Triazole tethered C5-curcuminoid-coumarin based molecular hybrids as novel antitubulin agents: Design, synthesis, biological investigation and docking studies

Article abstract of DOI:10.1016/j.ejmech.2016.03.050

Keeping in view the confines allied with presently accessible antitumor agents and success of C₅-curcuminoid based bifunctional hybrids as novel antitubulin agnets, molecular hybrids of C₅-curcuminoid and coumarin tethered by triazole ring have been synthesized and investigated for in-vitro cytotoxicity against THP-1, COLO-205, HCT-116 and PC-3 human tumor cell lines. The results revealed that the compounds A-2 to A-9, B-2, B-3, B-7 showed significant cytotoxic potential against THP-1, COLO-205 and HCT-116 cell lines, while the PC-3 cell line among these was found to be almost resistant. Structure activity relationship revealed that the nature of Ring X and the length of carbon-bridge (n) connecting triazole ring with coumarin moiety considerably influence the activity. Methoxy substituted phenyl ring as Ring X and two carbon-bridges were found to be the ideal structural features. The most potent compounds (A-2, A-3 and A-7) were further tested for tubulin polymerization inhibition. Compound A-2 was found to significantly inhibit the tubulin polymerization (IC₅₀ = 0.82 μM in THP-1 tumor cells). The significant cytotoxicity and tubulin polymerization inhibition by A-2 was further rationalized by docking studies where it was docked at the curcumin binding site of tubulin.

Full text of DOI:10.1016/j.ejmech.2016.03.050

Accepted Manuscript
Triazole tethered C -curcuminoid-coumarin based molecular hybrids as novel
5
antitubulin agents: Design, synthesis, biological investigation and docking studies
Harbinder Singh, Mandeep Kumar, Kunal Nepali, Manish K. Gupta, Ajit K. Saxena,
Sahil Sharma, Preet Mohinder S. Bedi
PII:
S0223-5234(16)30232-X
10.1016/j.ejmech.2016.03.050
EJMECH 8475
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 18 February 2016
Revised Date: 17 March 2016
Accepted Date: 18 March 2016
Please cite this article as: H. Singh, M. Kumar, K. Nepali, M.K. Gupta, A.K. Saxena, S. Sharma,
P.M.S. Bedi, Triazole tethered C -curcuminoid-coumarin based molecular hybrids as novel antitubulin
5
agents: Design, synthesis, biological investigation and docking studies, European Journal of Medicinal
Chemistry (2016), doi: 10.1016/j.ejmech.2016.03.050.
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ACCEPTED MANUSCRIPT
Triazole tethered C5-curcuminoid-coumarin based molecular hybrids as novel antitubulin agents: Design, synthesis, biological investigation
and docking studies
a,
a,
a
b
c
a,
a,
Harbinder Singh †, Mandeep Kumar †, Kunal Nepali , Manish K. Gupta , Ajit K. Saxena , Sahil Sharma *, Preet Mohinder S. Bedi *
a
Department of Pharmaceutical Sciences, Guru Nanak Dev University, Amritsar, Punjab-143005, India
Lloyd Institute of Management and Technology, Greater Noida, UP, India
b
c
Indian Institute of Integrative Medicine, Jammu, India
O
O
OCH3
O
O
OCH3
X
H CO
3
OCH3
X
O
Triazole as
Linker
O
OCH3
N
Hybridization
OCH3
N
)n
N
C -curcuminoid
5
N
N
(
O
N
O
OH
O
Coumarin
O
MOST POTENT COMPOUND
IC50 = 0.82 µM (THP-1)
= 1.55 µM (Tubulin)
O
O
O
O
n = 2, 3, 4, 5, 6
C -curcuminoid-coumarin hybrid
5

ACCEPTED MANUSCRIPT
Triazole tethered C -curcuminoid-coumarin based molecular hybrids as novel antitubulin
5
agents: Design, synthesis, biological investigation and docking studies
a,
a,
a
b
c
Harbinder Singh †, Mandeep Kumar †, Kunal Nepali , Manish K. Gupta , Ajit K. Saxena , Sahil
a,
a,
Sharma *, Preet Mohinder S. Bedi *
a
Department of Pharmaceutical Sciences, Guru Nanak Dev University, Amritsar, Punjab
1
43005, India
b
Lloyd Institute of Management and Technology, Greater Noida, UP, India
Indian Institute of Integrative Medicine, Jammu, India
c
Corresponding author: 1) Mr. Sahil Sharma
Department of Pharmaceutical Sciences,
Guru Nanak Dev University,
Amritsar, Punjab 143005, India
Mob no. +918427007614
2
) Dr. P.M.S. Bedi
Department of Pharmaceutical Sciences,
Guru Nanak Dev University,
Amritsar, Punjab 143005, India
Mob no. +919815698249
†
These authors contributed equally to this work.

ACCEPTED MANUSCRIPT
1
. Introduction
Many of the currently available antitumor drugs are unable to differentiate between normal and
neoplastic cells or to overcome primary or secondary resistance mechanisms evolved in the
tumor cells [1-4]. Thus there is a pressing need for new antitumor agents with high potency, less
toxicity in non-cancerous cells, and unique targets of action. Currently tumor therapy interfering
with a single biological molecule or pathway has been successfully utilized. However, there is
general belief that agents modulating more than one target or multiple sites on solitary target
could have superior efficacy compared to solo target drugs.
Therefore, modulating various targets or more than one site on distinct target simultaneously can
be achieved by the combination of several drugs with different targets or by single chemical
entity that could modulate several sites of a particular target. As a result, there is increasing
interest in the discovery of hybrids that concomitantly address more than one biological target
for tumor treatment [5-7]. Keeping in view the limitations of the molecules modulating one
target, these hybrids can give new dimensions to the molecules modulating more than one target
site and can thus be extremely beneficial for the society which at the moment is striving hard to
fight with this dreadful disease.
Tubulin is one of the most useful and strategic molecular targets for antitumor drugs. The
dynamic process of microtubule assembly and disassembly can be blocked by various agents that
bind to distinct sites in the β-tubulin subunit. By interfering with microtubule function, these
agents arrest cells in mitosis, eventually leading to cell death, by both apoptosis and necrosis [8-
1
0].
Triazole tethered bi-functional hybrids have been an area of focus for developing new cytotoxic
agents in the last several years. Recently published reports on the cytotoxic potential of these
hybrids attracted researchers worldwide towards their selection as a linker for the two
functionalities [11-14].
Curcumin, a naturally occurring polyphenolic compound has shown tremendous success in
various forms of therapeutic activities, among which, the chemotherapeutic effect in diverse
forms of tumor is well documented and the most successful one. But, its poor solubility,
absorption, bioavailability and rapid metabolism in human limit its clinical efficacy. To

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overcome such limitations, C -curcuminoids have been synthesized [15-18] and some of the
5
recently developed C -curcuminoid based molecular hybrids (1 and 2) (Fig. 1), exhibit potent
5
antitumor activity [13, 14], which further supports its insertion as a vital pharmacophore in bi-
functional molecular hybrids.
Coumarins, an elite class of naturally occurring compounds with promising therapeutic
perspectives [19-21]. Sufficient numbers of recent reports on the cell damage potential of
coumarin based molecules rationalize their inclusion in bi-functional molecular hybrids [22-24].
Therefore, the present study is targeted at the design (Fig. 2) and synthesis of triazole tethered
C -curcuminoid-coumarin bifunctional hybrids via simple synthetic procedures and evaluation of
5
these synthesized hybrids against a panel of human tumor cell lines using sulforhodamine B
assay. Current reports on C -curcuminoid and coumarin based antitumor molecules (2 and 3)
5
(Fig. 1) suggest tubulin as one of their primary targets [14, 24], therefore, hybrids exhibiting
significant cytotoxicity were further tested for in vitro tubulin polymerization inhibition. In vitro
microtubule polymerization inhibition by the most potent hybrid was also streamlined by
exploring its key binding interactions with amino acid residues of tubulin using docking studies.
2
. Results and discussion
2
.1. Chemistry
Target hybrids were synthesized via a sequence of reactions (Scheme 1) starting from vanillin,
where it was treated with acetone in the presence of 40% KOH at room temperature. The desired
product vinyldenacetone (VDA) was thus formed. VDA was then treated with propargyl bromide
in DMF as a solvent, in the presence of K CO at room temperature to get the desired product i.e.
2
3
propargylated vinyldenacetone (PVDA). PVDA was further treated with various substituted
aldehydes in methanol as a solvent and the presence of 5% NaOH at room temperature in order
to obtain the desired product wiz propargylated C -curcuminoid analogues.
5
Concurrently, 4-hydroxy coumarin was treated with diverse 1,2-dibromoalkanes in the presence
of K CO , in DMF as a solvent at room temperature to get the desired product (4-(2-
2
3
bromoalkoxy)-2H-chromen-2-one), which was further reacted with NaN in DMF as a solvent, at
3
room temperature to yield 4-(2-azidoalkoxy)-2H-chromen-2-one.

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These 4-(2-azidoalkoxy)-2H-chromen-2-ones were treated with propargylated C -curcuminoid
5
analogues in the presence of copper sulfate and sodium ascorbate, in DMF as a solvent, at room
temperature to gain the desired triazole linked C -curcuminoid-coumarin hybrids.
5
In view of our recent report on triazole linked mono carbonyl curcumin-isatin bi-functional
hybrids having antitubulin potential [14], only those C -curcuminoids have been synthesized and
5
tethered to coumarin through triazole which earlier showed significant cytotoxicity when linked
to isatin via triazole. Moreover, only those four cell lines i.e. THP-1, COLO-205, HCT-116 and
PC-3 out of eight cell lines, in the previous study, have been employed for biological
investigation which were sensitive towards triazole linked mono carbonyl curcumin-isatin bi-
functional hybrids.
2
.2. Bioassay
All the synthetics were investigated for in-vitro cytotoxicity against four human cancer cell lines
THP-1, COLO-205, HCT-116 and PC-3) using sulforhodamine B19 [25, 26]. The cells were
(
allowed to proliferate in the presence of test material for 48 h. All the synthesized hybrids were
screened against the cell lines at 50 µM. The hybrids showing percentage inhibition (Table 1) of
greater than 70% at least against one cell line were only evaluated at different concentrations and
IC values (Table 2) were calculated. In contrary to our previous report [14], among the four
5
0
cancer cell lines, THP-1, COLO-205 and HCT-116 were sensitive towards the designed hybrids
whereas PC-3 was found to be resistant. More surprisingly, among the three sensitive cell lines,
THP-1 cancer cell line instead of HCT-116 (most sensitive towards triazole linked mono
carbonyl curcumin-isatin bi-functional hybrids) was the most sensitive one towards these
hybrids. Compounds A-2, A-3 and A-7 displayed significant cytotoxicity with IC value ranging
5
0
from 0.82-4.68 µM, 2.34-6.78 µM and 4.48-9.95 µM respectively, against THP-1, HCT-116 and
COLO-205 cell lines. The most active hybrid A-2 with trimethoxy phenyl ring as Ring X (IC =
5
0
0
2
.82 µM) was almost three folds more active than A-3 (dimethoxy phenyl ring as Ring X, IC =
50
.34 µM), the second most potent hybrid against THP-1 cell line. Cytotoxicity results in Table 1
revealed an interesting structure activity relationship (Fig. 3) for these designed hybrids: (i)
methoxy substituted phenyl ring as Ring X remarkably enhances the cytotoxic potential
(compare A-1 with A-2 to A-6); (ii) placement of a hateroaryl ring such as furan and thiophene
in place of the unsubstituted phenyl ring as Ring X improved the activity profile (compare A-1

