Design, synthesis and antibacterial evaluation of novel 1,2,3-triazole derivatives incorporating 3′-deoxythymidine

Article abstract of DOI:10.3184/174751917X15094552081189

A series of novel 1,2,3-triazole derivatives incorporating 3′-deoxythymidine were designed, synthesised and characterised. Antibacterial activity against Escherichia coli and Staphylococcus aureus was evaluated for all of the synthesised compounds and com

Full text of DOI:10.3184/174751917X15094552081189

JOURNAL OF CHEMICAL RESEARCH 2017
VOL. 41 NOVEMBER, 645–649
RESEARCH PAPER 645
Design, synthesis and antibacterial evaluation of novel 1,2,3-triazole
derivatives incorporating 3′-deoxythymidine
Long-Fei Maoa, Gui-Qing Xua*, Bin Sunb, Yu-Qin Jianga, Wen-Pei Donga, Shu-Ting Zhanga,
Jia-Xuan Shena and Xing Hec
aSchool of Chemistry and Chemical Engineering, Henan Normal University, Henan Engineering Research Center of Chiral Hydroxyl Pharmaceutical, Collaborative
Innovation Center of Henan Province for Green Manufacturing of Fine Chemicals, Key Laboratory of Green Chemical Media and Reactions, Ministry of Education,
Xinxiang 453007, P.R. China
bCollege of Chemistry, Chemical Engineering and Materials Science, Shandong Normal University, Jinan 250014, P.R. China
cCollege of Chemistry, Nankai University, Tianjin 300071, P.R. China
A series of novel 1,2,3-triazole derivatives incorporating 3′-deoxythymidine were designed, synthesised and characterised. Antibacterial
activity against Escherichia coli and Staphylococcus aureus was evaluated for all of the synthesised compounds and compared
against standard antibiotic drugs, streptomycin sulfate and doxycycline, as controls. Some compounds showed potential antibacterial
activity towards E. coli, and the best minimum inhibitory concentration was 0.14 mM. Preliminary structure–activity relationships and
computational simulations were also studied.
Keywords: 1,2,3-triazole derivatives, 3′-deoxythymidine, antibacterial activity, computational simulations
Dihydrofolate reductase (DHFR) is an indispensable enzyme for
bacteria,anditisregardedasanimportantdrugtargetforantibacterial
agents. Dihydrofolate reductase inhibitors possess varying degrees
of antibacterial activity. Pyrimidine derivatives have exhibited
good antibacterial properties.1–3 Thymidine deoxyribose is one of
the nucleosides containing thymidine that may eventually interfere
with the synthesis of bacterial DNA. 1,2,3-Triazole derivatives have
attracted significant attention because of their multiple bioactivities,
such as antibacterial,4 antifungal,5,6 anticancer7 and anti-
inflammatory.8 Copper(I)-catalysed azide-alkyne cycloaddition
(CuAAC) reactions are a convenient and regiospecific approach
to the synthesis of 1,4-disubstituted triazoles9 and evoke greater
interest in the preparation of 1,2,3-triazole derivatives.10–12 Drug
resistance has become a massive health problem worldwide, and
new types of antibacterial agents are still needed.
and Staphylococcus aureus was evaluated, and the results reveal
that some compounds showed potential activity towards E. coli.
Preliminary structure–activity relationship studies are discussed
herein, and computational simulations of the synthesised
compounds with DHFR were also performed. Some of these studies
are the subject of a patent applied for in China (CN 106589027).
Results and discussion
The synthetic strategy for the preparation of the target molecules
is illustrated in Scheme 1. A nucleophilic substitution reaction
between substituted aniline 1 and propargyl bromide in acetone
yielded intermediate 2, which was subjected to the CuAAC
reaction with 3 to yield 4. Compounds 4a–r were obtained in
1
good yields, and their structures were confirmed by H NMR,
13C NMR and high-resolution mass spectrometry (HRMS).
The antibacterial activities against both Gram positive
bacteria (S. aureus) and Gram negative bacteria (E. coli) of all of
the synthesised compounds were evaluated using streptomycin
sulfate and doxycycline as controls. The values of the minimum
inhibitory concentration (MIC) are summarised in Table 1.
Building on our previous work,5,6,13 and in the search for
new antibacterial compounds, a series of novel 1,2,3-triazole
derivatives that incorporate 3′-deoxythymidine were designed,
synthesised and characterised. Furthermore, the antibacterial
activity of the synthesised compounds against Escherichia coli
O
HN
N
O
R2
R1
R2
R1
a
b
R2
R1
+
O
r
B
R3
NH2
R3
NH
R3
HN
H
O
N
1
2
N
4
N
R1
Br
R2
R3
Compd.
R1 R2
R3
Compd.
j
4
H
H
H
H
H
H
H
H
OC
H
H
H
H
F
N
OCH3
4a
4b
4c
4d
4e
4f
H
C
H
H
H
H
N
N
H
H
H
H
H
H
C
4k
4l
N
C
F
O2
F
l
H
C
3
3
4m
4n
4o
4p
4q
4r
H
C
H
O
O
H
H
3
H
OC
OCH33
H
H
H
H
O2
O2
3
H
O2
H
C
3
g
4
H
H
H
F
C
H
4h
4i
H
C
3
r3
H
B
3
Scheme 1 Synthesis and structures of compounds 4a–r.
