Fabrication of deoxycholic acid tethered α-cyanostilbenes as smart low molecular weight gelators and AIEE probes for bio-imaging

Article abstract of DOI:10.1016/j.steroids.2020.108659

Four novel deoxycholic acid tethered α-cyanostilbenes were designed, synthesized and characterized using detailed spectroscopic analysis. The synthesized deoxycholic acid tethered α-cyanostilbene derivatives formed stable gels with a variety of solvents,

Full text of DOI:10.1016/j.steroids.2020.108659

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Fabrication of Deoxycholic Acid tethered α-Cyanostilbenes as Smart Low
Molecular Weight Gelators and AIEE probes for Bio-imaging
Devesh S. Agarwal, Rajnish Prakash, Prabhat N. Jhad, Rajeev Sakhuja
PII:
S0039-128X(20)30084-2
DOI:
Reference:
https://doi.org/10.1016/j.steroids.2020.108659
STE 108659
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Steroids
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Revised Date:
Accepted Date:
11 October 2019
22 April 2020
14 May 2020
Please cite this article as: Agarwal, D.S., Prakash, R., Jhad, P.N., Sakhuja, R., Fabrication of Deoxycholic Acid
tethered α-Cyanostilbenes as Smart Low Molecular Weight Gelators and AIEE probes for Bio-imaging, Steroids
(2020), doi: https://doi.org/10.1016/j.steroids.2020.108659
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Fabrication of Deoxycholic Acid tethered α-Cyanostilbenes as Smart Low Molecular
Weight Gelators and AIEE probes for Bio-imaging
Devesh S. Agarwal,a Rajnish Prakash,b Prabhat N. Jhab and Rajeev Sakhuja *a
aDepartment of Chemistry, Birla Institute of Technology and Science, Pilani 333 031, Rajasthan, India
bDepartment of Biological Sciences, Birla Institute of Technology and Science, Pilani 333 031,
Rajasthan, India
O
OH
CN
HN
n
O
O
HO
n
O
N
H
Gelation Study of the
Synthesized Conjugates
O
H
O
H
N
N
H
N
N
O
H
O
O
H
N
O
H
N
N
H
N
O
H
O
Proposed Mechanism of Self-Assembly Process
Application of AIEE in bioimaging
AIEE Study of the
Synthesized Conjugates
Deoxycholic Acid tethered -Cyanostilbene as OrganoGelator and AIEE probe

Abstract
Four novel deoxycholic acid tethered α-cyanostilbenes were designed, synthesized and characterized
using detailed spectroscopic analysis. The synthesized deoxycholic acid tethered α-cyanostilbene
derivatives formed stable gels with a variety of solvents, such as xylene, toluene, mesitylene, decane,
dodecane etc. The stable gels showed lamellar sheet type structures stacked over each other, consisting of
entangled fibres as evident from SEM, TEM and Fluorescence Microscopy images; The synthesized
compounds exhibited AIEE behaviour in H2O/THF mixture, with the maximum emission observed in
70% H2O/THF fraction along with a bathochromic shift. A solvent thickening experiment was perform to
establish the mechanism of AIEE and the AIEE property was explored for bacterial bio-imaging. The
synthesized derivatized steroids proved their potential as multifunctional organic materials.
Keywords: Bile acids; Gelation; AIEE; Bio-imaging

Introduction
Stilbenes have attracted considerable interest in the past two decades towards investigating their
photophysical, photochemical and biological applications.[1-3] Affixing functionalities such as
alkyl chain, aromatic/heteroaromatic moieties, donor/acceptor groups and steroidal architectures
significantly influence their emission properties in solid and solution states.[4-6] Such
functionalized stilbenes have exhibited wide range of applications as self-assembling systems,[7-9]
fluorescent probes,[10] liquid crystals,[11] and optoelectronics.[12] Further, the steric effects in α-
cyanostilbene plays an important role in the enhancement of emission in aggregation state, while
the electronic effects offer great color tunability of AIEE luminogens to widen their emission
spectrum.[12-17] Thus, α-cyanostilbenes have been widely studied for their AIEE behaviour,[18-22]
and explored for various applications such as bio-imaging,[23,24] analyte detection,[13, 25-27]
explosive and biomolecule detection.[28,29]
Strikingly, a few conjugates containing a steroidal moiety appended to α-cyanostilbene have been
explored for AIEE and gelation behavior. For example, Kanvah and co-workers studied the AIEE
behaviour of the cholesterol appended α-cyanostilbene/diene hybrids (Fig. 1, I-II) that exhibited
weak emission in organic solvent, however, the emission increased 5-fold accompanied by a
strong bathochromic emission shift in aqueous media. In addition, the hybrids exhibited
intermolecular charge transfer (ICT) phenomena under aqueous condition in sodium cholate
matrix, and the compounds self-assembled to form organized nanostructures having distinct
morphologies.[2] Likewise, Zhao and co-workers reported cholesterol appended α-cyanostilbene
(Fig. 1, II) to be non-fluorescent in assembled state, and found that the emission of free molecules
was greatly enhanced upon UV irradiation, which was attributed to the conformational change
due to the photo-isomerization of the α-cyanostilbene unit to form capped and branched
nanotubes in assembled state. This unique feature has been utilized to detect H2O2 with good
selectivity and linear response.[30]
O
N
N
O
O
O
O
O
CN
CN
II
I
NC
O
H
N
O
N
H
O
III
Fig. 1 Previous reported steroid appended α-cyanostilbenes as AIEE probes and organogelator

