Synthesis and biological activities study of novel phthalimides and phenylpyrazolo[1,5-a]pyrimidines

Article abstract of DOI:10.1177/1747519821993433

Phosphodiesterase II (PDE2) is mainly distributed in brain and heart cells, and it is a potential therapeutic target for the treatment of central nervous system (CNS) diseases such as Alzheimer’s disease. Based on the structure of the existing PDE2 inhibitor BAY60-7550, a series of novel phthalimides and phenylpyrazolo[1,5-a]pyrimidines have been designed and prepared. Furthermore, after evaluating their inhibitory activity toward PDE2, compound 7-oxo-N-phenethyl-5-phenyl-4,7-dihydropyrazolo[1,5-a]pyrimidine-3-carboxamide is found to have the optimum inhibitory potential (IC₅₀: 1.82 ± 0.29 μM). Discovery Studio software used to simulate the structure–activity relationship between this compound and the PDE2 protein crystal 4HTX to illustrate the binding modes, which provides favorable guidance for the further development of effective PDE2 inhibitors.

Full text of DOI:10.1177/1747519821993433

993433CHL0010.1177/1747519821993433Journal of Chemical ResearchTang et al.
research-article2021
Research Paper
Journal of Chemical Research
2021: 1–7
© The Author(s) 2021
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Synthesis and biological activities
study of novel phthalimides and
phenylpyrazolo[1,5-a]pyrimidines
https://doi.org/10.1177/1747519821993433
DOI: 10.1177/1747519821993433
journals.sagepub.com/home/chl
Long Tang1,2 , Jianchun Jiang1, Guoqiang Song2,
Yajing Wang2, Min Wei1, Yijing Cao2, Xianfeng Huang2
and Xiaoqing Feng2
Abstract
Phosphodiesterase II (PDE2) is mainly distributed in brain and heart cells, and it is a potential therapeutic target for the
treatment of central nervous system (CNS) diseases such as Alzheimer’s disease. Based on the structure of the existing
PDE2 inhibitor BAY60-7550, a series of novel phthalimides and phenylpyrazolo[1,5-a]pyrimidines have been designed
and prepared. Furthermore, after evaluating their inhibitory activity toward PDE2, compound 7-oxo-N-phenethyl-5-
phenyl-4,7-dihydropyrazolo[1,5-a]pyrimidine-3-carboxamide is found to have the optimum inhibitory potential (IC50:
1.82 ± 0.29 μM). Discovery Studio software used to simulate the structure–activity relationship between this compound
and the PDE2 protein crystal 4HTX to illustrate the binding modes, which provides favorable guidance for the further
development of effective PDE2 inhibitors.
Keywords
inhibitors, phenylpyrazolo[1,5-a]pyrimidines, phosphodiesterase II, phthalimides
Date received: 2 November 2020; accepted: 20 January 2021
1Institute of Chemical Industry of Forest Products, CAF; National
Engineering Laboratory for Biomass Chemical Utilization; Key and
Introduction
Open Laboratory of Forest Chemical Engineering, SFA; Key Laboratory
of Biomass Energy Sources and Materials, Nanjing, P.R. China
2School of Pharmaceutical Engineering & Life Science, Changzhou
University, Changzhou, P.R. China
It is well known that cAMP and cGMP, as important sec-
ondary messengers, affect the physiological activities of
the human body, and, importantly, their imbalance in cell
concentration in the body may lead to a variety of
diseases.1–3 Previous studies have shown that the activity of
phosphodiesterases (PDEs) directly affects the expression
concentration of cAMP and cGMP in vivo.4 PDEs are a
large family of polygenes, including 11 different subtype
structures, which have different substrate specificities,
enzyme kinetics, and cell distribution. They also have simi-
lar cell structures, show their own homology, and deter-
mine the specificity of the substrate or inhibitor.5–7 Among
Corresponding authors:
Xiaoqing Feng, School of Pharmaceutical Engineering & Life Science,
Changzhou University, Room 614, Lexing Building, Science and
Education City, Wujin, Changzhou 213164, Jiangsu, P.R. China.
