Synthesis, Cytotoxicity and Antimicrobial Evaluation of New Coumarin-Tagged β-Lactam Triazole Hybrid

Article abstract of DOI:10.1002/cbdv.201900462

A series of coumarin-tagged β-lactam triazole hybrids (10a–10o) were synthesized and tested for their cytotoxic activity against MDA-MB-231 (triple negative breast cancer), MCF-7 (estrogen receptor positive breast cancer (ER+)) and A549 (human lung carcinoma) cancer cell lines including one normal cell line, HEK-293 (human embryonic kidney). Two compounds 10b and 10d exhibited substantial cytotoxic effect against MCF-7 cancer cell lines with IC₅₀ values of 53.55 and 58.62 μm, respectively. More importantly, compounds 10b and 10d were non-cytotoxic against HEK-293 cell lines. Structure–activity relationship (SAR) studies suggested that the nitro and chloro group at the C-3 position of phenyl ring are favorable for anticancer activity, particularly against MCF-7 cell lines. Furthermore, antimicrobial evaluation of these compounds revealed modest inhibition of examined pathogenic strains with compounds 10c and 10i being the most promising antimicrobial agents against Pseudomonas aeruginosa and Candida albicans, respectively.

Full text of DOI:10.1002/cbdv.201900462

Title: Synthesis, cytotoxicity and antimicrobial evaluation of new
coumarin-tagged β-lactam triazole hybrid
Authors: Sanjeev Dhawan, Paul Awolade, Prishani Kisten, Nosipho
Cele, Ashona-Singh Pillay, Sourav Taru Saha, Mandeep
Kaur, sreekanth b. Jonnalagaddas, and Parvesh Singh
This manuscript has been accepted after peer review and appears as an
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of the final Version of Record (VoR). This work is currently citable by
using the Digital Object Identifier (DOI) given below. The VoR will be
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to this Accepted Article as a result of editing. Readers should obtain
the VoR from the journal website shown below when it is published
to ensure accuracy of information. The authors are responsible for the
content of this Accepted Article.
To be cited as: Chem. Biodiversity 10.1002/cbdv.201900462
Link to VoR: http://dx.doi.org/10.1002/cbdv.201900462

10.1002/cbdv.201900462
Chemistry & Biodiversity
Synthesis, cytotoxicity and antimicrobial evaluation of new coumarin-tagged β-
lactam triazole hybrid
Sanjeev Dhawana, Paul Awoladea, Prishani Kistena,Nosipho Cele, Ashona-Singh Pillaya,
SouravTaru Sahab, Mandeep Kaurb, Sreekantha B. Jonnalagaddaa and Parvesh Singha*
aSchool of Chemistry and Physics, University of KwaZulu-Natal, P/Bag X54001, Westville,
Durban, South Africa-4000
bSchool of Molecular and Cell Biology, University of the Witwatersrand, Private Bag 3,
Johannesburg, WITS-2050, South Africa
*Email address for correspondence: singh4@ukzn.ac.za, parveshdurban@gmail.com
Abstract:
A series of coumarin-tagged β-lactam triazole hybrids (10a-o) were synthesized and tested for
their cytotoxic activity against MDA-MB-231 (triple negative breast cancer), MCF-7 (estrogen
receptor positive breast cancer (ER+) and A549 cells (human lung carcinoma) cancer cell lines
including a normal cell line, HEK-293 (human embryonic kidney). Two compounds, 10b and 10d
exhibited substantial cytotoxic effect against MCF-7 cancer cell-line with IC50 values of 53.55 and
58.62 µM, respectively. More importantly, compounds 10b and 10d were non-cytotoxic against
HEK-293 cells. Structure-activity relationship (SAR) studies suggested that the nitro and chloro
group at the C-3 position of phenyl ring are favorable for anticancer activity, particularly against
MCF-7 cell-line. Furthermore, antimicrobial evaluation of these compounds revealed modest
inhibiton of examined pathogenic strains with compounds 10c and 10i being the most promising
antimicrobial agents against Pseudomonas aeruginosa and Candida albicans respectively.
Keywords: β-Lactam, Coumarin, 1,2,3-triazole,MDA-MB-231, MCF7, HEK-293, Molecular
hybridization.
1
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Introduction
Cancer remains the most typical cause of death in many developed countries, with breast and lung
adenocarcinomas being the most prevalent forms.[1,2]The adverse effects of combination
chemotherapy, the most preferred method for cancer treatment, necessitates the development of
new anticancer agents with acceptable toxicity profile and improved efficacy.[3] Similarly, bacterial
infections pose an increasing community health issue and a consequential life-threatening risk to
human existence due to the rising resistance to current antibiotics.[4] According to the World Health
Organization (WHO) fact sheet 2018, approximately 500,000 people suffer from bacterial
infections caused by Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus,
Streptococcus pneumoniae and Salmonella species in over 22 countries. Therefore, there is an
imperative requirement to establish new antimicrobial agents with potent activity against resistant
microorganisms.[5]
Coumarin is a versatile scaffold that is widely distributed in nature with vast application in the
field of medicinal chemistry. The synthetic accessibility of the coumarin scaffold enables it to
become the perfect platform for structural modifications, and derivatisation.[6] Coumarin-based
compounds have displayed a broad spectrum of pharmacological activity such as anti-viral,[7]
anticancer,[8] anti-fungal,[9] anti-inflammatory[10] and anti-HIV.[11] Currently, a coumarin-based
VEGF expression and angiogenesis inhibitor, NM-3 is under phase I clinical trials.[12]
Nevertheless, only a few synthetic coumarin derivatives have been used to perform antimicrobial
characterisation, particularly those effective against gram-positive bacterial species.[13] Also, there
has been a resurgent interest for coumarin-based antibiotics since their isolations from
Streptomyces species. Notable examples include coumermycin, clorobiocin, as well as novobiocin
which were identified as potent bacterial DNA gyrase and topoisomerase inhibitor.[14] For instance,
2
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10.1002/cbdv.201900462
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Damu et al. reported some coumarins possessing potent antibacterial activity against methicillin
and ciprofloxacin-resistant S. aureus strains.[15] Similarly, Ghalehshahi et al. synthesized new
coumarin analogues with superior antimicrobial activity when compared to their parent
molecules.[16]
Furthermore, β-lactam antibiotics such as the classical cephalosporins, penicillins, non-classical
monobactams and carbapenams due to their synthetic accessibility and low hepatotoxicity profile
have played a vital role in treating bacterial infections.[17] So far, they remain the leading class of
agents used in the antimicrobial regimen and have significantly enhanced human health and life
expectancy.[18] However, pathogenic resistance has been a significant setback to the efficacious
clinical utility of β-lactams.[19] Current research efforts are focused on the synthetic modifications
of the β-lactam scaffold to afford compounds with enhanced antimicrobial and anticancer activity
while prevailing against β-lactamase induced resistance. β-Lactams execute their antimicrobial
properties via the disruption of interpeptide cross-links into bacterial peptidoglycan cell wall by
inhibiting bacterial transpeptidases, termed penicillin-binding proteins (PBPs), thereby curbing
rigidity of the bacterial cell wall.[20] Accordingly, Chen et al. reported that the β-lactam derivatives
display their anticancer activity by inhibiting DNA replication thus influencing DNA damage and
activating the apoptotic death program in human leukemic Jurkat T-cells. In addition, considering
the structural flexibility of β-lactams, they have the potential to be developed into anticancer agents
by various structural modifications.[21]
3
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Chemistry & Biodiversity
Figure 1.β-lactam tagged1,2,3triazole-based drugs in the market
Likewise, 1,2,3-triazole serve as an attractive heterocyclic group to connect two pharmacophores
and biologically active fragments into one molecule to get progressive multifunctional
compounds.[22-27] Moreover, the binding ability of 1,2,3-triazole framework with molecular targets
due to their rigidity, high dipole moment, stability and capability to form hydrogen bonding under
in vivo conditions has resulted in highly potent bioactive agents.[28,29]
In view of the aforementioned biological significance, we decided to fuse coumarin, -lactam, and
1,2,3-triazole pharmacophoric units into a single architecture using the concept of molecular
hybridisation.[30-33] The desired molecular conjugates were synthesised by using the Copper-
assisted 1,3-dipolar cycloaddition reaction between the alkyne-substituted coumarins and azido
lactams. All synthesized compounds were further tested in vitro against three cancer cell lines and
microbial strains.
