Green chemoselective synthesis of thiazolo[3,2-a]pyridine derivatives and evaluation of their antioxidant and cytotoxic activities

Article abstract of DOI:10.1016/j.bmcl.2009.08.046

The green chemoselective synthesis of thiazolo[3,2-a]pyridine derivatives was achieved in water via microwave-assisted three-component reactions of malononitrile, aromatic aldehydes and 2-mercaptoacetic acid with molar ratios of 2:1:1.5 and 2:2.2:1, respe

Full text of DOI:10.1016/j.bmcl.2009.08.046

Bioorganic & Medicinal Chemistry Letters 19 (2009) 5565–5568
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Green chemoselective synthesis of thiazolo[3,2-a]pyridine derivatives
and evaluation of their antioxidant and cytotoxic activities
a,
Feng Shi a, Chunmei Li a, Ming Xia b, Kangjie Miao a, Yanxia Zhao a, Shujiang Tu a, , Weifa Zheng
,
*
*
Ge Zhang a, Ning Ma a
a School of Chemistry and Chemical Engineering, Key Laboratory of Biotechnology for Medicinal Plant, Xuzhou Normal University, Xuzhou, Jiangsu 221116, PR China
b Department of Anesthesiology, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210093, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
The green chemoselective synthesis of thiazolo[3,2-a]pyridine derivatives was achieved in water via micro-
wave-assisted three-component reactions of malononitrile, aromatic aldehydes and 2-mercaptoacetic acid
with molar ratios of 2:1:1.5 and 2:2.2:1, respectively. These compounds were subject to the experiments of
antioxidant activity and cytotoxicity to carcinoma HCT-116 cells and mice lymphocytes. Nearly all of the
tested compounds possessed potent capacities for scavenging free radicals. In addition, most of these com-
pounds showed cytotoxicity to HCT-116 cells and mice lymphocytes with no selectivity. Of these, only
thiazolo[3,2-a]pyridine derivative 5d suggested selective cytotoxicity to tumor cell line HCT-116 cells.
Ó 2009 Elsevier Ltd. All rights reserved.
Received 12 June 2009
Revised 11 August 2009
Accepted 12 August 2009
Available online 15 August 2009
Keywords:
Thiazolopyridine
Green synthesis
Antioxidant activity
Cytotoxicity
Thiazolo[3,2-a]pyridines, containing two fused heterocyclic
motifs in one molecule, have many important bioactivities such
as inhibiting beta-amyloid production,1 potent CDK2-Cyclin A
proaches to synthesize this class of novel thiazolo[3,2-a]pyridines
1 are of theoretical and practical significance.
In recent years, microwave-assisted synthesis using water as
solvent has become an active topic because of the use of greener
and inexpensive solvents and energy efficiency, which is in well
accordance with the two prominent green chemistry principles.8
inhibitor,2
a
-glucosidase inhibitor,3 potential uterus stimulant,4
antibacterial and antifungal activities.5 5-Amino-7-aryl-6,8-dicy-
ano-3-oxo-2,3-dihydro-7H-thiazolo[3,2-a]pyridines 1 (Scheme 2),
representing a novel class of thiazolo[3,2-a]pyridine derivatives,
possess electron acceptors and donors and are assumed to scav-
enge free radicals. However, to the best of our knowledge, the
synthetic methods for this class of thiazolo[3,2-a]pyridines and
their bioactivities including antioxidant and cytotoxic activities
have not been well investigated.
Ph
Ph
NC
CONHR
S
CONHR
+
N
H
MeOH
NC
Cl
COOMe
+
H2N
N
CSNH2
CN
O
R = H, Me
Previous studies showed that the synthesis of the compounds
with similar structures can be conducted by multi-step reactions
or from complicated starting materials.6 A relatively simple synthe-
sis was achieved by the three-component reactions of benzylidene-
malononitrile, N-substituted thiocarbamoylacetamides and methyl
2-chloroacetate catalyzed by piperidine in methanol (Scheme 1).7
Unfortunately, synthesis of these compounds involves the use
of volatile and toxic organic catalyst and solvent. In addition, start-
ing materials of benzylidenemalononitrile and N-substituted thi-
ocarbamoylacetamides must be synthesized beforehand. This
leads to the elongation of reaction time and reduction in the yield
of target products. Thus attempts to develop green and facile ap-
Scheme 1.