Jiang and co-workers studied the gelation behaviour of cholesterol appended α-cyanostilbene
analogue (Fig. 1, III) that formed gels in various aromatic solvents such as xylene, mesitylene,
bromobenzene, 2-chlorobenzene, chlorobenzene, 1,2-dichlorobenzene with a CGC of 8.0-15.0
mg/mL. The gels were efficiently used for phase selective gelation of aromatic solvents and
absorption of rhodamine dye from water.[31,32] In striking analogy to cholesterol, bile acids are
another interesting architectures proven for the self-assembly process, which have been explored
towards the formation of organogels and hydrogels with varied applications.[33-36] The presence of
hydroxyl groups and a carboxylic acid group on the concave hydrophilic face and methyl groups
on the convex hydrophobic face favours the formation of molecular umbrellas or pockets with
star-shaped oligomers of bile acids.[37-39] Unfortunately, the development of photo-responsive bile
acid appended α-cyanostilbene analogues as AIEE probes and organogelators remains
unexplored. In continuation to our interest on bile acid based smart materials, [40,41] we plan to
affix α-cyanostilbene to deoxycholic acid skeleton, and study their AIEE behaviour and gelation.
Results and Discussion
From the outset of the proposed work, the synthesis commenced with the preparation of 3,4-
bis(alkyloxy)benzaldehyde (3a-d) by O-alkylating 3,4-dihydroxybenzaldehyde (1) with
appropriate bromoalkanes (2a-d) in acetone using K2CO3 at 65 ºC for 12 h (Scheme 1).
Thereafter, 3a-d on Knoevenagel condensation with 2-(4-nitrophenyl)acetonitrile (4) using
piperidine in ethanol afforded nitro-substituted bis(alkyloxy) α-cyanostilbenes (5a-d) in 89-93%
yields (Scheme 1). 5a-d on reduction with stannous chloride in THF/water at 80 ºC furnished
their corresponding amino-substituted α-cyanostilbenes (6a-d) in 80-87% yield. Finally, 6a-d on
coupling with deoxycholic acid (7) using EDC.HCl/HOBt/DMAP in DCM afforded DCA
tethered bis(alkyloxy) α-cyanostilbene hybrids (8a-d) in 81-85% yields (Scheme 1). All the
intermediates and final compounds were characterized on the basis of 1H NMR, 13C NMR and IR
spectroscopy.
K2CO3
CN
Piperidine
O2N
EtOH, Reflux, 4 h
n
CN
OHC
OH
OH
Br
OHC
O
n
Acetone, 65 °C, 12 h
O2N
n
O
O
5a: n = 6, 92%
5b: n = 7, 90%
5c: n = 11, 89%
5d: n = 17, 93%
n
O
4
1
2a-d
3a: n = 6, 82%
3b: n = 7, 85%
3c: n = 11, 90%
3d: n = 17, 91%
n
SnCl2
THF/H2O, 80 °C, 4 h
O
OH
CN
Deoxycholic Acid
CN
HN
7
H2N
n
O
n
n
O
EDC.HCl/HOBt/NEt3
DCM, rt, 18-24 h
O
6a: n = 6, 87%
6b: n = 7, 87%
6c: n = 11,80%
6d: n = 17, 85%
HO
O
8a: n = 6, 84%
8b: n = 7, 82%
8c: n = 11,85%
8d: n = 17, 81%
n
Scheme 1. Systematic route for the synthesis of DCA tethered α-cyanostilbene hybrids (8a-d)

Gelation Studies
The synthesized compounds (8a-d) were studied for their gelation behaviour in a variety of
solvents using inverted test tube method. To our delight, the synthesized molecules gelate a wide
range of solvents including octane, dodecane, decane, tetradecane, hexadecane, toluene, benzene,
xylene, mesitylene, iodobenzene, fluorobenzene, decanol, octanol, dimethylformamide (DMF)
and dimethyl sulfoxide (DMSO) (Fig. 2: representative gels of 8b and 8c). The minimum gel
concentration for the compounds varied from 1% to 16% w/v (Table 1).
Fig. 2 Inverted vials showing stable gels of 8b (left) and 8c (right) in different solvents (left to
right: xylene, toluene, mesitylene, decane, dodecane, tetradecane, hexadecane)
Table 1. Gelation studies of 8a-d in different solvents
Solvent
Xylene
Toluene
8a
G (1.6)
G (1.2)
G (1.2)
G (1.3)
G (1.3)
G (1.3)
G (1.2)
WG (10)
WG (12)
G (8)
G (7)
G (7)
G (9)
G (7)
S
8b
G (1.4)
G (1.3)
G (1.3)
G (1.3)
G (1.2)
G (1.2)
G (1.3)
WG (10)
WG (11)
G (7)
G (8)
G (9)
G (9)
G (8)
S
8c
G (1)
G (1)
G (1.1)
WG (1.1)
WG (1.1)
WG (1.1)
WG (1.1)
PS
8d
G (1)
G (1.3)
G (1.2)
WG (1.4)
WG (1.4)
WG (1.4)
WG (1.2)
PS
Mesitylene
Iodobenzene
Fluorobenzene
p-Cymene
tert-Butylbenzene
Hexane
Heptane
Octane
Decane
Dodecane
PS
PS
PS
PS
G (5)
G (6)
G (4)
G (5)
S
S
I
PS
G (5)
G (4)
G (4)
G (6)
S
S
I
PS
Teteradecane
Hexadecane
Dichloromethane
Chloroform
1,2-Dichloroethane
Methanol
S
I
PS
S
I
PS
Ethanol
PS
PS
PS
PS
isopropanol
tert-Butanol
Decanol
PS
PS
PS
PS
PS
S
S
PS
PS
PS
PS
PS
PS
S
S
PS
PS
PS
WG (8)
WG (6)
WG (10)
PS
WG (8)
WG (9)
WG (9)
PS
Lauryl alcohol
Acetonitrile
Tetrahydrofuran
N,N-Dimethylformamide
Dimethyl sulfoxide
S
S
WG (16)
WG (15)
WG (14)
WG (13)
G = gel formed at room temperature [minimum gel concentration (% w/v)]; WG = weak gel (unstable
above 20 ºC); PS = partially soluble (a part of it becomes soluble upon heating but re-precipitation was
observed after cooling to room temperature); S = soluble; I = insoluble (was not soluble at all even on
heating)

To get an insight into the microscopic morphology of the stable gels, Scanning Electron
Microscopy (SEM) and Transmission Electron Microscopy (TEM) images of the xerogels of 8c in
benzene, toluene and xylene were obtained. The SEM images of the xerogels revealed lamellar
sheets stacked over each other, while the TEM images further indicated the presence of fibres
entangled into each other, forming sheets (Fig. 3).
Fig. 3 SEM images of xerogels of 8c in benzene, toluene and xylene (a-c), and TEM images of xerogels
of 8c in benzene, toluene and xylene (d-f)

Fluorescence optical microscopic images also revealed a fibrillar structure of the wet gel of 8c in
dodecane, while the wet gel of 8c in toluene revealed a long entangled fibrous network (Fig. 4).
Fig. 4 Fluorescence optical microscopic images of wet gel of 8c in dodecane (a-b) and toluene (c-d) in
red and green filters
Mechanism of self-assembly process
To get an insight into self-assembly process, FT-IR analysis was performed on the dilute solution
of 8c in DCM, and its xerogel in toluene and decane. The IR spectrum of dilute solution of 8c in
DCM showed prominent bands at 3418 cm-1 (O-H/N-H stretching), 2214 cm-1 (C≡N stretching),
1666 cm-1 (C=O stretching of amide bond) and 1597 cm-1 (N-H bending) (Fig. 5). In striking
contrast, two well-separated bands at 3448 and 3456 cm-1 were observed for the O-H/N-H
stretching in the IR spectrum of xerogel of 8c in toluene (Fig. 5). This indicated the involvement
of O-H bond(s) of deoxycholic acid and N-H of amide functionality in hydrogen bonding in the
gel state. A red shift in C=O absorption band of amide functionality from 1666 cm-1 to 1659 cm-1
further supported the above indication. Unfortunately, no change was observed in C≡N stretching
of xerogel of 8c in toluene, and its dilute solution in dichloromethane (DCM). Likewise, in IR
spectrum of xerogel of decane, the O-H/N-H overlapped bands were red-shifted to 3474 cm-1
,
which is in concordance with the above inference of their active contribution in H-bonding (Fig.
5). Contentedly, the two antisymmetric and symmetric stretching vibrations bands of CH2
appeared at 2924 and 2854 cm-1 in the IR spectrum of dilute solution of 8c in DCM, indicating the
possibility of the gauche conformation of the two appended O-alkyl chains, while in xerogel of 8c
in decane, the band shifted to 2862 cm-1 after gelation (Fig. 5). This shift probably suggests
indicated the transformation of gauche to trans conformation of the O-alkyl chains to a well-
ordered arrangement, restricting their mobility in gel state.

