Synthesis and antimicrobial activities of some 6-methyl-3-thioxo-2,3- dihydro-1,2,4-triazine derivatives

Article abstract of DOI:10.1080/10426507.2012.755972

4-Arylidene-imidazole derivatives (4a,b) were readily prepared by reacting 4-am- ino-6-methyl-3-thioxo-2,3-dihydro[1,2,4]triazin-5(4H)-one (1) with 4-arylidene-2-phenyl- 4H-oxazol-5-one (2). Reaction of 1 with some aromatic aldehydes in presence of trieth

Full text of DOI:10.1080/10426507.2012.755972

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Synthesis and Antimicrobial Activities of Some 6-
Methyl-3-Thioxo-2,3-Dihydro-1,2,4-Triazine Derivatives
Ahmed A. El-Barbary a , Ashraf A. El-Shehawy b & Nabiha I. Abdo a
a Chemistry Department, Faculty of Science, Tanta University, Tanta, 31527, Egypt
b Chemistry Department, Faculty of Science, Kafr El-Sheikh University, Kafr El-Sheikh,
33516, Egypt
Accepted author version posted online: 02 Jan 2013.
To cite this article: Ahmed A. El-Barbary , Ashraf A. El-Shehawy & Nabiha I. Abdo (2013): Synthesis and Antimicrobial
Activities of Some 6-Methyl-3-Thioxo-2,3-Dihydro-1,2,4-Triazine Derivatives, Phosphorus, Sulfur, and Silicon and the Related
Elements, DOI:10.1080/10426507.2012.755972
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ACCEPTED MANUSCRIPT
Synthesis and Antimicrobial Activities of Some 6-Methyl-3-Thioxo-2,3-Dihydro-1,2,4-
Triazine Derivatives
Ahmed A. El-Barbary,[a] Ashraf A. El-Shehawy,[b] and Nabiha I. Abdo[a]
[a] Chemistry Department, Faculty of Science, Tanta University, Tanta 31527, Egypt
[b] Chemistry Department, Faculty of Science, Kafr El-Sheikh University, Kafr El-Sheikh 33516,
Egypt
Email:
Graphical Abstract
O
O
O
H3C
H3C
R
C
H
N NH2
S
H3C
N
S
N
N
N
N
N
H
1
N
N
H
S
N
H
N-S-attack
N-attack
O
H3C
N NH2
S
N
N
S-attack
4-Arylidene-imidazole derivatives (4a,b) were readily prepared by reacting 4-amino-6-
methyl-3–thioxo-2,3–dihydro[1,2,4]triazin-5(4H)-one (1) with 4-arylidene-2-phenyl-4H-
oxazol-5-one (2). Reaction of 1 with some aromatic aldehydes in presence of
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triethylphosphite exclusively afforded the corresponding aminophosphonates 5a-c. Reaction
of 1 with 3-phenyl-1H-quinazoline-2,4-dione (6a) and/or 3-phenyl-2-thioxo-2,3-dihydro-
1H-quinazolin-4-one (6b) gave 2-(6-methyl-5-oxo-3-thioxo-2,5-dihydro-3H-[1,2,4]triazin-
4-ylimino)-3-phenyl-2,3-dihydro-1H-quinazolin-4-one (7). Moreover, on treating 1 with 2-
phenylbenzo[d][1,3]thiazine-4-thione (8), 6-methyl-4-(2-phenyl-4-thioxo-4H-quinazolin-3-
yl)-3-thioxo-3,4-dihydro -2H-[1,2,4]triazine-5-one (9) was obtained in 65% yield. Reaction
of 1 with 4-sulfonylaminoacetic acid derivatives (10a,b) afforded the corresponding
sulfonamides (11a,b), respectively. Acid hydrolysis of 11a afforded 7-aminomethyl-3-
methyl[1,3,4]thiadiazole[2,3-c][1,2,4]triazin-4-one (12). 4-Amino-6-methyl-3-(morpholine-
4-ylsulfanyl)-4H-[1,2,4]triazin-5-one (14) was prepared by reacting compound 1 with
morpholine in presence of KI/I2, while 3,3' -bis(4-amino-6-methyl-5-oxo-triazinyl)disulfide
(16) was obtained by oxidation of 1 with lead tetraacetate. The antimicrobial activity of the
products was evaluated against Gram-positive and Gram-negative bacteria, as well as the
fungus Candida albicans.
Keywords: 1,2,4-Triazines, Imidazoles, Phosphonic acid ester, Quinazonlines, Antimicrobial
activity.
INTRODUCTION
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Nitrogen containing heterocyclic compounds have received considerable attention due to
their wide range of pharmacological activities.1,2 It is well known that substituted triazines
occupied a unique position in medicinal chemistry.3-6 Triazine derivatives have attracted
considerable pharmaceutical interest due to their antiproliferative activity,7 antiviral agent,8,9
antihypertensive agent,10 antitumor and in vitro supporting their anti-HIV activity,11,12
anticonvulsant13 and antileukemic.14
Furthermore, pyrimido-triazines have exhibited antibacterial activity.15 Antimicrobial
activities of 1,2,4-triazines became a subject of interest for researchers.16-19 The increasing
importance of triazine derivatives as intermediates for the synthesis of biologically active
compounds has led to continued development of new simple procedures for their synthesis.