Results and discussion
Chemistry
The general synthetic routes employed in this study are depicted in Schemes 1-2. A novel series
of triazole-linked coumarin-β-lactam (10a-o) molecular hybrids were prepared from 7-
(ethynyloxy)-4,5-dimethyl-2H-chromen-2-one (9a) and 7-(ethynyloxy)-4-methyl-2H-chromen-2-
one (9b), using various substituted azido lactams (6) in the presence of CuSO4.5H2O and sodium
ascorbate (as reducing agent) in Dichloromethane:Water (DCM:H2O) (8:2) at room
temperature.[34-35] The key starting materials 7-hydroxy-4,5-dimethyl-2H-chromen-2-one (8a) and
7-hydroxy-4-methyl-2H-chromen-2-one (8b) were prepared by the reaction of orcinol (7a) and
phloroglucinol (7b), respectively with ethyl acetoacetate in sulfuric acid (Pechmann reaction).[36]
7-(Ethynyloxy)-4,5-dimethyl-2H-chromen-2-one 9a, in turn, was prepared through the base-
4
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10.1002/cbdv.201900462
Chemistry & Biodiversity
promoted reaction of 7-hydroxy-4,5-dimethyl-2H-chromen-2-one 8a with propargyl bromide in
Dimethylformamide (DMF). Substituted azido lactams were prepared by using azido acetic acid 2
and an imine (5a-h) at 0-5oC.[37] The key azidoketene intermediate was generated in situ in the
presence of p-toluene sulfonyl chloride and triethylamine and subsequently converted to the
desired azido lactams.
The precise structure elucidation of all newly synthesised compounds was accomplished on the
basis of spectroscopic data (1H NMR, 13C NMR, COSY, HSQC, HMBC, IR and mass
spectrometry). The details of spectroscopic data are shown in experimental data whereas the salient
features of the representative compound are discussed here. For instance, the formation of
compound 10a was confirmed by a characteristic deshielded proton of its triazole ring that
resonated at δ 7.84 ppm in its proton NMR. Moreover, a doublet of doublet (dd) at δ 5.90 for H1,
a doublet at 6.27 ppm for H3 proton, and two singlets at δ2.33 and δ 2.17 ppm for methyl protons
also confirmed the structure of 10a. The HRMS spectrum of compound 10a also exhibited its
molecular ion peak of m/z 541.1850, and corroborated the proposed structure.
5
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10.1002/cbdv.201900462
Chemistry & Biodiversity
Scheme 1. Synthetic pathway to azido β-lactam derivatives
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10.1002/cbdv.201900462
Chemistry & Biodiversity
Scheme 2. Synthetic pathway to coumarin-β-lactam derivatives
Furthermore, the quaternary carbons in compound 10a were identified based on the heteronuclear
multiple bond correlation (HMBC) data, depicted in Figure 2. For instance, the carbon C-4a was
identified based on its correlation with the CH3-9 and CH3-10 protons. Similarly, the carbon C-2
was distinguished by its HMBC with H-3; C-4a and C-8 with H-6, C-7 and C-6 with H-8 and
finally C-6 was assigned based on its HMBC with CH3-9, and H-8.
7
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10.1002/cbdv.201900462
Chemistry & Biodiversity
Figure 2. HMBC (1H
Anticancer evaluation
13C) spectrum of 10a showing important correlations.
The cytotoxic efficacy of all synthesized compounds (10a-g, and 10h-o) was tested against three
cancer cell lines, namely MCF7 cells (human breast adenocarcinoma) (ER+), MDA-MB-231 cells
(human breast adenocarcinoma) (TNBC) and A549 cells (Human lung carcinoma), including one
normal cell-line HEK-293 (Human embryonic kidney), using an MTT (3-(4,5-Dimethylthiazol-z-
yl)-2,5-diphenyltetrazolium bromide) assay Table 1.
Fig 3 to 7 illustrate the cytotoxicity results of selected compounds, whereas the IC50 value, which
is the concentration required to inhibit 50% of cell viability by the test compounds, is summarised
in Table 1. As observed in Fig.3, compound 10b and 10d appeared to be active against the MCF7
cells when compared to MDA-MB-231 cells with IC50 values being 53.55 µM and 58.62 µM
respectively. Further testing of these compounds in HEK-293 cells did not display any toxicity
(IC50> 100 µM, Table 1) in normal cells, thus, making these compounds attractive for further
derivatization and testing. The remaining compounds of the series displayed no activity against
both breast cancer cell lines with IC50 values greater than 100 µM. Furthermore, all compounds
showed no toxicity (IC50> 100 µM) against A549 cells, and thus suggested their specificity towards
the breast carcinoma cells.
