Synthesis, characterization and antibacterial studies of some 1,2,4-triazole derivatives containing a 6-chloropyridin-3-yl methyl moiety

Article abstract of DOI:10.1515/znb-2006-0315

5-(6-Chloropyridin-3-yl methyl)-4-phenyl-1,2,4-triazole-3-thiol (2) and 5-substituted-4-phenyl-1,2,4-triazole-3-thiols (3) were synthesized. Alkylation and aminomethylation reactions of these triazoles were also carried out. Some of the newly synthesized

Full text of DOI:10.1515/znb-2006-0315

Synthesis, Characterization and Antibacterial Studies of Some 1,2,4-Tri-
azole Derivatives Containing a 6-Chloropyridin-3-yl methyl Moiety
Bantwala S. Hollaa, Channamata S. Prasannaa,b, Boja Poojarya, Mithun Ashoka,
Kottapalli S. Raob, and Kanakamajalu Shridharab
a Department of Chemistry, Mangalore University, Mangalagangotri-574199, Karnataka, India
b Rallis Research Centre, #21&22, PIA, Peenya II Phase, Bangalore-560058, Karnataka, India
Reprint requests to Dr. Boja Poojary. E-mail: bojapoojary@yahoo.com
Z. Naturforsch. 61b, 334 – 338 (2006); received April 18, 2005
5-(6-Chloropyridin-3-yl methyl)-4-phenyl-1,2,4-triazole-3-thiol (2) and 5-substituted-4-phenyl-
1,2,4-triazole-3-thiols (3) were synthesized. Alkylation and aminomethylation reactions of these tri-
azoles were also carried out. Some of the newly synthesized compounds were screened for their
antibacterial activities.
Key words: 6-Chloropyridin-3-yl methyl Moiety, Triazoles, Aminomethylation,
Antibacterial Activity, Serial Dilution Method
Introduction
5-(6-chloropyridin-3-yl methyl)-4-phenyl-3-alkylsulf-
anyl-1,2,4-triazoles (4) by reacting it with alkyl/aryl
halides using aqueous sodium hydroxide. Aminometh-
ylation of triazole (2) was carried out with formal-
dehyde and a secondary amine to yield 5-(6-chloro-
pyridin-3-yl methyl)-4-phenyl-2-morpholino/N-meth-
ylpiperazino-4-yl methyl-3H-1,2,4-triazole-3-thiones
(5) (Scheme 1).
Many pyridine derivatives find important applica-
tions in the field of agriculture, medicine and indus-
try [1 – 3]. Pyridine derivatives are also used as ef-
fective insecticides, fungicides and herbicides [4 – 6].
Triazoles gained increasing attention because of their
immense biological activities [7 – 9]. Further, the pres-
ence of 6-chloro-pyridin-3-yl methyl group is known
to enhance the biological activities of the mole-
cules [10]. Prompted by these findings and in continu-
ation of our studies on 3,6-disubstituted pyridine deriv-
atives [11, 12], it was contemplated to synthesize a se-
ries of new 1,2,4-triazoles containing 6-chloropyridin-
3-yl methyl moiety and screen them for their antibac-
terial activity.
On the other hand, 5-phenyl/benzyl-4-phenyl-3-(6-
chloropyridin-3-yl methyl)sulfanyl-1,2,4-triazoles (7)
and 2-{[(6-chloropyridin-3-yl methyl)methylamino]-
methyl}-5-phenyl/benzyl-4-phenyl-3H-1,2,4-triazole-
3-thiones (9) were prepared from 5-phenyl/benzyl-
4-phenyl-1,2,4-triazole-3-thiols (3) using 6-chloro-3-
chloromethyl pyridine (6) and (6-chloropyridin-3-yl
methyl) methylamine (8), respectively (Scheme 2).