Thus, we wish to report herein on the synthesis of 4-amino-6-methyl-3-thioxo-5-oxo-3,4-
dihydro-1,2,4-triazine and some of its derivatives for evaluating their antimicrobial activities.
RESULTS AND DISCUSSION
4-Amino-6-methyl-3-thioxo-2,3-dihydro[1,2,4]-triazin-5(4H)-one (1) was prepared
according to Dorrnow et al.20 On refluxing compound 1 with 4-arylidene-2-phenyl-4H-oxazol-5-
ones (2a,b), prepared according to recommended method,21 in glacial acetic acid, the
corresponding 4-(4-benzylidene- and 4-(4-methoxybenzylidene)-5-oxo-2-phenyl-4,5-
dihydroimidazol-1-yl)-6-methyl-3-thioxo-3,4-dihydro-2H-[1,2,4]triazin-5-ones (4a,b)
derivatives were obtained, instead of the expected products 3a,b (Scheme 1). The structure of
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compounds 4a,b were confirmed by each of Infrared (IR), 1H NMR and mass spectral analyses.
IR spectra of compounds 4a,b showed broad absorptions at 1631 cm-1 (C=O), 1694 cm-1 (C=O-
imidazolo) and 1641 cm-1 (C=O-triazine) and also showed the absence of the absorption due to
NH2 group. Mass spectrum of 4a showed a low intensity molecular ion peak (M+, <1%) at m/z =
389. Reacting 1 with some aromatic aldehydes in presence of triethylphosphite in glacial
acetic acid at 100oC afforded the corresponding 4-methoxyphenyl- and 4-
dimethylaminophenyl[6-methyl-5-oxo-3-thioxo-2,5-dihydro-3H-[1,2,4]triazin-4-yl-aminomethyl
]phosphonate (5a-c)22 (Scheme 1). IR spectra of compounds 5a-c showed absorption bands due
to the stretching frequency of C=C aromatic at 1511-1614 cm-1 and bands of P-O-alkyl at 1024-
1058 cm-1. 1H NMR spectrum of 5b showed signals at 2.18 (s, 3H, CH3-triazine), a triplet at
2.48-2.50 ppm (CH3), a quartet at 3.80 ppm (CH2), 3.90 (s, 1H, NH-CH-Ar), 8.50 ppm (s, 1H,
NH-triazine), 13.63 ppm (s, 1H, NH-CH-Ar). MS spectrum of 5a showed a low intensity pseudo
molecular ion peak (M++1, <1%) at m/z = 385, which confirmed the postulated structure. 31P
NMR spectrum of compound 5a showed a singlet peak at 68.0 ppm.23
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O
Ar
H3C
N
N
N
O
O
N
N
H
S
O
Ph
Ar
O
H3C
3a,b
N NH2
S
AcOH
N
O
+
N
Ar
N
O
Ph
2a,b
H
1
N
N
H3C
N
Ph
N
i) ArCHO
ii) (EtO)3P
N
H
S
4a,b
O
N
Ar O
N NH C P OEt
H3C
2,4: Ar= Ph (a)
H
N
OEt
5: Ar= Ph (a)
S
Ar= 4-Me-O-C6H4 (b)
Ar= 4-Me-O-C6H4 (b)
Ar= 4-Me2NC6H4 (c)
H
5a-c
Scheme 1
On heating compound 1 with 3-phenyl-1H-quinazoline-2,4-dione (6a)24 at 240oC for 4 h
and/or with 3-phenyl-2-thioxo-2,3-dihydro-1H-quinazolin-4-one (6b)24 at 225oC for 3 h, the
same product 2-(6-methyl-5-oxo-3-thioxo-2,5-dihydro-3H-[1,2,4]triazin-4-ylimino)-3-phenyl-
2,3-dihydro-1H-quinazolin-4-one (7) was obtained (Scheme 2). Product 7 seemed to be formed
via successive elimination of H2O and H2S. Its IR spectrum showed bands at 1616 cm-1 (C=O-
triazine), 1662 cm-1 (C=O-quinazoline), 3214 cm-1 (NH-triazine) and 3428 (NH-quinazoline).
