Synthesis, anti-inflammatory, analgesic, and antibacterial activities of some triazole, triazolothiadiazole, and triazolothiadiazine derivatives

Article abstract of DOI:10.1007/s12272-011-0802-z

This study is concerned with the synthesis of new 1,2,4-triazoles, 1,3,4-thiadiazoles, and 1,3,4- thiadiazines derivatives. Derivatives 3a-i were obtained by condensation of 4-amino-3-(4-pyridine)- 5-mercapto-1,2,4-triazole 1 with the appropriate aldehyde

Full text of DOI:10.1007/s12272-011-0802-z

Arch Pharm Res Vol 34, No 8, 1239-1250, 2011
DOI 10.1007/s12272-011-0802-z
Synthesis, Anti-inflammatory, Analgesic, and Antibacterial Activities of
Some Triazole, Triazolothiadiazole, and Triazolothiadiazine Derivatives
Mostafa A. Hussein1, Refaat M. Shaker2, Mohammed A. Ameen2, and Mohammed F. Mohammed2
1Department of Pharmaceutical Organic Chemistry, Faculty of Pharmacy, Assiut University, Assiut 71526, Egypt and
2Department of Chemistry, Faculty of Science, El-Minia University, El-Minia 61511, Egypt
(Received December 5, 2010/Revised January 3, 2011/Accepted January 4, 2011)
This study is concerned with the synthesis of new 1,2,4-triazoles, 1,3,4-thiadiazoles, and 1,3,4-
thiadiazines derivatives. Derivatives 3a-i were obtained by condensation of 4-amino-3-(4-pyri-
dine)-5-mercapto-1,2,4-triazole
1 with the appropriate aldehyde. Compounds 4a-i were syn-
thesized in a one pot reaction involving compounds 3a-i, formaldehyde, and morpholine.
Condensation of compound with the appropriate acids or 4-substituted phenacyl bromide
1
gave compounds 6a-d and 8a-f respectively. The chemical structures of the newly synthesized
derivatives were elucidated using different spectral and elemental methods of analysis. All
compounds were evaluated for their anti-inflammatory activity and the most potent deriva-
tives were tested for their analgesic activity using indomethacin as a reference drug. In addi-
tion, ulcerogenicity and LD50 for the most active compounds were evaluated. Moreover, the
antibacterial activities of the newly synthesized derivatives were investigated.
Key words: 1,2,4-Triazoles, Triazolothiadiazoles, Triazolothiadiazines, Anti-inflammatory, Anal-
gesic, Antibacterial
NSAIDs because they cause gastric erosions which
can cause stomach ulcers, and in extreme cases result
INTRODUCTION
Currently available non-steroidal anti-inflammatory in life threatening severe hemorrhage. Thus, the dis-
drugs (NSAIDs) alleviate pain by inhibiting the cyclo- covery of COX-2 provided the rationale for the de-
oxygenase (COX) enzyme which catalyzes the synthe- velopment of drugs devoid of GI effects while retaining
sis of prostaglandins. The NSAIDs prevent the prosta- clinical efficacy as anti-inflammatory agents. Therefore,
glandins from being synthesized, thereby reducing or the development of novel compounds having anti-
eliminating the pain (Karthikeyan et al., 2006). Addi- inflammatory and analgesic activity with improved
tionally, there are analgesics which are commonly as- safety and ideally, antimicrobial activities, is still a
sociated with anti-inflammatory drugs but have no necessity (Shivarama Holla et al., 2006).
anti-inflammatory effects; an example is paracetamol
(Kritsanida et al., 2002). There are at least two COX and the
Substituted 4-amino-1,2,4-triazol-5-thione derivatives
-bridged heterocycles derived from them have
N
isoforms - COX-1 and COX-2 (Kritsanida et al., 2002; been reported to possess antibacterial (Karthikeyan et
Karthikeyan et al., 2006). Constitutive COX-1 is re- al., 2006), antiviral (Karthikeyan et al., 2006), anti-
sponsible for providing cytoprotection in the gastro- cancer (Shivarama Holla et al., 2006), antitubercular
intestinal (GI) tract, whereas inducible COX-2 medi- (Walczak et al., 2004), analgesic (Amir and Shikha,
ates inflammation (Holla et al., 2005). Some inves- 2004), anti-inflammatory (Berk et al., 2001; Amir and
tigators are concerned about the long term usage of Shikha, 2004), anticonvulsant (Stillings et al., 1986),
and anti-depressant properties (Kane et al., 1988).
Correspondence to: Mostafa A. Hussein, Department of Phar-
maceutical Organic Chemistry, Faculty of Pharmacy, Assiut
University, Assiut 71526, Egypt
Tel: 2-12-743-2082, Fax: 2-88-233-2776
E-mail: mostafa1705@yahoo.com
Moreover, Mannich bases have been reported as poten-
tial biological agents. They find applications as antitu-
bercular (Joshi et al., 2004), antimalarial (Francisca et
al., 2004), vasorelaxing (Ferlin et al., 2002), anticancer
1239

1240
M. A. Hussein et al.
(Shivarama Holla et al., 2003), and analgesic agents catalyst. The reaction mixture was refluxed for 9 h,
(Malinka et al., 2005). Prompted by the aforementioned cooled, and the precipitated solid was filtered and
biological and pharmaceutical activities, and in conti- recrystallized from ethanol.
nuation of our previous work (Hamdy and Mostafa,
2006; Mohamed et al., 2006; Raafat, 2006; Raafat and 4-[Phenylmethylidene]amino-3-(4-pyridyl)-4,5-
Ali, 2006; Raafat and Mohammed, 2008), we have dihydro-1H-1,2,4-triazole-5-thione (3a)
attempted to develop new approaches for the synthe- M.p. 203-204oC, yield 65%; IR (KBr) νmax (cm−1): 1344
1
sis of polyfunctionally substituted heterocyclic com- (C=S), 1535 (C-N), 1470 (C=N), 3490 (NH). H-NMR
pounds with expected biological activity. This study (DMSO-d6):
δ (ppm) 8.20-9.50 (9H, m, aromatic protons),
presents the synthesis of a new series of Schiff and 10.75 (IH, s, CH), 13.85 (1H, s, triazole, NH). EI-MS
Mannich bases, containing 1,2,4-triazole, triazolothia- m/z: 280.92 (M+-1/20.5%), 281.15 (M+/0.6%), 192.93
diazoles, and triazolothiadiazines, and the screening (72.6%), 177.94 (100%), 105.01 (38.8%). Anal. Calcd.
of their anti-inflammatory, antibacterial, and analgesic (%) for C14H11N5S: C, 59.77; H, 3.94; N, 24.89; S,
activities. In addition, ulcerogenic and LD50 activities 11.40. Found: C, 59.87; H, 3.85; N, 24.90; S, 11.60.
were also investigated.
