Syntheses and biological activities of 6-aryl-3-(3-hydroxypropyl)-7H-1,2,4- triazolo[3,4-b][1,3,4]thiadiazines

Article abstract of DOI:10.3390/12030297

A series of 6-aryl-3-(3-hydroxypropyl)-7H-1,2,4-triazolo[3,4-b][1,3,4]- thiadiazines were synthesized by the reaction of 4-amino-3-(3-hydroxypropyl)-5- mercapto-1,2,4-triazole (1) with substituted ω-haloacetophenones. Their structures were confirmed by elemental analysis, IR, ¹H-NMR, and ¹³C-NMR. Tests of plant growth regulating effects showed that the title compounds display remarkable inhibitory activities on the growth of radish and wheat.

Full text of DOI:10.3390/12030297

Molecules 2007, 12, 297-303
molecules
ISSN 1420-3049
http://www.mdpi.org
Full Paper
Syntheses and Biological Activities of 6-Aryl-3-(3-hydroxy-
propyl)-7H-1,2,4-triazolo[3,4-b][1,3,4]thiadiazines
Jian-yu Jin 1, Li-xue Zhang 2,*, Xian-xing Chen 3, An-jiang Zhang 2 and Hai-le Zhang 4
1
College of Education, Wenzhou University, Wenzhou 325035, P.R. China
2
College of Chemistry and Materials Engineering, Wenzhou University, Wenzhou 325027, P.R.
China
3
College of Life and Environmental Sciences, Wenzhou University, Wenzhou 325027, P.R. China
4
Department of Molecular Biology & Chemistry, Scripps Research Institute, San Diego, CA 92037,
USA; E-mail: hailezh@scripps.edu
* Author to whom correspondence should be addressed; E-mail: zhanglixuelz@yahoo.com.cn; Tel: (+86)
577-88373073; Fax: (+86) 577-88339081
Received: 12 February 2007; in revised form: 27 February 2007 / Accepted: 28 February 2007 /
Published: 3 March 2007
Abstract: A series of 6-aryl-3-(3-hydroxypropyl)-7H-1,2,4-triazolo[3,4-b][1,3,4]-thia-
diazines were synthesized by the reaction of 4-amino-3-(3-hydroxypropyl)-5-
mercapto-1,2,4-triazole (1) with substituted ω-haloacetophenones. Their structures were
1
confirmed by elemental analysis, IR, H-NMR, and 13C-NMR. Tests of plant growth
regulating effects showed that the title compounds display remarkable inhibitory
activities on the growth of radish and wheat.
Keywords: Hydroxyl groups, 1,2,4-triazolo[3,4-b]-1,3,4-thiadiazine, NMR, plant growth
regulation.
Introduction
Triazoles and their heterocyclic derivatives represent an interesting class of compounds which
possess a wide range of biological activities, such as analgesic, anthelmintic, antitubercular, plant

Molecules 2007, 12
298
growth regulating, antiviral, antifungal and anticancer properties [1-6]. Attachment of an aliphatic side
chain containing a hydroxyl group to the 3-position of the fused heterocycle could bring about changes
in its solubility; moreover, the -OH group can be transformed into many other useful functional groups,
allowing preparation of numerous novel compounds. With all these considerations in mind, we have
synthesized a series of 6-aryl-3-(3-hydroxypropyl)-7H-1,2,4-triazolo[3,4-b][1,3,4]thiadiazines, then
1
confirmed their structures by elemental analysis, IR, H- and 13C-NMR, and determined the plant
growth regulating effects of all the title compounds and the key intermediate 1.
Results and Discussion
Synthetic route and improvement of synthetic methods
The synthetic route used is shown in Scheme 1. After preparing 4-amino-3-(3-hydroxypropyl)-5-
mercapto-1,2,4-triazole (1), the 3,6-disubstituted 1,2,4-triazolo[3,4-b][1,3,4]thiadiazines 2a~2j were
obtained through the reaction with the appropriate substituted phenacyl bromide (or chloride).
Scheme 1. Synthesis of the title compounds.
N
N
S
pyridine
HO
H2NHNCNHNH2
SH
N
O
O
NH2
1
1
2
O
C
N
N
11
9
3
R
CH2Br
8
HO
4
5
S
N
N
10
12
C2H5OH
7
6
2a~2j
R
Cl
R= 2a:
2b:
2c:
Cl
Cl
F
F
2f:
2e:
2h:
OCH3
2d:
2g:
F
CH3
NO2
2i:
Br
2j:

