Deep red emitting triphenylamine based coumarin-rhodamine hybrids with large Stokes shift and viscosity sensing: Synthesis, photophysical properties and DFT studies of their spirocyclic and open forms

Article abstract of DOI:10.1016/j.dyepig.2016.11.010

We designed and synthesized triphenylamine based and coumarin fused rhodamine hybrid dyes and characterized using ¹H, ¹³C NMR and HR-LCMS analysis. Both the newly synthesized hybrid dyes were found to show red shifted absorption as well as emissions and large Stokes shift (40–68 nm) as compared to the small Stokes shift (25–30 nm) of reported dyes Rhodamine B and 101. Photophysical properties of these dyes were studied in different solvents and according to the solvents acidity or basicity they preferred to remain in their spirocyclic or open form in different ratio. We studied the spirocyclic as well as open form derivatives of these dyes for their viscosity sensitivity in three different mixture of solvents i.e. polar-protic [EtOH-PEG 400], polar-aprotic [toluene-PEG 400] and non-polar-aprotic [toluene-paraffin]. They are found to show very high viscosity sensitivity in polar-protic mixture of solvents [EtOH-PEG 400] and hence concluded that both polarity as well as viscosity factor worked together for the higher emission enhancement rather than only viscosity factor. As these dyes showed very high viscosity sensitivity in their spirocyclic as well as open form, they can be utilized as viscosity sensors in visible as well as deep red region. We also correlated our experimental finding theoretically by using Density Functional theory computations.

Full text of DOI:10.1016/j.dyepig.2016.11.010

Accepted Manuscript
Deep red emitting triphenylamine based coumarin-rhodamine hybrids with large
Stokes shift and viscosity sensing: Synthesis, photophysical properties and DFT
studies of their spirocyclic and open forms
Shantaram Kothavale, Amol G. Jadhav, Nagaiyan Sekar
PII:
S0143-7208(16)30792-6
10.1016/j.dyepig.2016.11.010
DYPI 5579
DOI:
Reference:
To appear in: Dyes and Pigments
Received Date: 21 September 2016
Revised Date: 8 November 2016
Accepted Date: 8 November 2016
Please cite this article as: Kothavale S, Jadhav AG, Sekar N, Deep red emitting triphenylamine based
coumarin-rhodamine hybrids with large Stokes shift and viscosity sensing: Synthesis, photophysical
properties and DFT studies of their spirocyclic and open forms, Dyes and Pigments (2016), doi: 10.1016/
j.dyepig.2016.11.010.
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ACCEPTED MANUSCRIPT
Deep red emitting triphenylamine based coumarin-rhodamine hybrids with large Stokes
shift and viscosity sensing: Synthesis, photophysical properties and DFT studies of their
spirocyclic and open forms
Shantaram Kothavale, Amol G. Jadhav, Nagaiyan Sekar*
Department of Dyestuff Technology, Institute of Chemical Technology, Matunga,
Mumbai-400 019, India.
E-mail: n.sekar@ictmumbai.edu.in, nethi.sekar@gmail.com
Tel: +91 22 3361 2707; Fax: +91 22 3361 1020.
Graphical Abstract

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Deep red emitting triphenylamine based coumarin-rhodamine hybrids with
large Stokes shift and viscosity sensing: Synthesis, photophysical properties
and DFT studies of their spirocyclic and open forms
Shantaram Kothavale, Amol G. Jadhav, Nagaiyan Sekar*
Department of Dyestuff Technology, Institute of Chemical Technology, Matunga,
Mumbai-400 019, India.
E-mail: n.sekar@ictmumbai.edu.in, nethi.sekar@gmail.com
Tel: +91 22 3361 2707; Fax: +91 22 3361 1020.
Abstract
We designed and synthesized triphenylamine based and coumarin fused rhodamine hybrid dyes
1
and characterized using H, 13C NMR and HR-LCMS analysis. Both the newly synthesized
hybrid dyes were found to show red shifted absorption as well as emissions and large Stokes
shift (40-68 nm) as compared to the small Stokes shift (25-30 nm) of reported dyes Rhodamine B
and 101. Photophysical properties of these dyes were studied in different solvents and according
to the solvents acidity or basicity they preferred to remain in their spirocyclic or open form in
different ratio. We studied the spirocyclic as well as open form derivatives of these dyes for their
viscosity sensitivity in three different mixture of solvents i.e. polar-protic [EtOH-PEG 400],
polar-aprotic [toluene-PEG 400] and non-polar-aprotic [toluene-paraffin]. They are found to
show very high viscosity sensitivity in polar-protic mixture of solvents [EtOH-PEG 400] and
hence concluded that both polarity as well as viscosity factor worked together for the higher
emission enhancement rather than only viscosity factor. As these dyes showed very high
viscosity sensitivity in their spirocyclic as well as open form, they can be utilized as viscosity

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sensors in visible as well as deep red region. We also correlated our experimental finding
theoretically by using Density Functional theory computations.
Keywords: Coumarin rhodamine hybrids, solvatochromism, viscosity sensitivity, deep red
emitting dyes, Density functional theory (DFT).
Introduction
Fluorescent environment sensitive dyes such as molecular rotors are widely used to measure
viscosity in biological systems in place of conventional methods due to its advantages such as
small sample volume and high speed of the readout [1–3]. Hence currently fluorescent
molecular probes are the most suitable tools for real-time measurements of viscosity in both
micro- and macro systems [4,5]. Molecular rotors are a group of fluorescent compounds that
consists of an electron donor group in π-conjugation with an electron acceptor group which form
twisted intramolecular charge transfer (TICT) states after excitation. In such cases fluorescence
quenching via an internal non-radiative process that involves molecular rotation between the
donor and the acceptor (TICT) is observed which is hindered in highly viscous environment to
regain the fluorescence emission [6,7]. 9-Dicyanovinyl)-julolidine (DCVJ) and its different
derivatives have been extensively studied for their viscosity sensitivity which are weakly
fluorescent initially due to the fast rotation around the single bond interconnecting the julolidine
and dicyanovinyl subunits [8–13] . Freely rotating phenyl ring at meso position in BODIPY dye
is also utilized for application as viscosity sensors [14–17].
Though some highly fluorescent dyes like rhodamine, fluorescein, BODIPY and cyanine
derivatives have been widely used as fluorescent probes[18,19], labels[20–22], logic gates