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with A-8 and A-9); (iii) an enhanced effect was observed with the increased number of methoxy
substituents on phenyl ring as Ring X such as trimethoxy phenyl > dimethoxy phenyl >
monomethoxy phenyl (compare A-2 and A-3 with A-4, A-5 and A-6); (iv) placement of naphthyl
ring as Ring X behaved as a surrogate for dimethoxy substituted phenyl ring (compare A-3 with
A-7); (v) cytotxicity of hybrids with monomethoxy substituted phenyl ring as Ring X was found
similar to the heteroaryl ring substituted hybrids (compare A-4, A-5 and A-6 with A-8 and A-9).
Thus, the overall preference order of Ring X is established as follows: trimethoxy phenyl >
dimethoxy phenyl = naphthyl > monomethoxy phenyl = furan = thiophene > phenyl.
Additionally, the length of carbon-bridge connecting triazole ring with coumarin moiety
considerably influences the activity. With the increase in chain length of carbon-bridge,
cytotoxicity decreases significantly (compare A with B, C and D).
The most active compounds (A-2, A-3 and A-7) among the series were evaluated for their
inhibitory effects on tubulin polymerization as per the reported assay using a cytoskeleton
tubulin polymerization assay kit [27, 28]. Compound A-2 (most potent hybrid) with a trimethoxy
phenyl ring (Ring X) also displayed the most potent antitubulin activity with IC value of 1.55
5
0
µM. A-3 also exhibited significant inhibition of tubulin polymerization with an IC value of
5
0
2
.88 µM. A-7 with naphthyl ring (Ring X) was found to be endowed with weak inhibitory
potential for tubulin polymerization. The results of the in-vitro tubulin polymerization assay
clearly indicate that both A-2 and A-3 exert their cytotoxic effects through tubulin inhibition
(Table 3).
2
.3. Molecular docking
Docking study revealed that A-2 fits well at the interface of β1/α2 subunits of tubulin and
stabilized by various electrostatic interactions. Residues from the both subunits (β1and α2) were
involved in D-R interactions (Fig. 4). The trimethoxy-phenyl ring (ring A) gets positioned in a
cavity formed byArg2α2 (polar residue), Pro72β1, Ala99β1 and leu242α2 (a nonpolar residues).
Here, -NH function of Arg2α2 (H-bond donor) is involved in a hydrogen bond interaction with –
OCH (-O: H=bond acceptor; d= 2.995Å). The presence of three methoxy groups (-OCH ) in the
3
3
aromatic phenyl ring (ring A) provides a unique blend polarity and hydrophobicity to the
inhibitor A-2. Such functional group significantly increases the protein binding of inhibitors via
van der Waals and dispersion interactions [29]. The trimethoxy-phenyl group is also present in

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potent tubulin inhibitors such as colchicine, podophyllotoxin and combretastatin A-4. Therefore,
this can be suggested that the presence of trimethoxy-phenyl group in A-2 may be attributed for
its superior activity. The ring B was found to be sandwiched between two polar residues namely
Gln11β1 and Glu254α2. The –OCH group of ring B was oriented towards the side chain
3
carboxylate ionic head of Glu254α2 and involved H-bond interactions (d=2.404Å). Here, –OCH3
2
+
is H-bond donor while carboxylate oxygen is H-bond acceptor. The Mg has shown two polar
interactions with two oxygen linked at 3 and 4 position of ring B (ring B: 3–OCH d=2.377Å
3
,
and 4–OCH – function, d=2.510Å).The triazole ring (ring C) was located near the carboxylate
2
ionic head of Asp197β1.Here, this can be suggested that it is involved in polar interaction with
Asp197β1. The ring D/E (2H-chromen-2-one moiety) was located in a cavity formed by
Pro222β1, Thr223β1 and Thr224β1. Here, ring E has shown face-to-face π-π stacking interaction
with Tyr224β1 (d=3.479Å). The face-to-face π-π stacking interaction play important role in
binding of aromatic compounds with their target receptors and also contribute in their target
specificity [30].
The docking study was helpful to explain the plausible mechanism of A-2 mediated tubulin
inhibition. Microtubules are dynamic polymers of αβ-tubulin that form various cellular structures
including mitotic spindle for cell division. The microtubule-associated proteins (MAPs) and
motor proteins regulate the growth and shrinkage of microtubule by selective binding to distinct
conformations of polymerized or un-polymerized αβ-tubulin [31, 32]. The αβ-tubulin can adopt
distinct conformation which contributes to the polymerization dynamics of microtubules. The
diverse MAPs identify the distinct αβ-Tubulin conformations to build proper microtubule
networks. This can be suggested that binding of A-2 at the interface of β1-α2 heterodimer restrict
the tubulin to adopt MAP-recognizable conformations and thus inhibit the tubulin
polymerization into microtubule assembly. This results in G2/M arrest within the cell cycle and
eventually cell death [31-34].
3
. Conclusion
In continuation of our search for novel antitubulin bi-functional hybrids, C -curcuminoid-
5
coumarin hybrids were designed, synthesized and evaluated for cytotoxicity against a panel of
human cancer cell lines in the present study. Tubulin inhibitory potential of the most potent
compound A-2 (with trimethoxy phenyl ring as ring X and n = 2) was confirmed by in vitro

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tubulin assay which clearly indicates that A-2 exert its cytotoxic effect through tubulin
inhibition. The worth mentioning inhibition of tubulin by compound A-2 was also streamlined by
docking studies.
4
4
. Experimental section
.1. Materials and measurements
The reagents were purchased from Sigma-Aldrich, Loba and CDH, India and used without
further purification. All yields refer to isolated products after purification. Products were
1
13
characterized by comparison with authentic samples and by spectroscopic data ( H and
C
1
13
NMR, Elemental Analysis). H NMR and C NMR Spectra were recorded on Avance III HD
5
00 MHz BrukerBiospin Nuclear Magnetic Resonance Spectrometer. The spectra were measured
1
in CDCl and DMSO-d relative to TMS (0.00 ppm). In H NMR chemical shifts were reported
3
6
in δ values using tetramethylsilane as internal standard with number of protons, multiplicities (s-
singlet, d-doublet, t-triplet, q-quartet, m-multiplet, dd-double doublet) and coupling constants (J)
in Hz (Hertz) in the solvent indicated.
4
.2. General procedure for the synthesis of vinyldenacetone (VDA)
Vanillin (20 g) was dissolved in acetone (50 ml) in a round bottom flask and 40% KOH (5 ml)
was added to this reaction mixture. It was kept on stirring at room temperature. After the
completion of reaction (monitored by TLC), reaction mixture was acidified with dilute
hydrochloric acid until pH 7 was achieved. It was then poured in ice-cold water, filtered and
dried to get the desired product i.e. vinyldenacetone (VDA). The physical data of
vinyldenacetone is given below:
o
1
4
.2.1. (E)-4-(4-hydroxy-3-methoxyphenyl)but-en-2-one (VDA): Yield 72%; mp 71-73 C. H
NMR (CDCl , 500 MHz, δ, TMS = 0): 2.36 (s, 3H), 3.91 (s, 3H), 6.58 (d, J = 16.5 Hz, 1H), 6.92
3
1
3
(
d, J = 7.5 Hz, 1H), 7.04–7.08 (m, 2H), 7.45 (d, J = 16.5 Hz, 1H). C NMR (CDCl , 125 MHz,
3
δ, TMS = 0): 27.22, 55.84, 109.56, 114.99, 123.52, 124.82, 126.79, 144.03, 147.08, 148.51,
1
98.72.
4
.3. General procedure for the synthesis of propargylated vinyldenacetone (PVDA)

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VDA (1 eq) was dissolved in DMF, propargyl bromide (1 eq) and K CO (1.5 eq) were added.
2
3
Reaction mixture was kept on stirring at room temperature. After the completion of reaction
monitored by TLC), reaction mixture was poured on crushed ice and kept aside for some time.
Then it was filtered and dried to obtain the desired product i.e. propargylated vinyldenacetone
PVDA).The physical data of propargylated vinyldenacetone is given below:
(
(
4
.3.1. (E)-4-(3-methoxy-4-(prop-2-ynyloxy)phenyl)but-3-en-2-one (PVDA): Yield 74%; mp
o
1
7
4-77 C. H NMR (CDCl , 500 MHz, δ, TMS = 0): 2.38 (s, 3H). 2.55 (s, 1H), 3.92 (s, 3H), 4.81
3
(s, 2H), 6.63 (d, J = 16.5 Hz, 1H), 7.05 (d, J = 7.5 Hz, 1H), 7.08–7.14 (m, 2H), 7.47 (d, J = 16.5
1
3
Hz, 1H). C NMR (CDCl , 125 MHz, δ, TMS = 0): 27.38, 55.93, 56.62, 76.28, 77.94, 110.30,
3
1
13.75, 122.45, 125.71, 128.50, 143.28, 149.00, 149.83, 198.33.
4
.4. General procedure for the synthesis of propargylated C -curcuminoidanalogs
5
PVDA (1 eq) was dissolved in methanol, 3,4-dimethoxy benzaldehyde (1 eq) and 5% NaOH (0.5
ml) were added. Reaction mixture was kept on stirring at room temperature. After the
completion of reaction (monitored by TLC), reaction mixture was poured on crushed ice and
kept aside for some time. Then it was filtered and dried to obtain the desired product i.e.
propargylated C -curcuminoid analog (M-3).
5
4
.4.1 (1E, 4E)-1-(3-methoxy-4-(prop-2-ynyloxy)phenyl)-5-(3,4-dimethoxyphenyl)penta-1,4-
o
1
dien-3-one (M-3): Yield 71%; mp 76-78 C. H NMR (CDCl , 500 MHz, δ, TMS = 0): 2.88 (s,
3
1
7
8
1
H), 3.91 (s, 3H), 3.93 (s, 6H), 4.81 (s, 2H), 6.94 (d, J = 8.5 Hz, 1H), 7.03–7.08 (m, 3H), 7.23–
1
3
.25 (m, 4H), 7.66-7.70 (m, 2H). C NMR (CDCl , 125 MHz, δ, TMS = 0): 60.75, 61.38, 64.92,
3
1.90, 83.25, 114.94, 115.01, 115.55, 116.18, 118.70, 127.90, 128.14, 128.53, 129.01, 132.96,
33.72, 147.40, 147.81, 153.72, 154.00, 154.50, 156.12, 175.48, 193.29.
All the other propargylated C -curcuminoid analogs were synthesized by following the above
5
mentioned procedure, using various substituted aromatic aldehydes.
4
.5. General procedure for the synthesis of 4-(2-bromoalkoxy)-2H-chromen-2-one
4
-Hydroxy coumarin (1 eq) was dissolved in DMF, dibromoethane (1 eq) and K CO (1.5 eq)
2
3
were added. Reaction mixture was kept on stirring at room temperature. After the completion of
reaction (evident by TLC), reaction mixture was poured on crushed ice and kept aside for some