Molecules 2007, 12
299
The rates of ring-closure were closely related to the structures of the substituents R. When R
contained electron-withdrawing groups, the rates of ring-closure were increased because the
electron-cloud densities of the carbonyl carbons were lowered.
The syntheses of thiocarbohydrazide and 4-amino-3-(3-hydroxypropyl)-5-mercapto-1,2,4-triazole
(1) were carried out according to the literature methods [7,8], with some improvements. First, if the
reaction temperature of hydrazine hydrate and carbon disulphide was slowly elevated, formation of a
yellow-green side product (m.p.:138~140°C) was increased, so we quickly raised the reaction
temperature to 100~110°C after carrying out the reaction at room temperature for half an hour in order
to maximize the yield of thiocarbohydrazide. Second, some water was added to the mixture when
γ-butyrolactone was reacted with thiocarbohydrazide in pyridine. This water improved the solubility of
the reactants and resulted in an improved yield of compound 1. Finally, the solubility of the triazole
intermediate in hot water differed obviously from that in cold water, so the crude product was
recrystallized from water.
IR spectra of the title compounds
The absence of N-H, S-H and C=O absorptions in the IR spectra confirms that the
cyclocondensation to form the title compounds has taken place. The characteristic stretching vibrations
of the products are at 3405-3446 cm-1 (O-H, strong), 1635-1597 cm-1 (C=N), 1636, 1600, 1578, 1522,
1507, 1471 cm-1 (aromatic ring skeleton vibrations). The C-S-C bending vibrations are in the region
around 670 cm-1. The stretching vibration peaks of CH2 are observed at about 2920 cm-1.
1H-NMR spectra of the title compounds
In the 1H-NMR spectra of triazolothiadiazines 2a~2j, the characteristic downfield signal at δ 13.40
attributed to the -N=C-SH (-NH-C=S of the tautomer) in compound 1 is absent, as is a sharp signal at δ
5.49 attributable to the N-NH2 group in the parent triazole. In the spectra of the title compounds, the
signal at δ 1.80 is attributed to the CH2 of the hydroxypropyl group, and that at δ 3.60 is attributed to
the O-H group. Resonance peaks at δ 7.13~8.42 are attributed to the aromatic ring protons. As a result
of the overall action of inductive effect and magnetic anisotropic effect of the aromatic rings, the
chemical shift of the SCH2 group in the thiadiazine moiety appears downfield at δ 4.26~4.57.
13C-NMR spectra of the title compounds
13
The C-NMR spectra of compound 2h, for instance, exhibits the expected 11 absorption peaks,
i.e. δ 22.92, 123.95, 128.69, 139.47, 139.79, 148.99, 152.81, 153.73, 20.73 and 29.46 ppm, which are
assigned to CH2-CH2 and one at δ 59.76, which is attributed to the O-CH2 of the hydroxypropyl group.
Because of the existence of carbon-nitrogen double bonds, the chemical shifts of the three carbon
atoms in the triazole ring appear at δ 148.99, 152.81, 153.73, which conforms approximately with the
values reported in the literature [9]. All the benzene carbons of the title compounds exhibit chemical
shifts between 105.07~153.82, whereas the peaks with δ values near 23 ppm are attributed to the
S-CH2 in the thiadiazine rings.

Molecules 2007, 12
300
Biological Activity
The effects of the title compounds 1, 2a~2j on sprouting of wheat and radish seeds have been
investigated. After treating with solutions of 100 µg/mL and 10 µg/mL of the title compounds for 7
days, the germination percentages have been determined, and from the difference in length between
stems and radicles of seedlings treated with the title compounds and those treated with distilled water,
the plant growth regulating activities have also been calculated. A positive result represents a growth
increase, whereas a negative result implies an inhibition:
the length of sample’s stem/radicle － the reference’s length
× 100％
effect =
the length of reference’s stem/radicle
The results are shown in Table 1. From the data it is apparent that the newly synthesized
compounds 2a~2j show a more pronounced inhibiting effect than 4-amino-3-(3-hydroxypropyl)-5-
mercapto-1,2,4-triazole (1) on the growth of wheat and radish.
Table 1. Plant growth regulating activities of compounds 1, 2a~2j
(germination percentage % / effect (on stem) % / effect (on root) %).
Radish
Wheat
Compd.
10 ppm
100 ppm
10 ppm
100 ppm
1
80/25.6/22.8
20/-43.2/-50.9
-100%
85/9.7/-12.9
-100%
-100%
-100%
-100%
-100%
-100%
-100%
-100%
-100%
-100%
85/0/0
70/-20.6 /4.7
81.3/-68/-68.9
85/-94.7/-89.5
94.7/-74.6/-71.9
83.3/-65.9/-64.3
83.3/-91.3/-84.7
84.2/-87.9/-80.7
100/-92/-87.7
75/-35.4/-44.5
94.7/-84.3/-77.8
77.8/-87.9/-83.7
100/0/0
55/-23.8/-34.1
-100%
2a
2b
-100%
2c
20/-42.3/-73.8
35/-51.4/-45.6
5/-100/-91.3
10/-52.7/-84.7
10/-38.5/-66.2
60/-36.3/-42.1
20/-68.5/-72.7
25/-57.7/-74.2
85/0/0
-100%
2d
-100%
2e
-100%
2f
-100%
2g
-100%
2h
95/-96.6/-96.1
-100%
2i
2j
-100%
Reference
100/0/0
Experimental
General
All melting points were determined on an XT-4A apparatus. The IR spectra (KBr disks) were
1
recorded on a Bruker Quinox 55 spectrophotometer. The H-NMR and 13C-NMR spectra were
measured at 25°C on a Bruker Advance 300 spectrometer operating at 300 and 75 MHz, respectively
(DMSO-d6 solutions using TMS as internal reference). Elemental analyses were carried out with a
Flash-EA 1112 elemental analyzer. All the reagents used were AR grade.