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[23,24], light-emitting materials [25–28], and light-harvesting systems [29–32], they have the
serious disadvantage of very small Stokes shifts (typically less than 25 nm), which can lead to
serious self-quenching and fluorescence detection errors because of excitation backscattering
effects [33,34]. The large Stokes shift could reduce self-absorption as well as obtain high
resolution and low detection limits. Therefore, it remains highly desirable to explore new design
strategies for the development of deep red fluorescent dyes with great photo stability and large
Stokes shift while being used as ratiometric sensing platforms.
Similarly deep red emitting or NIR fluorescent probes are attractive for biological
applications because of minimum photo damage to biological samples, deep tissue penetration,
and minimum interference from the background auto fluorescence of biomolecules in the living
systems [35,36]. Rhodamine dyes due to their excellent photophysical properties, such as high
molar extinction coefficients, excellent fluorescence quantum yields, and great photo stability,
are widely used as fluorescent probes and molecular markers in chemistry and biology [37–39].
However, the absorption and emission wavelengths of most rhodamine derivatives are below 600
nm and exhibit a very small Stokes shift (rhodamine B: 19 nm) which is sometimes unsuitable
for biological applications due to self-absorption. Many efforts have been made to acquire
longer-wavelength rhodamine derivatives, including extending the conjugation of the xanthene
ring, replacing the central carbon with a nitrogen atom or introducing a cyano group on the
central carbon atom, as well as replacing the oxygen bridge atom by other elements, such as N,
C, S, and Se [40,41]. However, rhodamine analogues developed by these strategies are very
difficult to synthesize.
Hence currently trend to hybridize two totally different dyes with unique photophysical
properties is being rapidly accepted by researchers to obtain the hybrid dye with desired

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photophysical properties. For example, ESIPT-BODIPY [42,43], ESIPT-Coumarin [44,45],
BODIPY-Coumarin [46,47], BODIPY-Rhodamine [48,49] and Coumarin- Rhodamine [50] dyes
were synthesized successfully and found to show better photophysical properties than their
precursors. Recently P. Wang and coworkers synthesized fused coumarin-rhodamine hybrid dyes
and utilized them for bioimaging study in the NIR region. The new hybrid dyes not only retained
the advantages of coumarin and rhodamine individual derivatives in themselves but also found to
show larger Stokes shift and red shifted emission wavelengths [50].
In this work, we have successfully synthesized triphenylamine based two novel deep red
emitting coumarin-rhodamine hybrid fluorescent dyes. Environment friendly deep eutectic ionic
liquid i.e. NMP.HSO4 is used as the reaction media to obtain the coumarin fused rhodamine dyes
in good yield instead of using traditional highly concentrated acids. As TICT is responsible for
the viscosity sensitivity we designed the dyes so that triphenylamine can be used as main donor
from coumarin side and N,N diethyl aniline or julolidine as main donors from rhodamine side.
The pure dyes in their spirocyclic form were studied for their viscosity sensitivity in polar protic
[EtOH-PEG 400], aprotic polar [toluene-PEG 400] and aprotic non-polar [toluene-paraffin]
viscous media in the presence and absence of TFA. A substantial change in the emission
intensities in the visible as well as deep red region were observed for both the dyes particularly
in the polar viscous medium [EtOH-PEG 400]. Fig. 1 represents the structure of hybrid dyes
Cou-Rh-1 and Cou-Rh-2 in their spirocyclic as well as open form.
<< Please insert Fig. 1. Structure of the coumarin-rhodamine hybrid dyes Cou-Rh-1 and Cou-
Rh-2 >>
2. Experimental section
2.1 Materials and equipments

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All the reagents and solvents were purchased from S d Fine Chemical Limited, Mumbai,
India and used without further purification. The reactions were monitored by TLC using on 0.25
1
mm E-Merck silica gel 60 F254 precoated plates, which were visualized with UV light. H and
13C NMR spectra were recorded on a Varian Cary Eclipse Australia using TMS
(Tetramethylsilane) as an internal standard. High resolution liquid chromatography mass spectra
were recorded on 6550 iFunnel QTOF LC/MS spectrometer from Agilent Technologies. The
absorption spectra of the compounds were recorded on a PerkinElmer UV- visible
spectrophotometer Lambda 25. Fluorescence emission spectra were recorded on Varian Cary
Eclipse fluorescence spectrophotometer using freshly prepared solutions at the concentration of
1x10-6 mol L-1. Melting points were measured on standard melting point apparatus from Sunder
Industrial Product Mumbai and are uncorrected.
2.2 Computational Methods
Density Functional Theory (DFT) [51] was utilized to optimize ground state (S0) geometries of
all the compounds using the Gaussian 09 package and the popular hybrid functional B3LYP. The
B3LYP combines Becke’s three parameter exchange functional (B3)[52] with the nonlocal
correlation functional by Lee, Yang, and Parr (LYP) [53]. In both the methods DFT and time-
dependent density functional theory (TD-DFT), same basis set i.e. 6-31G (d) was used. The low-
lying first singlet excited states (S1) of the dyes were relaxed to obtain their minimum energy
geometries using the TD-DFT in their gas phase. By using TD-DFT at the same hybrid
functional and basis set, the vertical excitation energies and oscillator strengths were obtained for
the lowest 10 singlet-singlet transitions at the optimized ground state equilibrium geometries