Ar
N
CN
NC
MW, 90ºC /H2O
2:3:4 = 2:1:1.5
H2N
S
O
SH
CN
1
5
+
+
ArCHO
Ar
CN
2
COOH
CN
NC
3
4
MW, 100ºC/H2O
2:3:4 = 2:2.2:1
H2N
N
S
O
Ar
* Corresponding authors. Tel./fax: +86 516 83500365 (S.T.); +86 516 83403179
(W.Z.).
E-mail addresses: laotu2001@263.net (S. Tu), yyzw@xznu.edu.cn (W. Zheng).
Scheme 2.
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.08.046

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F. Shi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5565–5568
On the other hand, multi-component reactions (MCR) occupy an
outstanding position in organic and medicinal chemistry by virtue
of the intrinsic atom economy and selectivity, simpler procedures
and equipments, time and energy savings, as well as environmen-
tal friendliness.9 Hence, microwave-assisted multi-component
reactions in water has become a green and facile approach to the
synthesis of biologically important heterocyclic compounds.
As a continuation of our efforts on the green synthesis and po-
tential bioactivities of heterocyclic compounds,10 we report the
green chemoselective synthesis of thiazolo[3,2-a]pyridines 1 and
5 via microwave-assisted three-component reactions of malono-
nitrile 2, aromatic aldehydes 3 and 2-mercaptoacetic acid 4 with
different molar ratios in water (Scheme 2) and their antioxidant
and cytotoxic activities.
diation (MW) in water. Yet the end products of this reaction turned
out to be the mixture of compound 1b and 5b (Table 1, entry 1).
After conducting numerous experiments (Table 1, entries 2–8),
the molar ratio of the three reactants for exclusively synthesizing
compound 1b was optimized to be 2:1:1.5.
In order to optimize the reaction temperature, the reaction of 2
(2 mmol), 3b and 4 in a molar ratio of 2:1:1.5 was conducted in
water under MW at temperatures ranging from 70 °C to 110 °C,
with an increment of 10 °C. The results (Table 1, entries 6 and 9–
12) showed that the reaction at 90 °C coincided with the highest
yield of 1b.
Similar attempts were also made to exclusively synthesize
thiazolo[3,2-a]pyridine 5b using different molar ratio of the same
three reactants for synthesizing 1b, which resulted in the optimi-
zation of molar ratio being 2:2.2:1 and the reaction temperature
at 100 °C.
The synthesis of thiazolo[3,2-a]pyridine 1b was initiated by the
three-component reaction of malononitrile 2 (2 mmol), 4-bromo-
phenyl aldehyde 3b (1 mmol) and 2-mercaptoacetic acid
(1 mmol) with molar ratio of 2:1:1 at 90 °C under microwave irra-
4
A series of thiazolo[3,2-a]pyridine derivatives 1 and 5 were che-
moselectively synthesized (Scheme 2) under their respective opti-
mal reaction conditions and the results are summarized in Table 2.
Obviously, this protocol can be applied not only to aromatic alde-
hydes with electron-withdrawing groups, but also to those with
electron-donating groups in high yields. Therefore, this green
chemoselective approach has wide scope of applicability in synthe-
sizing thiazolo[3,2-a]pyridine derivatives.