Fig. 5 Comparative FT-IR spectra of 8c in solution and gel state
It has been well documented that α-cyanostilbene undergo E- and Z- photoisomerization in solution
state.[30] To comprehend the impact of affixing deoxycholic acid on the envisioned E- and Z-
photoisomerization of α-cyanostilbene in 8, UV-Visible studies were performed on a sample of 8c in
toluene after irradiating it with UV light (λ = 365 nm) for requisite intervals of time (Fig. 6). As evident
from its UV-Visible absorbance spectra, the absorbance band at 365 nm gradually decreased in intensity
as well as hypsochromically shifted to 350 nm, with an increase in the exposure time from 0 to 128
minutes. Subsequently, the band at 264 nm gradually increased with a bathochromic shift to 267 nm. The
decrease and the increase in the intensity/wavelength of the bands at 365 nm and 264 nm showed an
exponential dependence on the irradiation time, which established the phenomena of E- and Z-
photoisomerization in the synthesized conjugate in solution phase (Fig. 6).
Fig. 6 a) UV-Visible absorption spectra of 8c after irradiating its dilute solution in toluene with UV-light
(λ = 365 nm) at different intervals of time; b) Graph of UV-Visible absorbance bands at 264 nm and 365
nm with irradiation time to indicate the cross-over point

To reveal the aggregation pattern of gelator molecules in the gel phase, X-Ray Diffraction (XRD)
analysis of one of the xerogel of 8c (in toluene) was recorded (Fig. 7). The XRD diffractogram showed a
series of intense reflections at 5.44°, 7.42°, 13.28°, 14.78°, 17.18°, 18.74° and 21.82° in the 2θ = 10-30
region, accounting for the corresponding d spacing of 16.2 Å, 11.9 Å, 6.7 Å, 6.0 Å, 5.2 Å, 4.7 Å, 4.2 Å
and 3.6 Å, respectively. Among these, the reflection at 3.6 Å can be accounted for the distance between
two π-stacked stilbene units in the self-assembled state, while the reflections at 4.7 Å and 4.2 Å accounts
for the presence of hydrogen bonded β sheet-like arrangement of the gelator molecules in the assembled
gel state.[42-45]
Fig. 7 XRD diffractogram of the xerogel of 8c in toluene
Based on the FT-IR and XRD studies, it may be concluded that O-H groups of deoxycholic acid
play eminent role in strong intermolecular hydrogen bonding, favoring face-to-face amphiphilic
packing of deoxycholic acid units leading to elongated fibres. In addition, Z-configuration of α-
cyanostilbene further provides an additive effect for the
stacking between two encountered
휋→휋
aromatic clouds. Further, the hydrogen bonding between N-H and C=O groups of the amide
bonds between two parallelly-elongated fibrous units may lead to their stacking, resulting in
three-dimensional network of lamellar stacked sheets as evident from SEM and TEM images
(Fig. 8).[33, 46-49]

O
N
CN
O
H
H
O
H
O
O
N
H
O
H
O
H
O
H
N
O
N
H
N
N
N
H
N
N
O
N
H
O
H
O
H
O
FORMATION OF FIBERS DRIVEN BY O-H....O HYDROGEN BONDING AND  STACKING
O
H
O
H
O
H
N
O
N
H
N
N
N
H
N
N
O
N
H
O
H
O
H
O
O
H
O
H
O
H
N
O
N
H
N
N
N
H
N
N
O
N
H
O
H
O
H
O
FORMATION OF LAMELLAR SHEET DRIVEN BY C=O....H-N HYDROGEN BONDING
Fig. 8 Plausible mode of aggregation in the self-assembly process
Aggregation-induced emission Enhancement (AIEE) study
α-Cyanostilbenes are known to exhibit enhanced emission in the solid state due to AIEE. Thus,
we evaluated the AIEE property in THF/water mixture. In case of 8a, a significant increase in the
photoluminescence was observed on increasing the water fraction from 0% to 70%, however
thereafter a slow decrease with further increase in the water concentration was observed. The
emission intensity reached a maximum in 70% water/THF fraction, with a bathochromic shift in
the wavelength of 37 nm (i.e. from 433 nm to 470 nm) (Fig. 9a). Similar results were observed for
8b, where a bathochromic shift of 27 nm was evident from 433 nm to 460 nm (Fig. 9b). Likewise,
in case of 8c and 8d, bathochromic shifts of 35 nm and 34 nm, and significant increase in the
photoluminescence intensity were observed on increasing the water fraction from 0% to 70%,
respectively (Fig. 9c-d). Such drastic shifts in the emission could be attributed to the formation of
aggregated species that results in the confinement of the molecular motion. This structural rigidity
further results in enhanced radiative transitions.

Fig. 9 Photoluminescence intensity spectra of 24a-d in varying water/THF fraction (a-d) (Inset: Picture
showing the enhancement in PL intensity in increasing the water fraction from 0 to 90% from left to
right)
To further verify the enhancement emission is due to aggregation, solid state emission spectra of
synthesized compounds (8a-d) were recorded (Fig. 10). The bands in the emission spectra of 8a-d in solid
state were narrower and of high intensity as compared to that in 70% water/THF fraction, as elucidated
from their full width half maxima calculations (Table 2). This could be due to reduced intermolecular
molecular rotations and packing efficiency as earlier documented by others.
Fig. 10 Solid photoluminescence spectra of 8a-d