Reacting compound 1 with 2-phenyl benzo[d][1,3]thiazine-4-thione (8) afforded the
corresponding 6-methyl-4-(2-phenyl-4-thioxo-4H-quinazolin-3-yl)-3-thioxo-3,4-dihydro-2H-
[1,2,4]triazin-5-one (9) (Scheme 2). IR spectrum of compound 9 showed absorption bands at
1229 cm-1 (C=S-triazine) and at 1309 cm-1 (C=S-quinazoline). Reaction of 1 with
sulfonylaminoacetic acid derivatives 10a,b, prepared according to Okuda et al.25 in POCl3 at
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85oC gave the corresponding 4-methyl-N-[(3-methyl-4-oxo-4H-[1,3,4]thiadiazolo[2,3-c]triazin-
7-ylmethyl]benzenesulfonamide and 4-methyl-N-[(3-methyl-4-oxo-4H-[1,3,4]thiadiazolo[2,3-
c]triazin-7-ylphenylmethyl]benzenesulfonamide derivatives (11a,b), respectively, were obtained.
Hydrolysis of compound 11a with dilute hydrochloric acid afforded 7-aminomethyl-3-methyl-
[1,3,4]thiadiazolo[2,3-c][1,2,4]triazin-4-one (12)26 and 4-toluenesulfonic acid27 (Scheme 3). IR
spectra of compounds 11a,b showed the characteristic absorption peaks of NH groups at 3284
and 3422 cm-1 and SO2 group at 1160 cm-1, respectively. Moreover, their IR spectra showed the
absence of C=S group. 1H NMR of compound 11b showed singlet signals at 4.9 ppm (s, 1H, CH-
Ph) and 2.2 ppm (s, 3H, CH3-Ar). IR spectrum of compound 12 showed the existence of NH2 at
3248, (sym.) and 3171 cm-1, (antisym.).
H
S
N
X
S
N
Ph
H3C
N
N
H
O
N
S
S
Ph
N
O
6a,b
H
a, X= O
b, X= S
N
N
O
8
N
N
N
1
H3C
Fusion
Xylene, reflux
N
Ph
Ph
O
N
N
H
S
CH3
7
9
CO2H
POCl3, reflux
R
C
H
NH SO2
10a,b
10, 11; R= H (a)
R= Ph (b)
O
CH3
i) dil. HCl
ii) NaOH
H3C
R
N
N
C
H
NH2
HO3S
+
CH3
O
N
S
Reflux
N
H
R
C
H
H3C
N
S
N
HN SO2
N
12 (R = H)
N
H
11a,b
Scheme 2
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Treating 1 with morpholine in presence of potassium iodide and iodine at room
temperature afforded 4-amino-6-methyl-3-(morpholin-4-ylsulfanyl)-4H-[1,2,4]triazin-5-one
(14)28 in a low yield instead of the expected structure 13 (Scheme 3). Although iodine is not very
soluble in pure water, it can dissolve in water that contains I- (aq) ion.29 In aqueous solution,
iodine has the tendency to react with excess iodide anions to form the tri-iodide anion. The tri-
iodide ion can react with another iodine molecule to form the linear penta-iodide anion.30 The
reduction of water in the presence of K+ proceeds through the reductive formation of alkali metal
which brought about a decrease in yield of product 14.31 1H and 13C NMR spectra of compound
14 could not be obtained due to its insolubility in hot DMSO. IR spectrum of compound 14
showed band at 3426 cm-1 due to NH2 group and the absence of C=S group. Oxidation of 1 with
lead tetraacetate (LTA) in methylene chloride at room temperature afforded unexpectedly bis(4-
amino-6-methyl-5-oxo-1,2,4-triazinyl)disulfide (16) instead of the proposed structure 15
(Scheme 3). IR spectrum of compound 16 showed the existence of NH2 at 3202 cm-1 (sym.) and
3292 cm-1 (antisym.) and the absence of C=S group. MS spectrum of compound 16 showed a
molecular ion peak (M+, <1%) at m/z = 314.
O
N
O
N
O
CH3
H3C
H3C
N
N
N NH2
N
N
N
N
S
S
N
O
15
HN
O
13
Pb(CH3COO)4
CH2Cl2, r.t.
1
I2/KI, r.t.
O
O
O
H3C
N NH2
H3C
NH
S
CH3
2 H2N
N
N
N
N
S
O
N
N
N
S
N
N
14
16
Scheme 3
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The antimicrobial activity of the products was evaluated against Gram-positive and
Gram-negative bacteria, as well as the fungus Candida albicans. The results showed that
compound 1 has antimicrobial activities against Escherichia coli and Candida albicans but failed
to show any activity against other tested organisms (Pseudomonas aeruginosa, Aeromonas sp,
Klebsiella sp, Bacillus subtilis and Staphylococcus aureus). Compound 1 was added to the liquid
media containing the sensitive organisms to obtain different concentrations (0.2-1.6 mg/mL).
The results that are represented in Figures S 1 and S 2 (Supplemental Materials) indicated that
increase of sensitivity of the selected organisms was enhanced by increasing the concentration of
the compound [as indicated by the ratio of surviving cell number (M/C)] till 0.5 mg/mL. It was
noticed that the sensitivity was nearly stable above 0.5 mg/mL with no further inhibition growth.
Thus 0.5 mg/mL of this compound could be considered the minimum inhibitory concentration
(MIC) against the tested sensitive organisms32.