Reactions and conditions: (a) K2CO3, DMF, rt; (b) 3 (3′-azido-3′-deoxythymidine), CuSO4·5H2O, lascorbic acid sodium, tert-butanol, 50 °C
* Correspondent. E-mail: guiqingxu@163.com

646 JOURNAL OF CHEMICAL RESEARCH 2017
Table 1 Antibacterial activities of compounds 4a–r against E. coli and S. aureus in vitro
MIC/(mM)
MIC/(mM)
Compound
Gram negative
E. coli
0.321
Gram positive
S. aureus
0.642
>0.591
0.310
0.621
0.560
>0.560
0.309
0.299
Compound
Gram negative
E. coli
0.577
Gram positive
S. aureus
>0.577
>0.549
>0.615
0.615
4a
4b
4c
4d
4e
4f
4g
4h
4i
4k
4l
4m
4n
4o
4p
4q
4r
0.591
0.310
0.310
0.140
0.140
0.453
0.149
0.536
0.268
0.549
0.154
0.154
0.270
0.279
0.274
0.299
0.044
>0.541
>0.558
0.549
0.598
0.022
0.536
>0.536
Streptomycin sulfate
Doxycycline
4j
0.072
0.072
Table 2 Docking scores of compounds 4a–r with DHFR (PDB ID: 4X5F)
Docking score
Docking score
Compound
Compound
(kcal mol-1)
(kcal mol-1)
4a
4b
4c
4d
4e
4f
4g
4h
4i
−6.490
−6.167
−6.671
−6.625
−7.699
−6.739
−5.987
−7.449
−6.594
4j
4k
4l
4m
4n
4o
4p
4q
4r
−6.105
−6.141
−6.925
−7.405
−7.061
−5.977
−6.566
−6.784
−6.189
Fig. 1 Binding mode of compound 4e in the active site of DHFR.
The data in Table 1 reveal that none of the compounds had
an obvious effect on S. aureus, yet some exerted moderate
inhibition towards E. coli. Among them, 4e, 4f, 4h, 4m and 4n
showed potential activity, with MIC values of about 0.15 mM.
Obviously, different substituents on the benzene ring exhibited
different influences on the activity of compounds 4 against
E. coli. For example, the nitro group at R2 (4e and 4f) gave a
fivefold higher potency than that of compound 4k. We also
observed that fluoro substitution on the benzene ring, which is
more electronegative than other halogens like chloro or bromo,
is important in enhancing antibacterial activity compared to
other molecules in the series (4b, 4j and 4m). The presence
of chloro or bromo substitution on the benzene ring gave
comparatively less activity.
Fig. 2 Binding mode of compound 4f in the active site of DHFR.
In order to determine the antibacterial mechanism of the
synthesised compounds, computational simulation of the
compoundsandDHFRwasperformedusingthecrystalstructure
of DHFR complexed with folate and NADP+ from E. coli (PDB
ID: 4X5F). Computational modelling was performed with the
Glide program using standard precision scoring. The docking
scores of all compounds are listed in Table 2. The data showed
that four compounds with the best activities (4e, 4h, 4m and
4n) had the best scores. The binding mode of some compounds
with DHFR (PDB ID: 4X5F) is shown in Figs 1–3. As shown
in Fig. 1, compound 4e forms five hydrogen bonds at the active
site of the enzyme and a π–π stacking between the benzene ring
and HIP 45 (HIP is the protonated histidine). The pyrimidine
ring is oriented towards the hydrophobic core and forms
two hydrogen bonds with TYR100 and ALA7, respectively.
Compound 4f forms four hydrogen bonds (Fig. 2). The binding
mode of the pyrimidine ring and the nitro group is similar to
that in 4e. The π–π stacking could affect the docking score of
these two compounds. A comparison of the binding mode of 4b,
4j and 4m is shown in Fig. 3. The numbers of hydrogen bonds
formed by 4b and 4j with the active site of DHFR are only two
and three, respectively. Compound 4m forms five hydrogen
bonds and a π–π stacking between the triazole ring and HIP
45. Therefore, we anticipated that the antibacterial activity of
compound 4m against E. coli would be the greatest among the
three compounds, and the activity determination results are
consistent with this prediction.
In conclusion, a series of novel 1,2,3-triazole derivatives
that incorporate 3′deoxythymidine were designed and
synthesised. Some exerted moderate inhibition towards E. coli.
Computational simulations of the synthesised compounds with
DHFR were also performed, and the results showed a certain
degree of correlation with the antibacterial test results.
Experimental
All reagents were obtained from commercial sources and were used
without further purification unless otherwise stated. Acetone was
dried with Type 4A Linde molecular sieves and then distilled. All
reactions were monitored by thin-layer chromatography (TLC).