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time. Then it was filtered and dried to obtain the desired product i.e. 4-(2-bromoethoxy)-2H-
chromen-2-one. All the other 4-(2-bromoalkoxy)-2H-chromen-2-ones were synthesized by
following the above mentioned procedure, using various dibromoalkanes.
4
.6. General procedure for the synthesis of 4-(2-azidoalkoxy)-2H-chromen-2-one
4
-(2-bromoethoxy)-2H-chromen-2-one (1 eq) was dissolved in DMF and sodium azide (1 eq)
was added. Reaction mixture was kept on stirring at room temperature. After the completion of
reaction (evident by TLC), reaction mixture was poured on crushed ice and kept aside for some
time. Then it was filtered and dried. Crude product thus obtained was impure, therefore,
subjected to column chromatography, using chloroform as an eluent, to get the desired product
i.e. 4-(2-azidoethoxy)-2H-chromen-2-one (HCE).
o
1
4
.6.1 4-(2-azidoethoxy)-2H-chromen-2-one (HCE): Yield 91%; mp 92-94 C. H NMR
(
CDCl , 500 MHz, δ, TMS = 0): 3.77 (t, 2H, J = 5 Hz), 4.47 (t, 2H, J = 5 Hz), 5.68 (s, 1H), 7.28-
3
1
3
7
.35 (m, 2H), 7.57-7.60 (m, 1H), 7.89 (d, 1H, J = 7.5 Hz). C NMR (CDCl , 125 MHz, δ, TMS
3
=
0): 27.57, 30.90, 68.48, 90.89, 115.32, 116.80, 123.10, 124.06, 132.67, 153.35, 162.56, 164.90,
2
06.99.
All the other 4-(2-azidoalkoxy)-2H-chromen-2-ones were synthesized by following the above
mentioned procedure, using various 4-(2-bromoalkoxy)-2H-chromen-2-ones.
4
.7. General procedure for the synthesis of triazole linked C -curcuminoid-coumarin
5
hybrids
C -curcuminoid (1 eq) and 4-(2-azidoalkoxy)-2H-chromen-2-one (1 eq) were dissolved in DMF,
5
catalytic amount of copper sulphate and sodium ascorbate were added in it. Reaction mixture
was kept aside at room temperature. After the completion of reaction (evident by TLC), reaction
mixture was filtered on crushed ice to remove the excess of copper sulphate and sodium
ascorbate and then kept aside for some time. Then it was filtered and dried to obtain the desired
product i.e. triazole tethered C -curcuminoid-coumarin hybrids (A, B, C, D). The physical data
5
of all the synthesized bi-functional hybrids is given below:
4
.7.1 (1E,4E)-1-phenyl-5-(3-methoxy-(4(1-(2-(2-oxo-2H-chromen-4-yloxy)ethyl)-1H-1,2,3-
o
triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-1): Yield 81%; mp 174-176 C; MW

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1
5
5
49.57. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.81 (s, 3H), 4.65 (s, 2H), 4.91 (s, 2H),
6
.21 (s, 2H), 5.93 (s, 1H), 7.13-7.18 (m, 3H), 7.21-7.31 (m, 6H), 7.46-7.58 (m, 5H), 7.79-7.84
1
3
(
m, 2H), 8.31 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.90, 49.03, 56.01, 61.90,
6
6
1
1
7
8.36, 91.42, 110.88, 111.21, 112.17, 113.73, 115.31, 116.80, 123.18, 123.30, 123.71, 126.41,
28.01, 128.76, 135.20, 142.90, 143.23, 149.40, 149.57, 150.01, 151.51, 153.06, 162.06, 164.71,
88.69. Anal. Calcd for C H N O : C, 69.93; H, 4.95; N, 7.65; Found: C, 70.05; H, 4.75; N,
32
27
3
6
.71.
4
.7.2
(1E,4E)-1-(3,4,5-trimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)ethyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-2): Yield 89%;
o
1
mp 130-132 C; MW 639.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.71 (s, 3H), 3.79 (s,
6
3
H), 3.85 (s, 6H), 4.64 (s, 2H), 4.95 (s, 2H), 5.21 (s, 2H), 5.97 (s, 1H), 7.12 (s, 2H), 7.21-7.39
1
3
(
m, 7H), 7.64-7.73 (m, 4H), 8.30 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.93,
6
5
1
1
6
6.11, 56.55, 60.61, 62.09, 68.31, 91.58, 106.58, 111.39, 113.78, 115.39, 116.87, 123.25, 123.43,
24.30, 124.66, 125.84, 130.79, 133.27, 140.10, 143.05, 143.20, 149.70, 150.19, 153.19, 153.59,
61.89, 164.71, 188.63. Anal. Calcd for C H N O : C, 65.72; H, 5.20; N, 6.57; Found: C,
3
5
33
3
9
5.58; H, 5.40; N, 6.46.
4
.7.3 (1E,4E)-1-(3,4-dimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)ethyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-3): Yield 81%;
o
1
mp 150-152 C; MW 609.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.78 (s, 3H), 3.80 (s,
6
3
H), 3.83 (s, 3H), 4.62 (s, 2H), 4.94 (s, 2H), 5.20 (s, 2H), 5.93 (s, 1H), 7.02 (d, 1H, J = 7.5 Hz),
1
3
7
.19-7.38 (m, 9H), 7.64-7.70 (m, 4H), 8.41 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
0
): 48.93, 49.05, 56.07, 56.08, 61.95, 68.30, 91.48, 110.92, 111.25, 112.10, 113.68, 115.32,
16.85, 123.23, 123.36, 123.73, 124.21, 124.71, 127.96, 133.33, 142.95, 143.19, 149.44, 149.63,
50.05, 151.55, 153.12, 162.00, 164.75, 188.69. Anal. Calcd for C H N O : C, 66.99; H, 5.13;
1
1
3
4
31
3
8
N, 6.89; Found: C, 66.72; H, 5.28; N, 6.78.
4
.7.4
(1E,4E)-1-(4-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)ethyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-4): Yield 84%;
o
1
mp 165-167 C; MW 579.60. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.73 (s, 6H), 4.66 (s,
6
2
3
H), 4.94 (s, 2H), 5.23 (s, 2H), 5.95 (s, 1H), 7.04-7.15 (m, 5H), 7.19-7.26 (m, 5H), 7.34-7.45 (m,
13
H), 7.77-7.80 (m, 2H), 8.33 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.92,
6

ACCEPTED MANUSCRIPT
5
6.15, 61.97, 68.84, 91.54, 89.38, 110.67, 111.68, 113.55, 115.28, 117.58, 118.79, 119.81,
20.94, 121.58, 123.55, 125.60, 126.86, 128.47, 128.59, 128.79, 136.28, 142.47, 149.05, 150.55,
52.77, 162.04, 164.78, 188.67. Anal. Calcd for C H N O : C, 68.38; H, 5.04; N, 7.25; Found:
1
1
33
29
3
7
C, 68.48; H, 4.95; N, 7.37.
4
.7.5
(1E,4E)-1-(3-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)ethyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-5): Yield 84%;
o
1
mp 163-165 C; MW 579.60. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.72 (s, 3H), 3.79 (s,
6
3
7
6
1
1
5
H), 4.64 (s, 2H), 4.90 (s, 2H), 5.20 (s, 2H), 5.93 (s, 1H), 7.04-7.14 (m, 6H), 7.22-7.34 (m, 7H),
1
3
.64 (m, 2H), 8.39 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.89, 56.09, 61.91,
6
8.78, 91.49, 89.30, 110.65, 111.62, 113.50, 115.20, 117.52, 118.73, 119.74, 120.90, 121.54,
23.49, 125.56, 126.82, 128.43, 128.54, 128.77, 136.20, 142.40, 149.00, 150.52, 152.74, 162.05,
64.74, 188.63. Anal. Calcd for C H N O : C, 68.38; H, 5.04; N, 7.25; Found: C, 66.21; H,
3
3
29
3
7
.14; N, 6.68.
4
.7.6
(1E,4E)-1-(2-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)ethyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-6): Yield 84%;
o
1
mp 161-163 C; MW 579.60. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.70 (s, 6H), 4.62 (s,
6
2
8
1
1
H), 4.87 (s, 2H), 5.18 (s, 2H), 5.85 (s, 1H), 7.04-7.14 (m, 6H), 7.18-7.25 (m, 7H), 8.12 (s, 1H),
13
.39 (m, 2H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.96, 56.15, 61.96, 68.79, 91.39,
6
11.60, 114.24, 115.00, 117.54, 119.76, 120.98, 121.06, 121.56, 123.40, 125.56, 126.87, 127.49,
28.48, 128.56, 129.00, 142.44, 149.08, 149.56, 152.74, 157.76, 162.10, 164.79, 188.74. Anal.
Calcd for C H N O : C, 68.38; H, 5.04; N, 7.25; Found: C, 66.42; H, 5.18; N, 6.86.
3
3
29
3
7
4
1
.7.7
(1E,4E)-1-naphthyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)ethyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-7): Yield 85%; mp 140-142 C;
1
MW 599.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.83 (s, 3H), 4.67 (s, 2H), 4.94 (s,
6
2
H), 5.23 (s, 2H), 5.91 (s, 1H), 7.14-7.19 (m, 3H), 7.24-7.34 (m, 5H), 7.43-7.53 (m, 5H), 7.77-
1
3
7
.84 (m, 5H), 8.31 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.94, 49.00, 56.06,
6
6
1.97, 68.40, 91.46, 110.93, 111.26, 112.24, 113.79, 115.37, 116.87, 123.26, 123.37, 123.77,
24.00, 126.02, 126.34, 126.41, 128.01, 128.76, 128.80, 135.20, 142.96, 143.26, 149.47, 149.63,
50.11, 151.58, 153.16, 162.08, 164.75, 188.60. Anal. Calcd for C H N O : C, 72.11; H, 4.87;
1
1
3
6
29
3
6
N, 7.01; Found: C, 72.21; H, 4.77; N, 7.21.

