Design and synthesis of aminocoumarin derivatives as DPP-IV inhibitors and anticancer agents

Article abstract of DOI:10.1016/j.bioorg.2018.05.008

DPP-IV “a moonlighting protein” has immerged as promising pathway to control Type 2 diabetes as well as found to play key role in earlier stages of cancer. Here we have reported design, synthesis and applications of aminocoumarin derivatives as DPP-IV inhibitors. Compounds have been synthesized and studied for their DPP-IV inhibition activity. Three compounds have shown moderate inhibition at 100 μM concentration. All compounds were also screened for their anticancer activity against A549 (Lung cancer cell line), MCF-7 (Breast cancer cell line) using MTT assay. One of the compounds has shown very good anticancer activity with IC₅₀ value 24 ± 0.1 nM against A549 cell line.

Full text of DOI:10.1016/j.bioorg.2018.05.008

Bioorganic Chemistry 79 (2018) 277–284
Contents lists available at ScienceDirect
Bioorganic Chemistry
journal homepage: www.elsevier.com/locate/bioorg
Design and synthesis of aminocoumarin derivatives as DPP-IV inhibitors and
anticancer agents
⁎
Rina Soni, Shubhangi S. Soman
Department of Chemistry, Faculty of Science, The M. S. University of Baroda, Vadodara 390 002, India
A R T I C L E I N F O
A B S T R A C T
Keywords:
DPP-IV “a moonlighting protein” has immerged as promising pathway to control Type 2 diabetes as well as
found to play key role in earlier stages of cancer. Here we have reported design, synthesis and applications of
aminocoumarin derivatives as DPP-IV inhibitors. Compounds have been synthesized and studied for their DPP-
IV inhibition activity. Three compounds have shown moderate inhibition at 100 µM concentration. All com-
pounds were also screened for their anticancer activity against A549 (Lung cancer cell line), MCF-7 (Breast
cancer cell line) using MTT assay. One of the compounds has shown very good anticancer activity with IC50
value 24 ± 0.1 nM against A549 cell line.
3
7
-Aminocoumarin derivatives
-Amino-4-methylcoumarin derivatives
DPP-IV inhibition
Anticancer activity
1. Introduction
various substituted amines with or without glycine type linkages for
their DPP-IV inhibition activity (Fig. 2). From our group, coumarin
Dipeptidyl peptidase-IV (DPP-IV) is multifunctional protein (a
moonlighting protein) also known as cluster of differentiation 26
CD26) [1]. DPP-IV enzyme is expressed on the surface of most cell
types, which is associated with immune regulation, signal transduction
and apoptosis. DPP-IV has been found to have roles in several important
diseases, including melanoma, autoimmune disease and AIDS.
It is an intrinsic membrane glycoprotein and serine exopeptidase
that cleaves proline dipeptidase from the N-terminus of polypeptidase
based compounds 3–6 have been reported with moderate to very good
DPP-IV inhibition activity [11,12]. In continuation to our work on DPP-
IV inhibitors [11–14], we have designed compounds containing 3-
aminocoumarin and 7-amino-4-methylcoumarin attached to different
amines with small linkage as DPP-IV inhibitors (Fig. 2) and screened for
their DPP-IV inhibition as well as their anticancer activity.
(
2. Results and discussion
[
2–3]. Most common feature in the design of substrate like inhibitors is
the bulkier groups in P2 site is liked through glycine moiety to proline
mimic at P1 site, both of them bulkier group and proline mimic show
interaction and binding at the active site of the enzyme.
2.1. Chemistry
3-Amino coumarin 9 was prepared in two steps by Perkin reaction
of 2-hydroxy-benzaldehyde 7 with N-acetyl glycine to give compound 8
followed by deprotection using conc. HCl in ethanol [15]. 3-Amino-
coumarin 9 was failed to react with bromoacetamide derivatives under
DPP-IV plays a major role in glucose metabolism and different
classes of DPP-IV inhibitors (Fig. 1) have been marketed to treat Type 2
Diabetes such as Vildagliptin, Sitagliptin, etc [4–6]. DPP-IV also acts on
tumor immunity by modifying chemokines and cytokines, which are
playing important role in tumor microenvironment [7]. Thus, DPP-IV
has been involved in regulating cancer, although exact role of DPP-IV is
dependent on type of cancer also. DPP-IV has been explored to find its
relation in regulation and prevention of inflammation during develop-
ment stages of cancer related with metabolism [8–10].
There are only few reports on synthetic coumarin derivatives as
DPP-IV inhibitors to treat Type 2 diabetes. The pharmacological prop-
erties and applications of coumarins can be varied by varying type and
position of substitution on coumarin rings, based on this we have de-
veloped and screened series of 3- and 7-amino coumarins linked to
2 3 2 3
different conditions such as K CO , Cs CO , LiOH in DMF. So, 3-amino
coumarin 9 was reacted with bromoacetyl bromide to give 3-bromoa-
cetamide coumarin 10. Bromo group of 3-bromoacetamide coumarin
10 was replaced by different amines using base in DMF to give desired
3-substituted coumarin compounds 11a-f (Scheme 1). All synthesized
compounds were characterized by 1H NMR, C NMR, ESI-MS, IR and
etc.
13
The IR spectrum of compound 11c exhibited two strong bands at
−1
−1
3331 cm
spectively. Cyano group showed stretching frequency at 2241 cm
Carbonyl group of coumarin lactone ring exhibited stretching frequency
and 3298 cm
amide N-H stretching and vibrations re-
−
1
.
⁎
Corresponding author.
E-mail address: shubhangiss@rediffmail.com (S.S. Soman).
https://doi.org/10.1016/j.bioorg.2018.05.008
Received 23 March 2018; Received in revised form 8 May 2018; Accepted 10 May 2018
0045-2068/ © 2018 Elsevier Inc. All rights reserved.

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
Fig. 1. DPP-IV inhibitors as antidiabetic agents.
Fig. 2. Design of coumarin derivatives as DPP-IV inhibitors.
−
1
and that of amide carbonyl in range of 1689 cm−1. 1
at 1714 cm
H
bromoacetyl bromide to give bromoacetamide derivative of coumarin
16. Bromo group of 7-bromoacetamide-4-methyl coumarin 16 was re-
placed by different amines using base in DMF to give desired 7-sub-
stituted-4-methyl coumarin compounds 17a-l (Scheme 2). All synthe-
sized compounds were characterized by H NMR, C NMR, ESI-MS, IR
and etc.