1H NMR and 13C NMR spectra were measured on a Bruker 400 MHz
spectrometer operating at 400.13 and 100.61 MHz, respectively, or

JOURNAL OF CHEMICAL RESEARCH 2017 647
Synthesis of 4a–r; general procedure
(a)
(b)
(c)
3′-Azido-3′-deoxythymidine (compound 3, 1.0 mmol) and compound 2
(1.0 mmol) were added to a flask with tert-butanol (5 mL). The reaction
was catalysed with copper sulfate pentahydrate (0.1 mmol) and
sodium ascorbate (0.2 mmol) and stirred at 50 °C. On completion of
the reaction, the mixture was extracted with diluted dichloromethane
(15 mL × 3). The organic phase was washed successively with brine,
dried over sodium sulfate and concentrated in vacuo. The residue was
purified by recrystallisation using ethanol to yield compound 4 as a
yellow solid.
1-(5-(Hydroxymethyl)-4-(4-((phenylamino)methyl)-1H-1,2,3-
triazol-1-yl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-
dione (4a): Yield 92%; 1H NMR (600 MHz, DMSO-d6): δ 11.35
(s, 1H), 8.16 (s, 1H), 7.81 (s, 1H), 7.07 (t, J = 12.0 Hz, 2H), 6.65 (d,
J = 6.0 Hz, 1H), 6.55 (t, J = 6.0 Hz, 1H), 6.41 (t, J = 6.0 Hz, 1H), 6.04
(s, 1H), 5.35 (d, J = 6.0 Hz, 1H), 5.27 (s, 1H), 4.30 (d, J = 6.0 Hz,
1H), 4.19 (s, 1H), 3.70–3.59 (m, 2H), 2.71–2.62 (m, 2H), 1.81 (s, 3H);
13C NMR (151 MHz, DMSO-d6): δ 163.68, 150.37, 145.66, 143.64,
136.19, 128.96, 127.91, 122.48, 117.88, 116.79, 111.47, 109.57, 84.41,
83.76, 60.68, 60.08, 59.09, 37.06, 12.21; HRMS (ESI) m/z: calcd for
C19H23N6O4: 399.1775 [M+H]+; found: 399.1784.
1-(4-(4-(((2-Chlorophenyl)amino)methyl)-1H-1,2,3-triazol-1-
yl)-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (4b): Yield 94%; 1H NMR (600 MHz, DMSO-d6):
δ 11.38 (s, 1H), 8.15 (s, 1H), 7.81 (s, 1H), 7.25 (d, J = 12.0 Hz, 1H),
7.11 (t, J = 12.0 Hz, 1H), 6.78 (d, J = 12.0 Hz, 1H), 6.59 (t, J = 12.0
Hz, 1H), 6.42 (t, J1 = 6.0 Hz, J2 = 12.0 Hz, 1H), 5.92 (t, J1 = 6.0 Hz,
J2 = 12.0 Hz, 1H), 5.35 (t, J1 = 6.0 Hz, J2 = 12.0 Hz, 1H), 4.45 (d, J = 6.0
Hz, 2H), 4.18 (d, J = 12.0 Hz, 1H), 3.69–3.58 (m, 3H), 2.73–2.66 (m,
2H), 1.80 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ 163.69, 150.39,
145.66, 143.64, 136.19, 128.93, 127.92, 122.48, 117.88, 116.79, 111.47,
109.57, 84.39, 83.76, 60.68, 60.09, 59.09, 37.06, 12.23; HRMS (ESI)
m/z: calcd for C19H21ClN6O4Na: 455.1205 [M+Na]+; found: 455.1195.
1-(5-(Hydroxymethyl)-4-(4-((o-tolylamino)methyl)-1H-1,2,3-
triazol-1-yl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-
dione (4c): Yield 86%; 1H NMR (600 MHz, DMSO-d6): δ 11.38 (s,
1H), 8.12 (s, 1H), 7.81 (s, 1H), 6.98 (t, J1 = 6.0 Hz, J2 = 12.0 Hz, 2H),
6.59 (d, J = 12.0 Hz, 1H), 6.51 (t, J = 12.0 Hz, 1H), 6.42 (t, J1 = 12.0
Hz, J2 = 6.0 Hz, 1H), 5.41 (t, J = 6.0 Hz, 1H), 5.36–5.28 (m, 2H), 4.39
(d, J = 6.0 Hz, 1H), 3.70–3.58 (m, 2H), 2.73–2.57 (m, 2H), 2.11 (s, 3H),
1.80 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ 163.68, 150.40, 146.41,
145.95, 136.18, 129.70, 126.65, 122.41, 121.93, 116.07, 109.57, 109.42,
84.42, 83.82, 60.70, 59.06, 37.08, 17.61, 12.21; HRMS (ESI) m/z: calcd
for C20H24N6O4Na: 435.1751 [M+Na]+; found: 435.1752.