ACCEPTED MANUSCRIPT
4
1
.7.8 (1E,4E)-1-(furan-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)ethyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-8): Yield 80%; mp 155-157 C;
1
MW 539.54. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.80 (s, 3H), 4.60 (s, 2H), 4.95 (s,
6
2
1
1
1
H), 5.22 (s, 2H), 5.95 (s, 1H), 7.13-7.16 (m, 3H), 7.32-7.43 (m, 6H), 7.69-7.77 (m, 5H), 8.42 (s,
1
3
H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.95, 56.14, 61.99, 68.80, 91.58, 111.40,
6
11.60, 112.72, 115.20, 117.50, 119.73, 120.90, 121.5, 123.45, 125.55, 126.72, 128.58, 129.45,
39.00, 142.42, 145.90, 149.02, 149.70, 151.62, 152.76, 162.05, 164.68, 188.66. Anal. Calcd for
C H N O : C, 66.78; H, 4.67; N, 7.79; Found: C, 66.82; H, 4.59; N, 7.86.
30
25
3
7
4
1
1
4
8
1
1
.7.9 (1E,4E)-1-(thiophen-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)ethyl)-
H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (A-9): Yield 85%; mp 147-
o
1
49 C; MW 555.60. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 3.83 (s, 3H), 4.61 (s, 2H),
6
.96 (s, 2H), 5.25 (s, 2H), 5.97 (s, 1H), 7.09-7.14 (m, 3H), 7.36-7.45 (m, 6H), 7.67-7.77 (m, 5H),
1
3
.41 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 48.92, 56.12, 61.97, 68.82, 91.56,
6
11.41, 111.64, 112.76, 115.27, 117.55, 119.77, 120.95, 121.4, 123.46, 125.54, 126.77, 128.54,
29.44, 137.87, 142.43, 145.96, 149.06, 149.74, 151.67, 152.77, 162.03, 164.62, 188.65. Anal.
Calcd for C H N O S: C, 64.85; H, 4.54; N, 7.56; S, 5.77; Found: C, 64.92; H, 4.46; N, 7.67;
3
0
25
3
6
S, 5.62.
4
1
.7.10
(1E,4E)-1-phenyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)propyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-1): Yield 80%; mp 158-160 C;
1
MW 563.60. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.11 (s, 2H), 3.75 (s, 3H), 4.61 (s,
6
2
H), 4.92 (s, 2H), 5.25 (s, 2H), 5.94 (s, 1H), 7.15-7.21 (m, 3H), 7.27-7.36 (m, 6H), 7.43-7.53
1
3
(
m, 5H), 7.78-7.84 (m, 2H), 8.34 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 28.16,
6
4
8.98, 56.25, 63.49, 72.73, 86.14, 110.73, 111.70, 113.56, 115.27, 117.64, 118.84, 119.83,
20.96, 121.57, 123.49, 125.55, 126.93, 128.49, 128.82, 129.84, 135.29, 142.49, 149.07, 149.78,
50.25, 152.84, 162.08, 164.88, 188.74. Anal. Calcd for C H N O : C, 70.33; H, 5.19; N, 7.46;
1
1
3
3
29
3
6
Found: C, 70.43; H, 5.29; N, 7.32.
4
.7.11
(1E,4E)-1-(3,4,5-trimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)propyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-2): Yield 89%;
o
1
mp 117-119 C; MW 653.68. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.15 (s, 2H), 3.73 (s,
6
3
H), 3.81 (s, 3H), 3.82 (s, 6H), 4.65 (s, 2H), 4.95 (s, 2H), 5.21 (s, 2H), 5.88 (s, 1H), 7.03 (s, 2H),

ACCEPTED MANUSCRIPT
1
3
7
.14-7.21 (m, 7H), 7.65-7.81 (m, 4H), 8.31 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
0
): 28.60, 48.90, 56.20, 63.40, 72.60, 86.04, 103.97, 111.60, 115.20, 117.55, 119.72, 120.97,
21.53, 123.46, 125.56, 126.87, 128.45, 128.54, 129.50, 138.49, 142.44, 149.00, 149.70, 150.26,
50.70,152.75, 162.13, 165.23, 188.65. Anal. Calcd for C H N O : C, 66.15; H, 5.40; N, 6.43;
1
1
3
6
35
3
9
Found: C, 66.18; H, 5.30; N, 6.56.
4
.7.12
(1E,4E)-1-(3,4-dimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)propyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-3): Yield 78%;
o
1
mp 133-135 C; MW 623.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.14 (s, 2H), 3.75 (s,
6
3
H), 3.83 (s, 3H), 3.85 (s, 3H), 4.64 (s, 2H), 4.96 (s, 2H), 5.23 (s, 2H), 5.96 (s, 1H), 7.03-7.13
1
3
(
m, 6H), 7.22-7.34 (m, 6H), 7.62-7.67 (m, 2H), 8.41 (s, 1H). C NMR (d -DMSO, 125 MHz, δ,
6
TMS = 0): 28.06, 48.94, 56.20, 63.40, 72.66, 111.62, 115.26, 117.54, 119.70, 120.98, 121.56,
23.46, 125.56, 126.88, 128.44, 128.56, 142.45, 149.70, 150.22, 152.77, 162.00, 164.75, 188.69.
Anal. Calcd for C H N O : C, 67.41; H, 5.33; N, 6.74; Found: C, 66.59; H, 5.29; N, 6.79.
1
3
5
33
3
8
4
.7.13
(1E,4E)-1-(4-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)propyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-4): Yield 84%;
o
1
mp 150-152 C; MW 593.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.15 (s, 2H), 3.79 (s,
6
6
H), 4.63 (s, 2H), 4.92 (s, 2H), 5.21 (s, 2H), 5.92 (s, 1H), 7.14-7.23 (m, 5H), 7.26-7.33 (m, 5H),
1
3
7
.43-7.48 (m, 3H), 7.67-7.72 (m, 2H), 8.35 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
0
): 28.15, 48.95, 55.97, 56.27, 63.45, 72.67, 86.19, 110.74, 111.73, 113.56, 115.27, 117.64,
18.83, 119.84, 120.96, 121.57, 123.49, 125.59, 126.93, 128.49, 128.63, 129.84, 136.28, 142.50,
49.06, 149.78, 150.25, 152.84, 162.13, 164.87, 188.76. Anal. Calcd for C H N O : C, 68.79;
1
1
34
31
3
7
H, 5.26; N, 7.08; Found: C, 68.89; H, 5.16; N, 7.24.
4
.7.14
(1E,4E)-1-(3-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)propyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-5): Yield 80%;
o
1
mp 146-148 C; MW 593.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.13 (s, 2H), 3.77 (s,
6
6
7
5
1
H), 4.65 (s, 2H), 4.95 (s, 2H), 5.22 (s, 2H), 5.97 (s, 1H), 7.13-7.25 (m, 6H), 7.29-7.40 (m, 7H),
1
3
.71-7.80 (m, 2H), 8.42 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 28.12, 48.91,
6
5.90, 56.20, 63.44, 72.67, 86.09, 110.68, 111.64, 113.51, 115.21, 117.57, 118.78, 119.77,
20.90, 121.51, 123.44, 125.55, 126.88, 128.45, 128.56, 129.77, 136.20, 142.44, 149.00, 149.70,

ACCEPTED MANUSCRIPT
1
50.20, 152.77, 162.05, 164.81, 188.70. Anal. Calcd for C H N O : C, 68.79; H, 5.26; N, 7.08;
3
4
31
3
7
Found: C, 66.82; H, 5.31; N, 6.97.
4
.7.15
(1E,4E)-1-(2-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)propyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-6): Yield 83%;
o
1
mp 145-147 C; MW 593.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.16 (s, 2H), 3.73 (s,
6
6
H), 4.64 (s, 2H), 4.97 (s, 2H), 5.25 (s, 2H), 5.94 (s, 1H), 7.23-7.30 (m, 6H), 7.40-7.48 (m, 7H),
1
3
7
.79-7.84 (m, 2H), 8.40 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 28.06, 48.97,
6
5
6.20, 56.35, 63.45, 72.67, 86.06, 111.60, 114.23, 115.00, 115.23, 117.53, 119.70, 120.90,
21.06, 121.56, 123.40, 125.52, 126.80, 127.40, 128.04, 128.50, 129.02, 142.48, 149.08, 150.24,
52.70, 157.78, 162.08, 164.82, 188.78. Anal. Calcd for C H N O : C, 68.79; H, 5.26; N, 7.08;
1
1
3
4
31
3
7
Found: 68.60; H, 5.36; N, 7.00.
4
1
.7.16 (1E,4E)-1-naphthyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)propyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-7): Yield 80%; mp 124-126 C;
1
MW 613.66. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.14 (s, 2H), 3.77 (s, 3H), 4.63 (s,
6
2
5
4
1
1
7
H), 4.91 (s, 2H), 5.22 (s, 2H), 5.93 (s, 1H), 7.15-7.21 (m, 3H), 7.25-7.34 (m, 5H), 7.42-7.52 (m,
13
H), 7.75-7.82 (m, 5H), 8.34 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 28.20,
6
8.96, 49.05, 56.04, 61.94, 68.46, 91.52, 110.91, 111.27, 112.27, 113.83, 115.42, 116.94, 123.32,
23.43, 123.83, 124.06, 126.07, 126.39, 126.48, 128.04, 128.78, 128.86, 135.28, 142.99, 143.30,
49.53, 149.67, 150.19, 151.59, 153.21, 162.13, 164.79, 188.63. Anal. Calcd for C H N O : C,
3
7
31
3
6
2.42; H, 5.09; N, 6.85; Found: C, 72.49; H, 6.96; N, 6.95.
4
1
.7.17
(1E,4E)-1-(furan-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)propyl)-
o
H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-8): Yield 83%; mp 139-141 C;
1
MW 553.56. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.14 (s, 2H), 3.73 (s, 3H), 4.66 (s,
6
2
3
7
1
1
6
H), 4.95 (s, 2H), 5.23 (s, 2H), 5.91 (s, 1H), 7.19-7.25 (m, 4H), 7.43-7.49 (m, 7H), 7.80-7.84 (m,
1
3
H), 8.42 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 28.60, 48.95, 56.25, 63.45,
6
2.65, 86.05, 111.45, 111.63, 112.79, 115.20, 117.50, 119.75, 120.91, 121.56, 123.45, 125.55,
26.87, 128.49, 128.56, 129.43, 139.00, 142.40, 145.94, 149.06, 149.70, 150.24, 151.67, 152.77,
62.08, 164.82, 188.78. Anal. Calcd for C H N O : C, 67.26; H, 4.92; N, 7.59; Found: C,
3
1
27
3
7
7.37; H, 4.81; N, 7.61.