NMR of compound 11c showed signals for methylene protons from
pyrrolidine ring in the region of 2.04–2.41 ppm and 2.79–3.13 ppm,
while bridging methylene group adjacent to amide observed at
.51–3.64 ppm with germinal coupling of 16.8 Hz. Signal for proton at
chiral carbon next to cyano group was observed as multiplet at
.97–4.00 ppm. Aromatic protons were observed as multiplet from 7.30
to 7.54 ppm and singlet at 8.69 ppm. Amide proton was appeared as
singlet at 9.40 ppm. In C NMR compound 11c showed peaks in ali-
phatic carbons in region of 22.5–56.3 ppm and aromatic carbons from
16.4 to 150.1 ppm. Cyano group showed signal at 117.7 ppm, the
1
13
3
3
The IR spectrum of compound 17c exhibited strong band at
−
1
3286 cm
for amide N-H stretching. Cyano group showed stretching
1
3
−1
frequency at 2220 cm . Carbonyl group of coumarin lactone ring ex-
hibited stretching frequency at 1700 cm
in range of 1685 cm
−
1
and that of amide carbonyl
−1 1
1
. H NMR of compound 17c showed signals for
amide carbon was observed at 158.5 ppm while carbonyl carbon of
methylene protons from pyrrolidine ring in the region of 2.09–2.39 ppm
and 2.92–3.03 ppm. Methyl group was observed as doublet at 2.43 ppm
with coupling value of 0.8 Hz, while bridging methylene group adjacent
to amide observed at 3.56 ppm as singlet. Signal for proton at chiral
carbon next to cyano group was observed as multiplet at
3.89–3.92 ppm. Aromatic protons observed as a singlet at 6.23 ppm and
as a multiplet from 7.56 to 7.63 ppm. Amide proton appeared as singlet
at 8.84 ppm. 13C NMR of compound 17c showed peaks in aliphatic
region from 18.6 to 57.3 ppm and aromatic carbons from 107.2 to
154.2 ppm. Cyano group showed signal at 118.21 ppm, the amide
carbon observed at 161.0 ppm while carbonyl carbon of coumarin
lactone ring at 167.5 ppm. ESI-MS analysis of compound 17c gave
[M + H]+ peak at 312.1 confirming the formation of desired com-
pound.
coumarin lactone ring at 168.5 ppm. ESI-MS analysis of compound 11c
+
gave [M + H] peak at 297.7 confirming the formation of desired
compound.
Amino group of 3-Aminophenol 12 was protected as ethyl carba-
mate group using ethyl chloroformate to give compound 13. Compound
13 was reacted with ethyl acetoacetate under Pechmann condition to
give compound 14. 7-Amino-4-methyl coumarin 15 was obtained by
carbamate deprotection of compound 14 under acidic condition [16].
7-Amino-4-methyl coumarin 15 was reacted with bromoacetamide
derivatives under different conditions such as K CO , Cs CO , LiOH in
2
3
2
3
DMF. None of the conditions worked to give desired product as amino
group at 7th position was found to be less reactive. As modification of
designed compounds, 7-Amino-4-methyl coumarin 15 was reacted with
278

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
Scheme 1. Synthesis of 3-aminocoumarin derivatives.
Scheme 2. Synthesis of 7-amino-4-methylcoumarin derivatives.
2
2
.2. Biological evaluation
concentrations (Tables 1 and 2).
3-Aminocoumarin derivatives 11a–c with substituted pyrrolidine
.2.1. DPP-IV inhibition activity
Preliminary in vitro DPP-IV inhibition assay was performed to screen
were screened for their DPP-IV inhibition activity and compared with
compound 11d containing pyrrolidine group (Table 1). Similarly DPP-
IV inhibition activity of compound 11e with piperidine-2-carboxamide
was compared with compound 11f with piperidine group. Compound
compounds 11a–f and 17a–l to study the effect of different amides at
the P1 site on their inhibition potential at 25 µM and 100 µM
279

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
Table 1
moiety can affect the orientation and possible interaction of resulting
compound in binding pockets of DPP-IV enzyme. Keeping this in mind
compounds 17a–f were synthesized and compared for their in vitro DPP-
IV inhibition activity (Table 2). However, pyrrolidine-2carboxylate
containing compound 17a showed moderate activity with 33.80% in-
hibition as compared to compound 17d having pyrrolidine group with
DPP-IV inhibition activity of 3-aminocoumarin derivatives 11a–f.
% DPP-4 enzyme inhibitiona
Compound
2
5 µM
100 µM
1
1
1
1
1
1
1a
1b
1c
1d
1e
1f
8.82
21.87
24.17
21.80
28.13
35.08
6.30
2
1.61% inhibition at 100 µM concentration (Table 2). Replacement of
10.91
11.09
14.86
17.64
3.80
ester group to more stable and interactive amide and cyano groups in
compound 17b and 17c respectively, resulted in the drop of DPP-IV
inhibition activity. Any major change on DPP-IV inhibition activity was
not observed on enlargement of ring to piperdine in compounds 17e-f
Table 2).
As a part of further modification, compound 17g–l attached to
various heterocyclic rings were synthesized and screened for their ac-
tivity (Table 2). Compound 17g with 3-methylpiperidine showed
moderate activity with 42.1% DPP-IV inhibition. Further, change of
methyl position from 3rd to 4th in compounds 17h with 4-methylpi-
peridine and compound 17i with N-methylpiperazine, showed good
activity with 56.9% and 56.3% DPP-IV inhibition respectively. Re-
placement of N-methylpiperazine by N-phenylpiperazine group in
compound 17j and bulkier tetrahydroisoquinoline in compound 17l,
resulted in drop of activity with 46.6% and 43.6% DPP-IV inhibition
respectively. However, compound 17k with morpholine retained DPP-
IV inhibition activity with 54.4% inhibition. Compound 17i showed
moderate activity with 52.90% inhibition activity at 25 µM concentra-
tion as compared to standard drug Vildagliptin with 56.3% inhibition
activity at 0.1 µM concentration.
at 0.1µM
Vildagliptin
56.3%
(
a
DPP-IV inhibitory activity determined by fluorescence-based assay was
measured using Spectra Max fluorometer (Molecular Devices, CA). Values of %
inhibition are mean of three independent determinations at 25 and 100 μM
concentrations of the test samples.
Table 2
DPP-IV inhibition activity of 7-amino-4-methylcoumarin derivatives 17a–l.
% DPP-4 enzyme inhibitiona
Compound
% DPP-4 enzyme
Compound
inhibitiona
2
5 µM
100 µM
25 µM
100 µM
17a
17b
17c
17d
17e
17f
19.35
13.60
4.64
7.00
14.36
9.80
33.80
25.24
11.10
21.61
23.84
22.90
17g
17h
17i
17j
17k
17l
39.10
39.70
52.90
44.20
23.80
30.90
42.10
56.90
56.30
46.60
54.40
43.60
at 0.1µM
Vildagliptin 56.3%
2.2.2. Anticancer activity by MTT assay
Current chemotherapy for treating cancer is based on cytotoxic
a
DPP-IV inhibitory activity determined by fluorescence-based assay was
measured using Spectra Max fluorometer (Molecular Devices, CA). Values of %
inhibition are mean of three independent determinations at 25 an 100 μM
concentrations of the test samples.
drugs with low specificity towards cancer cells. Due to that it is ne-
cessary to develop more specific anticancer agents. Compounds 11a–f
and 17a–l were screened for their anticancer activity against A549
(
Lungs cancer cell line), MCF-7 (Breast cancer cell line) using MTT
assay (Table 3).