4-[4-Bromophenylmethylidene]amino-3-(4-pyri-
dyl)-4,5-dihydro-1H-1,2,4-triazole-5-thione (3b)
MATERIALS AND METHODS
Chemistry
M.p. 295-300oC, yield 80%; IR (KBr) νmax (cm−1): 1345
1
(C=S), 1467 (C=N), 3425 (NH), 1557 (C-N). H-NMR
Melting points were determined in open capillary (DMSO-d6):
δ (ppm) 8.20-9.80 (8H, m, aromatic pro-
tubes on an electrothermal melting point apparatus tons), 10.30 (1H, s, N=CH), 13.85 (1H, NH, triazole).
(Stuart Scientific, model SMPI) and all were uncor- EI-MS m/z: 360.60 (M+/6.7%), 362.63 (M+ +2/8.0%),
rected. Precoated silica gel plates (Kieselgel 0.25 mm, 177.81 (100%), 118.91 (42.8%). Anal. Calcd. (%) for
60 f-254, Merck) were used for TLC for monitoring of C14H10N5SBr: C, 46.68; H, 2.80; N, 19.44; S, 8.90. Found:
reactions, and the spots were visualized under a UV C, 46.98; H, 2.50; N, 19.84; S, 8.90.
lamb (Spectroline, model CM-10). Infrared spectra
were recorded as KBr disks on a Shimadzu spectro- 4-(4-Chlorophenylmethylidene) amino-3-(4-pyri-
photometer (IR-470) at the Faculty of Pharmacy, Assiut dyl)-4,5-dihydro-1H-1,2,4-triazole-5-thione (3c)
1
University. H-NMR spectra were run on an EM-360 M.p. 220-224oC, yield 75%; IR (KBr) νmax (cm−1): 1344
1
Varian 60 MHz NMR spectrometer at the Faculty of (C=S), 1450 (C=N), 3455 (NH), 1534 (C-N). H-NMR
Pharmacy, Assiut University, and DMSO-d6 (unless (DMSO-d6):
otherwise stated) was used as a solvent and TMS as tons), 10.30 (1H, s, N=CH), 13.85 (1H, s, NH triazole).
an internal standard (chemical shifts are taken in
EI-MS m/z: 314.63 (M+-1/6.4), 316.66 (M++1/2.7),
δ (ppm) 8.20-10.20 (8H, m, aromatic pro-
δ
ppm). Mass spectra were produced using a JEOL JMS 177.81 (46.60%), 136.83 (100%). Anal. Calcd. (%) for
600 mass spectrometer at the Microanalytical Center, C14H10N5SCl: C, 53.25; H, 3.19; N, 22.18; S, 10.15.
Assiut University, Egypt. Elemental analyses were Found: C, 53.35; H, 3.33; N, 22.25; S, 10.19.
done on a Perkin-Elmer 240 elemental analyzer at the
unit of microanalysis, Assiut University or Cairo Uni- 4-[4-Hydroxyphenylmethylidene]amino-3-(4-pyri-
versity, Egypt.
4-Amino-1,2,4-triazole-3-thiones were prepared M.p. 200-205oC, yield 70%; IR (KBr) νmax (cm−1): 1350
according a reported procedures (Raafat and Ali, (C=S), 1458 (C=N), 3450 (NH). 1H-NMR (DMSO-d6):
2006). Moreover, during this work, other groups re- (ppm) 7.50-9.60 (8H, m, aromatic protons), 10.20 (1H,
ported the synthesis and characterization of Schiff s, N=CH), 15.50 (1H, s, NH triazole). EI-MS m/z
base hydraones bearing a 3-(4-pyridyl)-5-mercapto- 297.47 (M+/1.5%), 281.17 (18.0%). Anal. Calcd. (%) for
dyl)-4,5-dihydro-1H-1,2,4-triazole-5-thione (3d)
δ
:
1,2,4-triazoles moiety (Khanmohammadi et al., 2008).
C14H11ON5S: C, 56.55; H, 3.73; N, 23.55; S, 10.78.
Found: C, 53.55; H, 3.74; N, 22.59; S, 10.80.
General method for synthesis of 4-[1-substituted
methylidene]amino-3-(4-pyridyl)-4,5-dihydro-
1H-1,2,4-triazole-5-thione compounds (3a-i)
4-[4-Nitrophenylmethylidene]amino-3-(4-pyridyl)-
4,5-dihydro-1H-1,2,4-triazole-5-thione (3e)
To a suspension of aryl aldehyde,
dioxane (10 mL), was added an equimolar amount of (C=S), 1469 (C=N), 3410 (NH). 1H-NMR (DMSO-d6):
3-(4-pyridyl)-4-amino-5-mercapto-1,2,4-triazole, in (ppm) 8.60-9.60 (8H, m, aromatic protons), 11.60 (1H, s,
the presence of a few drops of conc. sulfuric acid as a N=CH), 16.60 (1H, s, NH triazole). EI-MS m/z: 325.49
2
(0.005 mol) in M.p. 270-272oC, yield 88%; IR (KBr) νmax (cm−1): 1337
δ
1

Biological Activities of New Triazole Derivatives
1241
(M+-/13.9%), 280.63 (12.1%), 177.76 (100%). Anal. Calcd. dissolved in a mixture of ethanol and dioxane (2:1),
(%) for C14H10O2N6S: C, 51.53; H, 3.09; N, 25.75; S, followed by addition of formaldehyde (40%, 1.5 mL)
9.83. Found: C, 51.63; H, 3.00; N, 25.95; S, 9.95
and morpholine (1 mL). The reaction mixture was
stirred for 1-2 h, and kept over night at room tempera-
ture. The separated solid was collected by filtration,
4-(4-Dimethylamino)phenylmethylidene]amino-
3-(4-pyridyl)-4,5-dihydro-1H-1,2,4-triazole-5-thione dried, and crystallized from ethanol.
(3f)
M.p. 215-220oC, yield 80%; IR (KBr)
ν
max (cm−1):
−
1352 4-(Benzylidenamino)-2-(morpholinomethyl)-5-
1
(C=S), 1571 (C-N), 1460 (C=N), 3490 (NH). H-NMR (pyridine-4-yl)-2H-1,2,4-triazole-(4H)-thione (4a)
(DMSO-d6):
δ −1249
(ppm) 3.40 (6H, S, CH3), 7.30-9.50 (8H, M.p. 170-172oC, yield 75%; IR (KBr) νmax (cm−1):
m, aromatic protons), 10.02 (IH, s, CH), 15.90 (1H, s, (C=S), 1693 (CH=N), 1422 (C=N). 1H-NMR (CDCl3):
δ
triazole, NH). EI-MS m/e: 324.32 (M+/2.1%), 326.11 (ppm) 3.20 (4H, d, CH2), 4.00 (4H, d, CH2), 5.70 (2H, s,
(4.6%), 177.55 (100.0%). Anal. Calcd. (%) for C16H16N6S: CH2), 8.10-9.50 (9H, m, aromatic protons), 11.10 (1H,
C, 59.24; H, 4.97; N, 25.91; S, 9.88. Found: C, 59.34; s, CH). EI-MS m/z: 380.33 (M+/1.6%), 381.10 (20.8%),
H, 5.10; N, 25.90; S, 10.00.