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[54]. Optimized ground state structures were utilized to obtain the electronic absorption spectra,
including maximum absorption and oscillator strengths.
<< Please insert Scheme 1: Synthesis of coumarin-rhodamine hybrids Cou-Rh-1 and Cou-Rh-2
>>
2.3 Synthetic strategy
We synthesized methoxy triphenylamine intermediate 2 starting from aniline utilizing Buchwald
reaction condition, which was further de-methylated to acquire intermediate 3 by using
pyridine.HCl as de-methylating reagent. At the same time intermediate 10 was synthesized
starting from 2, 4, 6 tri-chlorophenol (9) using reported procedure [55]. Intermediate 3 was
reacted with intermediate 10 to get triphenylamine based 4-hydroxyl coumarin derivative (4).
We also synthesized keto-acid intermediates 6 and 8 following the reported procedures [56,57]
and treated with coumarin intermediate 4 in ionic liquid HNMP.HSO4 as reaction media to get
the desired coumarin-rhodamine hybrids Cou-Rh-1 and Cou-Rh-2 in their spirocyclic form.
HNMP.HSO4 act as reaction medium as well as acid catalyst for the synthesis of desired
coumarin fused rhodamine derivatives. We further converted the spirocyclic form of Cou-Rh-1
and Cou-Rh-2 dyes into their respective open form by treating them with MeOH.HCl solution,
in which they get immediately converted into their respective open form hydrochloride salt form.
This procedure was repeated twice and the solvent was evaporated on rotavapour to get shiny
reddish black colored open salt form derivatives of Cou-Rh-1 and Cou-Rh-2.
2.4 Synthesis and Characterization
2.4.1 Synthesis of 3-methoxy-N,N-diphenylaniline (2)

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Same procedure as reported from our group[58] was followed.
2.4.2 Synthesis of 3-(diphenylamino)phenol (3)
3-Methoxy-N,N-diphenylaniline 2 (16 g, 58.2 mmol) was dissolved in 80 g of pyridine. HCl and
o
heated to 200 C for 10 hrs. After cooling to room temperature, water was added in the reaction
mass and solid precipitated out was filtered, dried well and collected as crude product, which
was further purified on column chromatography using 10% ethyl acetate in hexane as the eluent
to get the pure product.
Yield: 12 g, (79 %); Melting point = 98 – 100 oC
1H NMR (500 MHz, CDCl3) δ 4.60 (s, 1H), 6.46 (dd, J = 8 and 2 Hz, 1H), 6.52 (t, J = 2 Hz, 1H),
6.65 (dd, J = 8 and 2 Hz, 1H), 7.01-7.07 (m, 2H), 7.09-7.11 (m, 4H), 7.23-7.27 (m, 4H), 13C
NMR (126 MHz, CDCl3) δ 109.4, 110.4, 116.1, 123.1, 124.7, 129.3, 130.1, 147.6, 149.3, 156.2.
Elemental analysis calcd (%); Mol. formula: C18H15NO (C: 82.73, H: 5.79, N: 5.36, O: 6.12;
found: C: 82.68, H: 5.81, N: 5.32, O: 6.08).
2.4.3 Synthesis of 7-(diphenylamino)-4-hydroxy-2H-chromen-2-one (4)
3-(Diphenylamino) phenol 3 (5 g, 19.1 mmol) and diphenylmalonate 8 (9.7 g, 21 mmol) were
added into anhydrous toluene (20 mL). The mixture was heated to reflux for overnight, and after
completion the reaction mass was cooled to room temperature. The solid precipitated out was
filtered as pure product.
Yield: 3.6 g (57 %) Melting point = 258 – 262 oC
1H NMR (500 MHz, DMSO-d6) δ 5.39 (s, 1H), 6.51-6.52 (d, J = 2.0, 1H), 6.72-6.45 (dd, J = 8.5,
2.5, 1H), 7.15-7.20 (m, 6H), 7.36-7.39 (m, 4H), 7.59-7.61 (d, J = 8.5, 1H), 12.20 (s, 1H). 13C
NMR (126 MHz, DMSO-d6) δ 88.6, 105.5, 108.9, 115.7, 124.6, 125.6, 126.6, 130.4, 146.1,

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151.8, 155.3, 162.5, 166.3, 168.8. Elemental analysis calcd (%); Mol. formula: C21H15NO3 (C:
76.58, H: 4.59, N: 4.25, O: 14.57; found: C: 76.53, H: 4.62, N: 4.19, O: 14.59).
2.4.4 General procedure for the synthesis of coumarin-rhodamine hybrids Cou-Rh-1 and
Cou-Rh-2
4-Hydroxy coumarin intermediate 4 was reacted with keto-acid intermediate 6 and 8 in H-
o
NMP.HSO4 (10 mL) at 90 C for 14 - 16 hours. After completion the reaction mixture was
cooled to room temperature and was poured into 100 mL ice-cold water and stirred for 15 min.
The product was extracted with chloroform (100 mL x 3); the organic layer was separated, dried
over anhydrous Na2SO4 and evaporated over rotary evaporator. The crude product was purified
by silica gel (100-200 mesh size) column chromatography using 1-5 % methanol in chloroform
to give the respective dyes Cou-Rh-1 and Cou-Rh-2.
Dye Cou-Rh-1:
Yield = 28 %, Melting point: 258 oC
1H NMR (500 MHz, CDCl3) δ 1.17 (t, J = 7.5 Hz, 6H), 3.34 (q, J = 7 Hz, 4H), 6.42 (dd, J = 9
and 2.5 Hz, 1H), 6.49 (d, J = 2.5 Hz, 1H), 6.57 (d, J = 9 Hz, 1H), 6.76 (d, J = 2 Hz, 1H), 6.95
(dd, J = 9 and 2.5 Hz, 1H), 7.12 (t, J = 7.5 Hz, 1H), 7.16-7.19 (m, 5H), 7.33-7.36 (m, 4H), 7.50-
7.5 (m, 2H), 7.89 (d, J = 8.5 Hz, 1H), 7.98 (d, J = 8 Hz, 1H).13C NMR (126 MHz, CDCl3) δ
12.5, 44.4, 80.2, 96.6, 97.4, 106.2, 110.2, 116.1, 122.3, 124.0, 124.7, 125.3, 126.2, 128.0, 128.9,
129.0, 129.7,134.0, 145.8, 149.3, 150.5, 152.5, 152.9, 154.7, 158.2, 159.3, 170.0. HRMS (ESI):
m/z calculated for (M + H) + C39H30N2O5 607.2155; found 607.2176.
Dye Cou-Rh-2:

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Yield = 23 %, Melting point: >300 oC.1H NMR (500 MHz, CDCl3) δ 1.75-1.93 (m, 4H), 2.42-
2.43 (m, 2H), 2.93 (t, J = 7 Hz, 2H), 3.11 (m, 2H), 3.17 (t, J = 7 Hz, 2H), 6.01 (s, 1H), 6.51 (d, J
= 2.5 Hz, 1H), 6.88 (dd, J = 9 and 2.5 Hz, 1H), 7.20-7.27 (m, 7H), 7.40-7.43 (m, 4H), 7.55-7.65
(m, 2H), 7.85 (d, J = 7.5 Hz, 1H), 7.94 (d, J = 9 Hz, 1H).13C NMR (126 MHz, CDCl3) δ 25.5,
26.1, 31.8, 35.8, 53.6, 54.2, 85.5, 99.7, 109.3, 110.4, 110.7, 111.9, 120.9, 124.7, 128.0, 129.1,
129.2, 129.8, 130.9, 131.5, 132.3, 134.4, 135.2, 139.8, 149.3, 149.9, 150.4, 157.5, 157.8, 159.2,
162.6, 163.8, 174.5. HRMS (ESI): m/z calculated for (M + H) + C44H30N2O5 631.2155; found
631.2174.
3. Result and discussion
3.1 Photophysical properties
We studied the absorption and emission properties of these coumarin-rhodamine hybrid
derivatives in their spirocyclic as well as open form in different solvents. The spirocyclic form of
these dyes was converted into their open form using MeOH.HCl solution. The solvent was
evaporated on rotavapour and the same procedure was repeated twice to get the fully open form
derivatives. During photophysical studies after dilution in organic solvents these dyes again
preferred to remain in their spirocyclic form, hence we added required amount of TFA till the
disappearance of the spirocyclic absorption peak to acquire their complete open form. As there
are two main chromophores i.e. coumarin and rhodamine, hence two distinct absorption and
emission peaks for both the moieties are expected, but due to their fused hybrid structures they
behave as coumarin dye in their spirocyclic form to show absorption peak around 371 nm and
emission peak around 430 nm. In their open form they behave as rhodamine dye and show
absorption peak at around 610 nm and emission peak at around 670 nm. In DMSO solvent

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specifically they preferred to remain in their spirocyclic form even after adding excess TFA.
They are colorless solid in their spirocyclic form and are reddish black shiny compounds in their
open form. In their spirocyclic as well as open form no much difference in the absorption and
emission λmax is observed between the Cou-Rh-1 and Cou-Rh-2 dyes (Fig. 2 and S1). In their
open form both Cou-Rh-1 and Cou-Rh-2 dyes exhibited comparatively higher Stokes shifts (40-
68 nm) as compared to the small Stokes shift of 25-30 nm of reported RH- B and RH-101 dyes
(Table 1). Due to their extended length of conjugation and fused structures they also showed
highly red shifted absorption and emission peaks than the reported RH-B and RH-101 dyes in
their open form (Fig. S2).
<< Please insert Fig. 2: Normalized absorption and emission spectra of dyes Cou-Rh-1 and Cou-
Rh-2 in their open form >>
As N,N diethyl amine donating group in Cou-Rh-1 dye is replaced by rigid julolidine donor in
Cou-Rh-2 dye, we expect red shift in their absorption and emission λmax but not much shift in
both spirocyclic and open form is observed as like in RH-B and RH-101 reported dyes where
when we go from N,N diethyl aniline to julolidine donor, red shift of 20 and 22 nm is observed in
their absorption as well as emission spectra respectively (Fig. 3). As the role of donating group is
not clearly differentiated in the case of these coumarin-rhodamine dyes, the coumarin side
triphenylamine donor may have taken part in the spirocyclic ring opening mechanism of
rhodamine dye to disturb the main donor-acceptor spirocyclic ring opening mechanism of
rhodamine. From the available absorption data of these dyes in their spirocyclic as well as open
form we calculated their oscillator strength (f) as well as transition dipole moment (µeg) by using