Table 1
Reaction conditions optimization for the synthesis of 1ba
Br
Br
Br
The structures of thiazolo[3,2-a]pyridines 1 and 5 were unam-
biguously characterized by IR, 1H NMR, 13C NMR and HRMS
(ESI).11 For example, in the 1H NMR spectrum of 1f, the two dou-
blets at 6.95 and 7.24 ppm with a coupling constant of 8.4 Hz were
assigned to the four protons of the p-methoxyphenyl group, which
were verified by a singlet of 3H at 3.78 ppm designated as the
methoxy group. The singlet at 7.32 ppm was identified as the
two protons of the amino group and that at 4.40 ppm was assigned
to the tertiary proton in the pyridine ring. Another singlet at
4.21 ppm was designated as the two methylene protons in the thi-
azole ring. In the 13C NMR spectrum of 1f, the pyridine carbons
showed chemical shifts at 158.8, 148.2, 85.4, 63.9 and 40.3 ppm,
while the carbonyl carbon and methylene carbon contained in
the thiazole ring were located at 172.5 and 33.2 ppm, respectively.
Besides, the carbons in the symmetric aromatic ring and the at-
tached methoxyl exhibited chemical shifts at 147.7, 134.3, 128.9
(2C), 114.1 (2C) and 55.1 ppm, while the two cyano-group carbons
were positioned at 118.8 and 116.4 ppm. The IR spectrum of com-
pound 1f showed strong absorbance at 3394 and 3284 cmÀ1 due to
the presence of NH2 group, two sharp absorptions at 2208 and
2197 cmÀ1 suggesting the existence of two CN groups and strong
absorption at 1734 cmÀ1 contributed by the C@O group.
CN
S
CN
NC
NC
SH
CN
+
MW
H2O
+
+
H2N
N
H2N
N
S
CN
COOH
4
CHO
3b
O
Br
2
O
5b
1b
Entry
2:3b:4b
T (°C)
1b:5bb
Yield of 1bc (%)
1
2
3
4
5
6
7
8
9
2:1:1
90
90
90
90
90
90
90
90
70
1.8:1
2.2:1
3.1:1
5.2:1
57
62
70
77
82
89
73
70
74
81
84
78
2:1:1.1
2:1:1.2
2:1:1.3
2:1:1.4
2:1:1.5
2:1:1.6
2:1:1.7
2:1:1.5
2:1:1.5
2:1:1.5
2:1:1.5
7.8:1
>99:1
11.3:1
11.1:1
10.6:1
>99:1
>99:1
11.6:1
10
11
12
80
100
110
a
All the reactions were carried out in the presence of 2 (2 mmol), 3b and 4 with
various molar ratios in 2 mL water under MW at the initial power of 100 W and the
maximum power of 250 W.
b
Molar ratios.
Isolated yields.
c
Table 2
Synthesis of 1 and 5 under microwave irradiationa
Entry
Product
Ar
Time (min)
Yieldb (%)
Mp (lit.) (°C)
1
2
3
4
5
6
7
8
9
10
11
12
13
1a
1b
1c
1d
1e
1f
1g
1h
5a
5b
5c
5d
5e
4-FC6H4
4-BrC6H4
2-ClC6H4
2,4-Cl2C6H3
C6H5
4-CH3OC6H4
3,4-OCH2OC6H3
4-CH3C6H4
4-NO2C6H4
4-BrC6H4
3-NO2C6H4
4-OH-3-NO2C6H3
2-Thienyl
6
6
6
7
8
8
9
8
7
6
6
6
7
87
89
83
80
81
82
85
83
87
89
84
87
82
267–269
292–294
234–236
269–271
238–240
236–238
271–273
265–267
258–260 (243–245)5b
272–273 (278–280)5b
278–279
269–271
247–248
a
All the synthesis of 1 were carried out in the presence of 2 mmol of 2, 1 mmol of 3 and 1.5 mmol of 4 in 2 mL water under MW at 90 °C with the initial power of 100 W and
the maximum power of 250 W. All the synthesis of 5 were carried out in the presence of 2 mmol of 2, 2.2 mmol of 3 and 1 mmol of 4 in 2 mL water under MW at 100 °C with
the initial power of 100 W and the maximum power of 250 W.
b
Isolated yields.