1-(5-(Hydroxymethyl)-4-(4-((p-tolylamino)methyl)-1H-1,2,3-
triazol-1-yl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-
dione (4d): Yield 89%;. 1H NMR (600 MHz, DMSO-d6): δ 11.38 (s,
1H), 8.14 (s, 1H), 7.81 (s, 1H), 6.97–6.88 (m, 2H), 6.55 (d, J = 12.0
Hz, 2H), 6.42 (t, J1 = 6.0 Hz, J2 = 12.0 Hz, 1H), 5.86 (s, 1H), 5.36–5.29
(m, 2H), 4.26 (s, 2H), 4.19 (d, J = 6.0 Hz, 1H), 3.70–3.58 (m, 2H),
2.73–2.58 (m, 2H), 2.14 (s, 3H), 1.81 (s, 3H); 13C NMR (151 MHz,
DMSO-d6): δ 163.68, 150.39, 146.15, 136.18, 129.26, 124.42, 122.41,
112.42, 109.57, 84.43, 83.82, 62.99, 60.68, 59.03, 37.06, 29.75, 28.60,
20.03, 13.96, 12.21; HRMS (ESI) m/z: calcd for C20H24N6O4Na:
435.1751 [M+Na]+; found: 435.1755.
Fig. 3 Binding modes of compounds containing a halogen
atom on the benzene ring in the active site of DHFR: (a) 4b;
(b) 4j; (c) 4m.
on a Bruker 600 MHz spectrometer operating at 600 and 151 MHz,
respectively, using deuterated dimethyl sulfoxide (DMSO-d6) as
solvent and TMS as internal standard. Chemical shifts are expressed
as δ ppm. High-resolution mass spectra were obtained on a Bruker
MicrOTOF-Q II mass spectrometer with an ESI source.
Synthesis of 2; general procedure
Compounds 2 were prepared according to the previously reported
method.14
A
mixture of propargyl bromide (20.0 mmol), 2-ethylaniline
(20.0 mmol), anhydrous potassium carbonate (40.0 mmol) and
potassium iodide (1.0 mmol) in N,N-dimethylformamide (35 mL) was
vigorously stirred at room temperature. After completion of the reaction,
as indicated by TLC, the reaction mixture was diluted with ethyl acetate
(40 mL) and washed with brine (20 mL × 3). The organic phases were
dried over Na2SO4 and concentrated in vacuo. The crude product was
purified by column chromatography on silica gel using ethyl acetate/
petroleum ether as eluent to afford 2a as a yellow oily liquid.
2-Ethyl-N-(prop-2-yn-1-yl)aniline (2a): Yield 87%; 1H NMR
(400 MHz, DMSO-d6): δ 7.05–7.01 (m, 1H), 6.98 (d, J = 8 Hz, 1H),
6.65 (d, J = 8 Hz, 1H), 6.62–6.58 (m, 1H), 5.36 (s, 1H), 3.90 (d,
J = 1.0 Hz, 2H), 3.00 (t, J = 4.0 Hz, 1H), 2.45 (q, J = 8.0 Hz, 2H), 1.12
(t, J = 8.0 Hz, 3H).
1- (5- (Hydroxymethyl) - 4- (4- (((2-methyl-3-nitrophenyl)
amino)methyl)-1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-
methylpyrimidine-2,4(1H,3H)-dione (4e): Yield 81%; 1H NMR
(600 MHz, DMSO-d6): δ 11.34 (s, 1H), 8.14 (s, 1H), 7.80 (s, 1H),
7.18 (t, J = 12.0 Hz, 1H), 6.97 (d, J = 12.0 Hz, 1H), 6.88 (d, J = 12.0
Hz, 1H), 6.41 (t, J = 12.0 Hz, 1H), 6.17 (t, J1 = 6.0 Hz, J2 = 12.0 Hz,
1H), 5.35–5.33 (m, 1H), 5.26 (t, J1 = 6.0 Hz, J2 = 12.0 Hz, 1H), 4.47 (d,
J = 12.0 Hz, 2H), 4.19 (d, J = 6.0 Hz, 1H), 3.69–3.58 (m, 2H), 2.71–2.59
(m, 2H), 2.14 (s, 3H), 1.80 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ
163.67, 151.42, 150.39, 147.26, 145.64, 136.16, 127.04, 122.51, 114.63,
112.84, 110.39, 109.57, 84.39, 83.81, 62.99, 60.69, 59.11, 37.08, 12.42,
12.20; HRMS (ESI) m/z: calcd for C20H23N7O6Na: 480.1602 [M+Na]+;
found: 480.1616.
Compounds 2b–r were obtained as pale yellow to reddish brown
liquids using the same procedure.