382.60 (5.2%), 99.56 (100%). Anal. Calcd. (%) for
C19H20N6OS: C, 59.98; H, 5.30; N, 22.09; S, 8.43.
Found: C, 59.99; H, 5.33; N, 22.30; S, 8.49.
4-[4-Methoxyphenylmethylidene]amino-3-(4-pyri-
dyl)-4,5-dihydro-1H-1,2,4-triazole-5-thione (3g)
M.p. 210-212oC, yield 55%; IR (KBr)
ν
max (cm−1):
(ppm) 4.50 methyl)-5-(pyridine-4-yl)-2H-1,2,4-triazole-3(4H)-
−1560 4-(4-Bromobenzylideneamino)-2-(morpholino-
1
(C=N), 3495 (NH). H-NMR (DMSO-d6):
δ
(3H, s, OCH3), 8.70 (dd, NCH), 8.20-9.40 (8H, m, aro- thione (4b)
matic protons), 14.20 (1H, s, triazole, NH). EI-MS m/ M.p. 180-182oC, yield 85%; IR (KBr)
ν
max (cm−1):
−
1475
z
: 310.86 (M+-1/1.0%), 278.91 (100.0%), 120.94 (77.5 (C=N), 1272 (C=S), 1587 (C-N). 1H-NMR (CDCl3):
δ (ppm)
%), 101.06 (74.70%). Anal. Calcd. (%) for C15H13N5S: 3.40 (4H, d, CH2), 4.10 (4H, d, CH2), 5.90 (2H, s, CH2),
C, 57.86; H, 4.21; N, 22.49; S, 10.30. Found: C, 57.96; 8.10-9.50 (8H, m, aromatic protons), 11.80 (1H, s, CH).
H, 4.42; N, 22.60; S, 10.20.
EI-MS m/z: 459.36 (M+/1.5%), 460.37 (M++1/0.2%),
461.44 (M++2/0.2%), 267.66 (1.7%), 180.67 (80.3%), 99.92
(100%). Anal. Calcd. (%) for C19H19N6OBrS: C, 46.68;
H, 4.17; N, 18.29; S, 6.98. Found: C, 46.98; H, 4.33; N,
18.34; S, 6.90.
3-(4-Pyridyl)-4,5-dihydro-1H-1,2,4-triazole-5-thione
3-(4-pyridyl)-4-[1-(2-thienyl)methylidene]amino-
4,5-dihydro-1
M.p. 255-257oC, yield 85%; IR (KBr) νmax (cm−1): 1348
(C=S), 1461 (C=N), 3500 (NH). 1H-NMR (DMSO-d6):
H-1,2,4-triazole-5-thione (3h)
δ
4-[(4-Chlorophenyl)methylidene]amino-1-(mor-
(ppm) 7.90-9.37 (7H, m, aromatic protons), 10.20 (1H, pholinomethyl)-3-(4-pyridyl)-4,5-dihydro-1H-1,2,4-
s, N=CH), 15.40 (1H, s, NH triazole). EI-MS m/z
:
triazole-5-thione (4c)
287.40 (M+/1.2%), 289.03 (9.5%), 177.06 (100.0%). Anal. M.p. 198-200oC, yield 80%; IR (KBr) νmax (cm−1):
−
1475
1
Calcd. (%) for C12H9N5S2: C, 50.16; H, 3.16; N, 24.37; (C=N), 1272 (C=S), 1587 (C-N). H-NMR (CDCl3):
S, 22.32. Found: C, 50.26; H, 3.36; N, 24.67; S, 22.66.
δ
(ppm) 3.20 (4H, d, CH2), 4.10 (4H, d, CH2), 5.90 (2H, s,
CH2), 8.10-9.50 (8H, m, aromatic protons), 11.80 (1H, s,
4-[1-(2-Furyl) methylidene] amino-3-(4-pyridyl)- CH). EI-MS m/z: 414.85 (M+/0.4%), 416.90 (M++2/
4,5-dihydro-1H-1,2,4-triazole-5-thione (3i)
0.1%), 100.02 (100%). Anal. Calcd. (%) for C19H19N6OClS:
−1392 C, 55.00; H, 4.62; N, 20.25; S, 7.73. Found: C, 55.05;
M.p. 350-352oC, yield 47%; IR (KBr) νmax (cm−1):
1
(C=S), 1461 (C=N), 3490 (NH). H-NMR (DMSO-d6):
δ
H, 4.75; N, 20.35; S, 7.87.
(ppm) 7.40-9.60 (7H, m, aromatic protons), 10.60 (1H,
s, N=CH), 15.40 (1H, s, NH triazole). EI-MS m/z
:
4-[4-Hydroxyphenyl) methylidene] amino-1-(mor-
271.37 (M+/1.8%), 272.06 (13.1%), 273.05 (4.8%). Anal. pholinomethyl)-3-(4-pyridyl)-4, 5-dihydro-1H-1,
Calcd. (%) for C12H9N5OS: C, 53.13; H, 3.34; N, 25.81; 2, 4-triazole-5-thione (4d)
S, 11.82. Found: C, 53.43; H, 3.74; N, 25.90; S, 12.01.
M.p. 195-196oC, yield 80%; IR (KBr) νmax (cm−1):
−1475
(C=N), 1272 (C=S), 1587 (C-N). 1H-NMR (CDCl3):
δ (ppm)
2.37 (4H, d, CH2), 3.67 (4H, d, CH2), 4.90 (2H, s, CH2),
8.10-9.50 (8H, m, aromatic protons), 9.50 (1H, s, CH),
14.10 (1H, s, OH). EI-MS m/z: 396.20 (M+/1.1%), 397.14
(21.7%), 398.13 (4.6%), 99.56 (100%). Anal. Calcd. (%)
General method for synthesis of 4-[1-4-substi-
tuted phenyl methylidene]amino-1-(morpho-
linomethyl)-3-(4-pyridyl)-4, 5-dihydro-1H-1, 2, 4-
triazole-5-thione (4a-i)
The appropriate compounds 3a-i (10 mmol) were for C19H19N5O2S: C, 57.56; H, 5.08; N, 21.20; S, 8.09.

1242
M. A. Hussein et al.
Found: C, 57.55; H, 5.35; N, 21.59; S, 8.30.
4-(Furan-2-yl)methyleneamino)-2-(morpholino-
methyl)-5-(pyridin-4-yl)-2H-1,2,4-triazole-3(4H)-
4-(4-Nitrobenzylideneamino)-2-(morpholinometh- thione (4i)
yl)-5-(pyridin-4-yl)-2H-1,2,4-triazole-3(4H)-thione
(4e)
M.p. 208-210oC, yield 40%; IR (KBr)
ν
max (cm−1):
(C=S), 1475 (C=N), 1595 (C-N). H-NMR (CDCl3):
1249 (ppm) 3.30 (4H, dd, CH2), 4.10 (4H, dd, CH2), 5.60 (2H,
(C=S), 1693 (CH=N), 1422 (C=N). 1H-NMR (CDCl3):
s, CH2), 7.60-8.40 (3H, m, CH furan), 8.50-9.40 (4H,
−1249
1
δ
M.p. 204-205oC, yield 80%; IR (KBr) max (cm−1):
ν −
δ
(ppm) 3.20 (4H, d, CH2), 4.20 (4H, d, CH2), 5.70 (2H, m, CH, pyridine), 11.70 (1H, s, CH). EI-MS m/z
:
s, CH2), 7.90-9.60 (8H, m, aromatic protons), 11.90 370.34 (M+/1.2%), 371.12 (21.4%), 372.12 (5.1%). Anal.