Table 2 Comparison of emission bands of 8a-d in 70% water/THF fraction and solid state
Compound
Emission in 70% water/THF state (nm)
Emission in solid state (nm)
8a
8b
8c
8d
470
460
468
467
490
490
498
498
To further support the AIEE, solution thickening experiment was carried out for 8c as a
representative example using PEG/THF mixture, as PEG is favorable for increasing the viscosity
of the mixture. Interestingly, on increasing the amount of PEG from 0 to 90%, an enhancement in
the emission band was observed, and the maximum emission was observed in 90% PEG fraction
(Fig. 11). Clearly, the emission intensity significantly enhanced on increasing the viscosity of the
solvent mixture probably due to restriction of intermolecular rotation (RIR), thus giving an
evidence for the AIEE property of the compound.
Fig. 11 a) Photoluminescence intensity graph of 8c in varying PEG/THF fraction (Inset: Picture showing
the enhancement in PL intensity in increasing the PEG fraction from 0 to 90% from left to right.); b)
Graph of photoluminescence intensity (λEx. = 365 nm) and PEG fraction
Application of AIEE property
In order to explore their application in the imaging of living cells, the synthesized compounds
(8a-d) were evaluated for their ability to stain the broad spectrum (gram +ve & gram –ve)
bacterial cells (Fig. 12). It became evident that the treatment of bacterial cells showed strong
fluorescence behavior. The staining ability of the compounds could be due to their ability to
penetrate the bacterial cell membrane. Using a Fluorescence Microscope, it was obvious that the
bacteria displayed bright fluorescence in the green channel (530–590 nm). Therefore, the resent
observation clearly supports the ability of the compounds as fluorescent probes for staining the
broad spectrum bacteria.

Fig. 12 Fluorescence images of gram +ve (S. Aureus) and gram –ve (Serratia) with compounds 24a-d
images in the green channel. The scale bar is 10 µM
Conclusion
In summary, we have successfully synthesized deoxycholic acid tethered α-cyanostilbene derivatives as
multifunctional materials. All the synthesized compounds were characterized using detailed spectroscopic
analysis. The synthesized deoxycholic acid tethered α-cyanostilbene derivatives formed stable gels with a
variety of solvents, such as xylene, toluene, mesitylene, decane, dodecane etc. The morphology of the
formed gels were in detailed characterized using SEM, TEM and Fluorescence Microscopy that showed
lamellar sheet type structures stacked over each other, consisting of entangled fibres. The synthesized
compounds also exhibited AIEE behaviour in Water/THF mixture. The maximum emission was observed
in 70% water/THF fraction with a bathochromic shift, and a solvent thickening experiment was carried
out to establish the mechanism of AIEE. The AIEE property of the hybrids was finally used for bacterial
bio-imaging, which established the multifunctional behavior of the synthesized organic materials.
Experimental
General materials and methods
All the chemicals were purchased from Sigma-Aldrich, Alfa Aesar, and Spectrochem India Pvt.
Ltd and used without further purification. The solvents used were purchased from Merck (India)
and were distilled and dried before use. Nuclear magnetic resonance spectra were recorded on
1
Bruker 400 spectrometer. The H NMR experiments were reported in δ units, parts per million
(ppm), and were measured relative to residual chloroform (7.26 ppm) or DMSO (2.5 ppm) in the
13
deuterated solvent. The C NMR spectra were reported in ppm relative to deuterochloroform
(77.0 ppm) or DMSO-d6 (39.5 ppm). All coupling constants J were reported in Hz. The following
abbreviations were used to describe peak splitting patterns when appropriate: s = singlet, d =
doublet, t = triplet, dd = doublet of doublet, m = multiplet and brs = broad singlet. Melting points
were determined on a capillary point apparatus equipped with a digital thermometer and are

uncorrected. Reactions were monitored by using thin layer chromatography (TLC) on 0.2 mm
silica gel F254 plates (Merck). High resolution mass spectra were recorded on an Agilent
Technologies 6545 Q-TOF LC/MS by using electrospray mode.
General procedure for the synthesis of 3,4-bis(alkyloxy)benzaldehydes 4a-d
To a stirred solution of 3,4-dihydroxybenzaldehyde (1) (1 mmol) in acetone (20 mL), K2CO3 (5
mmol) and appropriate bromo-alkane (2a-d) was added. The reaction mixture was refluxed for 12
h and the completion of the reaction was monitored by TLC. After completion, the solvent was
removed under reduced pressure, and subsequently water (100 mL) was added, and extracted
using ethyl acetate (100 mL x 2). The organic layer was dried over sodium sulphate and
concentrated under reduced to give crude product (3a-d), which was used for next step without
further purification.
General procedure for the synthesis of nitro-substituted bis(alkyloxy) α-cyanostilbenes 5a-d
To a stirred solution of 2-(4-nitrophenyl)acetonitrile (4) (1 mmol) in hot ethanol (20 mL), 3,4-
bis(alkyloxy)benzaldehyde (3a-d) (1 mmol) was added. After the reaction mixture became a clear
solution, a catalytic amount of piperidine (4 drops) was added, and the reaction turned dark red.
The reaction mixture was refluxed for 4 h and the completion of the reaction was monitored by
TLC. After the completion, the resultant precipitate was filtered to obtain crude product, which
was recrystallized using ethanol to obtain pure 5a-d.
(Z)-3-(3,4-Bis(heptyloxy)phenyl)-2-(4-nitrophenyl)acrylonitrile (5a): Yellow solid; yield: 92%
(2.62 g); mp: 97-98 C; 1H NMR (400 MHz, CDCl3) δ 8.31 (d, J = 8.9 Hz, 2H, HAr), 7.84 (d, J =
8.9 Hz, 2H, HAr), 7.75 (d, J = 2.1 Hz, 1H, HAr), 7.60 (s, 1H), 7.43 (dd, J = 8.4, 2.0 Hz, 1H, HAr),
6.96 (d, J = 8.5 Hz, 1H, HAr), 4.11 (dd, J = 11.3, 6.5 Hz, 4H, 2xO-CH2 Alkoxy), 1.93 – 1.85 (m, 4H,
2xO-β-CH2 Alkoxy), 1.56 – 1.47 (m, 4H, 2xCH2 Alkoxy), 1.44 – 1.28 (m, 12H, 2xCH2 Alkoxy), 0.96 –
13
0.85 (m, 6H, 2xMeAlkoxy); C NMR (100 MHz, CDCl3) δ 152.5, 149.1, 147.4, 145.5, 141.2,
126.3, 125.7, 125.5, 124.3, 118.0, 113.0, 112.5, 105.7, 69.3, 69.1, 31.8, 31.8, 29.1, 29.1, 29.1,
26.0, 25.9, 22.6, 14.1; IR (KBr, ῡ, cm-1) 2924, 2854, 2206, 1582, 1512, 1111, 849.
(Z)-3-(3,4-Bis(octyloxy)phenyl)-2-(4-nitrophenyl)acrylonitrile (5b): Yellow solid; yield: 90%
(2.51 g); mp: 99-100 C; 1H NMR (400 MHz, CDCl3) δ 8.32 (d, J = 8.9 Hz, 2H, HAr), 7.84 (d, J =
8.9 Hz, 2H, HAr), 7.74 (d, J = 1.9 Hz, 1H, HAr), 7.60 (s, 1H, =CH), 7.43 (dd, J = 8.4, 1.9 Hz, 1H,
HAr), 6.96 (d, J = 8.5 Hz, 1H, HAr), 4.11 (dd, J = 11.3, 6.5 Hz, 4H, 2xO-CH2 Alkoxy), 1.93 – 1.83
(m, 4H, 2xO-β-CH2 Alkoxy), 1.56 – 1.47 (m, 4H, 2xCH2 Alkoxy), 1.42 – 1.27 (m, 16H, 2xCH2 Alkoxy),
0.96 – 0.87 (m, 6H, 2xMeAlkoxy); 13C NMR (100 MHz, CDCl3); 152.5, 149.1, 147.4, 145.5, 141.3,
126.3, 125.7, 125.5, 124.4, 119.0, 113.1, 112.5, 105.7, 69.3, 69.1, 31.8, 29.4, 29.4, 29.3, 29.3,