ACCEPTED MANUSCRIPT
4
.7.18
(1E,4E)-1-(thiophen-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)propyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (B-9): Yield 83%;
o
1
mp 130-132 C; MW 569.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 2.16 (s, 2H), 3.76 (s,
6
3
7
5
1
1
4
H), 4.62 (s, 2H), 4.97 (s, 2H), 5.27 (s, 2H), 5.96 (s, 1H), 7.12-7.22 (m, 3H), 7.41-7.49 (m, 7H),
1
3
.82-7.88 (m, 4H), 8.40 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 28.64, 48.97,
6
6.29, 63.42, 72.61, 86.06, 111.42, 111.68, 112.80, 115.24, 117.55, 119.70, 120.93, 121.58,
23.42, 125.60, 126.90, 127.19, 128.20, 128.56, 137.83, 138.50, 142.45, 145.99, 149.10, 149.77,
50.30, 151.66, 152.80, 162.10, 164.94, 188.79. Anal. Calcd for C H N O S: C, 65.36; H,
3
1
27
3
6
.78; N, 7.38; S, 5.63; Found: C, 65.43; H, 4.68; N, 7.52; S, 5.53.
4
.7.19 (1E,4E)-1-phenyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)butyl)-1H-1,2,3-
o
triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-1): Yield 85%; mp 136-138 C; MW
1
5
77.63. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.83 (s, 2H), 2.07 (s, 2H), 3.86 (s, 3H),
6
4
.27 (s, 2H), 4.54 (s, 2H), 5.21 (s, 2H), 5.88 (s, 1H), 7.03-7.08 (m, 3H), 7.23-7.38 (m, 6H), 7.45-
1
3
7
.57 (m, 5H), 7.77-7.84 (m, 2H), 8.35 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0):
6
2
4.85, 28.00, 52.55, 56.27, 65.64, 72.67, 86.04, 111.65, 115.2, 117.53, 119.76, 120.93, 121.56,
23.45, 125.56, 126.40, 126.80, 128.09, 128.56, 128.74, 135.20, 142.40, 149.00, 149.75, 150.24,
52.78, 128.43, 162.12, 165.38, 188.55. Anal. Calcd for C H N O : C, 70.70; H, 5.41; N, 7.27;
1
1
3
4
31
3
6
Found: C, 70.80; H, 5.30; N, 7.38.
4
.7.20
(1E,4E)-1-(3,4,5-trimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)butyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-2): Yield 89%;
o
1
mp 96-98 C; MW 667.70. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.81 (s, 2H), 2.05 (s,
6
2
7
H), 3.71 (s, 3H), 3.82 (s, 3H), 3.85 (s, 6H), 4.25 (s, 2H), 4.50 (s, 2H), 5.20 (s, 2H), 5.89 (s, 1H),
1
3
.13 (s, 2H), 7.24-7.41 (m, 7H), 7.66-7.83 (m, 4H), 8.31 (s, 1H). C NMR (d -DMSO, 125
6
MHz, δ, TMS = 0): 25.49, 26.83, 49.46, 56.01, 56.51, 60.60, 62.09, 69.22, 91.03, 106.49,
1
1
11.18, 113.67, 115.67, 116.91, 123.33, 123.56, 124.25, 124.68, 125.83, 130.79, 133.82, 139.97,
43.09, 143.27, 149.65, 150.20, 153.20, 153.57, 162.14, 165.36, 188.64. Anal. Calcd for
C H N O : C, 66.56; H, 5.59; N, 6.29; Found: C, 66.46; H, 5.73; N, 6.15.
37
37
3
9
4
.7.21
(1E,4E)-1-(3,4-dimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)butyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-3): Yield 85%;
o
1
mp 112-114 C; MW 637.68. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.81 (s, 2H), 2.05 (s,
6

ACCEPTED MANUSCRIPT
2
H), 3.82 (s, 6H), 3.84 (s, 3H), 4.25 (s, 2H), 4.50 (s, 2H), 5.19 (s, 2H), 5.89 (s, 1H), 7.03 (d, 1H,
1
3
J = 7.5 Hz), 7.25-7.40 (m, 8H), 7.65-7.83 (m, 5H), 8.31 (s, 1H). C NMR (d -DMSO, 125 MHz,
6
δ, TMS = 0): 25.49, 26.83, 49.46, 56.04, 56.08, 69.21, 79.61, 91.48, 110.89, 111.17, 112.08,
1
1
6
13.68, 115.67, 116.91, 123.33, 123.46, 123.73, 124.26, 124.40, 124.67, 133.21, 142.92, 143.13,
49.44, 149.64, 150.12, 151.52, 153.20, 162.15, 165.36, 188.58. Anal. Calcd for C H N O : C,
3
6
35
3
8
7.81; H, 5.53; N, 6.59; Found: C, 67.92; H, 5.43; N, 6.68.
4
.7.22 (1E,4E)-1-(4-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)butyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-4): Yield 82%;
o
1
mp 126-128 C; MW 607.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.86 (s, 2H), 2.04 (s,
6
2
5
H), 3.80 (s, 3H), 3.82 (s,3H), 4.24 (s, 2H), 4.45 (s, 2H), 5.23 (s, 2H), 5.86 (s, 1H), 7.07-7.16 (m,
1
3
H), 7.20-7.27 (m, 5H), 7.38-7.42 (m, 3H), 7.75-7.80 (m, 2H), 8.32 (s, 1H). C NMR (d -
6
DMSO, 125 MHz, δ, TMS = 0): 25.45, 26.80, 49.43, 56.10, 56.08, 69.23, 79.64, 91.52, 110.67,
1
1
11.69, 113.59, 115.29, 117.62, 118.82, 119.79, 120.98, 121.58, 123.53, 125.59, 126.90, 128.49,
28.56, 129.58, 136.30, 142.49, 149.14, 149.77, 152.77, 162.19, 165.35, 188.57. Anal. Calcd for
C H N O : C, 69.18; H, 5.47; N, 6.92; Found: C, 69.31; H, 5.37; N, 7.06.
35
33
3
7
4
.7.23
(1E,4E)-1-(3-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)butyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-5): Yield 79%;
o
1
mp 124-126 C; MW 607.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.84 (s, 2H), 2.08 (s,
6
2
H), 3.80 (s, 3H), 3.82 (s, 3H), 4.22 (s, 2H), 4.48 (s, 2H), 5.23 (s, 2H), 5.86 (s, 1H), 7.12-7.19
1
3
(
m, 6H), 7.35-7.43 (m, 7H), 7.69-7.82 (m, 2H), 8.36 (s, 1H). C NMR (d -DMSO, 125 MHz, δ,
6
TMS = 0): 25.43, 26.79, 49.41, 56.05, 56.06, 69.18, 79.59, 91.46, 110.60, 111.62, 113.55,
1
1
6
15.23, 117.56, 118.78, 119.72, 120.90, 121.54, 123.47, 125.50, 126.85, 128.41, 128.51, 129.53,
36.26, 142.42, 149.06, 149.70, 152.73, 162.11, 165.32, 188.54. Anal. Calcd for C H N O : C,
3
5
33
3
7
9.18; H, 5.47; N, 6.92; Found: C, 67.98; H, 5.52; N, 6.83.
4
.7.24 (1E,4E)-1-(2-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)butyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-6): Yield 82%;
o
1
mp 122-124 C; MW 607.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.82 (s, 2H), 2.06 (s,
6
2
7
2
H), 3.81 (s, 6H), 4.21 (s, 2H), 4.44 (s, 2H), 5.22 (s, 2H), 5.85 (s, 1H), 7.15-7.23 (m, 6H), 7.39-
13
.50 (m, 7H), 7.65-7.76 (m, 2H), 8.34 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0):
6
5.41, 26.78, 49.40, 56.06, 56.07, 69.18, 79.59, 91.46, 111.66, 114.23, 115.20, 117.56, 119.72,