Email: fxqfw@163.com
Xianfeng Huang, School of Pharmaceutical Engineering & Life Science,
Changzhou University, Room 614, Lexing Building, Science and Educa-
tion City, Wujin, Changzhou 213164, Jiangsu, P.R. China.
Email: huangxf@cczu.edu.cn
Creative Commons Non Commercial CC BY-NC: This article is distributed under the terms of the Creative Commons
Attribution-NonCommercial 4.0 License (https://creativecommons.org/licenses/by-nc/4.0/) which permits non-commercial use,
reproduction and distribution of the work without further permission provided the original work is attributed as specified on the SAGE and
Open Access pages (https://us.sagepub.com/en-us/nam/open-access-at-sage).

2
Journal of Chemical Research
amines (benzylamine, phenethylamine, n-butylamine, n-hex-
ylamine, (tetrahydrofuran-2-yl)methanamine, and furfu-
rylamine) provided the desired phenylpyrazole[1,5-a]
pyrimidine PDE2 inhibitors 2a–f in moderate yields.
Bioassays
Protein expression and purification. In order to express the
catalytic domain (residues 580–941), the recombinant
pET15b-PDE2A plasmid was subcloned and purified by
the previously reported method.26 Subsequently, the plas-
mid was transferred into Escherichia coli strain BL 21
(CodonPlus) to grow in 2XYT medium at 37 °C until the
absorbance (A) at 600 nm was 0.6–0.8. Next, 0.1 mmol L−1
of isopropyl-β-D-1-thiogalactopyranoside was added to
induce the PDE2A protein expression at 15 °C for 20 h.
The next day, the bacteria were harvested at 8000 r/min at
4 °C, the supernatant was discarded, and the pellet was
saved. The lysis solution (20 mmol L−1 Tris 8.0, 300 mmol
L−1 NaCl, 15 mmol L−1 imidazole, and 1.0 mmol L−1 mer-
captoethanol) was added according to the bacterial weight
(1:5) to suspend the bacteria and the mixture was then soni-
cated. The centrifuged supernatant was mixed with a nickel
column. Then, the nickel column was washed with 10 mL
each of 5, 20, and 50 mmol L−1 imidazole solutions to elute
impurities and dialyzed overnight to remove excess imidaz-
ole with 3.0 mL of elution buffer containing 150 mmol L−1
of imidazole (20 mmol L−1 Tris 8.0, 50 mmol L−1 NaCl, 150
mmol L−1 imidazole, and 1.0 mmol L−1 mercaptoethanol).
The purified protein was detected by sodium dodecyl sul-
fate–polyacrylamide gel electrophoresis (SDS-PAGE), and
the protein concentration was measured by the bicincho-
ninic acid method.
Figure 1. The structure of BAY 60-7550.
them, PDE2 is a dual-substrate enzyme that can hydrolyze
cAMPand cGMP.Although cAMPand cGMPare expressed
in the brain, heart, liver, platelets, and T cells, the expres-
sion level is the highest in the brain regions,8–10 which indi-
cates that PED2 plays an important role in the emotional,
cognitive, and other behavior of humans.7,10,11 Therefore,
PDE2 inhibitors have the potential to treat a variety of dis-
eases, such as Alzheimer’s disease, angina pectoris,
vasodilatation, heart failure, hypertension, and arrhyth-
mia.12–22 However, due to poor pharmacological activity,
low blood–brain barrier permeability, and poor metabolic
stability, no PDE2 inhibitor has been approved for clinical
use. Therefore, the development of novel PDE2 inhibitors
is an important research goal.
BAY 60-7550 (Figure 1) is an imidazole–triazine com-
pound with the high selectivity and activity toward PDE2.
Moreover, it is often used as an important standard in PDE2
bioactivity tests to investigate other PDE2 inhibitors.23,24
Considering the structural–activity relationship between
BAY60-7550 and PDE2 protein (PDB ID: 4HTX, http://
www.rcsb.org),aseriesofphthalimideandphenylpyrazolo[1,5-
a]pyrimidine PDE2 inhibitors have been designed and pre-
pared. Furthermore, the IC50 values were determined by the
biological activity detection method. The structure–activity
relationship between the optimum product and 4HTX was
simulated by the Discovery Studio software to illustrate the
possible binding modes and for the further development of
effective PDE2 inhibitors.