11a containing pyrrolidine-2-carxoylate group showed poor activity
Pyrrolidine-3-carboxylate derivatives 11a showed moderate activity
against both tested cell lines. Replacement of ester group by carbox-
amide in compound 11b resulted in loss of activity against A549 cell
line, but increase of activity against MCF 7 cell line. Conversion of
amide to more stable cyano group on compound 11c resulted in mod-
erate activity against both tested cell lines. However, compound 11d
with pyrrolidine moiety showed good activity with IC50 values
with 21.87% DPP-IV inhibition at 100 µM. Change of ester group to
amide group in compound 11b resulted in slight increase in activity
with 24.17% DPP-IV inhibition at 100 µM, while replacement of amide
group by cyano group in compound 11c resulted in similar inhibition
activity with 21.80% DPP-IV inhibition at 100 µM concentration
(
Table 1). DPP-IV inhibition activity of compounds 11a-c having sub-
stituted pyrrolidine moiety can be easily compared with compound 11d
with pyrrolidine moiety with 28.13% inhibition for DPP-IV. Compound
1.80 ± 0.084 and 0.355 ± 0.0082 µM against A549 and MCF7 cell
lines respectively. Compound 11e with piperidine carboxamide showed
moderate activity against MCF7 cell line and poor activity against A549
cell line, reversal of activity was observed for compound 9f with pi-
peridine moiety (Table 3).
11e with piperidine-2-caroboxamide moiety showed moderate activity
with 35.08% DPP-IV inhibition as compared to compound 11f con-
taining piperidine moiety with 6.3% inhibition at 100 µM concentration
(
Table 1).
The change in amino position from 3rd to 7th position in coumarin
7-Amino coumarin derivatives 17a with ester and 17b with amide
group showed very poor anticancer activity, on other hand compound
Table 3
Anticancer activity against A549 (Lungs cancer cell line), MCF-7 (Breast cancer cell line) for compounds 11a–f and 17a–l.
Compound
IC50 µMa
Compound
IC50 µMa
A549
MCF7
A549
MCF7
1
1
1
1
1
1
1
1
1
1a
1b
1c
1d
1e
1f
7a
7b
7c
3.09 ± 1.03
24.3 ± 1.56
2.88 ± 0.85
1.80 ± 0.084
94.3 ± 4.05
2.53 ± 0.52
NA
43.08 ± 5.12
4.38 ± 0.84
11.13 ± 0.083
9.27 ± 1.02
5.40 ± 0.19
4.77 ± 0.045
0.355 ± 0.0082
2.77 ± 0.072
18.41 ± 1.32
430.09 ± 9.37
NA
17d
17e
17f
17g
17h
17i
17j
17k
17l
4.02 ± 0.095
0.097 ± 0.0023
19.49 ± 1.65
27.32 ± 2.11
9.34 ± 1.12
NA
2.43 ± 0.54
17.45 ± 1.65
0.024 ± 0.0001
4.80 ± .082
13.70 ± 1.78
3.35 ± 0.037
2.68 ± 0.037
42.24 ± 3.06
2.15 ± 0.093
2.80 ± 0.045
5.49 ± 0.082
4.85 ± 0.0389
4.43 ± 0.09
45.04 ± 1.02
Fluorouracil
a
IC50 values were determined based on MTT assay using GraphPad Prism software. NA = Not active.
280

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
1
7c with cyano and 17d with pyrrolidine moiety showed moderate
(45 mL) and conc. HCl (5 mL) was refluxed for 1 h to give clear solution.
The completion of reaction was checked by TLC. The reaction was
cooled to room temperature and concentrated on a rotavapor to give
residue. The residue was dissolved in water, filtered to remove in-
activity against both tested cell lines (Table 3). Piperidine carboxamide
containing compound 17e showed very good activity with IC50 value
0
.097 ± 0.0023 µM against A549 cell line, but showed moderate ac-
tivity against MCF7 cell line with IC50 value 13.70 ± 1.78. Compound
7f-h showed moderate to poor activity against both tested cell lines.
3
soluble impurity. The filtrate was neutralized with sat. NaHCO solu-
1
tion to give yellow solid. The solid was filtered washed with water,
dried and recrystallized from ethanol to give compound 9 as a yellow
Compound 17i remained inactive against A549 cell line and showed
moderate activity against MCF-7 cell line. Replacement of N-methyl
group of compound 17i by N-phenyl group in 17j, resulted in moderate
activity against both tested cell lines. However, on replacing N-phenyl
piperazine to morpholine in compound 17k resulted in the drop of
activity against both tested cell lines. Terahydroisoquinoline containing
compound 17l showed very good activity against A549 cell line with
IC50 value 0.024 ± 0.0001 µM as compared to standard drug fluor-
ouracil with IC50 value 11.13 ± 0.083 µM (Table 3).
1
solid. Yield: 61%; M.P.: 138–140 °C, Lit M.P.: 129 °C [15]. H NMR
(400 MHz, CDCl ): δ 4.28 (s, 2H), 6.73 (s, 1H), 7.20–7.24 (m, 1H),
3
7.27–7.33 (m, 3H).
4.2.3. Preparation of 2-bromo-N-(2-oxo-2H-chromen-3-yl)acetamide (10)
To
a solution of 3-aminocoumarin 9 (10.0 mmol) in di-
chloromethane (DCM) (25 mL) was added triethylamine (TEA)
(15.0 mmol) and stirred for 5–10 min. To this bromoacetyl bromide
(
12.0 mmol) was added dropwise over a period of 10 min. The resulting
3
. Conclusion
solution was stirred at room temperature for 2 h. The reaction mixture
filtered, washed with DCM, water and acetone to give compound 10 as
a pale white solid. The substituted bromoacetamide 10 thus obtained,
used directly for next step without any purification. Yield: 89%; M.P:
We have designed 3-aminocoumarin derivatives 11a–f and 7-
amino-4-methylcoumarin derivatives 17a–l for DPP-IV inhibition.
Compounds 11a–c and compounds 17a–c containing proline mimic,
showed poor DPP-IV inhibition activity at 100 µM concentration.
Compound 17i with N-methylpiperazine moiety, showed 52.90% of
DPP-IV inhibition at 25 µM concentration. DPP-IV inhibition plays a key
role for treating diabetes as well as cancer. All compounds were
screened for their anticancer activity using MTT assay. Interestingly,
compound 17l showed very good activity against A549 cell line with
IC50 value 24 ± 0.1 nM as compared to standard drug fluorouracil
with IC50 value 11.13 ± 0.083 µM. These studies provide insight for
designing of new potent DPP-IV inhibitors containing coumarin moiety.
208–210 °C; 1H NMR (400 MHz, DMSO‑d
): δ 4.29 (s, 2H), 7.33–7.37
(m, 1H), 7.39–7.41 (m, 1H), 7.50–7.55 (m, 1H), 7.74 (dd, J = 8.0,
1.6 Hz, 1H), 8.65 (s, 1H), 10.25 (s, 1H); 13C NMR (100 MHz, DMSO‑d
):
6
6
δ 30.28, 116.35, 119.79, 124.67, 124.95, 125.49, 128.57, 130.46,
150.29, 157.79, 167.03.