(1H, s, CH). EI-MS m/z: 424.73 (M+-1/0.2%), 325.75 Calcd. (%) for C17H18N6O2S: C, 55.12; H, 4.90; N,
(0.2%), 214.91 (0.1%). 99.9 (100%). Anal. Calcd. (%) for 22.69; S, 8.66. Found: C, 55.43; H, 4.95; N, 22.90; S,
C19H19N7O3S: C, 53.64; H, 4.50; N, 23.04; S, 7.54. 8.96.
Found: C, 53.63; H, 4.60; N, 23.45; S, 7.65.
General method for synthesis of 1,2,4-triazolo-
[3.4-b]-1,3,4-thiodiazole derivatives (6a-d)
To a mixture of corresponding 3-(4-pyridyl)-4-amino-
N,N-Dimethylamino-N'-[1-(morpholinomethyl)-3-
(4-pyridyl)-5-thioxo-4,5-dihydro-1H-1,2,4-triazol-
4-yl] iminoformamide (4f)
5-mercapto-1,2,4-triazole,
1518 ate carboxylic acid (0.52 mL, 0.01 mol), phosphorus
(C-N), 1273 (C=S), 1668 (CH=N), 1465 (CH2-O). H- oxychloride (10 mL) was added and the reaction mix-
NMR (CDCl3): (ppm) 3.30 (6H, s, CH3), 3.10 (4H, dd, ture was refluxed for 5 h. The mixture was concen-
1 (0.01 mol) and the appropri-
M.p. 168-169oC, yield 90%; IR (KBr)
ν
max (cm−1):
−
5
1
δ
CH2), 4.00 (4H, dd, CH2), 5.70 (2H, s, CH2), 7.20-9.50 trated under reduced pressure; ice-water was added
(8H, m, aromatic protons), 10.3 (1H, s, CH). EI-MS with vigorous stirring. The precipitate was filtered,
m/z: 423.54 (M+/1.9%), 424.29 (23.0%), 425.18 (5.2%), washed with 20% sodium bicarbonate solution and
99.05 (100%). Anal. Calcd. (%) for C21H25N7OS: C, water, dried, and recrystallized from dimethylform-
59.24; H, 5.95; N, 23.15; S, 7.57. Found: C, 59.34; H, amide/water mixture (1:1).
5.87; N, 23.50; S, 7.85.
6-Methyl-3-(pyridine-4yl)-[1,2,4]triazole[3,4-b][1,3,
4-[-1-(4-Methoxyphenyl)methylidene]amino-1-(mor-
pholinomethyl)-3-(4-pyridyl)-4,5-dihydro-1H-1,2,
4-triazole-5-thione (4g)
4]thiadiazole (6a)
M.p. 257-258oC, yield 95%; IR (KBr)
ν
max (cm−1):
(ppm) 2.90
M.p. 138-140oC, yield 45%; IR (KBr) max (cm−1):
ν −1249 (3H, s, CH3), 8.10-8.80 (4H, d, pyridine). EI-MS: m/z:
1
−1559
(C=N), 1259 (C-S). 1H-NMR (DMSO-d6):
δ
(C=S), 1556 (C-N), 1687 (CH=N), 1422 (C=N). H- 216.99 (M+/100%), 176.98 (0.2%). 104.01 (51.8%). Anal.
NMR (CDCl3): (ppm) 3.20 (4H, d, CH2), 3.90 (4H, d, Calcd. (%) for C9H7N5S: C, 49.76; H, 3.25; N, 32.24; S,
δ
CH2), 4.20 (3H, s, OCH3), 5.70 (2H, s, CH2), 7.50-8.40 14.76. Found: C, 49.99; H, 3.33; N, 32.30; S, 14.49.
(4H, dd, phenyl proton), 8.60-9.50 (4H, dd, pyridine
protons), 10.70 (1H, s, CH). EI-MS m/z: 409.86 (M+-1/ 6-Phenyl-3-(pyridin-4-yl)-[1,2,4]triazolo[3,4-b][1,3,
1.3%), 411.73 (0.1%), 310.69 (0.4%), 99.98 (100). Anal. 4]thiadiazole (6b)
Calcd. (%) for C20H22N6O2S: C, 58.52; H, 5.40; N, 20.47; M.p. 298-300oC, yield 60%; IR (KBr)
S, 7.81. Found: C, 58.86; H, 5.55; N, 20.60; S, 7.95.
ν
max (cm−1):
−1559
(C=N), 1259 (C-S). 1H-NMR (DMSO-d6):
δ (ppm) 8.40-
9.50 (9H, m, aromatic protons). EI-MS m/z: 278.99
(M+/100), 255.75 (7.1%), 120.98 (78.1%). Anal. Calcd.
(%) for C14H9N5S: C, 60.20; H, 3.25; N, 25.07; S, 11.48.
Found: C, 60.25; H, 3.26; N, 25.09; S, 11.55.
4-(Thienyl-2-yl)methyleneamino)-2-(morpholino-
methyl)-5-(pyridin-4-yl)-2H-1,2,4-triazole-3(4H)-
thione (4h)
M.p. 143-145oC, yield 85%; IR (KBr)
ν
max (cm−1):
−
1249
1
(C=S), 1475 (C=N), 1595 (C-N). H-NMR (CDCl3):
δ
3,6-Di(pyridin-4-yl)-[1,2,4]triazolo[3,4-b][1,3,4]thi-
(ppm) 3.01 (4H, dd, CH2), 3.90 (4H, dd, CH2), 5.60 (2H, adiazole (6c)
s, CH2), 7.60-8.20 (3H, m, CH thiophene), 8.50-9.40 M.p. 290-292oC, yield 80%; IR (KBr)
ν
max (cm−1):
−1559
(4H, m, CH, pyridine), 11.70 (1H, s, CH). EI-MS m/z
:
(C=N), 1259 (C-S). 1H-NMR (DMSO-d6):
δ (ppm) 8.20-
386.33 (M+/3.6%), 387.10 (22.4%), 388.09 (9.1%). Anal. 9.00 (8H, m, aromatic protons). EI-MS m/z: 279.91
Calcd. (%) for C17H18N6OS2: C, 52.83; H, 4.69; N, (M+/1.0%), 255.62 (57.9%), 191.72 (14.4%), 121.99
21.74; S, 16.56. Found: C, 52.27; H, 4.36; N, 21.85; S, (70.1%). Anal. Calcd. (%) for C13H8N6S: C, 55.70; H,
16.75.
2.88; N, 29.98; S, 11.44. Found: C, 55.89; H, 2.95; N,
29.99; S, 11.49.