Enzymatic assays
The enzymatic activities of 9 compounds were assayed
using Bio-cAMP as the substrate.1,26 Briefly, a reaction
mixture of 20 mmol L−1 of Tris HCl, 10 mmol L−1 of MgCl2,
0.5 mmol L−1 of DTT, 3H-cAMP or 3H-cGMP (20000-
40000 cpm 3H-cAMP or 3H-cGMP per assay ). and pro-
teins were incubated at 25 °C for 15min. The reaction was
then terminated by the addition of ZnSO4 and precipitated
out by Ba(OH)2. The compound was dissolved in with
DMSO, and the possible systematic errors of the method
were evaluated through the different contents of DMSO.
Finally, BAY60-7550, a typical PDE2 positive inhibitor,
was selected to determine its activity and to calculate the
accuracy of the IC50 verification method. The hydrolysis
rate of H-cAMP or H-cGMP expressed the inhibitory activ-
ity of the compound. Seven concentrations of inhibitors
were used for measuring the IC50 values. Each measure-
ment was repeated three times, and the IC50 values were
calculated by nonlinear regression. BAY 60-7550 was used
as the reference compound for PDE2 enzyme activity
detection.
Results and discussion
Synthetic pathways
The synthetic strategy toward phthalimide PDE2 inhibitors 1
is depicted in Scheme 1(a), employing ethyl furan and maleic
anhydride as the starting materials. Using 4-ethylisobenzo-
furan-1,3-dione (II) and amines, including 3,4-dimethoxy-
benzylamine, 3-aminopyrazole, and 3-(benzyloxy)aniline,
the desired phthalimide PDE2 inhibitors 1a–c were obtained
in good yields through an acylation process. Furthermore, the
synthetic strategy for phenylpyrazole pyrimidine PDE2
inhibitors 2 is shown in Scheme 1(b). As expected, the inter-
mediate (IV) could be easily prepared through a two-step
reaction involving cyclization and hydrolysis.25 Next, the
coupling reaction between intermediate (IV) and various
Analysis of biological activity results
The AlphaScreen kit method was used to detect the activity
of the nine synthesized compounds, which were diluted at

Tang et al.
3
(a)
O
O
O
O
O
O
RNH2
conc. H
0oC, 2 h
2SO4
R N
O
O
O
O
120oC, 4 h
ether, rt, overnight
O
O
O
II
I
1a-d
79%
95%
O
O
N
O
N
N
HN
N
O
O
O
O
O
O
1b
87%
Ph
1a
86%
1c
81%
(b)
O
O
O
H
N
O
O
HOAc
reflux, 12 h
O
HN
O
N
N
N
NH2
O
III 38%
KOH, EtOH, H2O/THF reflux, overnight
O
HO
N
N
O
H
R2NH
N
2, EDCl, HOBt
N
H
R2
DIEA, DMF
N
H
N
N
25oC, overnight
O
2a-f
IV
O
72%
O
O
O
N
N
N
N
N
N
N
H
N
N
H
Ph
H
Ph
N
N
N
H
O
H
H
O
O
2a
45%
2c
55%
O
2b
49%
O
N
O
N
N
O
N
N
O
N
N
H
N
H
N
H
N
H
N
O
N
H
O
H
O
2d
50%
2e
59%
2f
57%
Scheme 1. The synthetic pathway toward phthalimide and phenylpyrazolo[1,5-a]pyrimidine PDE2 inhibitors 1 and 2.
seven concentrations, and three sets of parallel experiments substituents did not improve their activity. From molecular
were performed. The IC50 values were obtained by fitting structure analysis, the phenylpyrazolo[1,5-a]pyrimidine
calculations using the software GraphPad Prism 7. The can be considered as a rigid structure, similar to the imida-
results are given in Table 1.