648 JOURNAL OF CHEMICAL RESEARCH 2017
1- (5- (Hydroxymethyl) - 4- (4- (((4-methyl-3-nitrophenyl)
amino)methyl)-1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-
methylpyrimidine-2,4(1H,3H)-dione (4f): Yield 94%; 1H NMR
(600 MHz, DMSO-d6): δ 11.34 (s, 1H), 8.19 (s, 1H), 7.81 (s, 1H),
7.18 (t, J1 = 18.0 Hz, J2 = 12.0 Hz, 2H), 6.92 (d, J = 12.0 Hz, 1H), 6.60
(t, J1 = 6.0 Hz, J2 = 12.0 Hz, 1H), 6.41 (t, J = 12.0 Hz, 1H), 6.17 (t,
J1 = 6.0 Hz, J2 = 12.0 Hz, 1H), 5.37–5.30 (m, 1H), 5.27 (t, J = 6.0 Hz,
1H), 4.36 (d, J = 6.0 Hz, 2H), 4.20 (d, J = 6.0 Hz, 1H), 3.71–3.59 (m,
2H), 2.74–2.59 (m, 2H), 2.33 (s, 3H), 1.81 (s, 3H); 13C NMR (151 MHz,
DMSO-d6): δ 163.67, 150.39, 149.41, 147.36, 145.22, 136.17, 132.97,
122.56, 118.84, 117.35, 109.57, 106.69, 84.41, 83.82, 60.66, 59.09,
38.29, 37.07, 18.56, 12.21; HRMS (ESI) m/z: calcd for C20H23N7O6Na:
480.1602 [M+Na]+; found: 480.1613.
1- (5- (Hydroxymethyl) -4- (4- (((2- (trifluoromethyl)phenyl)
amino)methyl)-1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-
methylpyrimidine-2,4(1H,3H)-dione (4l): Yield 93%; 1H NMR
(DMSO-d6): δ 11.34 (s, 1H), 8.11 (s, 1H), 7.80 (s, 1H), 7.39 (dd,
J1 = 12.0 Hz, J2 = 12.0 Hz, 2H), 6.89 (d, J = 12.0 Hz, 1H), 6.70 (t,
J = 12.0 Hz, 1H), 6.42 (t, J = 6.0 Hz, 1H), 6.00 (t, J = 6.0 Hz, 1H),
5.35 (dd, J1 = 6.0 Hz, J2 = 6.0Hz, 1H), 5.26 (t, J = 6.0 Hz, 1H), 4.49 (d,
J = 6.0 Hz, 2H), 4.19 (d, J = 6.0 Hz, 1H), 3.70–3.58 (m, 2H), 2.77–2.58
(m, 2H), 1.80 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ 164.19,
150.91, 145.96, 145.52, 136.67, 133.95, 126.72, 126.44, 124.63, 122.95,
115.93, 112.72, 110.08, 84.92, 84.34, 61.21, 59.66, 38.86, 37.60, 12.71;
HRMS (ESI) m/z: calcd for C20H21F3N6O4Na: 489.1474 [M+Na]+;
found: 489.1482.
1-(5-(Hydroxymethyl)-4-(4-(((2-hydroxyphenyl)amino)methyl)-
1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (4g): Yield 85%; 1H NMR (600 MHz, DMSO-d6):
δ 11.34 (s, 1H), 9.26 (s, 1H), 8.14 (s, 1H), 7.80 (s, 1H), 6.66–6.57 (m,
2H), 6.42 (d, J = 12.0 Hz, 1H), 5.34 (dd, J1 = 18.0 Hz, J2 = 12.0 Hz, 1H),
5.25 (t, J1 = 12.0 Hz, J2 = 6.0 Hz, 1H), 4.33 (s, 2H), 4.19 (d, J = 12.0
Hz, 1H), 3.70–3.58 (m, 2H), 2.75–2.57 (m, 2H), 1.81 (s, 3H), 1.24 (s,
2H); 13C NMR (151 MHz, DMSO-d6): δ 163.68, 150.39, 146.12, 144.15,
136.87, 136.19, 122.35, 119.58, 116.24, 113.40, 110.08, 109.56, 84.42,
83.81, 62.98, 60.69, 59.05, 37.06, 12.21; HRMS (ESI) m/z: calcd for
C19H23N6O5: 415.1724 [M+H]+; found: 415.1742.
1- (4- (4- (((2-Hydroxy-4-methylphenyl)amino)methyl) -1H-
1,2,3-triazol-1-yl)-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-
methylpyrimidine-2,4(1H,3H)-dione (4h): Yield 87%; 1H NMR
(600 MHz, DMSO-d6): δ 11.34 (s, 1H), 8.14 (s, 1H), 7.82 (s, 1H), 6.52
(s, 1H), 6.48–6.40 (m, 3H), 5.37–5.29 (m, 2H), 4.84 (s, 1H), 4.29 (s,
2H), 4.19 (d, J = 12.0 Hz, 1H), 3.69–3.59 (m, 3H), 2.75–2.58 (m, 2H),
2.10 (s, 3H), 1.81 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ 163.69,
150.40, 146.21, 144.23, 136.21, 134.48, 124.94, 122.36, 119.64, 114.52,
110.23, 109.56, 84.48, 83.82, 60.68, 59.71, 59.07, 37.06, 20.32, 12.20;
HRMS (ESI) m/z: calcd for C20H24N6O5Na: 451.1700 [M+Na]+; found:
451.1701.