Compounds 4, 5, 7, 9, 11a,b, 12, 14 and 16 were screened for their activities against the
test organisms namely E. coli, Ps. aeruginosa, Aeromonas sp, Klebsiella sp, B. subtilis and St.
aureus and C. albicans. The results that are listed in Table S 1 (Supplemental Materials) showed
that the antimicrobial activity was found to be affected on the structure of the tested derivatives
as well as on the tested organisms. However, compounds 5b, 5c, 7, 9, and 11b showed no
activity towards the tested microorganisms, this may be attributed to the presence of para-
substituted phenyl and bulky groups which may prevent these compounds to deal with the nature
of the pathogen. Other compounds showed variable inhibitory effects. None of the tested
compounds showed inhibitory activity against all the test organisms. However, the diameter of
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inhibition zone differed also according to the sensitivity of the test organism. It is interesting to
show that Bacillus subtilis showed good sensitivity to compounds 4a, 4b, 5a, 14 and 16. Among
the tested derivatives, compound 14 showed the best effect against the majority of the tested
organisms (Klebsiella sp, B. subtilis, E. coli, St. aureus and C. albicans). This is might be due to
the presence of unreacted free amino group beside, the S-morpholyl moiety in its structure.
Aeromonas sp and Ps. aeruginosa were resistant to compound.
EXPERIMENTAL
All melting points were uncorrected and performed by the open capillary melting point
apparatus. Microanalyses were performed by Microanalysis Unit, National Research Center,
Egypt. Infrared spectra were recorded with a Perkin-Elmer 1720 spectrometer. The NMR spectra
were recorded on a Bruker AC 200 FT NMR spectrometer at 250 MHz for 1H NMR, and 62.9
MHz for 13C NMR, Bruker 200 MHz and Bruker 90 MHz using TMS as internal standard
DMSO as a solvent. 31P NMR spectrum was recorded at 120 MHz on a Varian CFT-20
spectrometer. Chemical shifts (δ) are reported in parts per million (ppm) and signals are
expressed as s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), or br (broad). 31P
chemical shifts are related to 85% H3PO4. Mass spectra (MS) were recorded using electron
ionization (EI) on a Varian Mat 311A spectrometer and using fast atom bombardment (FAB) on
a Kratos MS 50 spectrometer.
Synthesis of 4a,b
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A mixture of compound 1 (1.0 g, 6 mmol) and 4-benzylidene-4H-oxazol-5-one (2a) (1.4 g, 6
mmol) and/or 4-(4-methoxybenzylidene)-2-phenyl-4H-oxazol-5-one (2b) (1.6 g, 6 mmol) was
refluxed in glacial acetic acid (35 mL) for about 25 h (TLC). The reaction mixture was cooled
and poured onto ice and the solid obtained was filtered and crystallized from methanol affording
4a,b.
4-(4-Benzylidene-5-oxo-2-phenyl-4,5-dihydroimidazol-1-yl)-6-methyl-3-thioxo-3,4-dihydro-
2H-[1,2,4]triazin-5-one (4a); Ar = Ph; Yield 2 g (75%); m.p. 123-25oC; IR (KBr): υ (cm-1):
3415 (NH), 3059-1565 (Harom), 1631 (C=O), 1477 C=N); 1H NMR (DMSO-d6, 250 MHz): δ 2.50
(s, 3H, CH3), 7.20 (s, 1H, Ph-CH=), 7.50-8.10 (m, 10H, Harom), 10.11 (s, 1H, NH) ppm; MS (EI):
m/z (%) = 389 (M+, <1), 356 (3), 312 (1), 212 (38), 93 (100); Anal. Calcd. for C20H15N5O2S: C,
61.68; H, 3.88; N, 17.98. Found: C, 61.53; H, 3.69; N, 17.90.
4-(4-(4-Methoxybenzylidene)-5-oxo-2-phenyl-4,5-dihydroimidazol-1-yl)-6-methyl-3-thioxo-
3,4-dihydro-2H-[1,2,4]triazin-5-one (4b); Ar = 4-MeOC6H4; Yield 1.5 g (71%); m.p. 158-
60oC; IR (KBr): υ (cm-1): 3231 (NH), 1694 (C=O-imidazolo), 1641 (C=O-triazine), 1504 (C=N),
825 (para-MeO-aryl); 1H NMR (DMSO-d6, 250 MHz): δ 2.49 (s, 3H, CH3-triazine), 3.80 (s, 3H,
OCH3), 7.09 (s, 1H, Ar-CH=), 7.30-8.40 (m, 9H, Harom) ppm; MS (EI): m/z (%) = 419 (M+, <1),
327 (4), 277 (2), 142 (38); Anal. Calcd. for C21H17N5O3S: C, 60.13; H, 4.09; N, 16.70. Found: C,
60.29; H, 3.90; N, 16.89.