F. Shi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5565–5568
5567
Table 3
Ar CHO
Free radicals scavenging capacities of compounds 1 and 5a
3
CN
CN
Ar
2
NC
S
Entry Compound
Ar
DPPH (%/mg)
OH (%/mg)
NC
C N
S
CN
1
2
3
4
5
6
7
8
9
1a
1b
1c
1d
1e
1f
1g
1h
5a
5b
5c
5d
4-FC6H4
4-BrC6H4
2-ClC6H4
2,4-Cl2C6H3
C6H5
4-CH3OC6H4
3,4-OCH2OC6H3
4-CH3C6H4
4-NO2C6H4
4-BrC6H4
3-NO2C6H4
4-OH-3-
880.99 7.00
67.65 3.08
811.85 3.92
708.64 23.77 486.36 6.01
884.94 32.87 431.82 15.91
654.55 12.03
250.00 19.42
570.45 29.81
2
4
-H2O
NH
O
SH
N
HOOC
NC
C
COOH
6
N
7
8
140.25 3.08
207.58 18.23
Ar
Ar
N
Ar
S
CN
444.44 13.17 431.06 15.96
858.77 16.85 609.85 28.60
387.65 23.63 297.73 12.03
107.65 6.68
45.93 2.96
NC
S
CN
NC
S
CN
NH
NC
C
N
N
NH2
N
10
11
12
125.00 7.87
56.82 6.82
O
O
O
9
1
108.64 12.60 201.52 15.13
NO2C6H3
2-Thienyl
Scheme 3.
13
14
5e
717.53 22.29 665.91 15.75
L
-Ascorbic
188.12 5.13
108.65 4.48
acidb
A plausible mechanism for the formation of compounds 1 is
suggested in Scheme 3. Firstly, the nucleophilic addition of 2-mer-
captoacetic acid 4 to malononitrile 2 yielded the intermediate 6,
which further gave thiazolinone derivatives 7 via intramolecular
dehydration. Then, the intermediate 7 underwent Michael addition
with arylidenemalononitrile 8 formed from Knoevenagel conden-
sation of malononitrile 2 and aldehyde 3 to give an open-chain
intermediate 9, which was subsequently intramolecular cyclized
and isomerized to afford the product 1. The formation mechanism
of compound 5 is similar to that of compound 1, which has been
demonstrated in literature.5b,12
In order to survey the possible biological activities of this class
of compounds, derivatives of thiazolo[3,2-a]pyridines 1 and 5 were
subject to the test of antioxidant activity and cytotoxicity to carci-
noma cell line HCT 116 (ATTC CCL 247) and mice lymphocytes. The
antioxidant activity is represented by their capacities for scaveng-
ing 1,1-diphenyl-2-picrylhydrazyl free radical (DPPH) and hydro-
xyl free radical (OH) using protocols described in our previous
works,10d,e and the results are summarized in Table 3. Nearly all
the tested compounds showed strong capacities for scavenging
a
The scavenging capacities were represented as percentage inhibition
(mean SD, n = 3) of the free radicals by 1 mg tested compound.
b
L-Ascorbic acid was used as a positive control and purchased locally with
purities higher than 98.5%. Other chemicals for bioassay of antioxidant activity are
obtained from Sigma.
Meanwhile these compounds also inhibited the proliferation of
mice lymphocytes up to 71.28%. In general, these compounds
inhibited the growth of mice lymphocytes in a rate higher than
they did to HCT-116 cells. Only compound 5d suggested higher
selective cytotoxicity to HCT 116 cells. Thus the simultaneously
existence of 4-OH and 3-NO2 in Ar group of 5d might be the neces-
sity for selectively inhibiting the growth of tumor cells.