zolopyrimidine in BAY 60-7550. Meanwhile, phenyl and
First, according to the IC50 values of phthalimide com- nitrogen-linked groups in phenylpyrazolo[1,5-a]pyrimi-
pounds 1a–c, we found that changing the nitrogen dine are similar to the dimethoxybenzene and alkylbenzene

4
Journal of Chemical Research
Table 1. Synthetic PDE2 inhibitors and their IC50 values.a,b
Entry
Compound
Structure
PDE2 IC50 (μM)
O
N
O
O
O
1
2
1a
1b
9.87 ± 0.43
6.89 ± 1.25
O
N
HN
N
O
O
N
O
O
Ph
3
4
5
6
1c
2a
2b
2c
>10
O
N
N
N
H
Ph
N
H
O
1.82 ± 0.29
2.62 ± 0.39
9.23 ± 0.65
O
N
N
N
H
Ph
N
H
O
O
N
N
N
H
N
O
H
O
N
N
N
N
N
H
N
H
O
7
8
9
2d
2e
2f
>10
O
N
O
N
H
N
H
O
5.67 ± 0.43
4.19 ± 0.52
O
N
O
N
H
N
H
O
aBAY60-7550 was used as the reference compound with an IC50 of 8.4 nM.
bIC50 values determined against 3H-cAMP.
groups in BAY60-7550. Next, we determined the IC50 val-
ues of compounds 2a–f and 7-oxo-N-phenethyl-5-phenyl-
4,7-dihydropyrazolo[1,5-a]pyrimidine-3-carboxamide (2a)
exhibits the optimum inhibitory potential toward PDE2
(IC50: 1.82 ± 0.29 μM).
Molecular docking
Discovery Studio software is a comprehensive molecular sim-
ulation platform. The working principle is based geometric
complementarity, chemical environment complementarity,

Tang et al.
5
Figure 2. A three-dimensional plan view of the BAY 60-7550–
PDE2 protein interaction active site.
Figure 3. A three-dimensional plan view of the 2a–PDE2
protein interaction active site.
and energy complementarity. If there is an interaction between
the ligand and the receptor, then molecular docking can be
evaluated and we can obtain the structure–activity relationship
and scoring results of the interaction between the molecule
and the protein.According to the results, we can judge whether
the binding between the molecule and the protein is good or
not, and further guide modifications of the compound struc-
ture. In this paper, a semi-flexible CDOCKER docking is used
to simulate the binding mode of the synthesized compound
and the PDE2 protein 4HTX.
confirm the structures of the compounds. All compounds
were tested for PDE2 enzyme activity with BAY60-7550 as
a reference and Bio-cAMP as the substrate. It was noted
that compound 2a has the optimum inhibitory potential
toward PDE2 (IC50: 1.82 ± 0.29 μM). In this work, the
molecular docking of the synthesized compound and the
PDE2 protein 4HTX was performed using a flexible dock-
ing software CDOCKER embedded in Discovery Studio
program. This research is expected to provide a basis for
the further development of effective PDE2 inhibitors.
According to the analysis of the structure–activity rela-
tionship of BAY60-7550 and PDE2 protein 4HTX, we
found that the carbonyl oxygen on the pyrimidinone ring
forms a hydrophilic region with GLN812 and GLN859.
The hydrophobic region is mainly due to the imidazotria-
zine skeleton, while the amino acids of ILe826, Phe862,
Leu770, Leu809, Tyr655, and Met847 can form π–σ, π–π
and π–alkyl stacking interactions with this core skeleton,
and promote the PDE2 effects of BAY60-7550 (Figure 2).
To further explain the inhibitory activity of compound
2a, it has been docked into the binding pocket of PDE2
Discovery Studio 2020 (Figure 3). The results show that
compound 2a can form hydrogen bond interactions with
Gln812, and π–π stacking interactions with Phe862,
ILe826, Leu809, Phe830, and Leu770. However, due to the
poor hydrogen bond interaction strength of Gln859 and the
weakening of the π–π stacking interactions, the inhibitory
activity of compound 2a is lower than that of BAY60-7550.
Although the inhibitory effect of compound 2a on PDE2 is
not as obvious as that of BAY60-7550, it has a certain
inhibitory effects on the PDE2 enzyme.