8
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Chemistry & Biodiversity
Figure 3. Representative graph comparing the percentage growth inhibition of A549, MCF7 and
MDA-MB-231 cells at selected concentrations of test compounds 10a, 10b, 10c and 10d. 40 µM
Plumbagin was used as a positive control. Data are mean ± standard deviation S.D. (n=3), where
*p<0.05, **p<0.01 and ***p<0.001 significant difference to untreated control.
Figure 4. Representative graph comparing the percentage growth inhibition of A549, MCF7 and
MDA-MB-231 cells at selected concentrations of test compounds 10e, 10f, 10g and 10h. 40 µM
Plumbagin was used as a positive control. Data are mean ± standard deviation S.D. (n=3), where
*p<0.05, **p<0.01 and ***p<0.001 significant difference to untreated control.
9
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Chemistry & Biodiversity
Figure 5. Representative graph comparing the percentage growth inhibition of A549, MCF7 and
MDA-MB-231 cells at selected concentrations of test compounds 10i, 10j, 10k and 10l. 40 µM
Plumbagin was used as a positive control. Data are mean ± standard deviation S.D. (n=3), where
*p<0.05, **p<0.01 and ***p<0.001 significant difference to untreated control.
Figure 6.Representative graph comparing the percentage growth inhibition of A549, MCF7 and
MDA-MB-231 cells at selected concentrations of test compounds 10m, 10n and 10o. 40 µM
10
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Chemistry & Biodiversity
Plumbagin was used as a positive control. Data are mean ± standard deviation S.D. (n=3), where
*p<0.05, **p<0.01 and ***p<0.001 significant difference to untreated control.
Figure 7. Representative graph comparing the percentage growth inhibition of HEK-293 cells at
selected concentrations of test compounds 10b and 10d. 40 µM Plumbagin was used as a positive
control. Data are mean ± standard deviation S.D. (n=3), where *p<0.05, **p<0.01 and ***p<0.001
significant difference to untreated control.
Structure-activity relationship (SAR) studies of the synthesized hybrids exhibited the dependency
of activity on the type of phenyl rings substitutions present on N-1 of the β-lactam ring attached
directly to triazole nucleus (Fig. 8). For instance, the meta-positioning of nitro (compound 10b)
and chloro (compound 10d) group increased both the potency (IC50<59 µM) and selectivity of
these conjugates against MCF7 cancer cell line. However, their structural analogues (10m and 10l)
bearing mono-methylated coumarin ring were found to be inactive (IC50>100 µM) against all the
tested cell lines. The unsubstituted phenyl conjugates (10a and 10h) also showed no cytotoxicity
(IC50>100 µM) against MDA-MB-231 and MCF7 cell lines. A summary of the result from the
SAR study is presented in Fig.8 below.
11
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Chemistry & Biodiversity
Figure 8. SAR study of phenyl substituted coumarin tagged 1,2,3-triazole with β-lactams as an anti-
cancer agent and antimicrobial agent.
Analyzing the outcomes in Table 1, we could illustrate some suitable conclusions: 1) introduction
of the nitro and chloro groups at meta position of the phenyl ring leads to anti-cancer activity of
compounds (10b and 10d) against MCF7 cells; 2) Conversely, there was a potency loss in
compound 10l and 10m (IC50 >100 µM), where similar groups attached to the phenyl ring were
introduced to the 4-methyl coumarin moiety, and 3) the positioning of electron-withdrawing
groups (EWG) at position-3 of phenyl moiety generally offered more potent compounds against
MCF7 cell-line.
12
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Chemistry & Biodiversity
Table 1: IC50 values of the synthesised compounds (10a-g/10h-o) againstA549 MCF7, MDA-
MB-231 and HEK-293lines.
Code
Structure
A549
(µM)
MCF-7
(µM)
MDA-MB-
231 cells
(µM)
HEK-293
(µM)
>100
>100
>100
>100
>100
-
10a
10b
>100
53.55
>100
>100
>100
>100
>100
-
10c
10d
>100
58.62
>100
>100
>100
-
-
10e
>100
>100
>100
>100
>100
>100
10f
-
10g
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Chemistry & Biodiversity
>100
>100
>100
>100
>100
10h
10i
-
-
>100
>100
>100
>100
>100
>100
>100
10j
-
-
10k
>100
>100
>100
10l
-
10m
>100
>100
>100
>100
>100
>100
-
10n
10o
-
>100
>100
>100
-
14
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Standard
Plumbagin
11.69
3.5
4.4
NA
Docking studies
The designed series of potential anti-breast cancer agents comprised 15 molecules with two distinct
scaffolds; i.e.10b and 10d. The biological evaluation of the series refined the screen to identify a
novel bioactive molecular hybrid encompassing critical pharmacophoric features which include a
β-lactam, a dimethylated coumarin moiety, a 1,2,3-triazole and a phenyl ring with a C-3
substitution by a strongly electron-withdrawing group. These characteristics were most prominent
and conserved between the most active inhibitors 10b and 10d. It was also unique to find that these
potential drug candidates selectively inhibited the MCF7 cell-line, which emphasizes the affinity
of the designed conjugate to target estrogen receptor-α (ER-α). Molecular docking of the most
active ligands in ER-α was performed to understand the binding interactions critical to induce a
pharmacological response.
Table 2: Docking scores of ligands in the binding pocket of ER-α
Docking score/
Ligand Identifier
RMSD i.b.
RMSD u.b.
(kcal/mol)
-10.9
-11.3
-10.7
-10.9
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
10a
10b
10c
10d
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Chemistry & Biodiversity
-11.0
-11.3
-11.0
-10.4
-10.7
-10.5
-10.6
-10.4
-10.3
-10.7
-9.7
0.000
0.000
1.906
1.719
0.000
0.000
0.000
2.116
1.611
0.000
1.740
0.000
10e
10f
0.000
3.708
2.542
0.000
0.000
0.000
3.747
2.615
0.000
2.474
10g
10h
10i
10j
10k
10l
10m
10n
10o
Compound 10b exhibited an extensive interaction with the binding domain of the receptor
(docking score = -11.3 kcal/mol, (Table 2) and was more potent than compound 10d (IC50 = 53.55
and 58.62 μM respectively). The interaction profile was primarily governed by Vander Waals and
hydrophobic interactions between residues Leu41, Ala45, Leu49, Trp78, Phe99, Ile119, Leu123,
Lys224, Val228 and Leu231 with the carbons of the coumarin ring, nitro-substituted phenyl ring
and methylene linked phenyl ring (bond distance = 3.92, 3.89, 3.71, 3.80, 3.66, 3.65, 3.94, 3.31,
3.26 and 3.93 Å, respectively). This was accompanied by two well-defined hydrogen bonds
between the protein donor residues Thr42 and Leu231 with the nitrogen of the triazole group and
the oxygen of the nitro moiety (bond distance = 3.99 and 2.96 Å; bond angle = 176.64˚ and 138.38˚,
respectively). As mentioned earlier, the presence of an electron withdrawing group,i.e. a nitro
group for compound 10b, at the C-3 position of the N-linked phenyl group has a direct impact on
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10.1002/cbdv.201900462
Chemistry & Biodiversity
the potency of the inhibitor. This is attributed to the formation of a unique electrostatic interaction,
a T-shaped π-stacking between residue Trp78 and the methylene linked phenyl ring (distance =
4.71 Å, angle = 63.45˚).