Synthesis of 5a-c
General procedure: To a stirred solution of compound 1 (1.0 g, 6 mmol) in glacial acetic acid
(30 mL), the desired aromatic aldehydes (benzaldehyde, 4-methoxy benzaldehyde, or 4-
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dimethylamino- benzaldehyde; 6 mmol) and triethylphosphite (1 mL, 6 mmol) were added and
the reaction mixture was heated under reflux at 100oC for 18-25 h (TLC). The reaction mixture
was concentrated to one quarter of its volume and then poured onto ice. The solid product thus
formed was filtered, washed by petroleum ether and then crystallized from methanol affording
the desired products 5a-c.22
[Phenyl-6-methyl-5-oxo-3-thioxo-2,3-dihydro-3H-[1,2,4]triazin-4-
ylaminomethyl]phosphonate (5a); Ar = Ph; Yield 1.2 g (55%); m.p. 218-20oC (25 h); IR
(KBr): υ (cm-1): 3423 (NH-broad), 1677 (C=O), 1029 (P-O-Et); 1H NMR (DMSO-d6, 250 MHz):
δ 0.91 (s, 3H, CH3-triazine), 1.10-1.12 (t, 6H, 2 x CH3-CH2-O, J = 5.0 Hz), 2.10 (s, 1H, N-NH),
3.90 (s, 1H, Ph-CH), 4.10 (q, 4H, 2 x CH3-CH2-O), 7.10-7.20 (m, 5H, Ph-), 13.20 (s, 1H, NH-
triazine) ppm; MS (EI): m/z (%) = 385 (M++1, <1), 313 (<1), 215 (1), 158 (100), 138 (C4H11O3P,
23.0); 31P NMR (DMSO-d6, 120 MHz): δ 68.0 (s, P, sym.) ppm; Anal. Calcd. for C15H21N4O4PS:
C, 46.87; H, 5.51; N, 14.58. Found: C, 46.71; H, 5.68; N, 13.78.
[(4-Methoxyphenyl)-6-methyl-5-oxo-3-thioxo-2,5-dihydro-3H-[1,2,4]triazin-4-
ylaminomethyl]phosphonate (5b); Ar = 4-MeOC6H4; Yield 1.8 g (70%); m.p. 247-49oC (20 h);
IR (KBr): υ (cm-1): 3212 (NH-amino), 3405 (NH-triazine), 1671 (C=O), 1024 (P-O-Et) 839 (p-
CH3O-aryl);1H NMR (DMSO-d6, 250 MHz): δ 2.18 (s, 3H, CH3-triazine), 2.48-2.50 (t, 6H, 2 x
CH3, J = 5.0 Hz), 3.80 (q, 4H, 2 x CH2, J = 6.1 Hz), 3.90 (s, 1H, NH-CH-Ar), 7.10-7.87 (m, 4H,
Harom), 8.50 (s, 1H, NH-triazine), 13.63 (s, 1H, NH-CH-Ar) ppm; MS (EI) m/z (%) = 414
(M+<1), 307 (11), 277 (14), 107 (8); Anal. Calcd. for C16H23N4O5PS: C, 46.37; H, 5.59; N,
13.52. Found: C, 46.20; H, 5.76; N, 13.72.
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[4-(Dimethylamino)phenyl)-6-methyl-5-oxo-3-thioxo-2,5-dihydro-3H-[1,2,4]triazin-4-
ylaminomethyl] phosphonate (5c); Ar = 4-Me2NC6H4; Yield 2.2 g (78%) [MeOH]; m.p. 268-
70oC (18 h); IR (KBr): υ (cm-1): 3183 (NH-amino), 3222 (NH-triazine), 1700 (C=O), 1058 (P-O-
Et), 809 (4-aminodimethyl-aryl); 1H NMR (DMSO-d6, 250 MHz): δ 0.95 (s, 3H, CH3-triazine),
1.10-1.12 (t, 6H, 2 x CH3-CH2-O, J = 5.0 Hz), 2.10 (s, 1H, N-NH), 2.80 (s, 6H, N-(CH3)2), 3.90
(s, 1H, Ph-CH), 4.11 (q, 4H, 2 x CH3-CH2-O), 6.60-6.80 (m, 4H, Ph-), 12.50 (s, 1H, NH-triazine)
ppm; MS (EI) m/z (%) = 427 (M+, <1), 307 (2), 291 (14), 137 (2); Anal. Calcd. for
C17H26N5O4PS: C, 47.77; H, 6.13; N, 16.38. Found: C, 47.87; H, 6.22; N, 16.29.