ACCEPTED MANUSCRIPT
1
20.91, 121.50, 123.43, 125.55, 126.88, 127.40, 128.47, 129.00, 142.42, 149.06, 149.72, 150.24,
52.70, 157.78, 162.11, 165.32, 188.54. Anal. Calcd for C H N O : C, 69.18; H, 5.47; N, 6.92;
1
3
5
33
3
7
Found: C, 69.28; H, 5.32; N, 6.98.
4
1
.7.25
(1E,4E)-1-naphthyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)butyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-7): Yield 82%; mp 102-104 C;
1
MW 627.69. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.81 (s, 2H), 2.08 (s, 2H), 3.87 (s,
6
3
7
0
1
1
H), 4.23 (s, 2H), 4.55 (s, 2H), 5.24 (s, 2H), 5.83 (s, 1H), 7.17-7.25 (m, 3H), 7.29-7.34 (m, 5H),
1
3
.41-7.50 (m, 5H), 7.76-7.81 (m, 5H), 8.32 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
): 24.85, 28.00, 52.55, 56.07, 65.64, 72.67, 86.04, 111.65, 115.2, 117.53, 119.76, 120.93,
21.56, 123.45, 124.03, 125.56, 126.05, 126.34, 126.40, 126.80, 128.09, 128.43, 128.87, 128.56,
28.74, 135.20, 142.40, 149.00, 149.75, 150.24, 152.78,162.12, 165.38, 188.55. Anal. Calcd for
C H N O : C, 72.71; H, 5.30; N, 6.69; Found: C, 72.81; H, 5.25; N, 6.74.
38
33
3
6
4
1
.7.26 (1E,4E)-1-(furan-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)butyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-8): Yield 82%; mp 119-121 C;
1
MW 567.59. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.84 (s, 2H), 2.04 (s, 2H), 3.82 (s,
6
3
H), 4.62 (s, 2H), 4.94 (s, 2H), 5.20 (s, 2H), 5.97 (s, 1H), 7.15-7.28 (m, 4H), 7.39-7.47 (m, 7H),
1
3
7
.72-7.80 (m, 3H), 8.33 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 25.44, 26.77,
6
4
9.41, 56.07, 56.08, 69.19, 79.57, 91.46, 111.43, 111.68, 112.77, 115.25, 117.50, 119.77, 120.90,
21.57, 123.44, 125.52, 126.88, 128.40, 128.56, 129.45, 139.00, 142.40, 145.99, 149.03, 149.72,
50.20, 151.67, 152.75, 162.12, 165.31, 188.52. Anal. Calcd for C H N O : C, 67.71; H, 5.15;
1
1
3
2
29
3
7
N, 7.40; Found: C, 67.80; H, 4.98; N, 7.55.
4
1
1
3
.7.27 (1E,4E)-1-(thiophen-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-yloxy)butyl)-
H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (C-9): Yield 84%; mp 111-
o
1
13 C; MW 583.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.87 (s, 2H), 2.06 (s, 2H),
6
.85 (s, 3H), 4.68 (s, 2H), 4.96 (s, 2H), 5.24 (s, 2H), 5.93 (s, 1H), 7.12-7.17 (m, 3H), 7.35-7.44
1
3
(
m, 7H), 7.75-7.83 (m, 4H), 8.37 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 25.48,
6
2
6.82, 49.48, 56.13, 56.18, 69.24, 79.64, 91.49, 111.48, 111.72, 112.79, 115.31, 117.56, 119.83,
20.97, 121.63, 123.47, 125.55, 127.11, 128.20, 128.56, 137.85, 138.50, 141.75, 146.04, 149.10,
49.78, 150.27, 151.73, 152.79, 162.15, 165.37, 188.57. Anal. Calcd for C H N O S: C, 65.85;
1
1
32
29
3
6
H, 5.01; N, 7.20; S, 5.49; Found: C, 65.92; H, 4.93; N, 7.29; S, 5.38.

ACCEPTED MANUSCRIPT
4
.7.28 (1E,4E)-1-phenyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-loxy)pentyl)-1H-1,2,3-
o
triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-1): Yield 81%; mp 122-124 C; MW
1
5
91.65. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.46 (s, 2H), 1.83-1.91 (m, 4H), 3.81 (s,
6
3
H), 4.18 (s, 2H), 4.45 (s, 2H), 5.12 (s, 2H), 5.89 (s, 1H), 7.06-7.10 (m, 3H), 7.20-7.31 (m, 6H),
1
3
7
.42-7.51 (m, 5H), 7.76-7.84 (m, 2H), 8.33 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
0
): 25.42, 26.73, 49.32, 56.03, 69.64, 79.67, 90.93, 111.69, 114.25, 115.14, 115.29, 117.59,
19.76, 120.95, 121.15, 121.57, 123.46, 125.58, 126.41, 128.04, 128.75, 135.20, 142.49, 149.10,
49.79, 150.29, 152.79, 157.78, 162.25, 165.48, 188.64. Anal. Calcd for C H N O : C, 71.05;
1
1
35
33
3
6
H, 5.62; N, 7.10; Found: C, 71.10; H, 5.52; N, 7.22.
4
.7.29
(1E,4E)-1-(3,4,5-trimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
yloxy)pentyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-2): Yield 87%;
o
1
mp 80-82 C; MW 681.73. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.45 (s, 2H), 1.85-1.93
6
(s, 4H), 3.71 (s, 3H), 3.82 (s, 3H), 3.85 (s, 6H), 4.19 (s, 2H), 4.43 (s, 2H), 5.18 (s, 2H), 5.87 (s,
1
3
1
H), 7.12 (s, 2H), 7.23-7.41 (m, 7H), 7.65-7.77 (m, 4H), 8.28-8.31 (m, 1H). C NMR (d -
6
DMSO, 125 MHz, δ, TMS = 0): 22.81, 27.74, 29.73, 49.70, 56.05, 56.50, 60.60, 69.62, 79.62,
9
1
0.95, 116.48, 111.18, 113.64, 115.69, 116.89, 123.26, 123.54, 124.24, 124.62, 125.82, 130.79,
33.17, 139.97, 143.08, 143.26, 149.44, 149.64, 150.21, 153.20, 153.57, 162.15, 165.38, 188.62.
Anal. Calcd for C H N O : C, C, 66.95; H, 5.77; N, 6.16; Found: C, 66.86; H, 5.88; N, 6.10.
3
8
39
3
9
4
.7.30
(1E,4E)-1-(3,4-dimethoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
loxy)pentyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-3): Yield 77%;
o
1
mp 99-101 C; MW 651.70. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.44 (s, 2H), 1.85-1.93
6
(m, 4H), 3.81 (s, 9H), 4.19 (s, 2H), 4.43 (s, 2H), 5.17 (s, 2H), 5.87 (s, 1H), 7.03 (s, 1H), 7.23-
1
3
7
.40 (m, 9H), 7.68-7.77 (m, 4H), 8.27-8.31 (m, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
0
): 25.42, 26.72, 49.40, 56.04, 56.06, 69.62, 79.62, 90.95, 110.89, 111.17, 112.07, 113.65,
15.69, 116.90, 123.26, 123.45, 123.72, 124.40, 124.63, 128.47, 133.17, 142.91, 143.12, 149.44,
49.64, 150.13, 151.52, 153.20, 162.15, 165.38, 188.57. Anal. Calcd for C H N O : C, 68.19;
1
1
37
37
3
8
H, 5.72; N, 6.45; Found: C, 68.30; H, 5.80; N, 6.55.
4
.7.31
(1E,4E)-1-(4-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
loxy)pentyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-4): Yield 86%;
o
1
mp 108-110 C; MW 621.68. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.44 (s, 2H), 1.82-
6

ACCEPTED MANUSCRIPT
1
5
.94 (m, 4H), 3.81 (s, 6H), 4.13 (s, 2H), 4.44 (s, 2H), 5.13 (s, 2H), 5.87 (s, 1H), 7.06-7.16 (m,
1
3
H), 7.21-7.28 (m, 5H), 7.36-7.42 (m, 3H), 7.73-7.80 (m, 2H), 8.31 (s, 1H). C NMR (d6-
DMSO, 125 MHz, δ, TMS = 0): 25.45, 26.77, 49.42, 56.03, 56.07, 69.69, 79.67, 90.90, 110.67,
1
1
11.66, 113.59, 115.27, 117.68, 118.75, 119.79, 120.97, 121.58, 123.48, 125.59, 126.94, 128.49,
28.64, 129.86, 136.32, 142.46, 149.09, 150.26, 172.84, 162.18, 165.47, 188.59. Anal. Calcd for
C H N O : C, 69.55; H, 5.67; N, 6.76; Found: C, 69.35; H, 5.78; N, 6.67.
36
35
3
7
4
.7.32
(1E,4E)-1-(3-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
loxy)pentyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-5): Yield 84%;
o
1
mp 106-108 C; MW 621.68. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.45 (s, 2H), 1.84-
6
1
7
.94 (m, 4H), 3.83 (s, 6H), 4.16 (s, 2H), 4.41 (s, 2H), 5.15 (s, 2H), 5.88 (s, 1H), 7.13 (s, 6H),
1
3
.33-7.40 (m, 7H), 7.68-7.77 (m, 2H), 8.25-8.30 (m, 1H). C NMR (d -DMSO, 125 MHz, δ,
6
TMS = 0): 25.44, 26.73, 49.45, 56.03, 56.05, 69.63, 79.65, 90.97, 110.67, 111.66, 113.56,
1
1
6
15.21, 117.59, 118.70, 119.74, 120.90, 121.51, 123.40, 125.52, 126.88, 128.45, 128.56, 129.77,
36.26, 142.42, 149.07, 150.20, 172.76, 162.17, 165.40, 188.58. Anal. Calcd for C H N O : C,
3
6
35
3
7
9.55; H, 5.67; N, 6.76; Found: C, 69.65; H, 5.58; N, 6.86.
4
.7.33 (1E,4E)-1-(2-methoxyphenyl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-
loxy)pentyl)-1H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-6): Yield 81%;
o
1
mp 105-107 C; MW 621.68. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.46 (s, 2H), 1.82-
6
1
7
.92 (m, 4H), 3.86 (s, 6H), 4.14 (s, 2H), 4.43 (s, 2H), 5.13 (s, 2H), 5.87 (s, 1H), 7.17 (s, 6H),
1
3
.28-7.37 (m, 7H), 7.61-7.73 (m, 2H), 8.26-8.31 (m, 1H). C NMR (d -DMSO, 125 MHz, δ,
6
TMS = 0): 25.41, 26.71, 49.39, 56.03, 56.05, 69.63, 79.65, 90.97, 111.67, 114.22, 115.08,
1
1
15.25, 117.55, 119.73, 120.91, 121.08, 121.50, 123.40, 125.52, 126.81, 127.40, 128.45, 128.56,
29.01, 142.46, 149.08, 149.77, 150.25, 152.77, 157.74, 162.18, 165.42, 188.59. Anal. Calcd for
C H N O : C, 69.55; H, 5.67; N, 6.76; Found: C, 69.66; H, 5.54; N, 6.89.
36
35
3
7
4
1
6
3
7
0
.7.34
(1E,4E)-1-naphthyl-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-loxy)pentyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-7): Yield 81%; mp 84-86 C; MW
1
41.71. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.47 (s, 2H), 1.82-1.93 (m, 4H), 3.84 (s,
6
H), 4.15 (s, 2H), 4.47 (s, 2H), 5.15 (s, 2H), 5.86 (s, 1H), 7.15-7.20 (m, 3H), 7.29-7.34 (m, 5H),
1
3
.42-7.49 (m, 5H), 7.77-7.83 (m, 5H), 8.33 (s, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS =
6
): 25.42, 26.73, 49.32, 56.03, 69.64, 79.67, 90.93, 111.69, 114.25, 115.14, 115.29, 117.59,