Biological Activities of New Triazole Derivatives
1243
6-(Pyridin-3-yl)-3-(pyridin-4-yl)-[1,2,4]triazolo[3,4-
b][1,3,4]thiadiazole (6d)
C=N, 1250 C-S. 1H-NMR (DMSO-d6):
s, methylene, protons), 7.60-8.90 (8H, m, aromatic pro-
1550 tons). EI-MS m/z: 372.55 (1.8%), 374.98 (4.7%), 99.05
δ (ppm) 8.40- (100.0%). Anal. Calcd. (%) for C15H10N5BrS: C, 48.40;
δ
(ppm) 4.40 (2H,
M.p. 262-265oC, yield 85%; IR (KBr) max (cm−1):
ν −
(C=N), 1259 (C-S). 1H-NMR (DMSO-d6):
9.50 (8H, m, aromatic protons). EI-MS m/z: 279.67 H, 2.71; N, 18.81; S, 8.61. Found: C, 48.67; H, 2.95; N,
(M+/100%), 255.41 (4.6%), 121.84 (73.3%). Anal. Calcd. 19.10; S, 8.72.
(%) for C13H8N6S: C, 55.70; H, 2.88; N, 29.98; S, 11.44.
Found: C, 55.95; H, 2.95; N, 30.00; S, 11.45.
6-(4-Methoxyphenyl)-3-(pyridin-4-yl)-7H-[1,2,4]tri-
azolo[3,4-b][1,3,4]thiadiazine (8e)
M.p. 215-227oC, yield 78%; IR (KBr) νmax (cm−1): 1675
General method for synthesis of 4-substitud-
phenyl-3-(pyridine-4-yl)-7H-(1,2,4)triazolo[3,4-b]-
[1,3,4]thiadiazine (8a-f)
C=N, 1250 C-S. 1H-NMR (DMSO-d6):
δ (ppm) 3.90 (3H,
s, OCH3), 4.60 (2H, s, methylene, protons), 7.20-9.00
A solution of 3-(4-pyridyl)-4-amino-5-mercapto-1,2,4- (8H, m, aromatic protons). EI-MS m/z: 323.55 (M+/
triazole,
1 (0.005 mol) and phenacyl bromide 7 (0.005 1.5%), 324.43 (17.5%), 325.58 (4.9%). Anal. Calcd. (%)
mol) in absolute ethanol (30 mL) was heated under for C16H13N5OS: C, 59.43; H, 4.05; N, 21.66; S, 9.92.
reflux for 2 h, cooled to room temperature, and then Found: C, 59.65; H, 4.35; N, 21.96; S, 10.10.
neutralized with ammonium hydroxide (35% v/v). The
solid was separated by filtration, dried, and recry- 6-(4-Fluorophenyl)-3-(pyridin-4-yl)-7H-[1,2,4]tria-
stallized from ethanol.
zolo[3,4-b][1,3,4]thiadiazine (8f)
M.p. 234-235oC, yield 80%; IR (KBr) νmax (cm−1): 1675
6-Phenyl-3-(pyridine-4-yl)-7H-[1,2,4]triazolo[3,4-b]
[1,3,4]thiadizaine (8a)
C=N, 1250 C-S. 1H-NMR (DMSO-d6):
δ (ppm) 4.60 (2H,
s, methylene, protons), 7.30-9.00 (8H, m, aromatic pro-
M.p. 268-270oC, yield 65%; IR (KBr) νmax (cm−1): 1690 tons). EI-MS m/z: 311.46 (0.2%), 312.11 (16.3%), 313.06
C=N, 1249 C-S. 1H-NMR (DMSO-d6):
δ (ppm) 4.80 (2H, (4.8%), 99.56 (100.0%). Anal. Calcd. (%) for C15H10N5FS:
s, methylene protons), 8.20-9.40 (9H, m, aromatic C, 57.87; H, 3.24; N, 22.49; S, 10.30. Found: C, 57.99;
protons). EI-MS m/z: 293.20 (M+/2.85%), 259.24 (2.3%), H, 3.22; N, 22.49; S, 10.30.
190.12 (7.9%). Anal. Calcd. (%) for C15H11N5S: C,
61.42; H, 3.78; N, 23.87; S, 10.93. Found: C, 61.44; H,
3.88; N, 23.99; S, 10.99.
Pharmacological screening
Male adult albino rats and mice were obtained from
the animal house, Faculty of Medicine, Assiut Uni-
versity. Indomethacin (Liometacin® vial, Nile Company),
carrageenan (Sigma), sodium carboxymethylcellulose
6-(4-Nitrophenyl)-3-(pyridine-4-yl)-7H-[1,2,4]tria-
zolo[3,4-b][1,3,4]thiadiazine (8b)
M.p. 200-202oC, yield 60%; IR (KBr) νmax (cm−1): 1675 (NaCMC) and normal saline were obtained from a
C=N, 1250 C-S. 1H-NMR (DMSO-d6):
s, methylene, protons), 8.70-9.80 (8H, m, aromatic pro- were suspended in 5% NaCMC in normal saline.
δ (ppm) 4.80 (2H, local market. The test compounds and reference drug
tons). EI-MS m/z: 338.14 (M+/3.1%), 339.15 (9.6%),
Animals were housed in separate cages, 6 animals
340.13 (0.2%), 293.21 (14.3%), 150.14 (100%). Anal. each, in temperature-controlled rooms at 25 ± 1oC.
Calcd. (%) for C15H10N6O2S: C, 53.25; H, 2.98; N, 24.84; Animals were allowed free access to food and water
S, 9.48. Found: C, 53.36; H, 3.12; N, 24.95; S, 9.55.
and maintained at a 12 h light/dark cycle. Work was
conducted in accordance with the internationally
6-(4-Chlorophenyl)-3-(pyridin-4-yl)-7H-[1,2,4]tri- accepted principles for laboratory animals' use and
azolo[3,4-b][1,3,4]thiadiazine (8c) care as found in the European Community Guidelines
M.p. 219-220oC, yield 60%; IR (KBr) νmax (cm−1): 1675 (Tan, 2004).
C=N, 1250 C-S. 1H-NMR (DMSO-d6):
(ppm) 4.40 (2H,
δ
s, methylene, protons), 7.50-8.90 (8H, m, aromatic
Anti-inflammatory activity
protons). EI-MS m/z: 327.27 (3.3%), 329.03 (36.8%),
The anti-inflammatory activity of 29 final compounds
150.34 (100.0%). Anal. Calcd. (%) for C15H10N5ClS: C, was determined according to the paw induced edema
54.96; H, 3.07; N, 21.37; S, 9.78. Found: C, 54.99; H, method (Nargund et al., 1994). Indomethacin was
3.17; N, 21.56; S, 9.95.
used as a comparator and reference drug. The test is
based on the pedal inflammation in rat paws induced
by subplantar injection of 0.2 mL carrageenan suspen-
sion (5% NaCMC) into the right hind paw of the rats.