29.1, 29.0, 26.0, 26.0, 22.7, 14.1; IR (KBr, ῡ, cm-1) 3479, 3387, 2913, 2847, 2206, 1605, 1520,
1142, 843.
(Z)-3-(3,4-Bis(dodecyloxy)phenyl)-2-(4-nitrophenyl)acrylonitrile (5c): Yellow solid; yield:
89% (2.32 g); mp: 104-105 C; 1H NMR (400 MHz, CDCl3) δ 8.32 (d, J = 8.9 Hz, 2H, HAr), 7.84
(d, J = 9.0 Hz, 2H, HAr), 7.74 (d, J = 2.1 Hz, 1H, HAr), 7.60 (s, 1H, =CH), 7.43 (dd, J = 8.5, 2.0
Hz, 1H, HAr), 6.96 (d, J = 8.5 Hz, 1H, HAr), 4.11 (dd, J = 11.3, 6.5 Hz, 4H, 2xO-CH2 Alkoxy), 1.93
– 1.84 (m, 4H, 2xO-β-CH2 Alkoxy), 1.55 – 1.47 (m, 4H, 2xCH2 Alkoxy), 1.40 – 1.24 (m, 32H, 2xCH2
13
Alkoxy), 0.95 – 0.86 (m, 6H, 2xMeAlkoxy); C NMR (100 MHz, CDCl3) δ 152.5, 149.1, 147.4,
145.5, 141.3, 126.3, 125.7, 125.5, 124.3, 118.0, 113.1, 112.5, 105.7, 69.3, 69.1, 32.0, 29.7, 29.7,
29.7, 29.7, 29.6, 29.4, 29.4, 29.1, 29.0, 26.0, 26.0, 22.7, 14.1; IR (KBr, ῡ, cm-1) 2916, 2847, 2206,
1582, 1512, 1111, 856.
(Z)-3-(3,4-bis(octadecyloxy)phenyl)-2-(4-nitrophenyl)acrylonitrile (5d): Yellow solid; yield:
93% (2.26 g); mp: 115-116 C; 1H NMR (400 MHz, CDCl3) δ 8.32 (d, J = 8.9 Hz, 2H, HAr), 7.84
(d, J = 8.8 Hz, 2H, HAr), 7.75 (d, J = 2.0 Hz, 1H, HAr), 7.60 (s, 1H, =CH), 7.43 (dd, J = 8.4, 2.0
Hz, 1H, HAr), 6.96 (d, J = 8.5 Hz, 1H, HAr), 4.11 (dd, J = 11.4, 6.4 Hz, 4H, 2xO-CH2 Alkoxy), 1.92
– 1.85 (m, 4H, 2xO-β-CH2 Alkoxy), 1.54 – 1.47 (m, 2xCH2 Alkoxy), 1.41 – 1.18 (s, 56H, 2xCH2
13
Alkoxy), 0.90 (t, J = 6.8 Hz, 6H, 2xMeAlkoxy); C NMR (100 MHz, CDCl3) δ 152.5, 149.1, 147.4,
145.5, 141.3, 126.3, 125.7, 125.5, 124.3, 118.0, 113.1, 112.5, 105.7, 69.3, 69.1, 32.0, 29.7, 29.7,
29.7, 29.6, 29.4, 29.4, 29.1, 29.0, 26.0, 26.0, 22.7, 14.2; IR (KBr, ῡ, cm-1) 2916, 2854, 2206,
1582, 1512, 1342, 1281, 1011, 849.
General procedure for the synthesis of amino-substituted α-cyanostilbenes 6a-d
To a stirred solution of 5a-d (1.89 mmol) in THF/H2O (20 mL : 5mL), SnCl2 (5.67 mmol) was
added and the reaction mixture was refluxed for 4 h. The completion of the reaction was
monitored by TLC. After the completion, the reaction mixture was cooled to 0 oC and the pH was
adjusted to 10 using saturated solution of sodium bicarbonate (30 mL). Subsequently, ethyl
acetate (20 mL) was added, and the reaction was filtered through celite and the organic layer was
separated. The organic layer was dried over sodium sulphate and concentrated under reduced to
give crude product, which was recrystallized using ethyl acetate/hexanes to give pure product 6a-
d.
(Z)-2-(4-Aminophenyl)-3-(3,4-bis(heptyloxy)phenyl)acrylonitrile (6a): Yellow solid; yield:
87% (1.63 g); mp: 113-114 C; 1H NMR (400 MHz, CDCl3) 1H NMR (400 MHz, CDCl3) δ 7.62
(d, J = 2.0 Hz, 1H, HAr), 7.48 (d, J = 8.6 Hz, 2H, HAr), 7.32 (dd, J = 8.4, 2.0 Hz, 1H, HAr), 7.29 (s,
1H), 6.91 (d, J = 8.4 Hz, 1H, HAr), 6.74 (d, J = 8.6 Hz, 2H, HAr), 4.14 – 4.03 (m, 4H, 2xO-CH2
Alkoxy), 3.88 (brs, 2H, NH2), 1.92 – 1.82 (m, 4H, 2xO-β-CH2 Alkoxy), 1.56 – 1.46 (m, 4H, 2xCH2