All newly synthesized compounds gave satisfactory
analysis for their nitrogen content. They also exhibited
characteristic spectral patterns pertaining to 6-chloro-
pyridin-3-yl moiety. In the IR spectra, the absorption
bands corresponding to C-Cl vibrations for these mole-
cules were observed in the region 725– 737 cm−1. In
Results and Discussion
6-Chloropyridin-3-yl acetic acid hydrazide (1) was
prepared from 6-chloro-3-chloromethylpyridine (6)
by the method reported by our group [11]. 6-
Chloro-3-chloromethyl pyridine (6) was also con-
verted into (6-chloropyridin-3-ylmethyl)methyl amine
(8) [13]. 5-Phenyl/benzyl-4-phenyl-1,2,4-triazole-3-
thiols (3) were synthesized from the respective acid hy-
drazides [14]. 6-Chloropyridin-3-yl acetic acid hydr-
azide (1) was reacted with phenyl isothiocyanate fol-
lowed by aqueous sodium hydroxide to obtain 5-(6-
chloropyridin-3-yl methyl)-4-phenyl-1,2,4-triazole-3-
thiol (2). The triazole (2) was then transformed into
1
the H NMR spectra, sharp singlets corresponding to
-CH2 group present in all compounds were seen at
δ ranging from 3.85 to 4.43. The characteristic sig-
nals due to three protons of 3,6-disubstituted pyridine
ring appeared as a quartet and two doublets. The proton
(Ha) in position-2 of pyridine ring was coupled with
the proton (Hm) in position-4 to give rise to a doublet
in the region δ = 7.88– 8.43 (Jam = 2.1 – 2.5 Hz). Sim-
c
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B. S. Holla et al. · 1,2,4-Triazole Derivatives
335
Scheme 1.
pound (1) were absent in the spectrum. In the IR spec-
tra of the alkylated products (4) as well as Mannich
bases (5), the NH/SH band was absent. The IR spectra
of compounds (7) and (9) also showed the absence of
NH/SH absorption bands which were found in the par-
1
ent triazoles. In the H NMR spectrum of triazole (2),
two singlets corresponding to the protons of NHNH2
group of parent hydrazide (1) were absent. The aro-
matic protons of the phenyl ring were seen as a mul-
tiplet in the region of δ = 7.16– 7.54 and the signal
of the SH proton was observed as a broad singlet at
δ = 13.75. The downfield shift of this signal indi-
cates the thiol-thione tautomerism in the triazole. This
downfield signal was absent in the 1H NMR spectra of
the alkylated product (4) and the Mannich base (5) thus
confirming their formation.
Scheme 2.
The mass spectra of triazole (2), its alkylated deriv-
ative (4a) and Mannich base (5a) were found to be
consistent with the assigned structures. Molecular ion
peaks for these compounds were observed at m/z 302,
316 and 401 corresponding to their molecular formu-
lae C14H11ClN4S, C15H13ClN4S and C19H20ClN5OS,
respectively. The molecular ion of compound (2) un-
derwent fragmentation to produce an ion at m/z 152
which corresponds to 6-chloropyridin-3-yl acetonitrile
cation (ClC5H3NCH2CN+). A peak at m/z 301 seen
in the mass spectrum of the compound (4a) was due
to the elimination of the methyl group from parent
ion during fragmentation. The molecular ion of Man-
nich base (5a) underwent characteristic fragmenta-
ilarly, proton (Hx) in the position-5 was coupled with
proton (Hm) in the position-4 to give rise to another
doublet in the region δ = 7.22– 7.48 (Jmx = 8.2 Hz),
while proton (Hm) in the position-4 was coupled with
other two protons to give rise to a doublet of dou-
blet in the region δ = 7.36 – 7.89 (Jam = 2.1 – 2.5 Hz,
Jmx = 8.2 Hz) in all compounds. These data support
the presence of a 6-chloropyridin-3-yl methyl moiety.
In the mass spectra, fragment corresponding to the 6-
chloropyridin-3-yl methyl cation was seen at m/z 126
which again supported the presence of a 6-chloropyr-
idin-3-yl methyl moiety in all newly synthesized com-
pounds.