Male adult albino rats (120-150 g) were divided into
6-(4-Bromophenyl)-3-(pyridin-4-yl)-7H-[1,2,4]tria-
zolo[3,4-b][1,3,4]thiadiazine (8d)
M.p. 225-228oC, yield 70%; IR (KBr) νmax (cm−1): 1675

1244
M. A. Hussein et al.
groups of 4 animals each and fed ad libitum with mm in length. All lesions of > 0.1 mm in length were
rodent’s chow and allowed free access to drinking summed to obtain the ulcer index and results are
water. The thickness of the rat paw was measured by listed in Table III.
a Veriner caliper (SMIEC) before and 1 h after injec-
tion of carrageenan to detect the inflammation in-
duced by carrageenan.
Acute toxicity (LD50)
The median lethal dose (LD50) of compounds 8e and
Test compounds at doses of 10 mg/kg in 5% NaCMC 8f was determined in mice by the graphical Litchfield
were injected i.p. into 3 different groups of rats. The method (O'Neil et al., 2001). Groups of male adult
control group received a vehicle (5% NaCMC), and the albino mice, (25-30 g, 4 mice/group) were injected i.p.
reference group received indomethacin i.p. at 0.03 with graded doses of the test compound. The percen-
mmol/kg. The difference between the thicknesses of tage mortality in each group of animals was determined
the 2 hind paws was taken as a measure of edema. 72 h after injection. Computation of LD50 was processed
The measurement was carried out at times of zero, by a graphical method and was found to be 275 and
0.5, 1, 2, 3, 4 and 5 h after injection of the test com- 300 mg/kg, for compounds 8e and 8f, respectively,
pounds, reference drug, or the vehicle. Results of anti- whereas the LD50 for indomethacin was 13 mg/kg (i.p.)
inflammatory activity of the test compounds and the (O'Neil et al., 2001).
reference drug are listed in Table I.
Antibacterial
The antibacterial activity of compounds
1, 3a-i, 4a-
Analgesic activity
Hot-plate test
i, 6a-d, and 8a-f was investigated in vitro against
The analgesic activity of 8a-f was determined in methicillin resistant Staphylococcus aureus (MRSA),
mice using the hot-plate method (Boyle et al., 1986), in Bacillus cereus Escherichia coli, and Klebsiella
,
comparison with indomethacin as a reference drug. pneumoniae (clinical isolates obtained from Infection
Male albino mice (30-35 g) were placed on a hot-plate Control Unit, Assiut Univeristy Hospital, Faculty of
apparatus for testing and the surface temperature Medicine, Assiut University) using the agar cup
was maintained at 55 ± 0.5°C. The reaction time in diffusion method for susceptibility screening, and the
seconds was taken as the time period from the instant 2-fold dilution method (William, 1977) for MIC deter-
the animal reached the hot plate until the moment the mination. Ampicillin was used as a reference drug,
animal licked its hind paw or jumped out within a and DMSO as the solvent.
Plexiglas cylinder placed on the hot-plate. The reac-
tion time was taken as the end point and the increase
in hot plate latency was taken as a measure of the
analgesic activity.
Agar cup diffusion method
Mueller-Hinton agar medium (MH) (Hi-Media, M
001), 38 g, was added to 1 L of distilled water, heated
Animals were randomly divided into groups of 4 to boiling to dissolve the ingredients completely, and
mice each and each animal was used once. Solutions sterilized by autoclaving at 121°C for 30 min. High
or suspensions of the test compounds and the refer- density inocula were prepared by diluting 3-5 well
ence drug in 5% NaCMC were injected i.p. at a dose isolated colonies grown overnight on selective media
level of 0.02 mmol/kg. Control animals were similarly in 5 mL of distilled water to prepare a suspension equi-
treated with 5% NaCMC. Testing was done at 0.5, 1, valent in density to 0.5 McFarland Barium Sulfate
2, 3, and 5 h after the injection. Mean licking time ± standard units with an average turbidity of 107 CFU/
S.E. was evaluated for each group and listed in Table mL.
II.
The sterile Petri dishes were seeded with 100 µL of
the microorganism; a specified amount of the molten
MH agar medium (45-50oC) was poured into the seed-
Ulcerogenic effect
The test was conducted according to a reported method ed Petri dishes to give a depth of 3-4 mm and allowed
(Boyle et al., 1986) using adult male albino rats. Male to solidify. Cylindrical plugs were removed from the
albino rats were fasted for 24 h. Tested compounds agar using a sterile cork borer. Tested compounds (20
8d
,
8e, and 8f or reference drug were administered
µ
L in DMSO), the blank solvent, and ampicillin sodium
orally to groups of 6 animals each. After 6 h, the (20
µmol/mL in DMSO) were added to the wells in
animals were sacrificed, their stomachs were removed triplicate. The seeded plates were incubated at 37°C
and gastric lesions on the mucosa were determined by for 24 h the average diameters of the inhibition zones
using a stereoscopic microscope (XJP-XSC-195-40X). were measured in millimeters (Table IV).
“Ulcer” was defined as at least 1 lesion that was 0.5

Biological Activities of New Triazole Derivatives
1245
Schiff bases 3a-i were obtained by the condensation
Minimum inhibitory concentration
The minimum inhibitory concentration (MIC) was of 4-amino-3-(4-pyridine)-5-mercapto-1,2,4-triazole
1
determined using the 2-fold dilution method (William, with appropriate aldehydes in the presence of a cata-
1977) for compounds having moderate to strong anti- lytic amount of concentrated sulphuric acid in reflux-
bacterial activity. The squares of inhibition zone ing dioxane (Scheme 1). The structures of the syn-
diameters were plotted against log concentrations of thesized compounds have been elucidated on the basis
1
the tested compounds. Extrapolation of the resulting of IR, H-NMR, mass spectral studies, and elemental
straight line to intersect with the log concentration analyses.
scale in the curve corresponded to log MIC, and MIC
was obtained as the antilog (Table IV).
IR data for compounds 3a-i revealed bands at 1344
cm−1 (C=S stretching) and the absence of an absorp-
tion in the region of ~2600-2550 cm−1 cited for an SH
group, proving that these compounds were in the
thionic form. In 1H-NMR spectra of compounds 3a-i in
DMSO-d6, the N=CH proton appeared at 10.75-15.40
RESULTS AND DISCUSSION
Chemistry
Recently, we have reviewed the chemistry of the 4- ppm as a singlet. Furthermore, the molecular ion peak
amino-1,2,4-triazole-3-thiones (Raafat and Ali, 2006). & base peak of the mass spectra of compounds 3a-i
The amino and mercapto groups are ready-made nu- were found to be in good agreement with the assigned
cleophilic centers for the synthesis of condensed hetero- structures.
cyclic rings. Therefore, these compounds can be con-
Compounds 4a-i were synthesized in a one pot multi-
sidered as useful tools in fusing to triazolothiadiazo- component Mannich reaction involving 3a-i, formal-
les or triazolothiadiazines. During this study, other dehyde, and morpholine (Scheme 1). Assignment of
groups reported the synthesis and characterization of the structures 4a-i was based on their elemental
Schiff base hydrazones bearing a 3-(4-pyridyl)-5-mer- analyses and spectroscopic data. The IR spectra of
capto-1,2,4-triazoles moiety (Khanmohammadi et al., compounds 4a-i showed an absorption band at 1587-
2008).