4.2.4. General procedure for the synthesis of compounds 11a–e
To a solution of compound 10 (0.5 g) and substituted amine (1.1 eq)
in DMF (20 mL) was added triethylamine (1.5 eq). The resulting mix-
ture was stirred at room temperature for 16 h and then poured into cold
water. The aqueous layer thus obtained was extracted using ethyl
acetate and/or dichloromethane (checked by TLC) and solvent evapo-
rated to give crude product. The crude compound was purified by
column chromatography.
4
. Experimental
4.1. Chemistry
Reagent grade chemicals and solvents were purchased from com-
4.2.5. Characterization data for compounds 11a-f
mercial supplier and used after purification. TLC was performed on
silica gel F254 plates (Merck). Acme’s silica gel (60–120 mesh) was
used for column chromatographic purification. All reactions were car-
ried out in nitrogen atmosphere. Melting points are uncorrected and
were measured in open capillary tubes, using a Rolex melting point
apparatus. IR spectra were recorded as KBr pellets on Perkin Elmer RX 1
4.2.5.1. (S)-ethyl
1-(2-oxo-2-((2-oxo-2H-chromen-3-yl)amino)ethyl)
pyrrolidine-2-carboxylate (11a). White solid, Yield: 49%; M.P:
82–84 °C; IR (KBr) 3180, 3155, 2983, 2920, 2754, 1747, 1718, 1697,
1624, 1606, 1537, 1359, 1209, 1033, 979, 758 cm
−1
1
;
H NMR
(400 MHz, CDCl ): δ 1.26 (t, J = 7.2 Hz, 3H), 1.92–2.10 (m, 3H),
3
2.24–2.27 (m, 1H), 2.64–2.67 (m, 1H), 3.21–3.25 (m, 1H), 3.34 (d,
J = 16.8 Hz, 1H), 3.52–3.56 (m, 1H), 3.62 (d, J = 17.2 Hz, 1H), 4.20
(q, J = 7.2 Hz, 2H), 7.28–7.34 (m, 2H), 7.42–7.44 (m, 1H), 7.51 (dd,
J = 7.6, 0.8 Hz, 1H), 8.70 (s, 1H), 10.07 (s, 1H); 13C NMR (100 MHz,
1
13
spectrometer. H NMR and C NMR spectral data were recorded on
Advance Bruker 400 spectrometer (400 MHz) with CDCl or DMSO‑d
3
6
as solvent and TMS as internal standard. J values are in Hz. Mass
spectra were determined by ESI-MS, using a Shimadzu LCMS 2020
apparatus. Elemental analyses were recorded on Thermosinnigan Flash
CDCl
119.90, 123.17, 124.02, 124.97, 127.69, 129.49, 150.18, 158.38,
171.05, 173.50; Anal. Calc. for C18 ; C, 62.78; H, 5.85; N,
3
): δ 14.23, 24.04, 29.57, 54.41, 58.53, 61.04, 65.71, 116.33,
11–12 series EA. All reactions were carried out under nitrogen atmo-
20 2 5
H N O
+
sphere. (S) ethyl pyrrolidine-2-carboxylate, (S) pyrrolidine-2-carbox-
amide, (S) pyrrolidine-2-carbonitrile and (S) piperidine-2-carboxamide
were prepared according to literature method [17].
8.13; found: C, 62.83; H, 5.89; N, 8.17%; ESI-MS: 345.0 [M + H] .
4.2.5.2. (S)-1-(2-oxo-2-((2-oxo-2H-chromen-3-yl)amino)ethyl)
pyrrolidine-2-carboxamide (11b). White solid, Yield: 52%; M.P:
202–204 °C; IR (KBr) 3387, 3309, 3200, 2970, 2872, 1718, 1687,
4
4
1
.2. Synthesis of 3-amino coumarin derivatives
−1
1
1633, 1539, 1352, 1220, 1209, 1105, 906, 758 cm
;
H NMR
.2.1. Preparation of N-(2-oxo-2H-chromen-3-yl)acetamide (8)
A mixture of 2-hydroxy-benzaldehyde 7 (9.15 mL, 86.87 mmol,
.0 eq), N-acetyl glycine (10.0 g, 86.87 mmol, 1.0 eq) and NaOAc
(400 MHz, CDCl ): 1.94–2.07 (m, 3H), 2.30–2.36 (m, 1H),
3
δ
2.54–2.60 (m, 1H), 3.33–3.40 (m, 3H), 3.66 (d, J = 16.4 Hz, 1H),
5.72 (br s, 1H), 6.88 (br s, 1H), 7.31–7.36 (m, 2H), 7.46–7.50 (m, 1H),
1
3
(
28.04 g, 341.88 mmol, 4.0 eq) into an acetic anhydride (40 mL) was
7.53 (dd, J = 7.6, 1.2 Hz, 1H), 8.67 (s, 1H), 9.29 (s, 1H); C NMR
(100 MHz, CDCl ): δ 24.50, 30.60, 55.28, 59.66, 68.28, 116.45, 119.67,
123.43, 123.77, 125.29, 127.86, 129.93, 150.08, 158.89, 169.60,
176.44; Anal. Calc. for C16 ; C, 60.94; H, 5.43; N, 13.33;
found: C, 61.02; H, 5.48; N, 13.45%; ESI-MS: 315.8 [M + H]
°
heated at 120 C for 5 h. The resulting brown suspension was allowed to
cool to room temperature. Resulting solid was diluted with cold water,
scratched, filtered, washed with water and ethyl acetate (15 mL). The
solid was recrystallized from ethanol to give compound 8 as light
yellow solid. Yield: 47%; M.P.: 208–210 °C, Lit M.P.: 203–204 °C [15].
3
17 3 4
H N O
+
.
4.2.5.3. (S)-2-(2-cyanopyrrolidin-1-yl)-N-(2-oxo-2H-chromen-3-yl)
4.2.2. Preparation of 3-amino-2H-chromen-2-one (9)
acetamide (11c). White solid, Yield: 49%; M.P: 122–124 °C; IR (KBr)
A suspension of compound 7 (5.0 g, 24.63 mmol) into ethanol
3331, 3298, 2956, 2820, 2241, 1714, 1689, 1627, 1514, 1367, 1147,
281

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
with water, dried and recrystallized from ethanol to give compound 14
as an off white solid. Yield: 85%; M.P.: 188–190 °C, Lit M.P.:
184–187 °C [16].