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reported expressions [59]. In general it is observed that both the dyes showed higher values of
oscillator strength and transition dipole moment in their open form as compared to the respective
spirocyclic form (Table 1).
<< Please insert Fig. 3: Normalized absorption and emission spectra of RH-B, RH-101, Cou-
Rh-1 and Cou-Rh-2 in their open form in chloroform solvent >>
<< Please insert Table 1: photophysical properties of Cou-Rh-1 and Cou-Rh-2 in their
spirocyclic and open form >>
To check out the effect of solvents in their spirocyclic and open form we studied the
photophysical properties of Cou-Rh-1 and Cou-Rh-2 dyes in both spirocyclic and open form
and it is observed that in both the cases quenching of fluorescence from non-polar to polar
solvents is observed (Fig. 4) . But the red shifted emissions observed in their spirocyclic form
suggest that these dyes behave as typical coumarin dyes in their spirocyclic form and as typical
rhodamine dyes in their open form. Also it can be predicted that the excited state is highly polar
in their spirocyclic form as compared to the ground state and vice versa in the case of their open
form derivatives. In their spirocyclic form for both Cou-Rh-1 and Cou-Rh-2 dyes in acetone
solvent typically red shifted emission λmax values (497 and 504 nm) and consequently very high
Stokes shift of 125 and 133 nm were observed respectively. The molar extinction coefficient
values are almost double for both the dyes when we go from spirocyclic to open form suggesting
that the energy gain process actually switched on only after the ring opening of the rhodamine
dye.

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<< Please insert Fig. 4: Absorption (a) and emission (b) spectra of Cou-Rh-1 in different
solvents in their spirocyclic form. >>
<< Please insert Table 2: Photophysical properties of Cou-Rh-1 dye in its spirocyclic form in all
solvents >>
Dye Cou-Rh-1 showed higher Stokes shift of 10-15 nm than Cou-Rh-2 dye in all the tested
solvents in their open form, but opposite trend is observed in their spirocyclic form where dye
Cou-Rh-2 showed comparatively higher Stokes shift than dye Cou-Rh-1 in all solvents (Table
2-3 and Table S1-S2). In their open form, both these dyes Cou-Rh-1 and Cou-Rh-2 showed
highest Stokes shift of 68 and 54 nm in chloroform solvent respectively. In their spirocyclic form
comparatively red shifted emissions were observed for dye Cou-Rh-2 (435-500 nm) than Cou-
Rh-1 dye (430- 476 nm), while the trend is opposite in their open form where dye Cou-Rh-1
(641-678 nm) exhibited red shifted emissions than dye Cou-Rh-2 (639-668 nm). From above
observations it is clear that dye Cou-Rh-2 showed better results in its spirocyclic form while dye
Cou-Rh-1 showed better results its open form. As very high quenching of fluorescence is
observed for dye Cou-Rh-1 in its spirocyclic form than dye Cou-Rh-2 (Fig. 4 and S3), the freely
rotating N,N diethyl aniline donating group may have get twisted in its excited state than the
rigid donor of Cou-Rh-2 dye. Also in their open form comparatively broader absorption peaks
were observed for dye Cou-Rh-2 than dye Cou-Rh-1 (Fig. 5 and S4).

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<< Please insert Fig. 5. Absorption (a) and emission (b) spectra of Cou-Rh-1 in all solvents
(open form) >>
<< Please insert Table 3: Photophysical properties of Cou-Rh-1 dye in its open form in all
solvents >>
3.2 Solvatochromism
In their spirocyclic form both Cou-Rh-1 and Cou-Rh-2 dyes shows red shift in their emission
λmax values from non-polar to polar solvents suggesting that the excited state is highly polar and
is stabilized by the polar solvents very well as compared to the non-polar solvents. This
observation further supported by the fact that very small variation in the absorption λmax values is
observed from non-polar to polar solvent suggesting that the ground state is comparatively less
polar than the excited state of these dyes. Fig. 6 represents normalized emission spectra of dye
Cou-Rh-1 in all solvents with red shift of 67 nm from non-polar to polar solvents. We correlated
the solvatochromism observed in these dyes by using Lippert-Mataga equation. It is a very
general equation for the effect of solvent properties such as dielectric constant and refractive
index on emission λmax values and does not account for specific solvent–fluorophore interactions,
e.g. hydrogen bonding. The linear relationship between Stokes shift and Lippert-Mataga function
for both dyes in their spirocyclic form (Fig. 7) suggests that solvent parameters such as dielectric
constant and refractive index are collectively responsible for the red shifted emissions in polar
solvents. As like other common rhodamine dyes no any red shift in their emission spectra for the
open form of these dyes was observed suggesting that in their open form as like excited state
ground state of these dyes is also stabilized by the polar solvents.

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<< Please insert Fig. 6. Normalized emission spectra of Cou-Rh-1 dye in all solvents (open
form) >>
<< Please insert Fig. 7. Lippert-Mataga plots of dyes Cou-Rh-1 and Cou-Rh-2 in their
spirocyclic form >>
3.3 Effect of solvent acidity and basicity on the ring opening mechanism of coumarin-
rhodamine hybrid dye Cou-Rh-1
When we recorded the absorption spectra of these coumarin-rhodamine hybrid dyes in their
spirocyclic as well as open form in different solvents, we found large variation in their molar
extinction coefficient both in spirocyclic as well as open form. Fig. 8 represents the absorption
spectra of Cou-Rh-1 dye in its open form in different solvent without addition of any acid. It can
be seen that it preferred to remain in its open form in MeOH, acetonitrile, acetone and
chloroform while in its spirocyclic form in DMF and as mixture of spirocyclic and open form in
toluene and hexane. As per their observed absorbance values (if absorbance 1 is considered as
100 % conversion) in MeOH the dye preferred to remain in its open form with 75.15 %
conversion and in spirocyclic form with 16.42 % which is the minimum value than any other
solvent in its spirocyclic form. Similarly in DMF it preferred to remain in its spirocyclic form
with 47.96 % conversion which is the highest value as compared to any other solvent and with
10.56 % it remains in its open form which is the lowest value for the dye in its open form. It is
well-known that the rhodamine derivatives are stable in their spirocyclic form in neutral or basic
conditions and in their open form in acidic conditions. We correlated these experimental finding