1-(4-(4-(((4-Bromophenyl)amino)methyl)-1H-1,2,3-triazol-1-
yl)-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (4i): Yield 91%; 1H NMR (600 MHz, DMSO-d6): δ
11.34 (s, 1H), 8.15 (s, 1H), 7.80 (s, 1H), 7.20 (d, J = 12.0 Hz, 2H), 6.60
(d, J = 12.0 Hz, 2H), 6.42–6.30 (m, 2H), 5.36–5.26 (m, 2H), 4.34–4.28
(m, 2H), 4.19 (d, J = 6.0 Hz, 1H), 3.67–3.58 (m, 2H), 2.80–2.60 (m,
2H), 1.80 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ 163.69, 150.39,
147.58, 145.53, 136.18, 131.31, 122.51, 114.17, 109.58, 106.65, 84.41,
83.83, 75.92, 66.33, 60.69, 59.09, 37.50, 37.07; HRMS (ESI) m/z: calcd
for C19H21BrN6O4Na: 499.0705 [M+Na]+; found: 499.0709.
1-(4-(4-(((2-Fluorophenyl)amino)methyl)-1H-1,2,3-triazol-1-yl)-5-
(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-2,4(1H,3H)-
dione (4m): Yield 88%;1H NMR (DMSO-d6): δ 11.34 (s, 1H), 8.15 (s, 1H),
7.81 (s, 1H), 7.04–6.93 (m, 2H), 6.79 (t, J = 12.0 Hz, 1H), 6.56 (dd, J1 = 12.0
Hz, J2 = 12.0 Hz, 1H), 6.42 (s, 1H), 5.93 (s, 1H), 5.36–5.26 (m, 2H), 4.40
(d, J = 6.0 Hz, 2H), 4.20 (s, 1H), 3.71–3.58 (m, 2H), 2.74–2.62 (m, 2H),
1.81 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ 164.20, 152.19, 150.91,
146.40, 136.68, 125.16, 122.98, 116.38, 114.70, 112.67, 110.08, 84.94,84.33,
61.19, 60.23, 59.58, 38.74, 37.57, 12.71; HRMS (ESI) m/z: calcd for
C19H21FN6O4Na: 439.1506 [M+Na]+; found: 439.1482.
1-(4-(4-(((4-Fluorophenyl)amino)methyl)-1H-1,2,3-triazol-1-
yl)-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (4n): Yield 91%; 1H NMR (DMSO-d6): δ 11.35
(s, 1H), 8.16 (s, 1H), 7.82 (s, 1H), 6.92 (t, J = 12.0Hz, 2H), 6.66–6.62
(m, 2H), 6.43 (s, 1H), 5.99 (s, 1H), 5.37–5.27 (m, 2H), 4.28–4.20 (m,
3H), 3.72–3.60 (m, 2H), 2.74–2.64 (m, 2H), 1.81 (s, 3H); 13C NMR
(151 MHz, DMSO-d6): δ 164.20, 155.70, 154.17, 150.91, 146.38,
145.56, 136.68, 122.98, 115.75, 115.60, 113.49, 110.10, 84.94, 84.34,
61.19, 59.58, 37.58, 12.72; HRMS (ESI) m/z: calcd for C19H21FN6O4Na:
439.1506 [M+Na]+; found: 439.1482.
1- (5- (Hydroxymethyl) -4- (4- (((2-methoxy-4-nitrophenyl)
amino)methyl)-1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-
methylpyrimidine-2,4(1H,3H)-dione (4o): Yield 87%; 1H NMR
(DMSO-d6): δ 11.35 (s, 1H), 8.17 (s, 1H), 7.83–7.81 (m, 2H), 7.59
(d, J = 6.0 Hz, 1H), 7.04 (t, J1 = 12.0 Hz, J2 = 6.0 Hz, 1H), 6.74 (d,
J = 12.0 Hz, 1H), 6.42 (t, J = 12.0 Hz, 1H), 5.37–5.27 (m, 2H), 4.55 (d,
J = 12.0 Hz, 2H), 4.20 (d, J = 6.0 Hz, 1H), 3.92 (s, 3H), 3.72–3.58 (m,
2H), 2.75–2.59 (m, 2H), 1.81 (s, 3H); 13C NMR (151 MHz, DMSO-d6):
δ 164.20, 150.90, 145.58, 145.20, 144.74, 136.67, 136.28, 123.11, 120.13,
110.10, 107.58, 104.95, 84.89, 84.34, 61.20, 59.68, 56.32, 38.16, 37.58,
12.70; HRMS (ESI) m/z: calcd for C20H23N7O7Na: 496.1557 [M+Na]+;
found: 496.1552.
1-(4-(4-(((2-Bromophenyl)amino)methyl)-1H-1,2,3-triazol-1-
yl)-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (4j): Yield 83%; 1H NMR (DMSO-d6): δ 11.34
(s, 1H), 8.14 (s, 1H), 7.81 (s, 1H), 7.40 (d, J = 8.0 Hz, 1H), 7.16 (d,
J = 8.0 Hz, 1H), 6.77 (d, J = 8.0 Hz, 1H), 6.53 (t, J = 8.0 Hz, 1H),
6.46–6.39 (m, 1H), 5.69 (t, J = 8.0 Hz, 1H), 5.39–5.23 (m, 2H), 4.45
(d, J = 8.0 Hz, 2H), 4.19 (s, 1H), 3.75–3.65 (m, 1H), 3.65–3.56 (m, 1H),
2.74–2.58 (m, 2H), 1.80 (s, 3H); 13C NMR (151 MHz, DMSO-d6): δ
164.2, 150.9, 146.0, 145.1, 136.7, 132.7, 129.0, 123.0, 118.0, 112.2, 110.1,
109.1, 84.9, 84.3, 61.2, 59.6, 39.1, 37.6, 12.7; HRMS (ESI) m/z: calcd for
C19H21BrN6O4Na: 499.0705 [M+Na]+; found: 499.0721.