Synthesis of 2-(6-methyl-5-oxo-3-thioxo-2,5-dihydro-3H-[1,2,4]triazin-4-ylimino)-3-phenyl-
2,3-dihydro-1H-quinazolin-4-one (7)
A mixture of compound 1 (1.0 g, 6 mmol) and 3-phenyl-1H-qunazoline-2,4-dione (6a; 1.4 g, 6
mmol) was heated at 240oC for 4 h (TLC). After cooling, the resulting crude solid product was
crystallized from ethanol to give 7. Yield 1.8 g (65%); m.p. 208-10oC; IR (KBr): υ (cm-1): 3462
(NH-triazine), 3208 (NH-quinazoline), 2961-2929 (CH-aliphatic), 1660 (C=O-triazine), 1616
1
(C=O-quinazoline), 1227 (C=S); H NMR (DMSO-d6, 250 MHz): δ 0.94 (s, 3H, CH3-triazine),
3.90 (s, 1H, NH-quinazoline), 6.60-6.70 (m, 4H, Ph-quinazoline), 7.20-7.50 (m, 5H, -Ph), 12.50
(s, 1H, NH-triazine) ppm; MS (EI) m/z (%) = 378 (M+, <1), 364 (5), 236 (15), 156 (6); Anal.
Calcd. for C18H14N6O2S: C, 57.13; H, 3.73; N, 22.21. Found: C, 56.92; H, 3.92; N, 21.94.
Similarly, when a mixture of compound 1 and 3-phenyl-2-thioxo-2,3-dihydro-1H-quinazoline-4-
one (6b; 1.5 g, 6 mmol) was heated at 225oC for 3 h, the same product 7 was obtained in 76%
yield (2.4 g); melting point and mixed melting point with 7 gave no depression.
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Synthesis of 6-methyl-4-(2-phenyl-4-thioxo-4H-quinazolin-3-yl)-3-thioxo-3,4-dihydro-2H-
[1,2,4]triazine-5-one (9)
A mixture of compound 1 (1.0 g, 6 mmol) and 2-phenylbenzo[d][1,3]thiazine-4-thione (8; 1.3 g,
6 mmol) in anhydrous xylene (25 mL) was heated under reflux for 13 h (TLC). The reaction
solvent was evaporated and the solid residue was crystallized from MeOH/H2O to give product
1
9. Yield 1.1 g (45%), m.p. 163-65oC; IR (KBr): υ (cm-1): 2961 (CH), 1679 (C=O); H NMR
(DMSO-d6, 250 MHz): δ 2.50 (s, 3H, CH3-triazine), 7.10-8.11 (m, 9H, Harom), 9.10 (s, 1H, NH)
ppm; MS (EI) m/z (%) = 379 (M+, <1), 378 (9), 237 (8), 143 (4); Anal. Calcd. for C18H13N5OS2 :
C, 56.97; H, 3.45; N, 18.46. Found: C, 57.13; H, 3.61; N, 18.62.
Synthesis of 11a,b
General procedure: To a stirred solution of compound 1 (1.0 g, 6 mmol) in phosphorus
oxychloride (15 mL), the desired 4-sulfonylaminoacetic acid derivative [(toluene-4-
sulfonylamino)acetic acid (10a) or phenyl(toluene-4-sulfonylamino)acetic acid (10b)] (6 mmol)
was added in small portions. The reaction mixture was heated under reflux at 85oC for 5 h
(TLC). After cooling to room temperature, sodium hydroxide solution (1%, 50 mL) was added
and the resulting solid product was filtered, washed with water, dried, and finally recrystallized
from MeOH to give 11a,b, respectively.
4-Methyl-N-[(3-methyl-4-oxo-4H-[1,3,4]thiadiazolo[2,3-c]triazin-7-ylmethyl]benzenesulfon
amide (11a): R = H; Yield 1.5 g (50%), m.p. 136-38oC; IR (KBr): υ (cm-1): 3421 (NH), 1628-
1597(Harom), 1696 (C=O), 809 (para-CH3-aryl); 1H NMR (DMSO-d6, 250 MHz): δ 0.90 (s, 3H,
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CH3-triazine), 2.30 (s, 3H, CH3-Ph), 3.21 (s, 2H, CH2-NH), 5.62 (s, 1H, -CH2-S), 7.50 (s, 1H,
NH-triazine), 7.49-7.50 (d, 2H, Ph-, J = 2.5 Hz), 7.79-7.80 (d, 2H, Ph-S, J = 2.5 Hz), 8.50 (s, 1H,
NH-CH2) ppm; MS (EI) m/z (%) = 353 (M+, <1), 199 (C6H9N5OS, 2), 198 (11), 170 (7); Anal.
Calcd. for C13H15N5O3S2: C, 44.18; H, 4.28; N, 19.82. Found: C, 44.40; H, 4.11; N, 20.05.
4-Methyl-N-[(3-methyl-4-oxo-4H-[1,3,4]thiadiazolo[2,3-c]triazin-
7yl)phenylmethyl]benzenesulfon amide (11b): R = Ph; Yield 3.2 g (80%), m.p. 146-48oC; IR
(KBr): υ (cm-1): 3284 (NH), 1721 (C=O), 812 (para-CH3-aryl);1H NMR (DMSO-d6, 250 MHz):
δ 2.23 (s, 3H, CH3-aryl), 2.50 (s, 3H, CH3-triazine), 4.90 (s, 1H, Ph-CH-), 7.20-7.60 (m, 9H,
Harom), 8.60 (s, 1H, NH) ppm; MS (EI) m/z (%) = 429 (M+, <1), 353 (13), 275 (3), 77 (17); Anal.