Experimental
General
1H NMR spectra were recorded on a BrukerBioSpin GmbH
spectrometer at 300 and 400 MHz, 13C NMR spectra were
recorded on a BrukerBioSpin GmbH spectrometer at 75
MHz; the coupling constants are given in Hz. High-
resolution mass spectrometry (HRMS) was performed
using an Agilent 6200 accurate-mass time-of-flight (TOF)
liquid chromatography (LC)/mass spectrometry (MS) sys-
tem with electrospray ionization (ESI). Thin-layer chroma-
tography (TLC) was performed on precoated silica gel
F-254 plates (25 mm × 75 mm, Shanghai Pinjia Chemical
Co., Ltd), and the samples were visualized with UV light.
High-performance liquid chromatography (HPLC) analysis
was performed using a SHIMADZU LC-20AB instrument.
Melting points were measured with an X4-A microscopic
melting point apparatus. All the starting materials and rea-
gents were purchased from commercial suppliers.
Synthesis of compounds (1a–c); general
Conclusion
procedures
In this study, two series of phthalimides (1a–c) and
phenylpyrazolo[1,5-a]pyrimidines (2a–f) have been
designed and synthesized as potential PDE2 inhibitors. 1H
NMR, 13C NMR, and mass spectrometry were used to
To a solution of maleic anhydride (30 mmol, 2.94 g) in
ether (30 mL) was added ethyl furan (30 mmol, 2.88 g) in a
dropwise manner, and the resulting mixture was stirred

6
Journal of Chemical Research
HRMS (ESI): m/z [M + H]+ calcd for C23H20NO3:
358.1365; found: 358.1369. Chromatographic purity:
97.1% (HPLC).
overnight at room temperature in the dark. The progress of
reaction was monitored by TLC until all the starting materi-
als had been consumed. The solution was filtered, and the
filter cake was washed with ether (60 mL) to afford I as a
white solid27 (5.52 g, 95%). Next, concentrated sulfuric
acid (180 mL) was added to a 250-mL round-bottom flask,
and compound I was slowly added. The temperature was
maintained at 0–5 °C with an ice–salt–water bath, and the
mixture was stirred for 2 h, during which time the solution
turned black/green. After completion of the reaction, the
solution was added dropwise to an ice–water mixture, and
a yellow flocculent solid was precipitated. This solid was
filtered and dried to give II as a white solid27 (4.34 g, 79%).
To a solution of compound II (17 mmol, 3 g) in acetic acid
(60 mL) was added the corresponding amine (19.3 mmol)
in a dropwise manner. The mixture was then heated to 120
°C, and the reaction progress was monitored by TLC
(PE:EtOAc = 3:1). After 4 h, when the reaction is over, the
reaction solution was evaporated under reduced pressure at
70 °C to give a black oily substance. NaHCO3 solution
(4%) was added dropwise until no more bubbles were gen-
Synthesis of compounds (2a–f); general
procedures
A mixture of 3-amino-4-ethoxycarbonylpyrazole (25 mmol,
3.82 g) and ethyl benzoyl acetate (25 mmol, 4.74 g) was
heated in acetic acid (15 mL) at 120 °C for 12 h. After the
completion of the reaction as evident by TLC, the solution
was cooled and filtered. The residue was washed with acetic
acid, filtered, and dried to give III as a beige crystalline
powder25 (38%). To a solution of compound III (1.2 g) in a
mixed solvent consisting of EtOH (32 mL), THF (16 mL),
and H2O (40 mL) was added KOH (10.16 mmol, 0.57 g) at
45 °C. After stirring for 10min, the solid was almost com-
pletely dissolved. After maintaining the temperature at 50
°C for 3 h, KOH (6.24 mmol, 0.35 g) was added again and
the temperature was raised to 80 °C. After 3 h, KOH (3.56
mmol, 0.2 g) was added, and the mixture was allowed to
erated. The product was then recrystallized from hot etha- react overnight. The reaction was stopped and the solution
nol to give the desired products 1a–c.
was then cooled to room temperature. HCl (1.0 mol L−1)
was slowly added dropwise to adjust the pH to weak acidity,
and a white crystalline solid was precipitated. After suction
filtration, the filter cake was washed with deionized water
and dried to obtain IV as a white crystal compound25 (72%).