23, 777, 759 cm 1; 1H NMR (400 MHz, CDCl
.23–2.30 (m, 1H), 2.33–2.41 (m, 1H), 2.79–2.86 (m, 1H), 3.08–3.13
−
): δ 2.04–2.14 (m, 2H),
9
2
3
(
m, 1H), 3.54 (d, J = 17.2 Hz, 1H), 3.62 (d, J = 16.8 Hz, 1H), 3.98 (dd,
J = 7.6, 2.4 Hz, 1H), 7.30–7.35 (m, 2H), 7.45–7.49 (m, 1H), 7.53 (dd,
1
3
J = 7.6, 1.2 Hz, 1H), 8.69 (s, 1H), 9.40 (s, 1H); C NMR (100 MHz,
CDCl ): δ 22.50, 30.03, 52.25, 54.23, 56.36, 116.41, 117.72, 119.67,
23.43, 123.81, 125.21, 127.86, 129.89, 150.15, 158.57, 168.55; Anal.
Calc. for C16 ; C, 64.64; H, 5.09; N, 14.13; found: C, 64.52; H,
4.3.3. Preparation of 7-amino-4-methyl-2H-chromen-2-one 15
3
2 4
A mixture of compound 14 (4.0 g, 16.194 mmol) into H SO : AcOH
1
(1:1, 15 mL) was refluxed at 104–106 °C for 1 h. The completion of
reaction was checked by TLC. The reaction was cooled to room tem-
perature and poured into ice cold water. The aqueous layer was allowed
to stand overnight to give solid. The solid was filtered, dissolved in
water. The aqueous layer was neutralized to pH 7 using dil. NaOH so-
lution to give solid. The solid was filtered washed with water, dried and
recrystallized from ethanol to give compound 15 as an off white solid.
15 3 3
H N O
+
5.18; N, 14.21%; ESI-MS: 297.7 [M + H]
.
4
(
.2.5.4. N-(2-oxo-2H-chromen-3-yl)-2-(pyrrolidin-1-yl)acetamide
11d). Off white solid, Yield: 80%; M.P: 126–128 °C; IR (KBr) 3289,
966, 2912, 2872, 2803, 1723, 1692, 1627, 1511, 1485, 1362, 1145,
): δ 1.88–1.91 (m, 4H),
.70–2.73 (m, 4H), 3.35 (s, 2H), 7.31–7.35 (m, 2H), 7.43–7.47 (m, 1H),
.53 (dd, J = 7.6, 1.2 Hz, 1H), 8.71 (s, 1H), 9.88 (s, 1H); C NMR
): δ 24.12, 54.58, 59.73, 116.35, 119.91, 123.23,
23.91, 125.04, 127.72, 129.54, 150.15, 158.55, 171.02; Anal. Calc. for
; C, 66.16; H, 5.92; N, 10.29; found: C, 66.02; H, 5.73; N,
2
9
2
7
−
1
1
1
23, 845, 773 cm ; H NMR (400 MHz, CDCl
3
Yield: 71%; M.P.: 218–220 °C, Lit M.P.: 220–224 °C [16]. H NMR
(400 MHz, DMSO‑d ): δ 2.29 (d, J = 0.8 Hz, 3H), 5.89 (d, J = 1.2 Hz,
6
1
3
1H), 6.11 (s, 2H), 6.39 (d, d, J = 2.0 Hz, 1H) 6.56 (dd, J = 8.4, 2.0 Hz,
1H), 7.39 (d, J = 8.4 Hz, 1H).
(
100 MHz, CDCl
3
1
C
15
H
16
N
2
O
3
4.3.4. Preparation
acetamide (16)
of
2-bromo-N-(4-methyl-2-oxo-2H-chromen-7-yl)
+
10.36%; ESI-MS: 273.1 [M + H] .
To a solution of 7-amino-4-methyl coumarin 15 (10.0 mmol) in di-
chloromethane (DCM) (25 mL) was added triethylamine (TEA)
(15.0 mmol) and stirred for 5–10 min. To this bromoacetyl bromide
(12.0 mmol) was added dropwise over a period of 10 min. The resulting
solution was stirred at room temperature for 2 hrs. The reaction mixture
filtered, washed with DCM, water and acetone to give compound 16 as
an off white solid. The substituted bromoacetamide 16 thus obtained,
4
2
.2.5.5. (S)-1-(2-oxo-2-((2-oxo-2H-chromen-3-yl)amino)ethyl)piperidine-
-carboxamide (11e). White solid, Yield: 58%; M.P: 204–206 °C; IR
(
KBr) 3387, 3327, 2945, 2922, 2870, 2854, 2823, 1728, 1666, 1627,
−1
1
1
510, 1487, 1375, 1363, 1321, 1141, 775, 758 cm
): 1.30–1.33 (m, 1H), 1.52–1.68 (m, 4H),
.76–1.79 (m, 1H), 2.26–2.32 (m, 1H), 2.99–2.93 (m, 2H), 3.02–3.05
;
H NMR
(
400 MHz, DMSO‑d
6
1
(
m, 1H), 3.25 (d, J = 16.8 Hz, 1H), 7.04 (s, 1H), 7.32–7.36 (m, 1H),
used directly for next step without any purification. Yield: 93%; M.P:
1
7
8
2
1
.41–7.43 (m, 2H), 7.49–7.53 (m, 1H), 7.73 (dd, J = 7.6, 1.2 Hz, 1H),
228–230 °C; H NMR (400 MHz, DMSO‑d
4.09 (s, 2H), 6.27 (d, J = 0.8 Hz, 1H), 7.45 (dd, J = 8.8, 2.0 Hz, 1H),
7.71–7.73 (m, 2H), 10.82 (s, 1H); 13C NMR (100 MHz, DMSO‑d
): δ
6
): δ 2.38 (d, J = 0.8 Hz, 3H),
1
3
.58 (s, 1H), 10.10 (s, 1H); C NMR (100 MHz, DMSO‑d
6
): δ 25.88,
5.75, 30.46, 52.73, 59.91, 65.18, 116.43, 119.99, 122.70, 124.36,
25.51, 128.34, 130.09, 150.14, 158.12, 170.83, 174.73; Anal. Calc. for
6
18.44, 30.74, 106.18, 113.04, 115.63, 115.92, 126.54, 142.25, 153.46,
154.02, 160.38, 165.97.
17 19 3 4
C H N O ; C, 62.00; H, 5.81; N, 12.76; found: C, 62.18; H, 5.94; N,
1
+
2.55%; ESI-MS: 330.10 [M + H]
.
4.3.5. General procedure for the synthesis of compounds 17a–l
4.2.5.6. N-(2-oxo-2H-chromen-3-yl)-2-(piperidin-1-yl)acetamide
To a solution of compound 16 (0.5 g) and substituted amine (1.1 eq)
(
2
7
11f). White solid, Yield: 85%; M.P: 154–156 °C; IR (KBr) 3257, 2943,
in DMF (20 mL) was added triethylamine (1.5 eq). The resulting mix-
ture was stirred at room temperature for 16 h and then poured into cold
water. The aqueous layer thus obtained was extracted using ethyl
acetate and/or dichloromethane (checked by TLC) and solvent evapo-
rated to give crude product. The crude compound was purified by
column chromatography.