Alkoxy), 1.42 – 1.29 (m, 12H, 2xCH2 Alkoxy), 0.92 – 0.85 (m, 6H, 2xMeAlkoxy); 13C NMR (100 MHz,
CDCl3) δ 150.7, 148.9, 147.1, 138.8, 127.1, 127.0, 125.1, 123.5, 119.0, 115.1, 113.0, 112.8,
108.5, 69.2, 69.1, 31.8, 31.8, 29.2, 29.2, 29.1, 29.1, 26.0, 26.0, 22.6, 14.1; IR (KBr, ῡ, cm-1) 3479,
3385, 2910, 2214, 1605, 1524. 1142, 843.
(Z)-2-(4-Aminophenyl)-3-(3,4-bis(octyloxy)phenyl)acrylonitrile (6b): Yellow solid; yield: 87%
(1.63 g); mp: 119-120 C; 1H NMR (400 MHz, CDCl3) δ 7.62 (d, J = 2.0 Hz, 1H, HAr), 7.48 (d, J
= 8.6 Hz, 2H, HAr), 7.32 (dd, J = 8.5, 2.0 Hz, 1H, HAr), 7.29 (s, 1H, =CH), 6.91 (d, J = 8.4 Hz,
1H, HAr), 6.74 (d, J = 8.6 Hz, 2H, HAr), 4.12 – 4.05 (m, 4H, 2xO-CH2 Alkoxy), 3.88 (brs, 2H, NH2),
1.91 – 1.83 (m, 4H, 2xO-β-CH2 Alkoxy), 1.54 – 1.47 (m, 4H, 2xCH2 Alkoxy), 1.24 – 1.39 (m, 16H,
13
2xCH2 Alkoxy), 0.96 – 0.87 (m, 6H, 2xMeAlkoxy); C NMR (100 MHz, CDCl3) δ 150.7, 148.9,
147.1, 138.8, 127.1, 127.0, 125.1, 123.5, 119.0, 115.1, 113.0, 112.9, 108.5, 69.2, 69.1, 31.8, 29.4,
29.4, 29.3, 29.3, 29.2, 29.2, 26.1, 26.0, 22.7, 14.1; IR (KBr, ῡ, cm-1) 3479, 3387, 2924, 2854,
2214, 1584, 1520, 1142, 833.
(Z)-2-(4-Aminophenyl)-3-(3,4-bis(dodecyloxy)phenyl)acrylonitrile (6c): Yellow solid; yield:
80% (1.53 g); mp: 90-91 C; 1H NMR (400 MHz, CDCl3) 7.62 (d, J = 2.1 Hz, 1H, HAr), 7.48 (d, J
= 8.6 Hz, 2H, HAr), 7.32 (dd, J = 8.5, 2.2 Hz, 1H, HAr), 7.29 (s, 1H), 6.91 (d, J = 8.4 Hz, 1H, HAr),
6.74 (d, J = 8.6 Hz, 2H, HAr), 4.14 – 4.03 (m, 4H, 2xO-CH2 Alkoxy), 3.88 (brs, 2H, NH2), 1.92 –
1.82 (m, 4H, 2xO-β-CH2 Alkoxy), 1.56 – 1.46 (m, 4H, 2xCH2 Alkoxy), 1.42 – 1.29 (m, 32H, 2xCH2
13
Alkoxy), 0.92 – 0.85 (m, 6H, 2xMeAlkoxy); C NMR (100 MHz, CDCl3) δ 150.7, 148.9, 147.1,
138.8, 127.1, 127.0, 125.1, 123.5, 119.0, 115.1, 113.0, 112.9, 108.5, 69.2, 69.1, 32.0, 29.7, 29.7,
29.7, 29.7, 29.7, 29.4, 29.4, 29.2, 29.2, 26.1, 26.0, 22.7, 14.2; IR (KBr, ῡ, cm-1) 3476, 3387, 2913,
2847, 2206, 1605, 1520, 1143, 843.
(Z)-2-(4-Aminophenyl)-3-(3,4-bis(octadecyloxy)phenyl)acrylonitrile (6d): Yellow solid; yield:
85% (1.64 g); mp: 93-94 C; 1H NMR (400 MHz, CDCl3) δ 7.62 (d, J = 2.1 Hz, 1H, HAr), 7.48 (d,
J = 8.6 Hz, 2H, HAr), 7.32 (dd, J = 8.5, 2.0 Hz, 1H, HAr), 7.29 (s, 1H, =CH), 6.91 (d, J = 8.4 Hz,
1H, HAr), 6.74 (d, J = 8.7 Hz, 2H, HAr), 4.12 – 4.05 (m, 4H, 2xO-CH2 Alkoxy), 3.88 (brs, 2H, NH2),
1.90 – 1.83 (m, 4H, 2xO-β-CH2 Alkoxy), 1.56 –1.46 (m, 4H, 2xCH2 Alkoxy), 1.40 – 1.16 (m, 56H,
13
2xCH2 Alkoxy), 0.90 – 0.85 (m, 6H, 2xMeAlkoxy); C NMR (100 MHz, CDCl3) δ 150.7, 148.9,
147.1, 138.8, 127.1, 127.0, 125.1, 123.5, 119.0, 115.1, 113.0, 11.8, 108.5, 69.2, 69.1, 32.0, 29.7,
29.7, 29.7, 29.7, 29.5, 29.4, 29.4, 29.2, 29.1, 26.1, 26.01, 22.7, 14.2; IR (KBr, ῡ, cm-1) 3479,
3387, 2910, 2847, 2214, 1605, 1520, 1142, 833.
General procedure for the synthesis of deoxycholic acid tethered α-cyanostilbenes 8a-d