The IR spectrum of triazole (2) showed an absorp- tion to give the base peak at m/z 100, which corre-
tion band at 3114 cm−1 indicating the presence of sponds to the molecular ion of N-methyl morpholine
NH/SH group. It could be seen that the characteristic cation. Compounds (7a) and (9a) also exhibited similar
absorption bands of hydrazide group of parent com- mass spectral fragmentation pattern. The molecular ion
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336
B. S. Holla et al. · 1,2,4-Triazole Derivatives
◦
Table 1. Antibacterial activity data of the newly synthesized
compounds.
and the incubation was carried out at 37 C for 16 –
18 h. Furacin was dissolved in dimethylformamide and
was used as a standard drug for comparison. The mini-
mum concentration at which there was no turbidity was
taken as the minimum inhibitory concentration (MIC
value).
Compound
Minimum inhibition concentration* (µg/ml)
E. coli P. aeruginosa S. aureus
B. subtilis
2
4a
4b
10
100
–
20
200
–
10
100
–
10
100
–
5a
5b
7a
7b
9a
9b
–
–
10
–
100
–
–
–
–
The antibacterial screening data indicate that among
the compounds tested, only triazole (2) and S-alkylated
derivative (7a) showed good antibacterial activities
against S. aureus and B. subtilis compared to Furacin.
50
20
–
200
–
100
10
–
100
–
200
10
–
100
–
Furacin
6
12.5
12.5
12.5
Experimental Section
* Index for antibacterial activity; diameter of the disc: 5 mm; amount
of the sample: 100 µg/ml; control(solvent): dimethyl formamide.
The melting points were determined by open capillary
method and are uncorrected. IR spectra were recorded in
KBr pellets on a Perkin-Elmer 882 IR spectrophotome-
ter. 1H NMR spectra were recorded in (DMSO-d6/CDCl3
or a mixture of them) on a Bruker AC-300F / amx 400
(300/400 MHz) NMR spectrometer using TMS as an in-
ternal standard. The mass spectra were recorded on a VG-
Micromass or Hewlett-Packard 6890-GC mass spectrometer
operating at 70 eV. Progresses of all the reactions and purity
of the products were monitored by TLC using ethyl acetate
peaks (M+) were found at m/z 378 and 421, respec-
tively. Compound (9a) being a Mannich base, showed
a base peak at m/z 169 correspondingto its characteris-
tic fragment [N-methyl-N-(6-chloropyridin-3-yl-meth-
yl)amino]methyl cation.
Antibacterial Activity
All the newly synthesized compounds were ini- as eluent and UV/iodine as visualizer.
tially screened for their in vitro antibacterial activ-
Preparation of 4-phenyl-5-(6-chloropyridin-3-yl methyl)-
4H-1,2,4-triazole-3-thiol (2)
ities against Escherichia coli, Pseudomonas aerugi-
nosa, Staphylococcus aureus and Bacillus subtilis ac-
cording to serial dilution method [15]. The compounds
were tested at a concentration of 200 µg/ml in di-
methylformamide against all organisms. Furacin was
used as standard drug for comparison of antibacterial
activity and solvent control was kept. The minimum
inhibitory concentrations (MIC values) of the above
compounds were determined. The results of antibac-
terial screening studies are reported in Table 1.
To a suspension of hydrazide (1.85 g, 0.01 mol) in dry
benzene (30 ml), phenyl isothiocyanate (1.35 g, 0.01 mol)
was added and refluxed for 5 h. The reaction mixture was
then cooled to room temperature. The solid obtained was fil-
tered and washed with light petrol. The resulting solid was
then dissolved in 2N sodium hydroxide solution (20 ml) and
heated on water bath for 2 h, cooled to room temperature and
filtered. The filtrate was neutralized with dilute hydrochloric
acid. The precipitate obtained was filtered, dried and recrys-
tallized.