924, 2860, 2804, 2785, 1722, 1680, 1624, 1506, 1361, 1145, 9455,
−
1 1
77, 758 cm ; H NMR (400 MHz, CDCl ): δ 1.48 (br s, 2H), 1.69–1.72
3
(
1
(
1
m, 4H), 2.55 (br s, 4H), 3.13 (s, 2H), 7.30–7.35 (m, 2H), 7.43–7.46 (m,
H), 7.51 (d, J = 7.6 Hz, 1H), 8.69 (s, 1H), 10.10 (s, 1H); 13C NMR
100 MHz, CDCl ): δ 23.63, 26.16, 54.99, 62.77, 116.35, 119.91,
22.99, 123.95, 125.01, 127.68, 129.48, 150.17, 158.49, 150.94;
; C, 67.12; H, 6.34; N, 9.78; found: C,
3
Anal. Calc. for C16
H
18
N
2
O
3
4.3.6. Characterization data for compounds 17a-l
+
6
4
4
7.32; H, 6.14; N, 9.92%; ESI-MS: 286.7 [M]
.
4.3.6.1. (S)-Ethyl
1-(2-((4-methyl-2-oxo-2H-chromen-7-yl)amino)-2-
oxoethyl)pyrrolidine-2-carboxylate (17a). White solid, Yield: 72%; M.P:
76–78 °C; IR (KBr) 3219, 3180, 2983, 1749, 1732, 1697, 1620, 1587,
.3. Synthesis of 7-substituted-4-methyl coumarin derivatives
−
1
1
535, 1423, 1392, 1367, 1261, 1151, 1037, 1020, 844, 821, 754 cm
;
1
.3.1. Preparation of ethyl (3-hydroxyphenyl)carbamate (13)
3
H NMR (400 MHz, CDCl ): δ 1.26 (t, J = 7.2 Hz, 3H), 1.89–1.96 (m,
3
-Aminophenol 12 (21.8 g, 20.0 mmol, 1.0 eq) into ethyl acetate
250 mL) was refluxed for 30 min. To this ethyl chloroformate (17.5 mL,
0.0 mmol, 1.0 eq) was added dropwise. The resulting mixture was
2H), 2.00–2.06 9 m, 1H), 2.25–2.31 (m, 1H), 2.41 (d, J = 0.8 Hz, 3H),
2.68 (q, J = 8.4 Hz, 1H), 3.28–3.32 (m, 1H), 3.41 (d, J = 17.6 Hz, 1H),
3.48–3.53 (m, 2H), 4.13–4.23 (m 2H), 6.18 (s, 1H), 7.53 (d, J = 8.8 Hz,
1H),7.63 (dd, J = 8.8, 2.0 Hz, 1H), 7.76 (d, J = 2.0 Hz, 1H), 10.31 (s,
(
2
refluxed for 1 h. The reaction mixture was cooled to room temperature,
filtered off and washed with ethyl acetate (2 × 50 mL). The filtrate was
concentrated on a rotavapor to give an oil, which was solidified on
standing. The solid was scratched in pet ether, filtered and dried to give
compound 13 as a white solid. Yield: 48%; M.P.: 92–94 °C, Lit M.P.:
1H); 13C NMR (100 MHz, CDCl
): δ 14.23, 18.63, 24.87, 30.73, 55.50,
60.05, 61.42, 66.79, 106.90, 113.17, 115.50, 115.87, 125.09, 141.51,
152.31, 154.31, 161.24, 170.39, 175.39; Anal. Calc. for C19 ; C,
63.68; H, 6.19; N, 7.82; found: C, 63.42; H, 6.04; N, 7.69%; ESI-MS:
359.0 [M + H]+
3
22 2 5
H N O
9
4–96 °C [16].
.
4
.3.2. Preparation of ethyl (4-methyl-2-oxo-2H-chromen-7-yl)carbamate
4.3.6.2. (S)-1-(2-((4-methyl-2-oxo-2H-chromen-7-yl)amino)-2-oxoethyl)
pyrrolidine-2-carboxamide (17b). Off white solid, Yield: 51%; M.P:
134–136 °C; IR (KBr) 3404, 3304, 2970, 2872, 1718, 1697, 1685,
(14)
To a cold solution of compound 13 (5.0 g, 27.624 mmol, 1.0 eq) into
−1 1
ethyl acetoacetate (4.18 mL, 33.149 mmol, 1.2 eq) was added a cold
solution of 70% ethanolic sulphuric acid (70 mL). The resulting mixture
was stirred at room temperature for 14 h. The reaction mixture was
poured into ice cold water to give solid. The solid was filtered, washed
1618, 1585, 1525, 1406, 1396, 1263, 1151, 1070, 944, 705 cm
; H
NMR (400 MHz, CDCl ): 1.92–2.03 (m, 3H), 2.26–2.32 (m, 1H), 2.42 (s,
3
3H), 2.58–2.64 (m, 1H), 3.41 (d, J = 16.4 Hz, 1H), 3.61 (d,
J = 16.8 Hz, 1H), 6.18 (br s, 2H), 6.99 (s, 1H), 7.52 (d, J = 8.8 Hz,
282

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
1
1
6
1
6
H), 7.72 (dd, J = 1.6 Hz, 1H), 7.83 (dd, J = 8.4, 2.0 Hz, 1H), 10.05 (s,
7.05; N, 8.91; found: C, 68.92; H, 6.91; N, 9.11%; ESI-MS: 314.9
1
3
[M + H]+
.
H); C NMR (100 MHz, CDCl
7.96, 107.09, 112.84, 115.86, 116.09, 125.20, 141.86, 153.16,
53.93, 161.77, 170.21, 177.80; Anal. Calc. for C17 ; C,
2.00; H, 5.81; N, 12.76; found: C, 62.13; H, 5.96; N, 12.91%; ESI-
3
): δ 18.67, 24.72, 31.32, 55.19, 59.74,
19 3 4
H N O
4.3.6.8. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-(4-methylpiperidin-1-yl)
acetamide (17 h). White solid, Yield: 61%; M.P: 176–178 °C; IR (KBr)
+
MS: 329.9 [M + H]
.