ACCEPTED MANUSCRIPT
1
1
19.76, 120.95, 121.15, 121.57, 123.46, 124.06, 125.58, 126.06, 126.34, 126.41, 128.04, 128.75,
28.84, 135.20, 142.49, 149.10, 149.79, 150.29, 152.79, 157.78, 162.25, 165.48, 188.64. Anal.
Calcd for C H N O : C, 72.99; H, 5.50; N, 6.55; Found: C, 72.89; H, 5.66; N, 6.39.
3
9
35
3
6
4
1
.7.35 (1E,4E)-1-(furan-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-loxy)pentyl)-1H-
o
,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-8): Yield 78%; mp 104-106 C;
1
MW 581.62. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.43 (s, 2H), 1.81-1.91 (m, 4H), 3.81
6
(s, 3H), 4.13 (s, 2H), 4.41 (s, 2H), 5.13 (s, 2H), 5.84 (s, 1H), 7.17-7.25 (m, 4H), 7.34-7.46 (m,
1
3
7
H), 7.66-7.78 (m, 3H), 8.26-8.31 (m, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 25.46,
6
2
6.73, 49.37, 56.04, 69.67, 79.69, 90.93, 111.40, 111.65, 112.78, 115.26, 117.54, 119.36, 119.70,
20.93, 123.44, 125.54, 126.84, 128.56, 129.45, 139.00, 142.48, 145.94, 149.06, 149.77, 150.20,
51.60, 152.70, 162.20, 165.46, 188.60. Anal. Calcd for C H N O : C, 68.15; H, 5.37; N, 7.22;
1
1
3
3
31
3
7
Found: C, 68.26; H, 5.19; N, 7.33.
4
1
.7.36 (1E,4E)-1-(thiophen-2-yl)-5-(3-methoxy-(4((1-(2-(2-oxo-2H-chromen-4-loxy)pentyl)-
o
H-1,2,3-triazol-4-yl)methoxy)phenyl)penta-1,4-dien-3-one (D-9): Yield 78%; mp 95-97 C;
1
MW 597.68. H NMR (d -DMSO, 500 MHz, δ, TMS = 0): 1.46 (s, 2H), 1.84-1.96 (m, 4H), 3.86
6
(s, 3H), 4.17 (s, 2H), 4.46 (s, 2H), 5.16 (s, 2H), 5.87 (s, 1H), 7.12-7.23 (m, 3H), 7.39-7.48 (m,
1
3
7
2
1
1
5
H), 7.76-7.84 (m, 4H), 8.28-8.32 (m, 1H). C NMR (d -DMSO, 125 MHz, δ, TMS = 0): 25.42,
6
6.70, 49.31, 56.01, 69.62, 79.63, 90.87, 111.37, 111.61, 112.73, 115.19, 117.47, 119.29, 119.65,
20.91, 123.39, 127.10, 128.14, 137.79, 138.46, 141.71, 145.89, 149.00, 149.67, 150.13, 151.55,
52.64, 162.11, 165.42, 188.58. Anal. Calcd for C H N O S: C, 66.32; H, 5.23; N, 7.03; S,
33
31
3
6
.36; Found: C, 66.21; H, 5.34; N, 6.92; S, 5.51.
4
.8. In-vitro cytotoxic assay
In vitro cytotoxicity against four human cancer cell lines was determined using 96-well tissue
culture plate. The cells were allowed to grow in carbon dioxide incubator (37ºC) for 24 h. Test
materials in complete growth medium (100 ml) were added after 24 h of incubation to the wells
containing cell suspension. The plates were stained with sulforhodamine B dye (0.4% in 1%
acetic acid, 100 ml) for 30 min. The plates were washed five times with 1% trichloroacetic acid
and then air-dried. The adsorbed dye was dissolved in Tris-HCl buffer (100 mL, 0.01 M, pH
1
0.4), and the plates were gently stirred for 10 min. The plates were further incubated for 48 h in

ACCEPTED MANUSCRIPT
a carbon dioxide incubator. The cell growth was stopped by gentle layering trichloroacetic acid
(50%, 50 ml) on top of the medium in all the wells. The plates were incubated at 4ºC for 1 h to
fix the cells attached to the bottom of the wells. The liquid of all the wells was gently pipette out
and discarded. The plates were washed five times with distilled water to remove trichloroacetic
acid, growth medium low molecular weight metabolites, serum on a mechanical stirrer. The
optical density (OD) was recorded on ELISA reader at 540 nm. The cell growth was determined
by subtracting the mean OD value of respective blank from the mean OD value of the
experimental set. Percent growth in presence of test material was calculated considering the
growth in the absence of any test material as 100%, and in turn, percent growth inhibition in
presence of test material was calculated [25, 26].
4
.9. In-vitro tubulin polymerization assay
A cytoskeleton tubulin polymerization assay kit was purchased from Labex India. For assessing
the tubulin polymerization inhibition potential, 100 mM stock solution of each compound was
made in molecular biology grade DMSO. Further dilutions (50 mM, 25 mM, 10 mM) of each
compound were made by diluting the stock solution. General tubulin buffer was prepared by
reconstituting the constituents (available in kit) in 10 mL distilled water. General Tubulin Buffer
(1 mL) was supplemented with 10 mL of 100 mM GTP solution and 10 mg Tubulin was
suspended in it and mixture was stored at 4ºC until used. Before starting the experiment 96 well
plate and Elisa plate reader were warmed at 37ºC as this temperature is required to achieve the
proper polymerization of tubulin. Tubulin polymerization buffer was prepared by mixing
General tubulin buffer (750 mL), Tubulin glycerol buffer (250 mL) and 100 mM GTP (10 mL).
1
0 mL General tubulin buffer was added in the wells, where first two wells were kept as control
(without any test compound), and different concentrations of compounds were subsequently
added in previously marked wells and plate was kept in plate reader for 2 min at 37ºC.
Meanwhile tubulin suspension was diluted with tubulin polymerization buffer and 100 mL of
diluted tubulin was added in each well. Plate was again kept in plate reader at 37ºC for 1 h and
absorbance was measured at 340 nm (Lee and Timasheff, 1977). Percentage inhibition and IC50
values were calculated according to the absorbance values obtained after 1 h [27, 28].
4
.10. Docking simulations

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The X-ray structure coordinates of tubulin bound with colchicine was obtained from protein data
bank (PDB entry: 1SA0; Resolution 3.5Å) [35]. The X-ray structure of tubulin consists of two
tubulin heterodimers in a curved complex capped by the stathmin-likedomain (SLD). In each
tubulin heterodimer, a colchicine molecule was found in complexed form at the interface of α
and β subunits. Chakraborti et al. investigated the tubulin-binding mode of few curcumin analogs
and reported that curcumin binds at the interface of β1-α2 heterodimer which is approximately
3
2 Å away from the colchicine-binding site [36].
To explore the tubulin binding mode of the synthesized compounds, we have docked A-2 at the
curcumin binding site of tubulin as reported by Chakraborti et al. The docking study was carried
out using the GOLD 5.3.0 software [37]. Chem score was applied as fitness function to rank
various docked conformations. Chem score estimates total free energy change that occurs on
ligand binding by considering parameters like hydrogen bond energies, atom radii and
polarisabilities, torsion potentials, hydrogen bond directionalities, lipophilicity, etc [38]. The 3D
conformation of A-2 was generated in ChemDraw software 2010. The 2D structure of A-2 was
drawn in ChemDraw Ultra (2010) and energy minimized using the MM2 force field in Chem 3D
Ultra software. The compound was docked ten times and conformation associated with highest
scoring value was considered to analyze various drug-receptor (D-R) interactions between A-2
and tubulin.
Conflict of interest
The authors confirm that this article content has no conflicts of interest.
Acknowledgements
Authors are grateful to University Grants Commission for providing funds under UPE Scheme to
carry out the research work.
References
[
1] R. Kaur, P. Kaur, S. Sharma, G. Singh, S. Mehndiratta, K. Nepali. Anti-cancer pyrimidines
in diverse scaffolds: A review of patent literature, Rec. Pat. Anticanc. Drug Discov. 10
(2015) 23-71.

ACCEPTED MANUSCRIPT
[
[
[
2] S. Sharma, S. Mehndiratta, S. Kumar, J. Singh, P.M.S. Bedi, K. Nepali. Purine analogues
as kinase inhibitors: A review, Rec. Pat. Anticanc. Drug Discov. 10 (2015) 308-341.
3] H. Singh, H. Singh, S. Sharma, P.M.S. Bedi. Chemotherapeutic potential of acridine
analogs: An ample review, Heterocycles 91 (2015) 2043-2085.
4] S. Sharma, J. Singh, R. Ojha, H. Singh, M. Kaur, P.M.S. Bedi, K. Nepali. Design
strategies, structure activity relationship and mechanistic insights for purines as kinase
inhibitors, Eur. J. Med. Chem. (2016) DOI: 10.1016/j.ejmech.2016.02.018.
[
5] K. Nepali, S. Sharma, M. Sharma, P.M.S. Bedi, K.L. Dhar. Rational approaches, design
strategies, structure activity relationship and mechanistic insights for anticancer hybrids,
Eur. J. Med. Chem. 77 (2014) 422-487.
[
[
6] K. Nepali, S. Sharma, D. Kumar, A. Budhiraja, K.L. Dhar. Anticancer hybrids- A patent
survey, Rec. Pat. Anticanc. Drug Discov. 9 (2014) 303-339.
7] S. Mehndiratta, S. Sharma, S. Kumar, K. Nepali. Molecular hybrids with anticancer
activity. In: Topics in Anti-Cancer Research, Eds. A. Rahman and K. Zaman. Patents
eBook Series. Bentham Science Publishers Ltd. 2015, Vol. 4, pp 383-454.
[
[
8] K. Nepali, R. Ojha, S. Sharma, P.M.S. Bedi, K.L. Dhar. Tubulin inhibitors: A patent
survey, Rec. Pat. Anticanc. Drug Discov. 9 (2014) 176-220.
9] R. Ojha, S. Sharma, K. Nepali. Anticancer agents targeting tubulin. In: Topics in Anti-
Cancer Research, Eds. A. Rahman and K. Zaman. Patents eBook Series. Bentham Science
Publishers Ltd. 2015, Vol. 4, pp 156-270.
[
10] S. Sharma, C. Kaur, A. Budhiraja, Kunal Nepali, M.K. Gupta, A.K. Saxena, P.M.S. Bedi.
Chalcone based azacarboline analogues as novel antitubulin agents: Design, synthesis,
biological evaluation and molecular modeling studies, Eur. J. Med. Chem. 85 (2014) 648-
6
60.
[
[
11] J. Singh, S. Sharma, A.K. Saxena, K. Nepali, P.M.S. Bedi. Synthesis of 1,2,3-triazole
tethered bifunctional hybrids by click chemistry and their cytotoxic studies, Med. Chem.
Res. 22 (2013) 3160-3169.
12] M. Sharma, S. Sharma, A. Buddhiraja, A.K. Saxena, K. Nepali, P.M.S. Bedi. Synthesis and
cytotoxicity studies of 3,5-diaryl N-acetyl pyrazoline-isatin hybrids, Med. Chem. Res. 23
(2014) 4337-4344.