A stock solution of concentration 100 µg/ml was
prepared by dissolving 5 mg of the test compound in
50 ml of dimethyl formamide. One loopful of an 18 h
broth culture was inoculated into 5 ml of nutrient broth
White crystals; m. p. 222 – 224 ◦C; yield 69%. – IR (KBr):
˜
ν = 3114 (N-H str.), 3043 – 2930 (C-H str.), 1581 – 1459
(C=N, C=C str.), 1423 (CH2 bend), 841 (Py-H bend), 723
(C-Cl str.), 699 (Ar-H bend) cm−1. -1H NMR (300 MHz,
DMSO-d6): δ = 3.85 (s, 2H, -CH2-), 7.16 – 7.54 (m, 5H,
Ar-H), 7.22 (d, 1H, Hx, J = 8.2 Hz), 7.36 (dd, 1H, Hm,
J = 2.5 Hz, 8.2 Hz), 7.88 (d, 1H, Ha, J = 2.5 Hz), 13.75 (s,
1H, NH/SH). -MS (VG, 70 eV): m/z (%) = 302 (100) [M+],
304 (36) [M++2], 152 (12) [ClC5H3NCH2CN+], 126 (26)
[ClC5H3NCH2], 77 (15) [C6H5+].
◦
and this was incubated at 37 C for 4 h. An assay was
prepared by diluting 4 h sub-culture in 1/1000 in nutri-
ent broth.
Nutrient broth (0.5 ml) was taken in tubes with la-
beled numbers 1 – 11 and 0.5 ml of the solution of
the test compound (100 µg/ml) was added to the first
tube no. 2. This process was repeated serially to ob-
tain the quantities indicated in each of the test tubes.
The eleventh tube was taken as control. A drop of di-
luted broth culture of the test organization (approxi-
mately 0.05 ml) was added to all the tubes using a ster-
General method of alkylation of 4-phenyl-5-substituted-
1,2,4-triazole-3-thiols (4,7)
To a mixture of 4-phenyl-5-substituted-1,2,4-triazole-3-
ilized Pasteur pipette. The solutions were mixed gently thiol (0.01 mol) and sodium hydroxide (0.01 mol) in water
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B. S. Holla et al. · 1,2,4-Triazole Derivatives
337
5a: Yellow powder; m. p. 143 – 144 C; yield 76.7%. –
◦
(25 ml), alkylating agent (0.01 mol) in methanol (10 ml) was
˜
added in drops. The contents were stirred for 8 h at room IR (KBr): ν = 3088 – 2807 (C-H str.), 1596 – 1467 (C=N,
temperature. The solid separated was filtered, washed with C=C str.), 1430 (CH2 bend), 1288 (C=S str.), 852 (Py-H
water and recrystallized from appropriate solvent to yield the bend), 734 (C-Cl str.), 707 (Ar-H str.) cm−1. – 1H NMR
title compound.
(300 MHz, DMSO-d6): δ = 2.72 (t, 4H, -CH2-N-CH2-, J =
4.3 Hz), 3.58 (t, 4H, -CH2-O-CH2-, J = 4.3 Hz), 3.90 (s, 2H,
-CH2-), 5.05 (s, 2H, N-CH2-N), 7.37 – 7.54 (m, 5H, Ar-H),
7.41 (d, 1H, Hx, J = 8.2 Hz), 7.57 (dd, 1H, Hm, J = 2.3 Hz,
8.2 Hz), 8.05 (d, 1H, Ha, J = 2.3 Hz). – MS(VG, 70 eV):
m/z (%) = 401 (46) [M+], 403 (16) [M++2], 302 (76)
[ClC5H3NCH2C7H6N3S+], 126 (24) [ClC5H3NCH2+], 100
(100) [C4H8O+].