Molecules 2007, 12
301
Preparation of 4-amino-3-(3-hydroxypropyl)- 3-(3-hydroxypropyl)-5-mercapto-1,2,4-triazole (1)
CS2 (13 mL, 0.22 mol) was added dropwise under stirring to a mixture of 85% hydrazine hydrate
(24 mL) and water (75 mL), then stirring was continued for 30 minutes at room temperature. The
reaction temperature was then rapidly raised to 100~110 °. After refluxing for 2h, the reaction mixture
was cooled in an ice-bath and filtered. The residue was washed with ethanol and recrystallized from
water to afford pure thiocarbohydrazide crystals, mp: 171~172 °C. The thiocarbohydrazide thus
prepared (1 mmol) was then added to a solution of γ-butyrolactone (1 mmol) in pyridine (20 mL) and
water (2 mL), and the mixture was refluxed for 6 h. After removing the solvent, the white solid formed
was recrystallized from water to give compound 1 in 76.6% yield; m.p.: 125~126 °C; Anal. Calc. (%)
for C5H10N4OS: C, 34.47; H, 5.79; N, 32.16. Found: C, 34.25; H, 5.70; N, 31.99; IR: 3390, 3309, 3189,
3096, 3044, 2925, 2775, 1615, 1570, 1508, 1326, 1056, 963 cm−1; 1H-NMR δ ppm: 1.73-3.45 (m，6H,
C-H), 4.53 (s, 1H, OH), 5.49 (s, 2H, NH2), 13.40 (s, 1H, NH);13C-NMR δ ppm: 21.28 (CH2), 28.88
(Ar-CH2), 59.94 (C-OH), 152.31 (C=N), 165.80 (C=S).
General method for the preparation of 6-aryl-3-(3-hydroxypropyl)-7H-1,2,4-triazolo[3,4-b][1,3,4]
thiadiazines 2a~2j
The appropriate ω-haloacetophenone (1 mmol) was added to a solution of compound 1 (1 mmol) in
75% C2H5OH (20 mL). The mixture was refluxed for 4 h. The solid obtained on cooling was filtered,
dried and recrystallized from 75% C2H5OH to give the title compounds. All the title compounds can be
dissolved in ethanol. The physical and spectral data of the title compounds 2a~2j are given in Tables
2~4.
Table 2. Physical data of compounds 2a~2j.
Anal./% Found (calc.)
H
Molecular
formula
Compd.
Colour Yield/% M.p./°C
C
N
2a
2b
2c
2d
2e
2f
C13H13ClN4OS
C13H12Cl2N4OS
C13H13FN4OS
C13H12F2N4OS
C13H14N4OS
white
yellow
white
yellow
white
white
white
brown
white
yellow
76.5
80.2
59.6
77.1
48.5
70.3
55.7
74.1
66.9
72.3
134~136
119~121
163~165
105~107
155~157
188~190
144~146
163~165
156~158
175~177
50.28(50.57) 4.09(4.24) 18.43(18.14)
45.31(45.49) 3.61(3.52) 16.05(16.32)
53.67(53.41) 4.70(4.48) 19.44(19.17)
50.19(50.32) 3.66(3.90) 18.37(18.05)
56.80(56.91) 5.36(5.14) 20.16(20.42)
55.37(55.25) 5.18(5.30) 18.69(18.41)
58.02(58.31) 5.72(5.59) 19.12(19.43)
48.64(48.89) 3.95(4.10) 22.25(21.93)
43.97(44.20) 3.98(3.71) 15.68(15.86)
65.42(65.12) 5.33(5.18) 15.84(15.99)
C14H16N4O2S
C14H16N4OS
2g
2h
2i
C13H13N5O3S
C13H13BrN4OS
C19H18N4OS
2j