A solution of compound IV (1.0 mmol, 0.25 g) in DMF (10
mL) was treated with EDCI (1.2 mmol, 0.23 g), HOBT (1.2
mmol, 0.16 g), and DIEA (1.2 mmol, 0.15 g). After 1 h, the
corresponding amine (1.2 mmol) was added dropwise, and
the mixture was stirred overnight at 25 °C. The solution was
transferred dropwise to the saturated NaHCO3 aqueous
solution, and a white flocculent solid was formed. The solid
was filtered and dried to give the desired product 2a–f.
2-(3,4-Dimethoxybenzyl)-4-ethylisoindoline-1,3-dione
(1a). Using 3,4-dimethoxybenzylamine as the starting
martial, the desired product 1a was obtained as a yellow
solid (0.19 g, 86%). M.p. 106–107 °C. 1H NMR (300 MHz,
DMSO-d6) δ 7.75-7.66 (m, 3H), 6.94-6.78 (m, 3H), 4.67 (s,
2H), 3.72 (s, 3H), 3.71 (s, 3H), 3.09-3.01 (q, J = 7.5 Hz,
2H), 1.21 (t, J = 7.5 Hz, 3H); 13C NMR (75 MHz, DMSO-
d6) δ 168.65, 168.04, 149.11, 148.65, 144.08, 135.55,
134.77, 132.54, 129.62, 128.00, 121.36, 120.23, 112.21,
112.07, 55.94, 55.87, 40.98, 24.23, 15.41; HRMS (ESI):
m/z [M + H]+ calcd for C19H20NO4: 326.1314; found:
326.1320. Chromatographic purity: 94.2% (HPLC).
7-Oxo-N-phenethyl-5-phenyl-4,7-dihydropyrazolo[1,5-a]
pyrimidine-3-carboxamide (2a). Using phenethylamine as
the starting martial, the desired product 2a was obtained as
a white solid (0.16 g, 45%). M.p. >300 °C. 1H NMR (400
MHz, DMSO-d6) δ 8.68 (s, 1H), 7.97 (s, 1H), 7.85-7.76 (m,
2H), 7.42 (t, J = 3.5 Hz, 3H), 7.30-7.22 (m, 5H), 6.07 (s,
4-Ethyl-2-(1H-pyrazol-3-yl)isoindoline-1,3-dione
(1b). Using 3-aminopyrazole as the starting martial, the
desired product 1b was obtained as a yellow solid (0.21 g,
87%). M.p. 169–170 °C. 1H NMR (300 MHz, DMSO-d6) δ
13.15 (s, 1H), 8.04-7.45 (m, 4H), 6.36 (d, J = 3.0 Hz, 1H),
1H), 3.64 (t, J = 6.5 Hz, 2H), 2.88 (t, J = 6.9 Hz, 2H); 13
C
3.13-3.04 (q, J = 9.0 Hz, 2H), 1.24 (t, J = 7.5 Hz, 3H); 13
C
NMR (75 MHz, DMSO-d6) δ 163.52, 162.81, 158.80,
157.81, 150.20, 142.68, 140.10, 139.37, 129.12, 128.91,
128.81, 127.09, 126.58, 102.00, 91.75, 36.31, 31.24;
HRMS (ESI): m/z [M + H]+ calcd for C21H19N4O2:
359.1430; found: 359.1433. Chromatographic purity:
100% (HPLC).
NMR (75 MHz, DMSO-d6) δ 167.61, 167.06, 144.55,
135.97, 135.20, 132.39, 130.67, 127.82, 123.00, 121.78,
103.18, 24.35, 15.41; HRMS (ESI): m/z [M + H]+ calcd
for C13H12N3O2: 242.0851; found: 242.0856. Chromato-
graphic purity: 96.5% (HPLC).