4a: White powder; m. p. 86 – 88 ◦C; yield 91%. –
˜
IR (KBr): ν = 3049 – 2931 (C-H str.), 1592 – 1455 (C=N,
C=C str.), 1422 (CH2 bend), 827 (Py-H bend), 735 (C-Cl
str.), 700 (Ar-H) cm−1. – 1H NMR (300 MHz, DMSO-
d6): δ = 2.56 (s, 3H, -CH3), 4.03 (s, 2H, -CH2-), 7.36 –
7.57 (m, 7H, Ar-H, Hx & Hm), 8.02 (d, 1H, HA, J =
2.0 Hz). – MS(VG, 70 eV): m/z (%) = 316 (100) [M+],
318 (38) [M++2], 301 (45) [M+-CH3], 281 (17) [M+-Cl],
269 (9) [M+-SCH3], 239 (8) [M+-C6H5+], 126 (66)
[ClC5H3NCH2+], 91 (42) [126-Cl].
◦
5b: Yellow powder; m. p. 119 – 121 C; yield 63.8%. –
˜
IR (KBr): ν = 3072 – 2827 (C-H str.), 1594 – 1463 (C=N,
C=C str.), 1444 (CH2 bend), 1299 (C=S str.), 825 (Py-H
4b: Yellow powder; m. p. 97 – 99 ◦C; yield 82.5%. –
bend), 727 (C-Cl str.), 702 (Ar-H str) cm◦−1
.
˜
IR (KBr): ν = 3054 – 2946 (C-H str.), 1591 – 1453 (C=N,
9a: White powder; m. p. 154 – 155 C; yield 72.4%. –
C=C str.), 1409 (CH2 bend), 838 (Py-H bend), 737 (C-Cl
˜
IR (KBr): ν = 3068 – 2831 (C-H str.), 1592 – 1454 (C=N,
str.), 703 (Ar-H bend) cm−1
.
C=C str.), 1420 (CH2 bend), 1288 (C=S str.), 849 (Py-H
◦
7a: Yellow powder; m. p. 113 – 116 C; yield 73.9%. – bend), 732 (C-Cl str.), 708 (Ar-H bend) cm−1. – 1H NMR
˜
IR (KBr): ν = 3057 – 2977 (C-H str.), 1591 – 1462 (C=N, (300 MHz, DMSO-d6): δ = 2.47 (s, 3H, CH3), 3.98
C=C str.), 1441 (CH2 bend), 835 (Py-H bend), 735 (C-Cl (s, 2H, -CH2-), 5.27 (s, 2H, N-CH2-N), 7.34 – 7.53 (m,
str.), 704 (Ar-H bend) cm−1. – 1H NMR (300 MHz, DMSO- 11H, Ar-H & Hx), 7.88 (dd, 1H, Hm, J = 2.3 Hz,
d6): δ = 4.43 (s, 2H, -CH2-), 7.19 – 7.55 (m, 10H, Ar-H), 8.2 Hz), 8.41 (d, 1H, Ha, J = 2.3 Hz). – MS(VG, 70 eV):
7.48 (d, 1H, Hx, J = 8.2 Hz), 7.89 (dd, 1H, Hm, J = 2.4 Hz, m/z (%) = 421 (18) [M+], 423 (7) [M++2], 295 (7) [M+-
8.2 Hz), 8.43 (d, 1H, Ha, J = 2.1 Hz). – MS(VG, 70 eV): ClC5H3NCH2+], 253 (11) [C6H5C8H6N3S+], 169 (100)
m/z (%) = 378 (100) [M+], 395 (32) [M++2], 126 (26) [ClC5H3NCH2N(CH3)CH2+], 126 (82) [ClC5H3NCH2+],
[ClC5H3NCH2+], 77 (23) [C6H5+].
91 (11) [126-Cl], 77 (17) [C6H5+].