1-(4-(4-(((2,4-Dimethoxyphenyl)amino)methyl)-1H-1,2,3-triazol-
1-yl)-5-(hydroxymethyl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
2,4(1H,3H)-dione (4p): Yield 95%;1H NMR (DMSO-d6): δ 11.35
(s, 1H), 8.16 (s, 1H), 7.82 (s, 1H), 6.97 (t, J = 12.0 Hz, 1H), 6.43 (dd,
J1 = 12.0, J2 = 6.0 Hz, 1H), 6.23 (dd, J1 = 12.0, J2 = 6.0 Hz, 2H), 6.14
(dd, J1 = 6.0, J2 = 6.0 Hz, 1H), 6.06 (t, J = 6.0 Hz, 1H), 5.37–5.28
(m, 2H), 4.30 (s, 2H), 4.21 (s, 1H), 3.66 (s, 5H), 3.37 (dd, J1 = 6.0,
J2 = 6.0 Hz, 2H), 2.76–2.62 (m, 2H), 1.81 (s, 3H); 13C NMR (151 MHz,
DMSO-d6): δ 164.2, 160.76, 150.91, 150.21, 146.48, 136.69, 130.03,
122.96, 110.09, 105.82, 102.10, 98.50, 84.95, 84.35, 61.19, 59.56,
55.08, 39.06, 37.59, 12.72; HRMS (ESI) m/z: calcd for C21H26N6O6Na:
481.1812 [M+Na]+; found: 481.1810.
1- (5- (Hydroxymethyl) -4- (4- (((4- (trifluoromethyl)phenyl)
amino)methyl)-1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-
methylpyrimidine-2,4(1H,3H)-dione (4q): Yield 84%; 1H NMR
(600 MHz, DMSO-d6): δ 11.36 (s, 1H), 8.17 (s, 1H), 7.80 (s, 1H),
7.40–7.38 (d, J = 12.0 Hz, 2H), 6.82–6.75 (m, 3H), 6.41 (t, J = 6.0 Hz,
1H), 5.36 (s, 2H), 4.38 (d, J = 12.0 Hz, 2H), 4.21 (s, 1H), 3.82–3.67 (m,
2H), 2.73–2.63 (m, 2H), 1.81 (s, 3H); 13C NMR (151 MHz, DMSO-d6):
δ 164.31, 151.67, 150.91, 145.72, 136.70, 128.43, 126.65, 124.84, 123.12,
112.07, 110.17, 84.93, 84.43, 61.19, 59.69, 38.38, 37.58, 12.64; HRMS
(ESI) m/z: calcd for C20H21F3N6O4Na: 489.1474 [M+Na]+; found:
489.1454.
1-(5-(Hydroxymethyl)-4-(4-(((2-nitrophenyl)amino)methyl)-
1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
1
2,4(1H,3H)-dione (4k): Yield 89%; H NMR (600 MHz, DMSO-d6):
δ 11.34 (s, 1H), 8.56 (t, J = 12.0 Hz, 1H), 8.23 (s, 1H), 8.08 (d,
J = 12.0 Hz, 1H), 7.80 (s, 1H), 7.53 (t, J = 12.0 Hz, 1H), 7.14 (d, J = 12.0
Hz, 1H), 6.72 (t, J = 12.0 Hz, 1H), 6.41 (t, J = 12.0 Hz, 1H), 5.37–5.32
(m, 1H), 5.27 (t, J = 12.0 Hz, 1H), 4.68 (d, J = 12.0 Hz, 2H), 4.19 (d,
J = 12.0 Hz, 1H), 3.70–3.58 (m, 2H), 2.74–2.58 (m, 2H), 1.80 (s, 3H);
13C NMR (151 MHz, DMSO-d6): δ 163.68, 150.39, 144.61, 144.41,
136.51, 136.17, 131.35, 126.16, 122.66, 115.64, 114.72, 109.58, 84.38,
83.83, 60.71, 59.24, 37.97, 37.06, 12.20; HRMS (ESI) m/z: calcd for
C19H21N7O6: 444.1671 [M+H]+; found: 444.1677.

JOURNAL OF CHEMICAL RESEARCH 2017 649
1-(5-(Hydroxymethyl)-4-(4-(((3-methoxyphenyl)amino)methyl)-
(No. 122107000014) and the Foundation of Henan
Educational Committee (14A350005, 15A150054, 16A350015,
5201039140120) for financial support.