The interaction network of compound 10d is also grounded by multiple hydrophobic interactions
between receptor residue Leu41, Ala45, Leu86, Leu123, Val228, Leu231 and Leu234 with the
phenyl and coumarin rings of the ligand (bond distance = 3.72, 3.74, 3.49, 3.91, 3.49, 3.85 and
3.43 Å, respectively). The differences in 10d and 10b binding landscapes are first observed with
the single hydrogen bond formation in the complex of 10d between amino acid Thr42 and the
nitrogen of the triazole (bond distance = 3.24 Å and bond angle = 137.56˚). The lack of the
additional stabilizing hydrogen bond could possibly be a reason for the slight discrepancy in the
efficacy of compound 10d (IC50 value ~53.62 μM less than compound 10b). However, the ligand
appears to compensate for the loss of the feature by forming a salt bridge to facilitate charge
transfer between residue Arg89 and the carboxylate of the coumarin ring at a bond distance of 5.38
Å. The electron-withdrawing group of compound 10d is a chlorine atom which when bound in the
catalytic domain of the receptor forms an electrostatic interaction with amino acid Val229 at a
distance of 3.22 Å (donor angle = 145.82˚ and acceptor angle = 126.15˚).
From Table 3 it is evident that the presence of the nitro group in compound 10b significantly alters
the lipophilicity of the inhibitor, whereby the more lipophilic the molecule, the more potent the
potential drug is against the MCF7 cell line. The total polar surface area of compound 10b is
greater than compound 10d, which may be a contributing feature to a more potent anti-breast
cancer agent. The inhibitory activity also appears to be affected by the ligands ability to form
hydrogen bond in the active site, thus greater number of acceptor pharmacophores will result in an
improved inhibitory effect.
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10.1002/cbdv.201900462
Chemistry & Biodiversity
The results observed from docking highlight the advantageous utilization of the coumarin-triazole-
β-lactam molecular hybrid in the future design and development of novel and efficient anti-breast
cancer chemotherapeutics.
Table 3: Physical properties of the most active compounds against MCF7 cell-line
Pharmacophore features
Molecula
Compoun clog TPSA Rotatabl
Ionisable
r Weight/
/ Å2
e bonds
Acceptor Donor
d ID
P
g.mol-1
groups/charg
s
s
e
3.89 136.28
4.96 90.46
8
7
563.56
553.01
8
6
0
0
1/-
1/-
10b
10d
Compound 10d
Compound 10b
Figure 9: Three-dimensional structure of compounds 10b and 10d in complex with ER-α.
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Antimicrobial Evaluation
In this study, five bacterial strains viz; a Gram-positive Methicillin-resistant Staphylococcus
aureus (MRSA; ATCC 43300) and four Gram-negative bacteria (Pseudomonas aeruginosa ATCC
27853, Klebsiella pneumonia ATCC 700603, Escherichia coli ATCC 25922, and Acinetobacter
baumannii ATCC 19606) and two fungal strains (Candida albicans ATCC 90028, and
Cryptococcus neoformans H99; ATCC 208821) were used to determine the antimicrobial activity
of the synthesized compounds. The disc diffusion method was only used to detect the antibacterial
activity of the test compounds at a concentration of 32 µg mL-1 with ≤1% DMSO. The results
showed that the test compounds were moderately active against MRSA, Klebsiella pneumonia,
Pseudomonas aeruginosa, Acinetobacter baumannii bacterial strains, and Candida albicans
fungal strain. Further, out of this, only three of the test compounds, the 10c, 10f, and 10g
compounds exhibited activity against MRSA while most of the test compounds showed moderate
activity in the other strains (Table 4). However, the test compounds did not show any antibacterial
activity against Escherichia coli whereas, compounds 10a, 10d, and 10e were found to be inactive
against Pseudomonas aeruginosa.
The three compounds that exhibited activity against MRSA, 10c, 10f, and 10g had values that
showed a maximum percentage inhibition 7.75, 7.92, and 8.94% respectively and other compounds
were found to be inactive against these strains. Further, the test compound 10i, 10j, 10k, and 10d
were found to be more active with percentage inhibition values of 21.65, 9.42, 6.24, and 14.96
against Candida albicans fungal strain (Table 4). However, all the test compounds were recorded
inactive against Cryptococcus neoformans.
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Table 4: Antibacterial and antifungal screening data of compounds 10a-g and 10h-o
Percentage inhibition (%) *
Bacterial Strain
Fungal Strian
Compound
Name
Sa
Ec
Kp
Pa
Ab
Ca
Cn
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
10a
10b
10c
-
10.44
16.18
-
7.75
18.97
-
-
-
-
14.96
4.35
10d
10e
7.92
-
6.91
17.78
7.8
-
10f
8.94
-
-
-
-
-
4.87
2.33
-
11.11
12.11
4.39
0.22
4.29
2.6
-
-
-
-
-
10g
10h
10i
-
-
-
-
-
-
-
-
21.65
9.42
6.24
-
5.74
10j
10k
3.41
7.32
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-
-
-
-
-
-
-
-
0.05
16.37
7.74
6.66
8.47
-
3.0
-
-
-
-
10l
-
-
-
-
-
-
10m
10n
10o
2.74
2.29
3.47
* All percentage inhibitions were determined at 32 µg/mL.Sa = Staphylococcus aureus, Ec = Escherichia coli, Kp =
Klebsiella pneumonia, Pa = Pseudomonas aeruginosa, Ab = Acinetobacterbaumannii, Ca = Candida albicans, Cn =
Cryptococcus neoformans; DMSO, negative control; Colistin and Vancomycin were used as standards for antibacterial
activity while Fluconazole was used as standard for antifungal activity.