7b: White powder; m. p. 170 – 171 ◦C; yield 72%. –
˜
9b: Paste, yield 64.3%. – IR (KBr): ν = 3059 – 2829
(C-H str.), 1596 – 1463 (C=N, C=C str.), 1422 (CH2 bend),
1291 (C=S str.), 855 (Py-H bend), 731 (C-Cl str.), 701 (Ar-H
˜
IR (KBr): ν = 3055 (C-H str.), 1597 – 1452 (C=N, C=C str.),
1421 (CH2 bend), 836 (Py-H bend), 725 (C-Cl str.), 702
str) cm−1
.
(Ar-H bend) cm−1
.
General method of aminomethylation of 4-phenyl-5-
substituted-1,2,4-triazole-3-thiols (5,9)
Acknowledgements
The authors are grateful to Professor Ananthakrishna,
Head, Department of Microbiology, KMC, Mangalore for
extending the facilities of the college for antibacterial screen-
ing. Authors are also thankful to Dr. M. S Mithyantha, Vice
President (R&D), Rallis India Limited, Bangalore and the
Management for providing facility and constant encourage-
ment.
4-Phenyl-5-substituted-1,2,4-triazole-3-thiol (0.01 mol)
was dissolved in ethanol (25 ml). Then formaldehyde (40%,
1.5 ml) and secondary amine (0.01 mol) were added to this
mass. The mixture was stirred and heated to reflux for 30 min
and left overnight at r. t. The resulting solid was collected by
filtration, washed with ethanol and recrystallized from appro-
priate solvent to yield the title compound.
[1] S. R. EI-Zemity, A. R. Mohammed, Pesticide Sci. 55,
1203 (1999).
[2] H. J. Roth, A. Klumenn, Hetrocycles, Chapt. 7, Phar-
maceutical Chemistry, Vol. I, p. 244, Ellis Horwood
Limited, Chichester (1988).
[3] J. Cheng, C. Zhang, E. D. Stevens, S. Izenwasser,
D. Wade, S. Chen, D. Paul, M. L. Trudell, J. Med.
Chem. 45, 3041 (2002).
[5] I. Atsushi, S. Keiichi, W. Tsumoru, E. Takayoshi,
N. Yoshitaka, WO Pat 2001066523 (2001); Chem.
Abstr. 135, 242142 (2002).
[6] Z. Yigang, L. Xin, H. Runqiu, C. Muru, L. Zaiguo,
Nongyaoxue Xuebao, 1(1) (1999) 93; Chem. Abstr.
135, 92358 (2002).
[7] G. S. Gadaginamath, S. A. Patil, Indian J. Heterocycl.
Chem. 9, 39 (1999).
[4] N. Sakamoto, T. Ishiwatari, N. Matsuuo, J. Pesticide
Sci. 25, 373 (2000).
[8] B. S. Vashi, D. S. Mehta, V. H. Shah, Indian J. Chem.
35B, 111 (1996).
Unauthenticated
Download Date | 2/13/17 1:08 PM

338
[9] R. H. Khan, R. A. K. Srivastava, R. C. Rastogi, Indian
B. S. Holla et al. · 1,2,4-Triazole Derivatives
Rao, K. Sridhara, U. G. Bhat, Indian J. Chem. 43B,
2170 (2004).
J. Chem. 24B, 883 (1985).
[10] G. Voss, G. Ramos, Chemistry of Crop Protection,
Wiley-VCH, Weinheim (2002).
[13] I. Katsuhiro, JP 05294932 (1993); Chem. Abstr. 120,
217304 (1994).
[11] B. Shivarama Holla, C. S. Prasanna, B. Poojary, K. S.
Rao, K. Sridhara, U. G. Bhat, Indian J. Chem. 43B, 864
(2004).
[14] H. Saad, Indian J. Chem. 35B, 980 (1996).
[15] R. Cruickshank, J. P. Marmion, R. H. A. Swain, Med-
ical Microbiology, Vol. II, p. 190, Churchill Living-
stone, London (1975).
[12] B. Shivarama Holla, C. S. Prasanna, B. Poojary, K. S.
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