Molecules 2007, 12
302
Table 3. IR data of compounds 2a~2j.
Compd.
IR (ν/cm-1)
2a
2b
2c
2d
2e
2f
3421 (OH), 2912 (CH2), 1633 (C=N), 1462 (N=C-S), 626 (C-S-C).
3436 (OH), 2921 (CH2), 1635 (C=N), 1465 (N=C-S), 630 (C-S-C).
3414 (OH), 2895 (CH2), 1606 (C=N), 1497 (N=C-S), 629 (C-S-C)
3417 (OH), 2975 (CH2), 1620 (C=N), 1474 (N=C-S), 630 (C-S-C).
3426 (OH), 2921 (CH2), 1635 (C=N), 1447 (N=C-S), 688 (C-S-C).
3446 (OH), 2979 (CH2), 1597 (C=N), 1469 (N=C-S), 650 (C-S-C).
3431 (OH), 2910 (CH2), 1624 (C=N), 1462 (N=C-S), 635 (C-S-C).
3405 (OH), 2929 (CH2), 1620 (C=N), 1467 (N=C-S), 688 (C-S-C).
3417 (OH), 2912 (CH2), 1630 (C=N), 1460 (N=C-S), 624 (C-S-C).
3417 (OH), 2902 (CH2), 1632 (C=N), 1462 (N=C-S), 690 (C-S-C).
2g
2h
2i
2j
Table 4. NMR data of compounds 2a~2j.
Compd.
1H-NMR (δ, ppm)
13C-NMR (δ, ppm)
2a
1.86~3.53 (m, 6H, C-H), 4.48 (s, 2H, S-CH2),
7.66~8.14 (m, 4H, Ar-H)
20.77, 22.93, 29.17, 59.71, 129.19, 129.40,
132.00, 137.10, 141.28, 153.78, 155.26
2b
1.80~3.51 (m, 6H, C-H), 4.26 (s, 2H, S-CH2),
4.53~4.56 (m, 1H, O-H), 7.61~7.86 (m, 3H,
Ar-H)
20.82, 25.83, 29.52, 59.82, 127.99, 129.76,
132.25, 132.51, 133.45, 136.14, 140.00, 153.66,
154.56
2c
1.87~3.53 (m, 6H, C-H), 4.47 (s, 2H, S-CH2),
7.41~8.14 (m, 4H, Ar-H)
20.75, 22.97, 29.27, 59.73, 116.38, 130.26,
141.20, 153.69, 155.28, 162.76, 166.09
2d
1.81~3.50 (m, 6H, C-H), 4.30 s, 2H, S-CH2),
7.28~7.93 (m, 3H, Ar-H)
21.02, 25.25, 29.67, 60.03, 105.42, 112.63,
119.83, 132.15, 140.09, 151.84, 153.82, 162.70,
165.87
20.98, 23.21, 29.35, 59.92, 127.79, 129.29,
132.46, 133.33, 141.68, 153.98, 156.61
2e
2f
1.86~3.53 (m, 6H, C-H), 4.49 (s, 2HS-CH2),
7.56~8.06 (m, 5H, Ar-H)
20.78, 22.69, 29.23, 55.57, 59.77, 114.52,
1.88~3.54 (m, 6H, C-H), 3.86 (s, 3H, CH3), 4.49
(s, 2H, S-CH2), 7.13~8.06 (m, 4H, Ar-H)
125.23, 129.48, 141.16, 153.50, 155.54,162.49
20.86, 21.00, 22.79, 29.58, 59.91,127.32,
2g
2h
2i
1.84~3.53 (m, 6H, C-H), 2.39 (s, 3H, CH3), 4.36
(s, 2H, S-CH2), 7.36~7.93 (m, 4H, Ar-H)
129.56, 130.76, 140.01, 142.00, 153.51, 154.45
20.73, 22.92, 29.46, 59.76, 123.95, 128.69,
139.47, 139.79, 148.99, 152.81, 153.73
1.85~3.54 (m, 6H, C-H), 4.46 (s, 2H, S-CH2),
8.24~8.42 (m, 4H, Ar-H)
20.75, 22.67, 29.48, 59.79, 125.51, 129.24,
131.95, 132.73, 139.83, 153.53, 153.58
1.84~3.54 (m, 6H, C-H), 4.38 (s, 2H, S-CH2),
4.54~4.57 (m, 1H, O-H), 7.77~7.96 (m, 4H,
Ar-H)
1.88~3.53 (m, 6H, C-H), 4.43 (s, 2H, S-CH2),
4.59 (s, 1H, O-H), 7.43~8.12 (m, 9H, Ar-H)
20.89, 22.83, 29.62, 59.92, 126.84, 127.12,
128.01, 128.28, 129.06, 132.46, 138.80, 139.97,
143.21, 153.59, 154.14
2j

Molecules 2007, 12
303
Acknowledgements
This work was financially supported by the Natural Science Foundation of Zhejiang Province,
China (Project No. M203149).
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Sample Availability: Not available.
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