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with the solvent acidity and basicity parameters proposed by Catalan [60] as well as their
dielectric constants as represented in Table 4. As expected in methanol, a solvent with highest
acidity (0.605) the dye Cou-Rh-1 preferred to remain mostly in its open form (75.15 %) and in
DMF, a solvent with highest basicity (0.613) it preferred to remain mostly in spirocyclic form
(47.96 %). Similar correlation is observed in the case of other solvents such as chloroform,
toluene and hexane. In acetone and acetonitrile interestingly the dye preferred to remain in open
form though these solvents are very negligible acidic (0.000, 0.044) and highly basic (0.475,
0.286) in nature, may be due to their very high dielectric constants (20.7 and 37.5 respectively).
<< Fig. 8. Absorption spectra of Cou-Rh-1 in different solvents in its open form in absence of
TFA. >>
<< Please insert Table 4. Variation of absorbance/percentage of dye Cou-Rh-1 with the
variation of acidity and basicity of different solvents. >>
3.4 Interconversion of spirocyclic to open form of Cou-Rh-1 and Cou-Rh-2 dyes with the
addition of TFA
Due to their unique structural properties rhodamine dyes are interconvertible into their
spirocyclic and open form in the presence of acid or base. Hence we studied the interconversion
mechanism of these coumarin rhodamine hybrid dyes from their spirocyclic to open form with
the addition of trifluoroacetic acid in toluene (Fig. 9 and S5). We used 10 uM solutions of Cou-
Rh-1 and Cou-Rh-2 dyes in their spirocyclic form in toluene solvent and tested their absorption
and emission spectra with the increased percentage of TFA in toluene. For both these dyes we

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observed decreased absorbance of their spirocyclic form at 382 nm and 375 nm with
simultaneous increased absorbance of their open form at 605 nm and 609 nm for Cou-Rh-1 and
Cou-Rh-2 dyes respectively. Similarly with the increased percentage of TFA in toluene, we
observed decreased emission intensity of their spirocyclic form at 450 and 466 nm with
simultaneous increased emission intensity of their open form at 653 and 648 nm when excited at
605 and 609 nm respectively. The respective absorption and emission spectra of Cou-Rh-1 and
Cou-Rh-2 dyes with the increased percentage of TFA in toluene are represented in Fig. 9 and S5
respectively. For both Cou-Rh-1 and Cou-Rh-2 dyes in their spirocyclic form with the increased
percentage of TFA slightly red shift in their absorption λmax (10 and 4 nm) as well as emission
λmax (35 and 23 nm) is observed respectively.
<< Please insert Fig. 9. Absorption and emission spectra of Cou-Rh-1 with the increased
percentage of TFA in toluene. >>
3.5 Effect of viscosity on absorption and emission of hybrid dyes Cou-Rh-1 and Cou-Rh-2
We studied the effect of viscosity and polarity of solvents on absorption as well as emission
spectra of rhodamine dyes Cou-Rh-1 and Cou-Rh-2 in their spirocyclic form both in absence
and presence of TFA, so that their viscosity sensitivity can be tested in both visible as well as
deep red region. For this purpose we choose three different mixture of solvents i.e. polar protic
[EtOH-PEG 400], polar aprotic [toluene-PEG 400] and non-polar aprotic [toluene-paraffin].
(1) Mixture of polar-protic solvents [EtOH-PEG 400]
For Cou-Rh-1 dye very high enhancement in emission intensity was observed as the percentage
of PEG was increased in ethanol from 0 to 98 %, both in presence as well as absence of TFA

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(10.14 and 6.41 fold respectively) (Fig. 10). At the same time very small variation in absorbance
is observed with the increased percentage of PEG in ethanol both in presence as well as absence
of TFA (Fig. S6). In the presence of TFA, new absorption peak at 600 nm for its open form is
observed and in the absence of TFA, absorbance was decreased after the addition PEG in ethanol
(Fig. S6). Interestingly in the presence of TFA, though the open form absorbance decreased after
the addition of PEG in ethanol, the spirocyclic form absorbance is increased. Also the emission
enhancement in the presence of TFA is higher as compared to that in the absence of TFA and no
any enhancement of emission intensity of the open form emission peak is observed in the
presence of TFA. As the spirocyclic form absorption and emission λmax mostly deals with the
coumarin part, the carbonyl oxygen atom of coumarin may have protonated to initiate more
effective donor-acceptor charge transfer in the presence of TFA.
<< Please insert Fig. 10. Emission spectra of Cou-Rh-1 in EtOH: PEG mixture in presence (left)
and absence (right) of TFA. >>
In the case of Cou-Rh-2 dye also we studied the change in absorption as well as emission
intensity with the increased percentage of PEG in ethanol in the presence or absence of TFA. If
we see the absorption spectra of this dye in the presence and absence of TFA with the increased
percentage of PEG in ethanol, same scenario as in Cou-Rh-1 dye is observed (Fig. S7). In the
emission spectra, the enhancement in emission intensity was observed with the increased
percentage of PEG in ethanol both in presence and absence of TFA, but opposite to the dye Cou-
Rh-1, it shows enhancement of emission intensity of the open form emission peak in the deep
red region in the presence as well as absence of TFA. It indicates that this dye is comparatively
more stable in its open form and the spirocyclic form gets easily converted into the open form