To the stirred solution of deoxycholic acid (7) (1.3 mmol) in DCM (20 mL), triethyl amine (3.2
o
mmol) was added at 0 C, and subsequently EDC.HCl (1.7 mmol) and HOBt (1.3 mmol) were
o
added. The reaction mixture was stirred for 15 min. at 0 C, after which, (Z)-2-(4-aminophenyl)-
3-(3,4-bis(alkoxy)phenyl)acrylonitrile (6a-d) (1.4 mmol) was added. The reaction was stirred at
room temperature for 18-24 h and its completion of the reaction was monitored by TLC. After
the completion, the reaction mixture was diluted with water and extracted with DCM (3 x 20 mL).
The organic layer was separated, dried over anhydrous sodium sulfate and evaporated under
reduced pressure to give crude product, which was recrystallized using ethyl acetate/hexanes to
yield pure 8a-d.
(4R)-N-(4-((Z)-2-(3,4-Bis(heptyloxy)phenyl)-1-cyanovinyl)phenyl)-4-((3R,10S,12S,13R,17R)-
3,12-dihydroxy-10,13-dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-
1
yl)pentanamide (8a): Yellow solid; yield: 84% (2.29 g); mp: 130-131 C; H NMR (400 MHz,
CDCl3) δ 7.98 (s, 1H, NHAmide), 7.68 – 7.63 (m, 3H), 7.62 – 7.57 (m, 2H), 7.39 (s, 1H), 7.35 (dd,
J = 8.5, 1.8 Hz, 1H), 6.91 (d, J = 8.5 Hz, 1H), 4.12 – 4.04 (m, 4H, 2xO-CH2 Alkoxy), 4.02 – 3.99
(m, 1H, H-12DCA), 3.68 – 3.60 (m, 1H, H-3DCA), 2.49 – 2.10 (m, 6H), 2.03 – 1.77 (m, 12H), 1.74 –
1.46 (m, 12H), 1.42 – 1.27 (m, 16H), 1.02 (d, J = 5.8 Hz, 3H, Me-19DCA), 0.94 – 0.87 (m, 9H,
13
Me-21DCA& 2xMeAlkoxy), 0.70 (s, 3H, Me-18DCA); C NMR (100 MHz, CDCl3) δ 172.3 (C=O),
151.2, 148.9, 141.3, 138.8, 130.3, 126.6, 126.3, 124.0, 119.9, 48.7, 113.0, 112.7, 107.6, 73.3 (C-
12DCA), 71.8 (C-3DCA), 69.2 (O-CH2 Alkoxy), 69.0 (O-CH2 Alkoxy), 48.2, 46.8, 46.5, 42.1, 36.4, 36.0,
35.3, 35.1, 34.2, 34.1, 34.1, 33.7, 31.8, 31.8, 31.2, 30.5, 29.2, 29.1, 29.1, 28.6, 27.6, 27.2, 26.2,
26.0, 26.0, 23.7, 23.2, 22.6 (C-19DCA), 17.5 (C-21DCA), 14.1 (2xCH3 Alkoxy), 12.8 (C-18DCA);
+
HRMS (ESI): m/z [M+H]+ calcd for Chemical Formula: C53H79N2O5 823.5984 found : 823.6009;
[α]20 = +21 (c 1.0, MeOH); IR (KBr, ῡ, cm-1) 3433, 3294, 2924, 2862, 2206, 1666, 1594, 1034,
D
841.
(4R)-N-(4-((Z)-2-(3,4-Bis(octyloxy)phenyl)-1-cyanovinyl)phenyl)-4-((3R,10S,12S,13R,17R)-
3,12-dihydroxy-10,13-dimethylhexadecahydro-1H-cyclopenta[a]phenanthren-17-
1
yl)pentanamide (8b): Yellow solid; yield: 82% (2.19 g); mp: 128-129 C; H NMR (400 MHz,
DMSO-d6) δ 10.09 (s, 1H, NHAmide), 7.81 (s, 1H), 7.71 (d, J = 8.4 Hz, 2H), 7.68 – 7.60 (m, 3H),
7.48 (d, J = 7.9 Hz, 1H), 7.07 (d, J = 8.4 Hz, 1H), 4.53 – 4.47 (m, 1H, OHDCA), 4.25 – 4.20 (m,
1H, H-12DCA), 4.04 – 3.94(m, 4H, 2xO-CH2 Alkoxy), 3.80 (brs, 1H, OHDCA), 3.35 – 3.32 (m, 1H, H-
3DCA), 2.39 – 1.89 (m, 4H), 1.87 – 1.64 (m, 12H), 1.58 – 1.35 (m, 12H), 1.32 – 1.10 (m, 22H),
0.97 (d, J = 5.0 Hz, 3H, Me-19DCA), 0.90 – 0.79 (m, 9H, Me-21DCA & 2xMeAlkoxy), 0.60 (s, 3H,
13
Me-18DCA); C NMR (100 MHz, DMSO-d6) δ 172.4 (C=O), 151.1, 148.6, 141.5, 140.4, 128.9,
126.9, 126.3, 124.0, 119.6, 119.0, 114.0, 113.5, 107.0, 71.5 (C-12DCA), 70.4 (C-3DCA), 68.8 (O-

CH2 Alkoxy), 68.7 (O-CH2 Alkoxy), 47.9, 46.6, 46.4, 42.1, 36.7, 36.1, 35.6, 35.5, 34.3, 34.3, 33.4,
31.7, 30.7, 29.2, 29.1, 27.7, 27.4, 26.6, 26.1, 24.0, 23.5, 23.3, 22.6 (C-19DCA), 17.6 (C-21DCA),
14.4 (2xCH3 Alkoxy), 12.9 (C-18DCA); HRMS (ESI): m/z [M+Na]+ calcd for Chemical Formula:
C55H82N2O5Na+ 873.6116 found : 873.6173; [α]20D = +92 (c 1.0, MeOH); IR (KBr, ῡ, cm-1) 3502,
3294, 3178, 2924, 2854, 2206, 1666, 1597, 1041, 841.
(4R)-N-(4-((Z)-2-(3,4-bis(dodecyloxy)phenyl)-1-cyanovinyl)phenyl)-4-
((3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethylhexadecahydro-1H-
cyclopenta[a]phenanthren-17-yl)pentanamide (8c): Yellow solid; yield: 85% (1.38 g); mp:
1
140-141 C; H NMR (400 MHz, DMSO-d6 + CDCl3) δ 10.02 (s, 1H, NHAmide), 7.75 – 7.68 (m,
3H), 7.66 – 7.58 (m, 3H), 7.45 (d, J = 7.8 Hz, 1H), 7.00 (d, J = 8.6 Hz, 1H), 4.48 – 4.42 (m, 1H,
OHDCA), 4.18 – 4.15 (m, 1H, H-12DCA), 4.03 – 3.95 (m, 4H, 2xO-CH2 Alkoxy), 3.80 (brs, 1H,
OHDCA), 3.36 – 3.33 (m, 1H, H-3DCA), 2.38 – 2.05 (m, 4H), 1.99 – 1.51 (m, 22H),1.41 – 1.32 (m,
10H), 1.28 – 1.22 (m, 30H), 0.98 (d, J = 5.8 Hz, 3H, Me-19DCA), 0.88 – 0.80 (m, 9H, Me-21DCA
&
13
2xMeAlkoxy), 0.60 (s, 3H, Me-18DCA); C NMR (100 MHz, DMSO-d6 + CDCl3) δ 172.4 (C=O),
151.1, 148.7, 141.2, 140.4, 129.0, 126.9, 126.2, 124.1, 119.6, 118.9, 113.9, 113.4, 107.2, 71.6 (C-
12DCA), 70.5 (C-3DCA), 68.9 (O-CH2 Alkoxy), 68.7 (O-CH2 Alkoxy), 47.9, 46.6, 46.4, 42.1, 36.7, 36.1,
35.6, 35.6, 35.3, 34.3, 33.9, 33.4, 31.9, 30.1, 29.7, 29.6, 29.6, 29.6, 29.3, 29.3, 29.2, 29.2, 29.1,
29.0, 27.7, 27.5, 26.5, 26.1, 24.0, 23.5, 23.3, 22.6 (C-19DCA), 17.6 (C-21DCA), 14.4 (2xCH3 Alkoxy),
+
12.9 (C-18DCA); HRMS (ESI): m/z [M+H]+ calcd for Chemical Formula: C63H99N2O5 963.7549
found : 963.7585; [α]20 = +13 (c 1.0, MeOH); IR (KBr, ῡ, cm-1) 3502, 3294, 2924, 2854, 2206,
D
1659, 1597, 1041, 841.
(4R)-N-(4-((Z)-2-(3,4-Bis(octadecyloxy)phenyl)-1-cyanovinyl)phenyl)-4-
((3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethylhexadecahydro-1H-
cyclopenta[a]phenanthren-17-yl)pentanamide (8d): Yellow solid; yield: 81% (1.82 g); mp:
1
138-139 C; H NMR (400 MHz, CDCl3) δ 7.62 (dd, J = 8.6, 2.3 Hz, 1H), 7.52 (d, J = 2.4 Hz,
1H), 7.39 – 7.35 (m, 2H), 7.19 (s, 1H), 7.16 (s, 1H), 6.90 (d, J = 8.6 Hz, 1H), 6.66 – 6.62 (m, 2H),
3.98 – 3.91 (m, 4H, 2xO-CH2 Alkoxy), 3.84 – 3.74 (m, 1H, H-12DCA), 3.58 – 3.50 (m, 1H, H-3DCA),
2.66 – 2.52 (m, 2H), 2.50 – 2.34 (m, 2H), 1.94 – 1.56 (m, 22H), 1.52 – 1.32 (m, 20H), 1.28 – 1.14
(m, 44H), 0.98 (d, J = 5.7 Hz, 3H, Me-19DCA), 0.88 – 0.78 (m, 9H, Me-21DCA & 2xMeAlkoxy),
0.64 (s, 3H, Me-18DCA); 13C NMR (100 MHz, CDCl3) δ 172.0 (C=O), 152.0, 147.3, 140.1, 137.4,
128.2, 127.1, 127.0, 124.8, 118.5, 115.1, 113.1, 109.5, 73.2 (C-12DCA), 71.9 (C-3DCA), 68.8 (O-
CH2 Alkoxy), 48.3, 47.4, 46.6, 42.1, 36.5, 36.1, 35.2, 35.1, 34.1, 33.7, 31.9, 30.9, 30.5, 29.7, 29.7,
29.7, 29.7, 29.4, 29.4, 29.1, 28.7, 27.5, 7.1, 26.2, 25.9, 23.7, 23.2, 22.7 (C-19DCA), 17.4 (C-
21DCA), 14.1 (2xCH3 Alkoxy), 12.8 (C-18DCA); HRMS (ESI): m/z [M+Na]+ calcd for Chemical