Calcd. for C19H19N5O3S2: C, 53.13; H, 4.46; N, 16.31. Found: C, 52.98; H, 4.27; N, 16.23.
Synthesis of 7-aminomethyl-3-methyl-[1,3,4]thiadiazolo[2,3-c][1,2,4]triazin-4-one (12)
A suspension of compound 11b (0.5 g, 3 mmol) in aqueous HCl (30%, 35 mL) was heated under
reflux for 4 h. The reaction mixture was cooled, filtered and the filtrate was neutralized with
NaOH solution (10%, 25 mL). The precipitate formed was filtered, dried, and recrystallized from
methanol to afford 12.25 The filtrate was neutralized by dil. HCl to afford a white precipitate of
4-toluenesulphonic acid (m.p., 108-10oC; yield 45%) (Lit., m.p., 103-5oC).27
12: Yield 0.9 g (40%), m.p. 239-40oC; IR (KBr): υ (cm-1): 3248 (NH2, sym.), 3171 (NH2,
antisym.), and 1697 (C=O); 1H NMR (DMSO-d6, 250 MHz): δ 0.90 (s, 3H, CH3-triazine), 2.60
(s, 2H, CH2-NH2), 4.90 (s, 2H, -NH2), 5.54 (s, 1H, -CH-S), 7.90 (s, 1H, NH) ppm; MS (EI) m/z
(%) = 199 (M+, <1), 198 (7), 170 (2), 156 (5); Anal. Calcd. for C6H9N5OS: C, 36.17; H, 4.55; N,
35.15. Found: C, 36.42; H, 4.67; N, 35.42.
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Synthesis of 4-amino-6-methyl-3-(morpholin-4-yl-sulfanyl)-4H-[1,2,4]triazin-5-one (14)
To a stirred suspension of compound 1 (1.0 g, 6 mmol) in distilled water (10 mL), a mixture of
NaOH solution (0.39 g in 30 mL water, 8 mmol) and morpholine (0.52 g, 6 mmol) was added. A
solution of iodine (1.5 g, 6 mmol) and potassium iodide (1.0 g, 6 mmol) in distilled water (20
mL) was then added dropwise to the above reaction mixture and stirred at room temperature for
6 h. The resulting crude solid product thus formed was filtered, dried, and crystallized from DMF
to afford 14.28 Yield 0.5 g (10%); m.p. 108-10oC; IR (KBr): υ (cm-1): 3426 (NH-broad), 2920-
2852 (CH-aliphatic), 1631 (C=O); 1H NMR (DMSO-d6, 250 MHz): δ 2.35 (s, 3H, CH3-triazine),
2.75-2.81 (t, 2 x CH2-N-morpholine, J = 12.5 Hz), 3.45-3.50 (t, 4H, 2 x CH2-O-morpholine, J =
12.5 Hz), 5.10 (s, 2H, -N-NH2) ppm; MS (EI) m/z (%) = 243 (M+, <1), 242 (11), 156 (5), 86 (2);
Anal. Calcd. for C8H13N5O2S: C, 39.49; H, 5.39; S, 13.18. Found: C, 39.78, H, 5.68; S, 13.57.
Synthesis of 3,3'-bis(4-amino-6-methyl-5-oxo-triazin-yl)disulfide (16)
To a stirred solution of 1 (1.0 g, 6 mmol) in anhydrous methylene chloride (40 mL), lead
tetraacetate (2.6 g, 6 mmol) was added in portions over 15 min. The reaction mixture was stirred
at room temperature for 5 h. The lead salts formed were filtered and washed with methylene
chloride (10 mL). The filtrate was evaporated and the residual solid was crystallized from
ethanol to give 16. Yield 1.2 g (55%); m.p. 150-52oC; IR (KBr): υ (cm-1): 3292 (NH2, sym.),
3202 (NH2, antisym.), 1666 (C=O), 1499 (C=N); 1H NMR (DMSO-d6, 250 MHz): δ 2.40 (s, 6H,
2 x CH3-triazine), 4.93 (s, 4H, 2 x -N-NH2) ppm; MS (EI) m/z (%) = 314 (M+, <1), 281 (2), 249
(49), 160 (<1), 105 (100); Anal. Calcd. for C8H10N8O2S2: C, 30.57; H, 3.21; N, 35.65; S, 20.40.
Found: C, 30.82; H, 3.33; N, 35.92; S, 20.50.
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ACKNOWLEDGMENT
Authors are very thankful to Prof. A. R. El-Shanshoury (Microbiology Unit, Botany Department,
Faculty of Science, Tanta University, Tanta, Egypt) for his effort in the experimental work and
discussion of the antimicrobial part. The authors also are thankful to the Danish International
Development Agency (DINIDA) for their support.
REFERENCES
[1] Ahmada, M. R.; Sastrya, V. G.; Prasada, Y. R.; Khanb, M. H. R.; Banob N.; Anwarc, S. Eur.