All the tested compounds were found to be less active than they used standard drugs (Colistin and
Vancomycin for bacterial and Fluconazole for fungal strains).
Conclusions
Novel coumarin-β-lactam-triazole molecular hybrids were synthesized using the copper(I)-
catalyzed Huisgen 1,3-dipolar cycloaddition reaction. The biological evaluation of these
compounds three cancer cell lines (MDA-MB-231, MCF7, A549) and one control cell line HEK-
293 revealed two potent compounds (10b and 10d)against MCF-7 cancer cells with no toxicity
against the normal cells. Further, the antimicrobial studies concluded that the compounds bearing
iodo and methyl groups displayed moderate antimicrobial activity against Pseudomonas
aeruginosa and Candida albicans strains. Thus, further modification and structure derivatization
of these compounds may result in the introduction of selective anti-breast cancer agents.
21
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10.1002/cbdv.201900462
Chemistry & Biodiversity
Experimental section
Chemistry
The progress of all reactions was monitored by pre-coated Merck silica gel 60F-254 TLC plates, using
254 nm UV light for visualisation. 1H NMR and 13C NMR spectra were on Bruker 400 and 600 MHz
NMR spectrometer. Chemical shifts given in δ values (ppm) using tetramethylsilane as internal
standard referenced to the residual solvents signal of CDCl3 at δ 7.26 ppm for 1H and δ 77.39 ppm for
13C and residual signal for DMSO at δ 2.50 ppm for 1H and δ 39.50 ppm for 13C. Chemical shift values
expressed in parts per million (ppm) downfield from TMS and J values in Hertz. Splitting patterns
indicated as s: singlet, d: doublet, t: triplet, m: multiplet, dd: double doublet, and br: broad peak.
Resonance assigned by chemical shift, a multiplicity of resonance, signal intensities and H-H coupling
constants. Mass spectra were recorded using high-resolution mass spectrometer (HRMS). The IR
spectra recorded on an FTIR spectrophotometer. Chromatographic separations were carried out on
60:120 silica gel using EtOAc and hexane mixture as eluent.
General Procedure for the synthesis of 7-hydroxy-4,5-dimethyl coumarin (8a) and 7-hydroxy-4-
methyl-2H-chromen-2-one (8b)
In two different 50 mL round-bottom flasks, equimolar quantities of 5-methylbenzene-1,3-diol
(7a) (1.0 eq), ethyl acetoacetate (1.0 eq) for the synthesis of compound 8a and 3-hydroxy phenol
(7b) (1.0 eq), ethyl acetoacetate (1.0 eq) for compound 8b, were added to stirring concentrated
sulfuric (H2SO4) at 0-5°C and stirred for 2.0 hrs. The reactions were monitored by the TLC. After
reaction completion, the reaction mixtures were poured into the crushed ice with continuous
stirring and then filtered, washed with water and recrystallised with ethanol to get 7-hydroxy-4,5-
dimethyl-2H-chromen-2-one (8a) and 7-hydroxy-4-methyl-2H-chromen-2-one (8b). The pure
products were obtained in 83-85% yield. m.p. Found: 260-262°C (8a) and 186-188°C (8b).
22
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10.1002/cbdv.201900462
Chemistry & Biodiversity
General procedure for the synthesis of procedure for the preparation of β-lactam Coumarin
conjugates (10a-g and 10h-o)
To a stirred solution of 7-(ethynyloxy)-4,5-dimethyl-2H-chromen-2-one (9a) and 4-methyl-7-
(prop-2-yn-1-yloxy)-2H-chromen-2-one (9b) (1 mmol for 9a and 9b) in DCM:H2O (8:2),
theazidoβ-lactam(6a-h) (2 mmol for 9a and 9b) was added, followed by the addition of copper
sulphate (0.055 mmol for 9a and 9b) and sodium ascorbate (0.13 mmol for 9a and 9b) at room
temperature. The progress of the reaction was monitored using TLCuntil the disappearance of
starting material was observed then, and water was added (15 mL) to the reaction mixture and
extracted with chloroform (50 mL). Combined organic layers was dried with anhydrous sodium
sulfate and concentrated under reduced pressure to result in the isolation of a crude productwhich
was purified by silica gel chromatography using 4:6 (EtOAc:hexane) for (10a-g) and (10h-o)
compounds.
(Z)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-1-phenyl-4-
styrylazetidin-2-one (10a)
0
Yield-76%; mp 213-215 C; Cream amorphous solid; IR (neat) (νmax/cm-1): 2933, 1755, 1713,
1600; 1H NMR (400 MHz, CDCl3) δ 7.84 (s, 1H, triazole-CH), 7.54 (d, J = 7.9 Hz, 2H, ArH), 7.38
(t, J = 7.8 Hz, 2H. ArH), 7.28 (s, 1H, ArH), 7.21 (dd, J = 16.6, 7.3 Hz, 4H, ArH), 7.14 (d, J = 6.3
Hz, 2H, ArH), 6.77 (d, J = 16.0 Hz, 1H, Vinyl-CH), 6.73 (s, 1H, Coumarin-CH), 6.64 (s, 1H,
Coumarin-CH), 6.27 (d, J = 5.4 Hz, 1H, Vinyl-CH), 5.90 (dd, J = 14.8, 8.5 Hz, 1H, lactam-CH),
5.27 – 5.24 (m, 1H, lactam-CH), 5.18 (s, 2H, CH2), 2.33 (s, 3H, CH3), 2.17 (s, 3H, CH3). 13C NMR
(101 MHz, CDCl3) δ 160.84, 158.53, 156.23, 155.20, 153.91, 143.06, 143.17, 137.80, 136.84,
134.79, 129.45, 129.00, 128.73, 126.70, 125.48, 123.64, 119.69, 117.44, 113.60, 110.86, 108.40,
23
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10.1002/cbdv.201900462
Chemistry & Biodiversity
108.32, 67.19, 62.18, 60.06, 24.15, 21.93. HRMS of [C31H26N4O4 + Na]+ (m/z): 541.1852; Calcd:
541.1850.