1697 cm−1, indicating the presence of –C=N, while
Scheme 1. Synthesis of compounds 3a-i and 4a-i

1246
M. A. Hussein et al.
bands observed at 1249-1375 cm−1 could be attributed protons.
to the –C=S functional group. The 1H-NMR spectrum
of compound 4a showed the -NCH2N- protons reson-
ated as a singlet at 5.70 integrating for two protons.
The -CH2-O-CH2- protons of the morpholine residue 8a-f were evaluated for their in vivo anti-inflamma-
appeared as a triplet at 4.10, while the -CH2-N-CH2- tory effect by the carrageenan induced paw edema
protons of the morpholine residue resonated as a bioassay in rats using indomethacin as a reference
triplet at 3.30 in addition to the aromatic protons at drug (Nargund et al., 1994), and the results were pre-
8.10-9.50, and a singlet at 5.9 ppm (N=CH-). In MS sented as the time course and percentage of edema in-
Anti-inflammatory activity
The synthesized compounds 1, 3a-i, 4a-i, 6a-d, and
δ
δ
δ
δ
δ
data of compound 4a, a peak observed at m/z 100 was hibition at a dose of 0.03 mmol/kg and time intervals
explained due to the formation of a morpholinomethyl of 0.5, 1, 2, 3, 4, and 5 h after injection (Table I).
radical cation, thus confirming the formation of Mannich
bases 4a-i. The triazolothiadiazole derivatives 6a-d activity of compounds
have been synthesized by the condensation of com- activity at the 1 h post-injection time point, and the
pound with the appropriate acids in the presence activity declined with time, indicating short acting
of phosphorus oxychloride (Scheme 2). Structures of compounds. Compounds and 3a-i exhibited ~50-111%
6a-d were confirmed by 1H-NMR, IR, MS spectral data, edema inhibition compared to indomethacin, and
and elemental analyses. IR spectra of 6a-d are devoid compounds 3e 3g, and 3i were the most active among
of absorption bands due to –SH (-C=S) and –NH2 this series.
stretching frequencies of parent compound clearly On the other hand, compounds 4a
The results showed that the anti-inflammatory
1
and 3a-i revealed maximum
1
5
1
,
1
, 4d, 4e, 4f, 4h, and
indicated the formation of the target compounds. The 4i showed maximum activity at 1 h post-injection,
1H-NMR spectrum of compound 6a, for example, with ~50-130% edema inhibition compared to indo-
showed a singlet at
the triazolothiadiazole ring. On the other hand, carrying Cl, Br, and NMe2 moieties, respectively re-
treatment of with 4-substituted phenacyl bromide in vealed maximum activity at 4 h post-injection and
δ 2.90 ppm (3H) of the (C6-CH3) of methacin. Furthermore, compounds 4b, 4c, and 4g
1
absolute ethanol containing potassium carbonate showed ~84-100% of the inhibition exhibited by in-
resulted in cyclocondensation, giving the correspond- domethacin; a result that could be attributed to the
ing triazolothiadiazines 8a-f (Scheme 2). Structures of structural similarities to that moieties in indomethacin.
8a-f were established on the basis of elemental Compounds 6a-d having CH3, C6H5, 4-pyridyl, and 3-
analyses and spectral data. IR data of 8a-f showed no pyridyl, respectively showed anti-inflammatory activity
absorption bands assignable to NH2 and NH groups. of ~20-89% that of indomethacin at 5 h post-injection.
1H-NMR spectra of 8a-f exhibited a singlet signal for In addition, results of compounds 8a-f (R = H, NO2,
SCH2 at
δ 4.8-4.6 ppm, in addition to aryl and pyridyl Cl, Br, OMe, and F) showed maximum activities at
Scheme 2. Synthesis of compounds 6a-d and 8a-f

Biological Activities of New Triazole Derivatives
1247
Table I. Inhibitory effect of compounds
1,
3a-i
,
4a-i
,
6a-d
,
8a-f, and indomethacin on carrageenan induced paw edema
in rats at 0.03 mmol/kg
% edema inhibition
2 h
Compound
No.
0 h
0.5 h
1 h
3 h
4 h
5 h
Control
−
−
−
−
−
−
−
Indomethacin
2.27
2.27
0.25
3.02
3.25
2.27
5.54
4.79
2.77
5.29
2.77
0.00
4.27
5.29
2.27
0.50
2.27
5.29
2.27
2.27
0.76
3.02
4.03
4.79
3.53
0.25
4.79
3.02
2.27
0.76
41.65
18.12
6.82
52.55
26.00
48.90
47.90
48.80
48.91
58.51
47.23
58.30
37.02
58.30
42.55
44.91
47.66
68.51
42.55
26.80
63.62
48.00
68.51
64.04
63.83
53.00
47.23
68.51
21.06
57.02
58.51
52.55
64.04
63.73
16.56
40.21
47.60
48.00
48.00
50.88
40.00
53.67
43.19
53.67
37.74
46.69
43.19
63.73
37.74
16.56
69.39
36.17
63.73
69.81
64.36
48.43
53.25
53.25
27.46
58.10
65.41
63.73
69.81
67.16
16.21
32.70
37.89
38.11
42.53
22.11
26.74
48.21
37.68
48.21
37.47
47.79
47.68
63.58
37.16
16.21
69.26
36.84
63.58
74.95
64.21
64.00
37.26
53.05
27.16
63.58
69.47
67.16
74.95
69.04
16.74
37.66
22.59
28.30
32.43
22.59
27.20
17.15
22.38
17.15
27.82
58.58
58.08
63.81
27.82
16.74
69.46
21.00
63.81
75.10
69.66
58.58
27.20
63.81
27.62
63.81
75.00
69.04
75.10
84.85
17.43
27.87
22.20
28.26
22.61
23.24
17.43
17.84
23.02
17.84
38.80
48.55
48.59
58.92
28.42
17.43
64.52
21.58
58.92
75.31
75.10
48.96
17.43
64.11
27.87
68.51
75.00
84.85
75.31
1
3a
3b
3c
3d
3e
3f
3g
3h
3i
4a
4b
4c
4d
4e
4f
4g
4h
4i
6a
6b
6c
6d
8a
8b
8c
8d
8e
8f
35.71
36.71
28.71
48.24
35.76
42.12
24.47
42.12
42.35
47.65
48.00
41.65
42.35
18.12
52.71
29.41
41.65
53.89
53.00
53.41
35.76
52.48
25.65
38.12
35.29
41.65
53.89
the 5 h interval, giving ~32-100% of the inhibition Cl, Br, OMe, and F), indicated that 8a and 8d
shown by indomethacin, while compounds 8d 8e, and hibited ~76-145% activity compared to indomethacin
8f were the most active molecules with a long duration as a reference drug at the 5 h interval. Moreover, com-
-f ex-
,
of action.
pounds 8c-e were the most active compounds when
they contained -Cl, -Br, or -OMe moieties, respec-
p
p
p
tively in their structures.