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even in the presence of protic solvents such as ethanol. Both in the presence and absence of TFA
almost same amount of enhancement in emission intensity was observed (4.02 and 3.98 fold
respectively) (Fig. 11). As this dye shows good enhancement of emission intensity in its open
form at above 600 nm, it can be utilized as viscosity sensor in the deep red region.
<< Please insert Fig. 11. Emission spectra of Cou-Rh-2 in EtOH: PEG mixture in presence (left)
and absence (right) of TFA. >>
(2) Mixture of polar-aprotic solvents [toluene-PEG 400]
In toluene-PEG mixture also as like ethanol-PEG mixture the absorption spectra of Cou-Rh-1
and Cou-Rh-2 dyes shows decrease in absorbance of its open form and increase in absorbance of
spirocyclic form in the presence of TFA and decrease in absorbance of spirocyclic form in the
absence of TFA (Fig. S8 and S10). In their emission spectra, Cou-Rh-1 dye again found more
stable in its spirocyclic form as compared to the Cou-Rh-2 dye and shows emission peak only
for its spirocyclic form. In the absence of TFA, for both Cou-Rh-1 and Cou-Rh-2 dyes
quenching of emission intensity of its spirocyclic emission peak with the increased percentage of
PEG in toluene was observed (Fig. S9 and S11). In the presence of TFA, both Cou-Rh-1 and
Cou-Rh-2 dyes shows sudden increase in emission intensity after the addition of 30 % PEG in
ethanol, but after that no much enhancement in emission intensity is observed with the increased
percentage of PEG in toluene. Overall no any good emission enhancement was observed in this
mixture of solvents as compared to the ethanol-PEG mixture.
(3) Mixture of non-polar-aprotic solvents [toluene-paraffin]

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We also studied the absorption and emission spectra of Cou-Rh-1 and Cou-Rh-2 dyes in non-
polar aprotic mixture of solvents i.e. toluene-paraffin in the presence of TFA. A very good
enhancement in emission intensity as like EtOH-PEG mixture is observed for both the dyes,
where dye Cou-Rh-1 again showed emission enhancement in its spirocyclic form only and dye
Cou-Rh-2 showed emission enhancement in both spirocyclic as well as open form (Fig. 12 and
S12). For their spirocyclic form emission peak, comparatively higher emission enhancement of
dye Cou-Rh-1 (4.21 fold) is observed than dye Cou-Rh-2 (2.18 fold). Dye Cou-Rh-2 showed
emission enhancement in both spirocyclic as well as open form. It showed emission
enhancement of 3.56 fold for its open form at 650 nm and hence can be utilized as deep red
emitting viscosity sensor. For this dye, when up to 90 % of paraffin in toluene was added highest
emission intensity for the open form emission peak is observed, but after that i.e. up to 98 % of
paraffin in toluene, spirocyclic form showed highest emission enhancement. In contrast to the
above two solvent mixtures studied, in the toluene-paraffin mixture, absorbance of both
spirocyclic and open form of both Cou-Rh-1 and Cou-Rh-2 dyes was decreased with the
increasing percentage of paraffin in ethanol.
<< Please insert Fig. 12. Absorption (left) and emission (right) spectra of Cou-Rh-1 in toluene:
paraffin mixture in presence of TFA >>
In short, highest emission enhancement in EtOH-PEG mixture than toluene-paraffin mixture was
observed. Hence it can be predicted that in the case of EtOH-PEG mixture both polarity as well
as viscosity factors are responsible while in the case of toluene-paraffin mixture only viscosity
factor is responsible for the emission enhancement and the toluene-PEG mixture is totally
rejected as very low emission enhancement was observed in this mixture of solvents. Overall,

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comparatively higher emission enhancement for Cou-Rh-1 dye than Cou-Rh-2 dye was
observed, but the Cou-Rh-1 dye can be used as viscosity sensor only in the visible region, while
Cou-Rh-2 dye can be used as viscosity sensor in both visible as well as deep red region.
3.6
Fluorescence quantum yield (Фf) and life time measurements
Absolute quantum yield of Cou-Rh-1 and Cou-Rh-2 dyes are calculated in toluene solvent in
the presence and absence of TFA. Absolute quantum yield and fluorescence lifetime values are
presented in Table 5. As we calculated the absolute quantum yield of their spirocyclic form, we
found higher values in the absence of TFA which are decreased after the addition of TFA,
because some amount of dye get converted into their open form also. In their spirocyclic form,
dye Cou-Rh-1 (15.1 %) shows higher quantum yield than Cou-Rh-2 (5.8 %). At the same time,
the fluorescence lifetime is higher for Cou-Rh-2 dye (4.97 ns) than Cou-Rh-1 dye (2.07 ns).
Fluorescence Lifetime bi-exponential curves indicate the existence of two species at the
equilibrium may be due to the mixture of their spirocyclic and open form (Fig. 13). We also
studied the lifetime measurement of Cou-Rh-1 dye in different viscosity mixture of solvents and
it is found that as the ratio of PEG 400 increases i.e. viscosity increases lifetime is also increases
(Table 6). The lifetime measured in toluene solvent was 0.85 ns and it was increased up to 1.93
ns for toluene-PEG (2:98) mixture.
<< Please insert Table 5. Fluorescence quantum yield and life time measurements of dyes Cou-
Rh-1 and Cou-Rh-2 >>
<< Please insert Table 6. Lifetime measurement of Cou-Rh-1 dye in different composition of
PEG 400 in toluene >>