3
1
1
2
294, 2949, 2922, 2858, 2810, 1720, 1708, 1689, 1618, 1572, 1529,
−
1 1
506, 1417, 1406, 1138, 866, 839 cm ; H NMR (400 MHz, CDCl ): δ
3
4
7
.3.6.3. (S)-2-(2-cyanopyrrolidin-1-yl)-N-(4-methyl-2-oxo-2H-chromen-
-yl)acetamide (17c). Off white solid, Yield: 54%; M.P: 146–148 °C; IR
.00 (d, J = 6.4 Hz, 3H), 1.29–1.32 (m, 2H), 1.70–1.71 (m, 3H),
.26–2.29 (m, 2H), 2.43 (d, J = 0.8 Hz, 3H), 2.87–2.90 (m, 2H), 3.13
(
KBr) 3286, 3111, 3097, 2956, 2821, 2220, 1685, 1624, 1595, 1417,
(
s, 2H), 6.21 (d, J = 0.8 Hz, 1H), 7.55–7.57 (m, 2H), 7.63 (d,
−
1 1
1
2
2
400, 1234, 1166, 862, 815, 752 cm
.09–2.19 (m, 2H), 2.26–2.39 (m, 2H), 2.43 (d, J = 0.8 Hz, 3H),
.92–3.03 (m, 2H), 3.56 (s, 1H), 3.89–3.92 (m, 1H), 6.23 (d,
;
H NMR (400 MHz, CDCl
3
):
13
J = 0.8 Hz, 1H), 9.55 (s, 1H); C NMR (100 MHz, CDCl
2
1
C H N O
18 22 2 3
1
3
): δ 18.63,
1.79, 30.09, 34.62, 54.40, 62.33, 106.82, 113.43, 115.29, 116.09,
25.27, 140.95, 152.15, 154.36, 161.04, 169.53; Anal. Calc. for
; C, 68.77; H, 7.05; N, 8.91; found: C, 68.82; H, 7.38; N,
1
3
J = 0.8 Hz, 1H), 7.56–7.63 (m, 3H), 8.84 (s, 1H); C NMR (100 MHz,
CDCl
3
): δ18.64, 22.96, 30.22, 53.02, 54.44, 57.28, 107.21, 113.68,
15.61, 116.48, 118.21, 125.37, 140.47, 152.17, 154.25, 160.98,
67.54; Anal. Calc. for C17 ; C, 65.58; H, 5.50; N, 13.50;
+
0.18%; ESI-MS: 314.9 [M + H] .
1
1
17 3 3
H N O
+
4.3.6.9. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-(4-methylpiperazin-1-yl)
acetamide (17i). Off white solid, Yield: 57%; M.P: 142–144 °C; IR (KBr)
found: C, 65.81; H, 5.78; N, 13.34%; ESI-MS: 312.1 [M + H]
.
3
1
2
294, 2935, 2816, 1712, 1693, 1618, 1573, 1529, 1406, 1168, 1139,
4
.3.6.4. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-(pyrrolidin-1-yl)
−
1 1
072, 1016, 866, 839 cm
H), 2.35 (s, 3H), 2.42 (d, J = 1.2 Hz, 3H), 2.52 (br s, 2H), 2.55 (br s,
3
; H NMR (400 MHz, CDCl ): δ 2.12 (br s,
acetamide HCl salt (17d). Off white solid, Yield: 47%; M.P: 200–202 °C;
IR (KBr) 3230, 3182, 2997, 1724, 1689, 1620, 1591, 1388, 1242, 1145,
1
4
J = 1.2 Hz, 1H), 7.75–7.81 (m, 2H), 10.46 (br s, 1H), 11.53 (s, 1H);
NMR (100 MHz, DMSO‑d
1
1
−
1
1
4H), 3.19 (s, 2H), 6.21 (d, J = 1.2 Hz, 1H), 7.54–7.61 (m, 2H), 7.61 (s,
H), 9.42 (s, 1H); 13C NMR (100 MHz, CDCl
): δ 18.68, 45.97, 53.38,
55.17, 61.80, 106.82, 113.47, 115.32, 116.19, 125.35, 140.77, 152.24,
54.29, 161.06, 168.89; Anal. Calc. for C17 ; C, 64.74; H, 6.71;
N, 13.32; found: C, 64.96; H, 7.08; N, 13.59%; ESI-MS: 315.9
M + H]+
018, 866, 839, 779 cm ; H NMR (400 MHz, DMSO‑d
6
): δ 1.96 (br s,
1
3
H), 2.40 (s, 3H), 3.36 (br s, 4H), 4.34 (s, 2H), 6.30 (s, 1H), 7.55 (d,
13
C
1
21 3 3
H N O
6
): δ 18.47, 23.26, 54.54, 56.32, 106.37,
13.12, 115.75, 116.03, 126.62, 141.83, 153.54, 153.99, 160.37,
64.95; Anal. Calc. for C16 ; C, 67.12; H, 6.34; N, 9.78;
[
.
18 2 3
H N O
+
found: C, 67.36; H, 6.77; N, 10.02%; ESI-MS: 286.7 [M + H]
.
4
.3.6.10. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-(4-phenylpiperazin-1-
4
.3.6.5. (S)-1-(2-((4-methyl-2-oxo-2H-chromen-7-yl)amino)-2-oxoethyl)
yl)acetamide (17j). White solid, Yield: 65%; M.P: 238–240 °C; IR (KBr)
3
1
J = 0.4 Hz, 3H), 2.82–2.85 (m, 4H), 3.26 (s, 2H), 3.30–3.32 (m, 4H),
6
7
piperidine-2-carboxamide (17e). White solid, Yield: 60%; M.P:
2
1
298, 2933, 2820, 1710, 1695, 1618, 1572, 1504, 1413, 1394, 1251,
02–204 °C; IR (KBr) 3423, 3309, 3290, 2937, 2858, 2835, 2796,
−1 1
143, 1070, 873, 840, 750 cm ; H NMR (400 MHz, CDCl ): δ 2.43 (d,
3
−
1 1
697, 1678, 1618, 1518, 1400, 1145, 875, 848, 721 cm
; H NMR
(
(
(
400 MHz, CDCl
3
): 1.25–1.27 (m, 1H), 1.56–1.65 (m, 4H), 1.79–1.76
.22 (d, J = 0.4 Hz, 1H), 6.91–6.94 (m, 1H), 6.98 (d, J = 8.4 Hz, 2H),
m, 1H), 2.23 (t, J = 9.0 Hz, 1H), 2.39 (s, 3H), 2.85–2.96 (m, 3H), 3.30
d, J = 16.0 Hz, 1H), 6.26 (s, 1H), 7.14 (s, 1H), 7.54 (d, J = 8.4 Hz,
.28–7.34 (m, 2H), 7.54–7.58 (m, 2H), 7.68 (d, J = 1.6 Hz, 1H), 9.41
13
(
1
1
3
s, 1H); C NMR (100 MHz, CDCl ): δ 18.63, 49.48, 53.57, 61.98,
1
H), 7.63 (s, 1H), 7.22 (d, J = 8.4 Hz, 1H), 7.79 (s, 1H), 10.16 (s, 1H);
06.96, 113.56, 115.31, 116.26, 116.32, 120.32, 125.32, 129.25,
40.72, 150.88, 152.11, 154.36, 160.98, 168.65; Anal. Calc. for
1
3
C NMR (100 MHz, CDCl
6.64, 106.42, 112.82, 115.63, 116.00, 126.29, 142.22, 153.53,
54.04, 160.46, 150.08, 175.50 ; Anal. Calc. for C18 ; C,
2.96; H, 6.16; N, 12.24; found: C, 63.62; H, 6.48; N, 12.45%; ESI-
3
): δ 18.46, 23.23, 24.97, 29.67, 52.85, 60.36,
6
1
6
22 23 3 3
C H N O ; C, 70.01; H, 6.14; N, 11.13; found: C, 70.32; H, 5.93; N,
1
H
21
N
3
O
4
+
1.41%; ESI-MS: 378.0 [M + H]
.