4-Ethyl-2-[3-(benzyloxy)phenyl]-isoindoline-1,3-dione
(1c). Using 3-(benzyloxy)aniline as the starting martial,
the desired product 1c was obtained as a yellow solid (0.30
g, 81%). M.p. 80–81 °C. 1H NMR (300 MHz, DMSO-d6) δ
7.87-7.68 (m, 3H), 7.53-7.27 (m, 6H), 7.17-7.07 (m, 2H),
7.05-7.01 (m, 1H), 5.12 (s, 2H), 3.13-3.06 (q, J = 7.5 Hz,
2H), 1.25 (t, J = 7.5 Hz, 3H); 13C NMR (75 MHz, DMSO-
d6) δ 167.83, 159.02, 144.35, 137.24, 135.79, 134.99,
133.48, 132.55, 130.05, 128.94, 128.42, 128.32, 127.99,
121.65, 120.49, 114.91, 114.77, 69.95, 24.33, 15.45;
N-Benzyl-7-oxo-5-phenyl-4,7-dihydropyrazolo[1,5-a]
pyrimidine-3-carboxamide (2b). Using benzylamine as the
starting martial, the desired product 2b was obtained as a
1
white solid (0.17 g, 49%). M.p. >300 °C. H NMR (400
MHz, DMSO-d6) δ 9.08 (s, 1H), 8.01 (s, 1H), 7.91-7.82 (m,
2H), 7.39-7.32 (m, 8H), 6.10 (s, 1H), 4.56 (s, 2H); 13C
NMR (75 MHz, DMSO-d6) δ 163.37, 158.89, 157.54,
150.28, 142.65, 140.09, 139.06, 129.50, 129.02, 128.86,
128.08, 127.47, 126.98, 101.84, 91.65, 42.81; HRMS

Tang et al.
7
(ESI): m/z [M + H]+ calcd for C20H17N4O2: 345.1273; Declaration of conflicting interests
found: 345.1278. Chromatographic purity: 100% (HPLC).
The author(s) declared no potential conflicts of interest with
respect to the research, authorship, and/or publication of this
article.
N-Butyl-7-oxo-5-phenyl-4,7-dihydropyrazolo[1,5-a]
pyrimidine-3-carboxamide (2c). Using n-butylamine as
the starting martial, the desired product 2c was obtained
Funding
1
as a white solid (0.17 g, 55%). M.p. >300 °C. H NMR
The author(s) disclosed receipt of the following financial support
for the research, authorship, and/or publication of this article: This
work was supported by the National Natural Science Foundation
of China (no. 81803471).
(400 MHz, DMSO-d6) δ 8.70 (s, 1H), 7.98 (d, J = 7.4 Hz,
2H), 7.93 (s, 1H), 7.52-7.34 (m, 3H), 6.07 (s, 1H), 3.32 (t,
J = 10.0 Hz, 2H), 1.59-1.49 (m, 2H), 1.43-1.40 (m, 2H),
0.93 (t, J = 7.2 Hz, 3H); 13C NMR (75 MHz, DMSO-d6)
δ 163.44, 158.91, 157.72, 150.15, 142.61, 139.41, 129.55,
128.93, 127.03, 102.19, 91.65, 37.93, 32.09, 20.25, 14.16;
HRMS (ESI): m/z [M + H]+ calcd for C17H19N4O2:
311.1430; found: 311.1435. Chromatographic purity:
100% (HPLC).
ORCID iD
Long Tang
https://orcid.org/0000-0001-9252-7122
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N-Hexyl-7-oxo-5-phenyl-4,7-dihydropyrazolo[1,5-a]
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1
as a white solid (0.17 g, 50%). M.p. >300 °C. H NMR
(400 MHz, DMSO-d6) δ 8.72 (s, 1H), 8.01 (d, J = 7.47
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3.35 (t, J = 12.0 Hz, 2H), 1.59-1.55 (m, 2H), 1.46-1.42
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C
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HRMS (ESI): m/z [M + H]+ calcd for C19H23N4O2:
339.1743; found: 339.1749. Chromatographic purity:
98.5% (HPLC).
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white solid (0.20 g, 59%). M.p. >300 °C. 1H NMR (400
MHz, DMSO-d6) δ 9.03 (s, 1H), 8.15 (d, J = 7.1 Hz, 2H),
7.97 (s, 1H), 7.46 (d, J = 8.5 Hz, 3H), 6.14 (s, 1H), 3.99
(m, 1H), 3.91 (d, J = 7.9 Hz, 1H), 3.76-3.50 (m, 3H),
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339.1379; found: 339.1384. Chromatographic purity:
100% (HPLC).
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graphic purity: 100% (HPLC).
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