Analgesic activity
The potent long acting compounds with regard to
their anti-inflammatory activity (8a-f) were tested for
their analgesic activities using mean licking time in
Ulcerogenic effect
The occurrence of gastrointestinal (GI) damage
the hot-plate test, and indomethacin as a reference (bleeding and/or ulceration) is probably one of the
drug (Boyle et al., 1986). The effect was compared at most prevalent and serious side effects associated
the 0.02 mmol/kg dose level with a control group with the use of nonsteroidal anti-inflammatory drugs
injected with the vehicle, and results were cited in (NSAIDs) such as indomethacin (Nargund et al.,
Table II.
Results for the analgesic activity of 8a
1994). Observation of the gastrointestinal mucosa for
(R2=H, NO2, the presence of lesions following oral administration of
-f

1248
M. A. Hussein et al.
Table II. Analgesic activity of compounds (8a-f) and indomethacin at 0.02 mmol/kg
The average reaction time (sec) at different time intervals after compound administration
Compound
No.
5 h
3 h
2 h
1 h
1/2 h
Control
27.0 ± 0.16
28.0 ± 1.29
29.0 ± 1.0
28.1 ± 0.74
28.6 ± 0.74
Indomethacin
29.2 ± 0.47**
22.2 ± 0.90**
22.2 ± 0.90**
24.2 ± 1.48**
38.1 ± 0.70**
42. 5± 0.40*
23.2 ± 0.48**
25.8 ± 1.10**
21.4 ± 1.90**
26.6 ± 1.90**
28.21 ± 1.4**
32.3 ± 1.70**
34.9 ± 0.90**
22.2 ± 1.40**
20.7 ± 2.20**
18.6 ± 2.20**
21.6 ± 2.20**
24.1 ± 1.10**
26.3 ± 1.70**
34.5 ± 0.90**
19.1 ± 1.10**
14.6 ± 0.75*
13.8 ± 1.30**
17.8 ± 1.30**
18.5 ± 0.70**
21.0 ± 1.30**
22.2 ± 0.10**
14.5 ± 0.70**
10.6 ± 0.75**
29.8 ± 0.18**
11.8 ± 0.80**
12.3 ± 0.80**
14.3 ± 0.90**
15.2 ± 1.80**
10.2 ± 0.8**
8a
8b
8c
8d
8e
8f
Values are the mean ± S.E. of 5 observations
*Significant difference at < 0.05 vs control value (Student’s-test)
**Significant difference at
< 0.01 vs control value (Student's-test)
Note: In this Table, a space on both sides of the ± sign in the data was inserted.
p
p
graded doses (10, 30, and 50 mg/kg) of the test com-
However, compound 8d was found to cause 33%,
pounds as well as the reference drug was conducted to 67%, and 100% ulceration at doses of 10, 30 and 50
examine for ulcerogenic effects in rats (Boyle et al., mg/kg, respectively when compared to indomethacin
1986). Both the frequency of ulceration (expressed as which showed 67% and 100% ulceration at doses of 10
a ratio of ulcerated animals) and the severity of ul- and 30 mg/kg, respectively.
ceration (expressed as an ulcer index) were used for
comparison of the tested compounds and indomethacin. contained a triazolothiadiazine (
Compounds 8d 8e, and 8f, that exhibited potent was the most active derivative with regards to the
It should be mentioned that compound 8e, which
p-Cl) in its structure,
,
analgesic and anti-inflammatory profiles in the pre- anti-inflammatory and analgesic activities, and dis-
mentioned animal models, were evaluated for their played the highest safety due its structural similarity
ulcerogenic effects in rats. Results are recorded in to indomethacin.
Table III.
The tested compounds 8e and 8f showed superior
Acute toxicity (LD50)
GI safety profiles, because they provided 100% protec-
The median lethal dose (LD50) of compounds 8a and
tion in the population of the tested animals at an oral 8e was also determined in mice according to a re-
dose of 10 mg/kg. At doses of 30 and 50 mg/kg, they ported method (Boyle et al., 1986), and found to be
elicited 83% and 50% protection, respectively.
275 and 300 mg/kg (i.p.) respectively, whereas that of
indomethacin was 13 mg/kg (i.p.) (O'Neil et al., 2001).
Table III. Ulcerogenic effects of compounds 8d, 8e, and 8f
in comparison to indomethacin
Antibacterial activity
Compounds
1, 3a-h, 4a-h, 6a-d, and 8a-f were
Ratio of
ulcerated
animals
Compound.
No.
Dose
mg/kg
Ulcer index
(mean ± S.E.)
tested for their antibacterial activity in vitro against
MRSA, Bacillus cereus as representatives of the Gram
positive strains, and Escherichia coli and Klebsiella
pneumoniae as representatives of the Gram negative
strains (clinical isolates obtained from Infection
Control Unit, Assiut University Hospital, Faculty of
Medicine, Assiut University) using the agar cup
diffusion method (William, 1977) for susceptibility
screening, and the 2-fold dilution method for MIC
determination. Ampicillin was used as a reference
drug (20 µmol/mL), and DMSO was used as a solvent
control.
10
30
50
4/6
6/6
Not tested
1.75 ± 0.23
2.95 ± 0.16
−
Indomethacin
10
30
50
2/6
4/6
6/6
1.10 ± 0.35
1.35 ± 0.58
4.00 ± 0.35
8d
8e
8f
10
30
50
0/6
1/6
3/6
0.00
0.95 ± 0.25
1.75 ± 0.17
10
30
50
0/6
1/6
3/6
0.00
1.15 ± 0.25
1.35 ± 0.34
Results from the tested compounds
, and 8a-f, showed higher antibacterial activity than
ampicillin against the organisms used, and especially
1, 3a-i, 4a-i, 6a-
d

Biological Activities of New Triazole Derivatives
1249
Table IV. Antibacterial activity of the test compounds 1,
most active against the tested organisms.
3a-i
,
4a-i
,
6a-d
In vitro activity-inhibition zone in mm (MIC)
Staphylococcus Bacillus Escherichia Klebsiella
, 8a-f, and ampicillin (20 µmol/mL)
ACKNOWLEDGEMENTS
Com-
pound
The authors would like to express their gratitude
to Prof. Dr. Mahran Shaker Abdelrahman, Dept of
Pharmacology and to Assist. Lecturer Helal F. Hetta,
Dept. of Microbiology, Faculty of Medicine, Assiut
University for carrying out the pharmacological and
the antibacterial screening, respectively.
aureus
cereus
coli
pneumoniae
1
19
23 (65)
20
31 (20)
−
18 (70)
12
18
28 (25)
10
21 (60)
13
20
18 (79)
12
18 (79)
11
3a
3b
3c
3d
3e
3f
−
29 (23)
−
31 (20)
16 (135)
11
23 (65)
11
16 (130)
13
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3g
3h
3i
4a
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4c
4d
4e
4f
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−
−
10
−
−
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