Formula: C75H122N2O5Na+ 1153.9246 found : 1153.9289; [α]20D = +7 (c 1.0, MeOH); IR (KBr, ῡ,
cm-1) 3433, 3294, 2924, 2862, 2206, 1666, 1594, 1034, 841.
Procedure for gelation for compounds 8a-d
In a screw cap vial, gelator (10 mg-20 mg) was added in a solvent (1 mL) and heated or swirled in
an oil bath until all the solid was completely dissolved. In most of the solvents, all gelators
becomes soluble at room temperature upon gentle swirling and heating was not required. The
vials were left undisturbed at room temperature. The gelation (G) was confirmed when no flow
was observed after inversion of the vial, if the formed gel was stable at lower temperature (>20˚C)
but unstable at room temperature (20˚C) it was confirmed as weak gel (WG), if the gelator was
partially soluble at room temperature it was confirmed as partial soluble (PS), if a clear solution
(>70 mg) was obtained the state was marked as soluble (S). Some formed gels were found to be
transparent while others were found to be opaque. The minimum gelation concentration (MGC)
was determined by measuring the minimum amount of gelator required for the formation of the
gel at room temperature.[50]
Scanning Electron Microscopy Analysis (SEM)
Morphology of the freeze dried gels were determined using Scanning Electron Microscope (SEM)
from Carl Zeiss (Ʃigma VP). The powder samples were spread over the carbon tape and coated
with Au-Pd alloy for 2 min. The gel sample was spread over a silicon wafer and dried completely
followed by coating with Au-Pd alloy for 2 min. Gels were dried using a labcon lyophillizer.
Transmission Electron Microscopy Analysis (TEM)
The TEM analysis were performed by placing a small amount of gel in xylene and toluene at
MGC on the copper grid and dried at room temperature for 24 h. Images were taken on JEOL-
2010F TEM operating with electron beam of energy of 200 kV.
FT-IR Analysis
FT-IR was taken on ABB Bomen MB 3000 FTIR for gelator and freeze dried gels, using KBr
disk technique.
XRD Analysis
XRD analysis of the xerogel was recorded on the Rigaku Ultima IV fully automatic high
resolution X-ray diffractometer system with Theta-Theta (θ-θ) Goniometer.
Fluorescence Microscopic Study
A small amount of gel obtained from respective solvents at minimum gel concentration was
placed on a glass coverslip slide and was kept undisturbed under dust free environment for slow
evaporation of the solvent and finally dried under vaccum to obtain xerogel which was placed

under the epi-Fluorescence Microscope (Olympus-BX41, Olympus, Japan) with a red and green
filter using 100X objective lens and 10 X eyepiece lens.
Procedure for bio-imaging
The compounds were tested for its bio-imaging application against the gram –ve bacterium
Serratia marcescens and gram +ve bacterium Staphylococcus aureus. The bacterial strain were
cultured using Luria-Bertani medium (Hi-media, India) at 37 °C on a rotary shaker with 150 rpm.
The fluorescent staining was performed using the previously described procedure with slight
modifications (Moyes, 2009). The bacterial cultures were grown up to mid-log phase and
incubated with 50 µg/mL concentration of compounds at 37 °C for 30 min. Following incubation,
5 µL of each bacterial culture was used for preparing the smears on the glass slide. The smears
were fixed using the 95% ethanol for 5 min and washed by 1 X PBS buffer (Phosphate buffer
saline, pH 7.2). The smears were air-dried and covered with glass cover-slips. The fluorescent
images of bacteria was captured using the epi-Fluorescence Microscope (Olympus-BX41,
Olympus, Japan) at 100X objective lens and 10 X eyepiece lens.
Conflicts of interest
There are no conflicts to declare.
Acknowledgements
The author RS would like to acknowledge UGC and BITS Pilani for research funding. The author
DA is thankful to BITS Pilani, for providing fellowship. We also like to thank central NMR
facility, BITS Pilani and financial support from DST under FIST program [Project: SR/FST/CSI-
270/2015] for HRMS facility. The authors would also like to thank Materials Research Centre
(MRC), MNIT, Jaipur for TEM analysis.
References
[1] A.K. Vasu, M. Radhakrishna, S. Kanvah, Self-Assembly Tuning of α-Cyanostilbene Fluorogens:
Aggregates to Nanostructures, J. Phys. Chem. C 121(40) (2017) 22478-22486.
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Highlights


Four novel deoxycholic acid tethered α-cyanostilbenes were synthesized and characterized.
The synthesized deoxycholic acid tethered α-cyanostilbene derivatives formed stable gels with a
variety of solvents.


The stable gels showed lamellar sheet type structures stacked over each other, consisting of
entangled fibres.
The synthesized compounds also exhibited AIEE behaviour in H2O/THF mixture and the AIEE
property was explored for bacterial bio-imaging.