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<< Please insert Fig. 13 Lifetime measurement spectra of Cou-Rh-1 (a) and Cou-Rh-2 (b) in
THF and toluene respectively. >>
3.7 Photo stability study of Cou-Rh-1 for its spirocyclic and open form
Photo stability study of dye Cou-Rh-1 both for its spirocyclic as well as open form was carried
out at various time intervals under UV light. For this purpose their 5 µM solutions in toluene
solvent were irradiated at 254 nm up to 150 min. The absorption and emission spectra of this
dye, both in spirocyclic as well as open form were recorded at each 30 min. time interval. Fig. 14
represents the absorption and emission spectra of Cou-Rh-1 dye for its spirocyclic form at each
30 min time interval, while Fig. S13 represents the absorption and emission spectra of this dye
for its open form at each 30 min. time interval. Emission spectra of Cou-Rh-1 were recorded by
exciting at 375 nm for its spirocyclic form and at 588 nm for its open form. We found this dye
highly photo stable both in its spirocyclic as well as open form under UV light, as no remarkable
change in its absorbance as well as emission intensity for both the forms was observed.
<<Please insert Fig. 14. Absorption (a) and emission (b) spectra of dye Cou-Rh-1 (spirocyclic
form) after the irradiation of UV light (254 nm) at different time intervals in toluene solvent (λex :
375 nm). >>
3.8 Optimized structures of Cou-Rh-1 and Cou-Rh-2 dyes in their spirocyclic and open
form by DFT method
To correlate our experimental finding theoretically we optimized the structures of Cou-Rh-1 and
Cou-Rh-2 dyes in their spirocyclic and open form using Density Functional Theory (DFT)[51]
with Gaussian 09 package and the popular hybrid functional B3LYP. The basis set used was 6-

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31G (d) in both DFT and time-dependent density functional theory (TD-DFT) method. The
computationally derived vertical excitation energies and oscillator strengths of these dyes were
obtained for the lowest 10 singlet-singlet transitions using TD-DFT method at the same hybrid
functional and basis set [54]. Fig. 15 represents the frontier molecular orbital profile of Cou-Rh-
1 and Cou-Rh-2 dyes in their spirocyclic and open form displaying electronic distributions in
their HOMO and LUMO energy levels in toluene and chloroform solvent respectively.
<< Please insert Fig. 15. HOMO-LUMO FMO diagrams of Cou-Rh-1 and Cou-Rh-2 dyes in
their spirocyclic and open form in toluene and chloroform solvent respectively at ground state.
>>
In their spirocyclic form, for Cou-Rh-1 dye electronic density at HOMO level is situated on
coumarin side triphenylamine donor while for Cou-Rh-2 dye it is situated on rhodamine side
julolidine donor highlighting different donating ability of N,N diethyl aniline and julolidine
donor. At LUMO level for both the dyes in their spirocyclic form most of the electron density is
situated on lactone ring of rhodamine and some part is spread towards coumarin core. In their
open form at HOMO level for both the dyes the electron density is situated at central coumarin
fused xanthene core while at LUMO level it is mostly situated on carboxyl group substituted
phenyl ring of rhodamine and no any overlap of electron densities between HOMO and LUMO
energy level is observed. As complete charge transfer is observed for these dyes in their open
form as compared to the partial charge transfer observed in their spirocyclic form, good
interaction between the donor and acceptor is expected in the open form of these dyes.
We also calculated the energy difference between HOMO and LUMO energy levels for both
Cou-Rh-1 and Cou-Rh-2 dyes in their both spirocyclic as well as open form. In general lower

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energy difference is observed in their open form (2.37 and 2.14 eV) as compared to their
respective spirocyclic form (3.67 and 3.58 eV). Also in between Cou-Rh-1 and Cou-Rh-2, the
dye with better donating group i.e. Cou-Rh-2 shows comparatively lower energy difference
between HOMO and LUMO than Cou-Rh-1. When we go from spirocyclic to open form,
energies of HOMO level are increased rather than decreasing the energies of LUMO level. The
HOMO-LUMO energy diagrams of Cou-Rh-1 and Cou-Rh-2 in their spirocyclic and open form
is represented in Fig. 16. We compared the experimental absorption λmax values with the
computationally derived vertical excitations and a minimum deviation between them in their
spirocyclic as well as open form is observed (Table 7). We also compared the experimentally
calculated and computationally derived oscillator strength (f) values and the computationally
derived values are lower than the respective experimental one in both spirocyclic as well as open
form.
<< Please insert Fig. 16. HOMO-LUMO energy diagrams of Cou-Rh-1 and Cou-Rh-2 dyes in
their spirocyclic and open form chloroform solvent. >>
<< Please insert Table 7. Comparative experimental and computational photophysical
parameters of Cou-Rh-1 and Cou-Rh-2 dyes in their spirocyclic (toluene) and open form
(chloroform). >>
4
Conclusion
By using H-NMP.HSO4 as novel reaction media for the synthesis of rhodamine core we
synthesized two novel triphenylamine based coumarin-rhodamine hybrid derivatives. A detail
photophysical study of these dyes in their spirocyclic and open form is presented in a
comparative approach. We found red shifted absorption and emission peaks for these dyes in

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their open form than the reported RH-B and RH-101 dyes with comparatively higher Stokes
shift also. We calculated the presence of spirocyclic and open form of Cou-Rh-1 dye in different
solvents and are correlated with the acidity or basicity as well as dielectric constant of the
respective solvent. The ring opening mechanism of both the dyes from spirocyclic to open form
is also presented with the addition of TFA in toluene. Viscosity sensitivity of these dyes was
studied in different mixture of solvents and both the dyes are found to show very good emission
enhancement in protic-polar mixture of solvents (EtOH-PEG) after increasing the ratio of
polyethylene glycol (PEG) in ethanol. Similar enhancement in emission intensity in aprotic-non-
polar mixture of solvent (toluene-paraffin) was also observed but the former was showing better
results. Hence it can be concluded that when polarity and viscosity works together better results
were obtained than when only viscosity works independently. As viscosity sensitivity of these
dyes was studied in both spirocyclic and open form they can be used as viscosity sensors in the
visible as well as deep red region. We also found better support to our experimental observations
with the computationally obtained results.
Acknowledgements
Shantaram Kothavale and Amol Jadhav are grateful to UGC for research support under UGC-
SRF fellowship. We are also thankful to the Sophisticated Analytical Instrument Facility (SAIF),
IIT, Mumbai, India for the recording of mass and HR-MS spectra.
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