(Z)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-1-(3-
nitrophenyl)-4-styrylazetidin-2-one (10b)
Yield-80%; mp 188-190 0C; Yellow amorphous solid; IR (neat) (νmax/cm-1): 1755, 1603, 1488;1H
NMR (400 MHz, CDCl3) δ 7.85 (s, 1H, triazole-CH), 7.53 (d, J = 7.6 Hz, 2H, ArH), 7.37 (t, J =
7.4 Hz, 2H, ArH), 7.20 (dd, J = 16.6, 7.1 Hz, 4H, ArH), 7.13 (s, 2H, ArH), 6.80 – 6.67 (m, 2H,
ArH), 6.62 (s, 1H, Coumarin-CH), 6.26 (d, J = 4.9 Hz, 1H, Vinyl-CH), 5.89 (t, J = 11.4 Hz, 2H,
lactam-CH), 5.27 (d, J = 6.0 Hz, 1H, lactam-CH), 5.19 (s, 2H, CH2), 2.33 (s, 3H, CH3), 2.17 (s,
3H, CH3). 13C NMR (101 MHz, CDCl3) δ 160.86, 158.55, 156.23, 155.18, 153.94, 143.07,
137.80,136.84, 134.79, 129.45, 129.00, 128.73, 126.70, 125.47, 123.66, 119.71, 117.43, 113.58,
110.85, 108.39, 108.32, 77.37, 77.05, 76.73, 67.19, 62.18, 60.07, 24.15, 21.93. HRMS of
[C31H25N5O6 + Na]+(m/z): 586.1703; Calcd: 586.1796.
(Z)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-1-(3-
methoxyphenyl)-4-styrylazetidin-2-one (10c)
0
Yield-83%; mp 198-200 C; White amorphous solid; IR (neat) (νmax/cm-1): 1715, 1606, 1488;1H
NMR (400 MHz, CDCl3) δ 7.84 (s, 1H, triazole-CH), 7.58 – 7.40 (m, 2H, ArH), 7.27 – 7.20 (m,
3H, ArH), 7.13 (dd, J = 7.7, 1.6 Hz, 2H, ArH), 6.94 – 6.89 (m, 2H, ArH), 6.76 (d, J = 3.2 Hz, 1H,
Vinyl-CH), 6.73 (s, 1H, Coumarin-CH), 6.65 (s, 1H, Coumarin-CH), 6.25 (d, J = 5.4 Hz, 1H,
Vinyl-CH), 5.93 (d, J = 0.9 Hz, 1H, Coumarin-CH), 5.87 (dd, J = 16.0, 7.3 Hz, 1H, lactam-CH),
5.26 – 5.22 (m, 1H, lactam-CH), 5.22 (s, 2H, CH2), 3.81 (s, 3H,Ar-OCH3), 2.36 (s, 3H, CH3), 2.21
(d, J = 0.9 Hz, 3H, CH3). 13C NMR (101 MHz, CDCl3) δ 160.84, 157.91, 157.13, 156.24, 155.20,
153.90, 143.05, 137.73, 134.80, 130.26, 128.98, 128.73, 126.68, 123.59, 119.82, 118.85, 114.61,
24
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10.1002/cbdv.201900462
Chemistry & Biodiversity
113.61, 110.87, 108.32, 67.22, 62.19, 60.11, 55.51, 24.16, 21.94. HRMS of [C32H28N4O5 +
Na]+(m/z): 571.1957; Calcd: 571.1953.
(Z)-1-(3-chlorophenyl)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-
triazol-1-yl)-4-styrylazetidin-2-one (10d)
Yield-86%; mp 188-190 0C; Cream amorphous solid; IR (neat) (νmax/cm-1): 1714, 1608, 1383; 1H
NMR (400 MHz, CDCl3) δ 7.83 (s, 1H, triazole-CH), 7.47 (d, J = 8.6 Hz, 2H, ArH), 7.33 (d, J =
8.6 Hz, 2H, ArH), 7.23 (d, J = 6.9 Hz, 3H, ArH), 7.13 (d, J = 6.7 Hz, 2H, ArH), 6.79 – 6.70 (m,
2H, ArH), 6.62 (s, 1H, Coumarin-CH), 6.26 (d, J = 5.4 Hz, 1H, Vinyl-CH), 5.87 (dd, J = 16.4,
6.8Hz, 2H, lactam-CH), 5.26 – 5.21 (m, 1H, lactam-CH), 5.19 (s, 2H, CH2), 2.33 (s, 3H, CH3),
2.17 (s, 3H, CH3). 13C NMR (101 MHz, CDCl3) δ 160.83, 158.43, 156.19, 155.19, 153.87, 143.08,
138.13, 135.32, 134.60, 130.69, 129.57, 129.16, 128.78, 126.72, 119.26, 118.67, 113.61, 110.89,
108.39, 108.32, 67.38, 62.17, 60.28, 24.15, 21.93. HRMS of [C31H25ClN4NaO4 + Na]+ (m/z):
575.1462; Calcd: 575.1448.
(Z)-1-(4-bromophenyl)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-
triazol-1-yl)-4-styrylazetidin-2-one (10e)
0
Yield-78%; mp 217-219 C; Cream amorphous solid; IR (neat) (νmax/cm-1): 1756, 1716, 1605,
1488;1H NMR (400 MHz, CDCl3) δ 7.82 (s, 1H, triazole-CH), 7.44 (dd, J = 27.3, 8.7 Hz, 4H,
ArH), 7.28 – 7.18 (m, 3H, ArH), 7.13 (d, J = 6.7 Hz, 2H, ArH), 6.77 (s, 1H, Coumarin-CH), 6.72
(d, J = 3.8 Hz, 2H, Vinyl-CH)., 6.62 (s, 1H, Coumarin-CH), 6.25 (d, J = 5.4 Hz, 1H, Vinyl-CH),
5.87 (dd, J = 16.3, 7.1 Hz, 1H, lactam-CH), 5.27 – 5.21 (m, 1H, lactam-CH), 5.19 (s, 2H, CH2),
2.33 (s, 3H, CH3), 2.17 (s, 3H, CH3). 13C NMR (101 MHz, CDCl3) δ 160.82, 158.49, 156.20,
155.19, 153.87, 143.07, 138.14, 135.79, 134.60, 132.49, 129.16, 128.78, 126.72, 123.64, 119.24,
25
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10.1002/cbdv.201900462
Chemistry & Biodiversity
118.97, 118.33, 113.61, 110.88, 108.38, 108.32, 67.38, 62.17, 60.26, 24.15, 21.94. HRMS of
[C31H25BrN4O4 + Na]+ (m/z): 619.0957; Calcd: 619.0952.
