the experiment, evidencing that
TD-DFT calculations provide excellent results, able to reproduce
even small bathocrhomic shifts. These shifts are related to a desta-
bilization/stabilization of the H/L and a decrease of the H–L gap.
The appearance of a new band at ca. 700 nm in the absorption
spectrum of indigo solution at high concentrations has been inves-
tigated by simulating the absorption spectra of different size clus-
ter aggregates. Stable dimer and trimer aggregates in vacuo and in
solution have been computed, revealing that aggregation takes
place with the formation of hydrogen bonded structures. For the
studied indigo aggregates a series of new electronic transitions
are computed. These transitions, retrieved at lower energies with
respect to the monomer ones, can be directly related to the exper-
imental band at 700 nm, thus confirming this band as the signature
of aggregation. Moreover, the optimized aggregates recall the indi-
go crystal structure, their intermolecular hydrogen bonds being
similar the interactions present in the solid state.
The IR and Raman spectra have been measured and computed,
finding a good agreement between theory and experiment. Normal
modes have been analyzed by means of potential energy distribu-
tion allowing to perform an unambiguous assignment to the func-
tional groups. Only a few experimental bands remain not assigned
or present important differences with the computed values. Due to
the similarity between the optimized aggregates and the crystal in-
digo structure, the effects of the hydrogen bonds present in the so-
lid on the vibrational properties have been evaluated by computing
the Raman and IR spectra on the optimized dimer aggregate. Re-
sults point out the main differences regarding the NH and CO
stretching, that are shifted to shorter wavelengths, and modes
involving C@C bond, tightened with aggregation. Some of the
uncertainties and inaccuracies in the assignment were improved
by computing the vibrational spectra of the dimer.
This work gathers important information for the comprehen-
sion of the spectroscopic properties of indigo, one of the most stud-
ied natural dyes present in nature. The computational strategy
described and successfully used in this study, can be extended to
the investigation of the optical and vibrational properties of differ-
ent dyes of interest in the art heritage field, thus complementing
the experimental results.
Acknowledgements
This work supported by the project Eu-ARTECH (CT-2004-
506171) within the 6th FP of the EC and the European Union
FP7-Research Infrastructure programme under the CHARISMA pro-
ject (GA228330).
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.molstruc.2010.11.046.
51
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