It has been reported that most natural products inhibiting the
growth of tumor cells also possess antioxidant activity13, and anti-
oxidant natural products are believed to be the pharmaceuticals
able to treat or prevent oxidative stress-induced cancers.14 In our
study, however, high antioxidant activity of thiazolo[3,2-a]pyri-
dine derivatives 1 and 5 does not coincide with their selective cyto-
toxicity to tumor cell line HCT-116, which seems to indicate that
highly antioxidant activity of these compounds does not guarantee
their selective cytotoxicity to tumor cells.
This study has achieved the green chemoselective synthesis of
thiazolo[3,2-a]pyridine derivatives and examined their in vitro anti-
oxidant and cytotoxic activities. Further experiments are needed to
determine the selective cytotoxicity to other tumor cell lines.
Regardless, thisstudy does providea green shortcut for thesynthesis
of thiazolo[3,2-a]pyridine derivatives with high chemoselectivity,
environmental friendliness, short reaction time and high yields of
target products. More importantly, bioassay of these compounds re-
sulted in the finding of thiazolo[3,2-a]pyridine derivative 5d with
DPPH and OH compared with those of the positive control L-Ascor-
bic acid. The top three for scavenging DPPH radical are compounds
1e, 1a and 1h with phenyl, 4-fluorobenzyl and 4-methylbenzyl as
Ar group, respectively. While the top three for scavenging OH are
compounds 5e, 1a and 1h with 2-thienyl, 4-fluorobenzyl and 4-
methylbenzyl as Ar group, separately. It is noted that the substitu-
ents of Ar group in these potent antioxidant compounds include
both electron-withdrawing and electron-donating group. These
aromatic groups possibly play as electron receptors and donors
that underlie their antioxidant activities.
The cytotoxic assay11 (Table 4) showed that the tested
compounds inhibited proliferation of HCT 116 cells up to 63.8%.
Table 4
Cytotoxicity of compounds 1 and 5a
Entry
compoundb
Ar
Inhibition rate on HCT-116 (%)
Inhibition rate on lymphocytes (%)
1
2
3
4
5
6
7
8
9
10
11
12
13
1a
1b
1c
1d
1e
1f
1g
1h
5a
5b
5c
5d
5e
4-FC6H4
4-BrC6H4
2-ClC6H4
2,4-Cl2C6H3
C6H5
4-CH3OC6H4
3,4-OCH2OC6H3
4-CH3C6H4
4-NO2C6H4
4-BrC6H4
3-NO2C6H4
4-OH-3-NO2C6H3
2-Thienyl
47.77 2.85
43.55 0.88
54.06 2.54
29.93 4.66
50.84 1.92
50.17 2.61
51.78 2.57
47.42 2.35
35.45 1.87
56.65 2.07
52.99 1.61
41.22 1.84
63.81 4.22
66.88 0.88
69.59 3.57
60.36 3.81
55.35 6.79
64.84 2.97
70.60 2.08
62.02 3.35
63.77 4.76
51.69 1.83
71.28 0.84
60.35 4.40
5.31 0.26
55.42 3.31
a
The cytotoxicity was represented as percentage inhibition (mean SD, n = 3) of HCT-116 cells and lymphocytes.
The concentration of the tested compounds is 1 mg/mL.
b

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F. Shi et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5565–5568
4. Manhi, F. M.; Soliman, G. A. Bull. Fac. Pharm. 1993, 31, 265; Chem. Abstr. 1993,
121, 50029.
5. (a) El-Hag Ali, G.; Khalil, A.; Lamphon, R.; El-Maghraby, A. Phosphorus, Sulfur
Silicon Relat. Elem. 2005, 180, 1909; (b) El-Maghraby, A. A.; El-Hag Ali, G. A. M.;
Ahmed, A. H. A.; El-Gaby, M. S. A. Phosphorus, Sulfur Silicon Relat. Elem. 2002,
177, 293.
potent antioxidant activity and selective tumor cytotoxicity, which
gains some insights into the synthesis of the compounds able to in-
hibit tumor growth without evident side-toxicities.