(Z)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-1-(4
iodophenyl)-4-styrylazetidin-2-one (10f)
0
Yield-87%; mp 212-214 C; White amorphous solid; IR (neat) (νmax/cm-1): 1757, 1717, 1605,
1487;1H NMR (400 MHz, CDCl3) δ 7.83 (s, 1H, triaozle-CH), 7.67 (d, J = 8.3 Hz, 2H, ArH), 7.35
– 7.18 (m, 5H, ArH), 7.12 (d, J = 6.5 Hz, 2H, ArH), 6.74 (d, J = 16.9 Hz, 2H, Vinyl-CH), 6.62
(s,1H, Coumarin-CH), 6.25 (d, J = 5.2 Hz, 1H, Vinyl-CH), 5.92 – 5.82 (m, 2H, lactam CH,
Coumarin-CH), 5.26 – 5.21 (m, 1H, lactam-CH), 5.19 (s, 2H, CH2), 2.33 (s, 3H, CH3), 2.17 (s, 3H,
13
CH3). C NMR (101 MHz, CDCl3) δ 160.81, 158.53, 156.20, 155.20, 153.85, 143.26, 143.06,
138.40, 138.11, 136.41, 134.60, 129.15, 128.78, 126.72, 123.61, 119.21, 113.62, 110.89, 108.31,
89.06, 67.37, 62.17, 60.18, 24.15, 21.94. HRMS of [C31H25IN4NaO4 + Na]+ (m/z): 667.0818;
Calcd: 667.0809.
(Z)-3-(4-(((4,5-dimethyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-4-styryl-1-
(p-tolyl)azetidin-2-one (10g)
0
Yield-81%; mp 220-222 C; White amorphous solid; IR (neat) (νmax/cm-1): 1753, 1713, 1610,
1492; 1H NMR (400 MHz, CDCl3) δ 7.84 (s, 1H, triazole-CH), 7.41 (d, J = 8.3 Hz, 2H, ArH), 7.25
– 7.06 (m, 7H, ArH), 6.76 (s, 1H, Coumarin-CH), 6.72 (s, 1H, Coumarin-CH), 6.62 (s, 1H, ArH),
6.24 (d, J = 5.4 Hz, 1H, Vinyl-CH), 5.86 (dd, J = 14.6, 5.8 Hz, 2H, lactam-CH), 5.25 – 5.20 (m,
13
1H, lactam-CH), 5.18 (s, 2H, CH2), 2.33 (d, J = 3.2 Hz, 6H, 2 CH3), 2.17 (s, 3H, Ar-CH3). C
NMR (101 MHz, CDCl3) δ 160.84, 158.26, 156.24, 155.19, 153.92, 143.13, 143.04, 137.65,
135.30, 134.84, 134.39, 129.91, 128.95, 128.71, 126.68, 123.62, 119.80, 117.38, 113.59, 110.84,
26
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10.1002/cbdv.201900462
Chemistry & Biodiversity
108.39, 108.31, 67.17, 62.18, 59.96, 24.15, 21.94, 21.01. HRMS of [C32H28N4NaO4 + Na]+ (m/z):
555.2008; Calcd: 555.1995.
(Z)-3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-1-phenyl-4-
styrylazetidin-2-one (10h)
0
Yield-78%; mp 172-174 C; Brown amorphous solid; IR (neat) (νmax/cm-1): 1759, 1709, 1611,
1
1387; H NMR (400 MHz, CDCl3) δ 7.87 (s, 1H, triazole-CH), 7.52 (d, J = 7.8 Hz, 2H, ArH),7.44
– 7.32 (m, 3H, ArH), 7.24 – 7.14 (m, 4H, ArH), 7.11 (dd, J = 6.6, 2.8 Hz, 2H, ArH), 6.87 – 6.79
(m, 2H, Coumarin-CH), 6.75 (d, J = 16.0 Hz, 1H, Vinyl-CH), 6.25 (d, J = 5.4 Hz, 1H, Vinyl-CH),
6.12 (s, 1H, Coumarin-CH), 5.86 (dd, J = 16.0, 7.4 Hz, 1H, lactam-CH), δ 5.25 (dd, J = 14.4, 8.1
Hz, 1H, lactam-CH), 5.20 (s, 2H, CH2), 2.36 (s, 3H, CH3). 13C NMR (101 MHz, CDCl3) δ 161.16,
160.99, 158.53, 155.05, 152.43, 143.44, 137.80, 136.87, 134.89, 129.42, 128.85, 128.65, 126.75,
125.66, 125.42, 123.70, 119.82, 117.44, 114.07, 112.29, 112.15, 102.12, 67.26, 62.04, 60.02,
18.65. HRMS of [C30H24N4O4 + Na]+ (m/z): 527.1695; Calcd: 527.1680.
(Z)-3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-4-styryl-1-(p-
tolyl)azetidin-2-one (10i)
0
Yield-76%; mp 198-200 C; White amorphous solid; IR (neat) (νmax/cm-1): 1757, 1711, 1610,
1387; 1H NMR (400 MHz, CDCl3) δ 7.86 (s, 1H, triazole-CH), 7.43 – 7.38 (m, 3H, ArH), 7.22 –
7.19 (m, 3H, ArH), 7.16 (d, J = 8.1 Hz, 2H, ArH), 7.11 (dd, J = 6.7, 2.9 Hz, 2H, ArH), 6.85 – 6.79
(m, 2H, Coumarin-CH), 6.73 (d, J = 16.0 Hz, 1H, Vinyl-CH), 6.23 (d, J = 5.4 Hz, 1H, Vinyl-CH),
6.13 (s, 1H, Coumarin-CH), 5.83 (dd, J = 16.0, 7.2 Hz, 1H, lactam-CH), 5.21 (dd, J = 4.0, 3.4 Hz,
13
1H, lactam-CH), 5.19 (s, 2H, CH2), 2.37 (s, 3H, CH3), 2.32 (s, 3H, CH3). C NMR (101 MHz,
CDCl3) δ 161.17, 161.00, 158.25, 155.05, 152.43, 143.39, 137.65, 135.24, 134.93, 134.42, 129.89,
27
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10.1002/cbdv.201900462
Chemistry & Biodiversity
128.80, 128.63, 126.73, 125.65, 123.69, 119.91, 117.38, 114.07, 112.29, 112.14, 102.13, 67.24,
62.04, 59.91, 21.00, 18.65. HRMS of [C31H26N4O4 + Na]+ (m/z): 541.1852; Calcd: 541.1846.