ACCEPTED MANUSCRIPT
[
[
13] S.K. Kandi, S. Manohar, C.E.V. Gerena, B. Zayas, S.V. Malhotra, D.S. Rawat. C5-
curcuminoid-4-aminoquinoline based molecular hybrids: design, synthesis and mechanistic
investigation of anticancer activity, New J. Chem. 39 (2015) 224-234.
14] S. Sharma, M.K. Gupta, A.K. Saxena, P.M.S. Bedi. Triazole linked mono carbonyl
curcumin-isatin bifunctional hybrids as novel anti tubulin agents: Design, synthesis,
biological evaluation and molecular modeling studies, Bioorg. Med. Chem. 23 (2015)
7
165-7180.
[
[
15] Y. Ai, B. Zhu, C. Ren, F. Kang, J. Li, Z. Huang, Y. Lai, S. Peng, K. Ding, J. Tian, Y.
Zhang. Discovery of new monocarbonyl ligustrazine−curcumin hybrids for intervention of
drug-sensitive and drug-resistant lung cancer, J. Med. Chem. 59, (2016) 1747-1760.
16] S.M. Bayomi, H.A. El-Kashef, M.B. El-Ashmawy, M.N.A. Nasr, M.A. El-Sherbeny, N.I.
Abdel-Aziz, M.A.-A. El-Sayed, G.M. Suddek, S.M. El-Messery, M.A. Ghaly. Synthesis
and biological evaluation of new curcumin analogues as antioxidant and antitumor agents:
Molecular modeling study, Eur. J. Med. Chem. 101 (2015) 584-594.
[
[
17] Q. Li, J. Chen, S. Luo, J. Xu, Q. Huang, T. Liu. Synthesis and assessment of the
antioxidant and antitumor properties of asymmetric curcumin analogues, Eur. J. Med.
Chem. 93 (2015) 461-469.
18] P.R. Baldwin, A.Z. Reeves, K.R. Powell, R.J. Napier, A.I. Swimm, A. Sun, K. Giesler, B.
Bommarius, T.M. Shinnick, J.P. Snyder, D.C. Liotta, D.Kalman. Monocarbonyl analogs of
curcumin inhibit growth of antibiotic sensitive and resistant strains of Mycobacterium
tuberculosis, Eur. J. Med. Chem. 92 (2015) 693-699.
[
[
[
[
[
19] S. Sandhu, Y. Bansal, O. Silakari, G. Bansal. Coumarin hybrids as novel therapeutic
agents, Bioorg. Med. Chem. 22 (2014) 3806-3814.
20] C. Kontogiorgis, A. Detsi, D. Hadjipavlou-Litina. Coumarin-based drugs: a patent review
(2008 - present), Expert Opin. Ther. Patents 22 (2012) 437-454.
21] M. Asif. Overview of diverse pharmacological activities of substituted coumarins:
Compounds with therapeutic potentials, Am. J. Curr. Org. Chem. 1 (2015) 1-16.
22] A. Thakur, R. Singla, V. Jaitak. Coumarins as anticancer agents: A review on synthetic
strategies, mechanism of action and SAR studies, Eur. J. Med. Chem. 101 (2015) 476-495.
23] R. Pingaew, A. Saekee, P. Mandi, C. Nantasenamat, S. Prachayasittikul, S. Ruchirawat, V.
Prachayasittikul. Synthesis, biological evaluation and molecular docking of novel

ACCEPTED MANUSCRIPT
chalcone-coumarin hybrids as anticancer and antimalarial agents, Eur. J. Med. Chem. 85
(
2014) 65-76.
24] Yi. Ma, Y. Zhou, C. Xie, D. Chen, J. Li. Novel microtubule-targeted agent 6-chloro-4-
methoxyphenyl) coumarin induces G2-M arrest and apoptosis in HeLa cells. Acta
[
[
(
Pharmacologica Sinica 33 (2012) 407-417.
25] A. Monks, A.D. Scudiero, P. Skehan, R. Shoemaker, K. Paull, D. Vistica, C. Hose, J.
Langley, P. Cronise, A. Vaigro-Wolff, M. Gray-Goodrich, H. Campbell, J. Mayo, M.
Boyd, Feasibility of a high-flux anticancer drug screen utilizing a diverse panel of human
tumor cell lines in culture, J. Natl. Cancer Inst. 83 (1991) 757-766.
[
[
[
26] P. Skehan, R. Storeng, D. Scudiero, A. Monks, J. Mc Mohan, D. Vistica, J.T. Warren, H.
Bokesch, S. Kenney, M.R. Boyd, New colorimetric cytotoxicity assay for anticancer-drug
screening, J. Natl. Cancer Inst. 82 (1990) 1107-1112.
27] G.R. Pettit, B. Toki, D.L. Herald, P. Verdier-Pinard, M.R. Boyd, H. Hamel, R.K. Pettit,
Antineoplastic agents. 379. Synthesis of phenstatin phosphate 1a, J. Med. Chem. 41 (1998)
1
688-1695.
28] M.L. Shelanski, F. Gaskin, C.R. Cantor, Microtubule assembly in the absence of added
nucleotides, Proc. Natl. Acad. Sci. 70 (1973) 765-768; c) J.C. Lee, S.N. Timassheff, In
vitro reconstitution of calf brain microtubules: Effects of solution variables, Biochemistry
1
6 (1977) 1754-1764.
[
29] S. Kumar, S. Mehndiratta, K. Nepali, M.K. Gupta, S. Koul, P.R. Sharma, A.K. Saxena,
K.L. Dhar. Novel indole-bearing combretastatin analogues as tubulin polymerization
inhibitors, Org. Med. Chem. Lett. 3 (2013) 3-13.
[
[
[
30] S.K. Burley, G.A. Petsko. Aromatic-aromatic interaction: a mechanism of protein structure
stabilization, Science. 229 (1985) 23-28.
31] L.D. Belmont, T.J. Mitchison. Identification of a protein that interacts with tubulin dimers
and increases the catastrophe rate of microtubules, Cell. 84 (1996) 623-631.
32] P. Ayaz, X. Ye, P. Huddleston, C.A. Brautigam, L.M. Rice. A TOG:αβ-tubulin complex
structure reveals conformation-based mechanisms for a microtubule polymerase, Science.
3
37 (2012) 857-860.

ACCEPTED MANUSCRIPT
[
33] H. Lodish, A. Berk, S.L. Zipursky, P. Matsudaira, D. Baltimore, J. Darnell. Molecular Cell
Biology, 4th edition. New York: W. H. Freeman; 2000. Section 19.2, Microtubule
Dynamics and Associated Proteins.
[
[
34] G.J. Brouhard, L.M. Rice. The contribution of α β-tubulin curvature to microtubule
dynamics, J. Cell. Biol. 207 (2014) 323-334.
35] R.B. Ravelli, B. Gigant, P.A. Curmi, I. Jourdain, S. Lachkar, A. Sobel, M. Knossow.
Insight into tubulin regulation from a complex with colchicine and a stathmin-like domain,
Nature. 11 (2004) 198-202.
[
36] S. Chakraborti, L. Das, N. Kapoor, A. Das, V. Dwivedi, A. Poddar, G. Chakraborti, M.
Janik, G. Basu, D. Panda, P. Chakrabarti, A. Surolia, B. Bhattacharyya. Curcumin
recognizes a unique binding site of tubulin, J. Med. Chem. 54 (2011) 6183-6196.
37] GOLD 5.3.0. Cambridge Crystallographic Data Centre: Cambridge, U.K. (2015).
38] M.D. Eldridge, C.W. Murray, T.R. Auton, G.V. Paolini, R.P. Mee. Empirical scoring
functions: I. The development of a fast empirical scoring function to estimate the binding
affinity of ligands in receptor complexes, J. Comput. Aided. Mol. Des. 11 (1997) 425-445.
[
[

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Captions
Fig. 1. C -curcuminoid based molecular hybrids (1 & 2) and coumarin derivative with cell
5
damage potential (3)
Fig. 2. Design strategy for C -curcuminoid-coumarin hybrids
5
Scheme 1. Synthesis of triazole linked C -curcuminoid-coumarin hybrids. Reagents and
5
conditions: (a) Acetone, 40% KOH, 2 hrs, stir, rt; (b) Propargyl bromide, K CO , DMF, 2 hrs,
2
3
stir, rt; (c) Aldehydes, 5% NaOH, MeOH, 2 hrs, stir, rt; (d) Dibromoalkane, K CO , DMF, 2 hrs,
2
3
stir, rt; (e) NaN , DMF, 1 hr, stir, rt; (f) Sodium ascorbate, CuSO , DMF, 15 mins, rt.
3
4
Table 1 Various substituted bi-functional hybrids with their %age inhibition against eight human
cancer cell lines at 50 µM
Table 2 IC values of active hybrids against sensitive human cancer cell lines
5
0
Fig. 3. Structure activity relationship
Table 3 Tubulin polymerization inhibition potential of most active hybrids
Fig. 4. (a) Docked conformation of compound A-2 (cyan) at the interface of β1-α2 subunits of
tubulin complexed with colchicine (red) (PDB id 1SA0); (b) Residues involved in drug-receptor
interactions between A-2 and tubulin (A-2: carbon atoms are shown in green; only hydrogen
which is involved in H-bond interactions are shown)

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Table 1
Various substituted bi-functional hybrids with their %age inhibition against eight human cancer cell lines at 50 µM
O
X1
X4
H CO
X2
X3
3
O
X5
(
)
nN
O
O
N
N
O
CODE
X1
X2
X3
X4
X5
n
Percentage growth inhibition (50 µM)
HCT-116
Colon)
COLO-205
(Colon)
THP-1
PC-3
(
(Leukemia) (Prostate)
A-1
A-2
A-3
A-4
A-5
A-6
A-7
H
H
H
H
H
H
H
H
H
2
2
2
2
2
2
56
89
82
71
70
70
83
54
86
80
68
69
67
85
61
99
90
78
77
75
89
10
22
19
15
13
11
20
OCH3
OCH3
H
OCH3
OCH3
OCH3
H
OCH3
H
H
H
H
H
H
H
OCH3
H
H
OCH3
H
H
O
H CO
3
O
N
N
O
N
O
O
A-8
72
68
78
14
O
H CO
O
3
O
N
N
O
N
O
O