6. Abbas-Temirek, H. H.; Harb, A. A.; Fandy, R. F.; Abo-Bakr, A. M. Assiut U. J. Chem.
2006, 35, 17; Chem. Abstr. 2006, 148, 495885.; Yakout, E. M. A. Egypt. J. Chem.
2003, 45, 1029; Chem. Abstr. 2003, 141, 314106.; (c) Elmoghayar, M. R. H.; El-
Agamey, A. A.; Nasr, M. Y. A.; Sallam, M. M. M. J. Heterocycl. Chem. 1984, 21,
1885.
7. Krauze, A.; Popelis, J.; Duburs, G. Tetrahedron 1998, 54, 9161.
8. Dallinger, D.; Kappe, C. O. Chem. Rev. 2007, 107, 2563.
9. (a) Ramon, D. J.; Miguel, Y. Angew. Chem., Int. Ed. 2005, 44, 1602; (b) Weber, L.;
Illgen, K.; Almstetter, M. Synlett 1999, 366; (c) Li, G.; Wei, H. X.; Kim, S. H.;
Carducci, M. D. Angew. Chem., Int. Ed. 2001, 40, 4277.
Acknowledgments
We are grateful for financial support by Natural Science Foun-
dation of China (Nos. 20672090 and 200810102050), the Prelimin-
ary Item of Xuzhou Normal University on NSFC (No. 08XLY04), the
Science & Technology Foundation of Xuzhou municipal govern-
ment (No. XM08C027) and Qing Lan Project (08QLT001).
10. (a) Tu, S.; Cao, X.; Hao, W.; Zhang, X.; Yan, S.; Wu, S.; Han, Z.; Shi, F. Org. Biomol.
Chem. 2009, 7, 557; (b) Tu, S.; Zhang, X.; Han, Z.; Cao, X.; Wu, S.; Yan, S.; Hao,
W.; Zhang, G.; Ma, N. J. Comb. Chem. 2009, 11, 428; (c) Wei, P.; Zhang, X.; Tu, S.;
Yan, S.; Ying, H.; Ouyang, P. Bioorg. Med. Chem. Lett. 2009, 19, 828; (d) Zheng,
W.; Zhang, M.; Zhao, Y.; Wang, Y.; Miao, K.; Wei, Z. Bioresour. Technol. 2009,
100, 1327; (e) Zheng, W.; Zhang, M.; Zhao, Y.; Miao, K.; Jiang, H. Bioresour.
Technol. 2009. doi:10.1016/j.biortech.2009.04.027.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2009.08.046.
11. The experimental procedures of synthesis and cytotoxic assay as well as the
spectroscopic data of the synthesized compounds are available in the
Supplementary data.
References and notes
1. Mullan, M. J.; Paris, D.; Bakshi, P. WO 2008070875, 2008; Chem. Abstr. 2008,
12. Kambe, S.; Saito, K.; Sakurai, A.; Midorikawa, H. Synthesis 1981, 531.
13. Anderson, R. A.; Broadhurst, C. L.; Polansky, M. M.; Schmidt, W. F.; Khan, A.;
Flanagan, V. P.; Schoene, N. W.; Graves, D. J. J. Agric. Food Chem. 2004, 52, 65.
14. Zheng, W.; Zhao, Y.; Zhang, M.; Wei, Z.; Miao, K.; Sun, W. Med. Mycol. 2009.
doi:10.1080/13693780802653933.
149, 45259.
2. Vadivelan, S.; Sinha, B. N.; Irudayam, S. J.; Jagarlapudi, S. A. R. P. J. Chem. Inf.
Model. 2007, 47, 1526.
3. Park, H.; Hwang, K. Y.; Oh, K. H.; Kim, Y. H.; Lee, J. Y.; Kim, K. Bioorg. Med. Chem.
2008, 16, 284.