1H-1,2,3-triazol-1-yl)tetrahydrofuran-2-yl)-5-methylpyrimidine-
1
2,4(1H,3H)-dione (4r): Yield 93%; H NMR (DMSO-d6): δ 11.36 (s,
1H), 8.14 (s, 1H), 7.82–7.79 (m, 1H), 7.64 (dd, J1 = 12.0, J2 = 6.0 Hz,
1H), 7.19 (t, J = 6.0 Hz, 1H), 6.71 (d, J = 6.0 Hz, 1H), 6.46–6.38 (m,
2H), 5.35 (t, J = 6.0 Hz, 3H), 4.44 (d, J = 6.0 Hz, 2H), 4.21 (d, J = 6.0
Hz, 1H), 3.76–3.60 (m, 5H), 2.77–2.58 (m, 2H), 1.81 (s, 3H); 13C NMR
(151 MHz, DMSO-d6): δ 164.3, 150.9, 147.4, 145.9, 139.3, 136.7, 129.8,
123.1, 119.1, 111.4, 110.2, 85.3, 84.9, 84.4, 61.2, 59.7, 39.3, 37.6, 12.7;
HRMS (ESI) m/z: calcd for C20H24N6O5Na: 451.1706 [M+Na]+; found:
451.1726.
Electronic Supplementary Information
The H NMR spectra of 2a is available through: http://
ingentaconnect.com/content/stl/jcr/2017/00000041/00000011/
art00007
Received 15 September 2017; accepted 19 October 2017
Paper 1705001
https://doi.org/10.3184/174751917X15094552081189
Published online: 17 November 2017
Antibacterial activity
The antibacterial activity of the synthetic compounds against
E. coli and S. aureus was assessed using a broth dilution assay to
determine the MIC values. Triplicate analyses were performed
for each of the standard strains. Stock cultures were preserved
at −70 °C using glycerol 10% (v/v) as the cryoprotectant, and
the strains were grown in tryptic soy broth (TSB) at 37 °C for
18 h.
MIC was determined using the standard serial dilution
method, according to the guidelines of the Clinical Laboratories
Standard Institute (CLSI).15 Compounds 4a–r were dissolved in
DMSO. The MIC values were determined using streptomycin
sulfate and doxycycline as positive controls. The results are
summarised in Table 1 and expressed as molar concentrations
(mM) for the discussion of their structure–activity relationships.
References
1
2
3
S. Baluja, N. Kacchadia and S. Chanda, Pharm. Chem. J., 2012, 46, 117.
A. Quintás-Cardama and J. Cortes, Future Oncol., 2008, 4, 611.
K.N. Mohana, B.N. Prasanna Kumar and L. Mallesha, Drug Inven. Today,
2013, 5, 216.
4
5
6
7
8
9
K. Dabak, Ö. Sezer, A. Akar and O. Anaç, Eur. J. Med. Chem., 2003, 38,
215.
G.Q. Xu, J.L. Zhao, Y.Q. Jiang, P. Zhang and W. Li, J. Chem. Res., 2016,
40, 269.
G.Q. Xu, S. Mao, L.F. Mao, Y.Q. Jiang, P. Zhang and W. Li, Z. Naturforsch
B, 2016, 71, 953.
Y.C. Duan, Y.C. Ma, E. Zhang, X.J. Shi, M.M. Wang, X.W. Ye and H.M.
Liu, Eur. J. Med. Chem., 2013, 62, 11.
S. Shafi, M.M. Alam, N. Mulakayala, C. Mulakayala, G. Vanaja, A.M.
Kalle, R. Pallu and M.S. Alam, Eur. J. Med. Chem., 2012, 49, 324.
V.V. Rostovtsev, L.G. Green, V.V. Fokin and K.B. Sharpless, Angew. Chem.
Int. Ed., 2002, 41, 2596.
Computational simulation
10 B. Garudachari, A.M. Isloor, M.N. Satyanarayana, H.-K. Fun and G.
Hegde, Eur. J. Med. Chem., 2014, 74, 324.
11 G. Surineni, P. Yogeeswari, D. Sriram and S. Kantevari, Med. Chem. Res.,
2015, 24, 1298.
12 C.P. Kaushik, K. Kumar, K. Lal, B. Narasimhan and A. Kumar, Monatsh.
Chem., 2016, 147, 817.
13 Y.Q. Jiang, B.Q. Ren, X.M. Lv, W.W. Zhang, W. Li and G.Q. Xu, J. Chem.
Res., 2015, 39, 243.
The X-ray crystal structure of the DHFR complexed with
folate and NADP+ from E. coli (PDB ID: 4X5F) was obtained
from a protein data bank in PDB format as starting point. The
synthesised compounds tested in this study were docked to
E. coli DHFR using the Glide program.
Acknowledgements
14 K.C. Majumdar and S. Ganai, Tetrahedron Lett., 2013, 54, 6192.
15 Clinical and Laboratory Standards Institute (CLSI), Methods for dilution
antimicrobial susceptibility tests for bacteria that grow aerobically;
approved standard, 7th edn. CSLI document M7-A7. Clinical and
Laboratory Standards Institute, Wayne, Pennsylvania, USA, 2012.
The authors acknowledge the Innovative Talents Program
of Henan Province (No. 164100510015), the Production-
Learning-Research Cooperation Project of Henan Province