J. Chem. 2012, 3, 94-98.
[2] Gautam, N.; Ajmera, N.; Gupta, S.; Gautam, D. C. Eur. J. Chem. 2012, 3, 106-111.
[3] Shaikh, B. M.; Chobe, S. S.; Konda, S. G.; Khandare, N. T.; Chavan, S. A.; Dawane, B. S.
Der Chem. Sin. 2010, 1, 86-88.
[4] Abdel‐Monem, W. R. Eur. J. Chem. 2010, 1, 168-172.
[5] Wrobleski S.T., Lin S., Hynes J. Jr., Wu H., Pitt S., Shen D.R., Zhang R., Gillooly K.M.,
Shuster D.J., McIntyre K.W., Doweyko A.M., Kish K.F., Tredup J.A., Duke G.J., Sack J.S.,
McKinnon M., Dodd J., Barrish J.C., Schieven G.L., Leftheris K.,. Bioorg. Med. Chem. Lett.
2008, 18, 2739-2744.
[6] Gahtori, P.; Ghosh, S. K. J. Enz. Inhib. Med. Chem. 2012, 27, 281-293.
[7] Krauth, F.; Dahse, H. M.; Rüttinger, H. H.; Froberg, P. Bioorg. Med. Chem. 2010, 18, 1816-
1821.
16
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[8] Pandey, V. K.; Yadav, S.; Joshi, M. N.; Bajpai, S. K. Biol. Mem. 2000, 26, 48-51.
[9] Bishnoi, A.; Saxena, R. Indian J. Hetero. Chem. 2001, 11, 47-50.
[10] Gottlieb, T. B.; Katz, F. H.; Chidsey, C. A. Circulation 1972, 45, 571-582.
[11] Unsalan, S.; Rollas, S. Indian J. Chem. B-Org. 2007, 46B, 185-191.
[12] Abdel-Rahman, R. M. Pharmazie 2001, 56, 18-22.
[13] Kumar, A.; Mukerjee, S. K.; Bhattacharya, S. K. Pharmazie 1983, 38, 66-67.
[14] Katsoulas, A.; Rachid, Z.; Brahimi, F.; McNamee, J.; Jean-Claude, B. J. Leukemia. Res.
2005, 29, 693-700.
[15] Wamhoff, H.; Tzanova, M. Arkivoc 2003, (ii), 98-102 and references therein.
[16] Baker, S. R.; Holdrege, C. T.; McGregor, D. N. J. Antibiot. 1979, 32, 468-471.
[17] El-Agrody, A. M.; Abdel-Latif, M. S.; El-Hady, N. A.; Fakery, A. H.; Bedair, A. H.
Molecules 2001, 6, 519-527.
[18] Modzelewska-Banachiewicz, B.; Kowalski, C.; Ziółkowska, G.; Banachiewicz, J. Annales
UMCS, sec DD 60, 2005, 2.
[19] Rauf, A.; Sharma, S.; Gnagal, S. Arkivoc 2007, (xvi), 137-147.
[20] Dornow, A.; Menzel, H.; Marx, P. Chem. Ber. 1964, 97, 2173-2178.
[21] Buck, J. S.; Ide, W.S. Organic Synthesis II, John Wiley & Sons, Inc. New York, 1964, 436-
453.
[22] Lu, S. M.; Chen, R. Y. Org. Prep. Proc. Int. 2000, 32, 302-306.
[23] Newallis, P. E.; In Mark, V. C.; Dungan, H.; Crutchfield, M. M.; Vanr, J. R. Topics in
Phosphorus Chemistry, Interscience New York, 1967, 5, 374.
[24] Ram, V.; Srimal, R. C. J. Prakt. Chem. 1990, 332, 629-639.
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[25] Okuda, J.; Nagaki, K.; Miwa, I.; Yashiro, T. Y. Chem. Pharm. Bull., 1982, 30, 3244-3254.
[26] Bader, H.; Downer, J. J. Chem. Soc. 1953, 1636-1640.
[27] Mann, F. G.; Saunders, B. C. Practical Organic Chemistry, Longman Group Ltd., London,
1975, 463-464.
[28] Fournier, E. U.S. Patent 1971, 3, 558, 775, Jan. 26; Chem. Abstr. 1972, 76, 1496316.
[29] 31st International Chemistry Olympiad Preparatory Problems, Bangkok, Thailand, July
1999.
[30] Diosady, L.L. Iodide, lodate, and Iron Phase 2 Final Report, Toronto ON Canada, M5S
1A4, July 1998.
[31] Shono, T; Matsumura, Y; Hayashi, J; Usui, M; Yamen, S-I; Inoue, K. Acta Chem.
Scand.1983, B 37, 491-498.
[32] Pridham, T. G.; Shotwell, O. L.; Stodola, F. H.; Lindenfelser, L. A.; Benedict, R. G.;
Jackson, R. W. Phytopathology 1956, 46, 575-581.
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