+
MS: 366.10 [M + Na]
.
4.3.6.11. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-morpholinoacetamide
(
2
1
17 k). White solid, Yield: 49%; M.P: 178–180 °C; IR (KBr) 3323, 3190,
4
.3.6.6. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-(piperidin-1-yl)acetamide
941, 2837, 1718, 1701, 1624, 1587, 1535, 1465, 1413, 1390, 1232,
(17f). White solid, Yield: 55%; M.P: 158–160 °C; IR (KBr) 3290, 2928,
−1 1
072, 1053, 1004, 885, 860, 815 cm
3
; H NMR (400 MHz, CDCl ): δ
2
8
2
854, 2798, 1724, 1699, 1618, 1566, 1525, 1413, 1230, 1151, 1068,
−
1
1
2.40 (s, 3H), 2.64 (br s, 4H), 3.18 (s, 2H), 3.80 (br s, 4H), 6.18 (s, 1H),
.50–7.55 (m, 2H), 7.65 (s, 1H), 9.34 (s, 1H); 13C NMR (100 MHz,
CDCl ): δ 18.67, 53.79, 62.44, 66.99, 106.92, 113.47, 115.36, 116.22,
125.33, 140.67, 152.26, 154.26, 161.03, 168.54 ; Anal. Calc. for
; C, 63.56; H, 6.00; N, 9.27; found: C, 63.62; H, 5.68; N,
44, 815, 810, 707 cm ; H NMR (400 MHz, CDCl
3
): δ 1.51–1.52 (br s,
7
H), 1.64–1.70 (m, 4H), 2.42 (d, J = 1.2 Hz, 3H), 2.56 (br s, 4H), 3.11
3
(
s, 2H), 6.20 (d, J = 1.2 Hz, 1H), 7.53–7.54 (m, 2H), 7.64 (br s, 1H),
1
3
9
6
1
6
.56 (s, 1H); C NMR (100 MHz, CDCl
2.73, 106.80, 113.37, 115.30, 116.06, 125.27, 140.96, 152.20,
54.34, 161.05, 169.53; Anal. Calc. for C17 ; C, 62.98; H,
.71; N, 9.33; found: C, 62.71; H, 6.89; N, 9.57%; ESI-MS: 300.9
3
): δ 13.62, 23.54, 26.27, 54.93,
16 18 2 4
C H N O
9
+
.45%; ESI-MS: 302.9 [M + H] .
20 2 3
H N O
+
[
M + H]
.
4.3.6.12. 2-(3,4-dihydroisoquinolin-2(1H)-yl)-N-(4-methyl-2-oxo-2H-
chromen-7-yl)acetamide (17 l). White solid, Yield: 69%; M.P:
178–180 °C; IR (KBr) 3281, 2993, 2904, 2829, 1720, 1676, 1624,
4
.3.6.7. N-(4-methyl-2-oxo-2H-chromen-7-yl)-2-(3-methylpiperidin-1-yl)
−
1 1
acetamide (17 g). White solid, Yield: 58%; M.P: 158–160 °C; IR (KBr)
1577, 1531, 1413, 1394, 1217, 1180, 873, 852, 746 cm
; H NMR
3
1
261, 2924, 2808, 1701, 1681, 1616, 1573, 1525, 1413, 1400, 1330,
(400 MHz, CDCl ): δ 2.42 (s, 3H), 2.93–2.96 (m, 2H), 3.02–3.04 (m,
3
−1
1
143, 1068, 842, 769 cm
;
H NMR (400 MHz, CDCl
3
): δ 1.00 (d,
2H), 3.37 (s, 2H), 3.85 (s, 2H), 6.21 (s, 1H), 7.07 (d, J = 7.2 Hz, 1H),
7.18–7.25 (m, 3H), 7.48–7.55 (m, 2H), 7.70 (d, J = 2.0 Hz, 1H), 9.47
J = 6.4 Hz, 3H), 1.61.1.64 (m, 1H), 1.74–1.76 (m, 3H), 1.92–1.97 (m,
1
6
H), 2.20–2.26 (m, 1H), 2.41 (s, 3H), 2.76–2.83 (m, 2H), 3.10 (s, 2H),
(s, 1H); 13C NMR (100 MHz, CDCl
3
) 18.62, 29.25, 51.53, 56.11, 61.88,
1
3
.19 (s, 1H), 7.54 (m, 2H), 7.63 (s, 1H), 9.55 (s, 1H); C NMR
): δ 18.62, 19.36, 25.55, 31.45, 31.95, 54.43, 62.09,
2.47, 106.75, 113.35, 115.27, 116.05, 125.28, 140.96, 152.21,
54.33, 161.05, 169.56; Anal. Calc. for C18 ; C, 68.77; H,
107.07, 113.50, 115.43, 116.20, 125.22, 126.11, 126.58, 126.76,
128.90, 133.37, 133.68, 140.78, 152.12, 154.33, 161.02, 168.99;
(
100 MHz, CDCl
3
6
1
Anal. Calc. for C21
72.62; H, 5.48; N, 8.33%; ESI-MS: 349.2 [M + H] .
20 2 3
H N O ; C, 72.40; H, 5.79; N, 8.04; found: C,
+
H
22
2
N O
3
283

R. Soni, S.S. Soman
Bioorganic Chemistry 79 (2018) 277–284
4
4
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emission wavelength of 450–465 nm. Human recombinant DPP-IV en-
zyme procured from Enzo Life Science (batch no BML-SE434-9091),
substrate, H-Gly-Pro-AMC procured from Enzo life science (batch no.
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4
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(
order of addition of reagents: assay buffer, enzyme, solvent/inhibitor
and finally substrate). Both the enzyme and 96-well plate were in-
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5
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3
60 nm and emission filter at 460 nm at sensitivity of 45.
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4
.4.2. MTT assay
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[
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay4
as per standard protocol. Cells were plated in a 96-well plate (1 × 10
cells/well) and incubated overnight in 100 µl DMEM media supple-
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(
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7
5, 100 µM conc. and incubated further for 48 h. 20 µl of MTT solution
(
(
5 mg/mL in PBS) was added and further plate was incubated for 4 h.
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Supernatant solution was removed and the blue formazan was dissolved
in 100 µl of acidified isopropanol. The absorbance was measured using
microplate reader at 570 nm (Metertech Σ960)
Cell viability (%) = (average absorbance of treated groups/average
absorbance of control group) × 100%. IC50 values were calculated
using GraphPad Prism.
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One of the authors (RS) is thankful to Department of Science &
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no. SR/WOS-A/CS-1028/2014 under Women Scientist Scheme to carry
out this work. Authors are thankful to Ms. Shweta Umar, Prof. B. Suresh
for their help with activity. Authors are also thankful to The Head,
Department of Chemistry and Department of Zoology, Faculty of
Science, The M. S. University of Baroda for providing laboratory facil-
ities, Zydus Research Centre, Ahmedabad, for the ESI-MS analyses.
[
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