(Z)-1-(4-bromophenyl)-3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-
yl)-4-styrylazetidin-2-one (10j)
0
Yield-79%; mp 224-226 C; Cream amorphous solid; IR (neat) (νmax/cm-1): 1758, 1710, 1611,
1386; 1H NMR (400 MHz, CDCl3) δ 7.85 (s, 1H, triazole-CH), 7.52 – 7.45 (m, 2H, ArH), 7.41 (dt,
J = 5.3, 3.3 Hz, 3H, ArH), 7.22 (dd, J = 5.0, 1.8 Hz, 3H, ArH), 7.12 (dd, J = 6.5, 3.0 Hz, 2H, ArH),
6.83 (dd, J = 7.6, 2.2 Hz, 2H, Coumarin-CH), 6.74 (d, J = 16.0 Hz, 1H, Vinyl-CH), 6.25 (d, J =
5.5 Hz, 1H, Vinyl-CH), 6.13 (d, J = 1.1 Hz, 1H, Coumarin-CH), 5.84 (dd, J = 16.0, 7.5 Hz, 1H,
lactam-CH), 5.24 – 5.21 (m, 1H, lactam-CH), 5.20 (s, 2H, CH2), 2.37 (d, J = 1.0 Hz, 3H, CH3).
13C NMR (101 MHz, CDCl3) δ 161.22, 160.96, 158.48, 152.49, 143.51, 138.14, 135.81, 134.68,
132.47, 129.01, 128.70, 126.77, 125.68, 123.71, 119.31, 118.98, 118.29, 114.10, 112.30, 112.18,
102.11, 7, 67.45, 62.01, 60.21, 18.66. HRMS of [C30H23BrN4O4 + Na]+ (m/z): 605.0800; Calcd:
605.0778.
(Z)-1-(4-iodophenyl)-3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-
yl)-4-styrylazetidin-2-one (10k)
0
Yield-83%; mp 238-240 C; Cream amorphous solid; IR (neat) (νmax/cm-1): 1758, 1710, 1611,
1385; 1H NMR (400 MHz, CDCl3) δ 7.84 (s, 1H, triazole-CH), 7.67 (dd, J = 9.2, 2.1 Hz, 2H, ArH),
7.45 – 7.37 (m, 1H, ArH), 7.29 (d, J = 8.8 Hz, 2H, ArH), 7.22 (dd, J = 5.0, 1.8 Hz, 3H, ArH), 7.11
(dd, J = 6.5, 2.9 Hz, 2H, ArH), 6.83 (dd, J = 7.0, 2.3 Hz, 2H, Coumarin-CH), 6.73 (d, J = 16.0 Hz,
1H, Vinyl-CH), 6.24 (d, J = 5.5 Hz, 1H, Vinyl-CH), 6.13 (d, J = 0.9 Hz, 1H, Coumarin-CH), 5.83
(dd, J = 16.0, 7.5 Hz, 1H, lactam-CH), 5.22 (d, J = 6.7 Hz, 1H, lactam-CH), 5.20 (s, 2H, CH2),
13
2.37 (d, J = 0.8 Hz, 3H, CH3). C NMR (101 MHz, CDCl3) δ 160.96, 158.51, 138.39, 138.11,
28
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10.1002/cbdv.201900462
Chemistry & Biodiversity
136.43, 134.67, 129.01, 128.70, 126.77, 125.68, 123.68, 119.26, 119.23, 114.11, 112.31, 112.18,
102.12, 89.03, 77.34, 77.02, 76.70, 67.44, 62.01, 60.12, 18.66. HRMS of [C30H23IN4NaO4 + Na]+
(m/z): 653.0662; Calcd: 653.0652.
(Z)-1-(3-chlorophenyl)-3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-
yl)-4-styrylazetidin-2-one (10l)
Yield-80%; mp 236-238 0C;White amorphous solid; IR (neat) (νmax/cm-1): 1758, 1710, 1611, 1388;
1H NMR (400 MHz, CDCl3) δ 7.84 (s, 1H, triazole-CH), 7.50 – 7.45 (m, 2H, ArH), 7.44 – 7.38
(m, 1H, ArH), 7.35 – 7.31 (m, 2H, ArH), 7.24 – 7.20 (m, 3H, ArH), 7.12 (dt, J = 7.7, 3.8 Hz, 2H,
ArH), 6.86 – 6.80 (m, 2H, Coumarin-CH), 6.74 (d, J = 16.0 Hz, 1H, Vinyl-CH), 6.25 (d, J = 5.5
Hz, 1H, Vinyl-CH), 6.13 (d, J = 1.0 Hz, 1H, Coumarin-CH), 5.84 (dd, J = 15.9, 7.4 Hz, 1H, lactam-
CH), 5.23 (d, J = 6.5 Hz, 1H, lactam-CH), 5.20 (s, 2H, CH2), 2.37 (d, J = 1.0 Hz, 3H, CH3). 13C
NMR (101 MHz, CDCl3) δ 160.95, 158.42, 152.39, 143.52, 138.11, 135.35, 134.69, 130.65,
129.55, 129.01, 128.70, 126.77, 125.67, 123.64, 119.34, 118.67, 114.11, 112.34, 112.13, 102.13,
77.34, 77.02, 76.70, 67.44, 62.03, 60.23, 18.65. HRMS of [C30H23ClN4NaO4 + Na]+ (m/z):
561.1306; Calcd: 561.1299.
(Z)-3-(4-(((4-methyl-2-oxo-2H-chromen-7-yl)oxy)methyl)-1H-1,2,3-triazol-1-yl)-1-(3-
nitrophenyl)-4-styrylazetidin-2-one (10m)
Yield-87%; mp 214-2160C; White crystalline solid; IR (neat) (νmax/cm-1): 1759, 1709, 1611,
1388;1H NMR (400 MHz, CDCl3) δ 8.31 (t, J = 2.0 Hz, 1H), 8.03 (dd, J = 8.1, 1.6 Hz, 1H), 1H δ
7.91 (d, J = 9.4 Hz, 1H, ArH), 7.87 (s, 1H, triazole-CH)., 7.56 (t, J = 8.2 Hz, 1H, ArH), 7.42 (d, J
= 9.3 Hz, 1H, ArH), 7.25 – 7.19 (m, 3H, ArH), 7.14 (dd, J = 6.5, 2.8 Hz, 2H, ArH), 6.85 (d, J =
2.8 Hz, 1H, Vinyl-CH), 6.82 (d, J = 5.1 Hz, 2H, Coumarin-CH), 6.31 (d, J = 5.0 Hz, 1H, Vinyl-
CH), 6.13 (d, J = 1.0 Hz, 1H, Coumarin-CH), 5.88 (dd, J = 15.9, 7.7 Hz, 1H, lactam-CH), 5.32
29
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