European Journal of Medicinal Chemistry 63 (2013) 854e868

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European Journal of Medicinal Chemistry

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Mini-review

Triazolothiadiazoles and triazolothiadiazines e Biologically attractive

scaffolds
Imtiaz Khan a, *, Aliya Ibrar a, Naeem Abbas b
a
Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan
b
Department of Chemistry, University of Gujrat, Hafiz Hayat Campus, Gujrat 50700, Pakistan

a r t i c l e i n f o a b s t r a c t

Article history: Contemporary medicinal chemistry faces diverse challenges from several directions, including the need
Received 18 October 2012 for both potency and specificity of any therapeutic agent. Therefore, in the present perspective, the
Received in revised form triazolothiadiazoles and triazolothiadiazines with broad spectrum biological profile have matured into
6 January 2013
indispensable heterocyclic scaffolds. This review article is an effort to summarize medicinal chemistry
Accepted 8 January 2013
Available online 5 March 2013
investigations over the last decade, in search for new N-bridged heterocycles which can be a rich source
of promising biological activities.
Ó 2013 Elsevier Masson SAS. All rights reserved.
Keywords:
Heterocycles
Inhibitors
Synthesis
Biological activities

1. Introduction antiviral, diuretics, CNS-stimulant, PDE4 inhibitors and hypogly-

cemic agents [5e14]. New analogs of triazolothiadiazoles and tri-
The design, synthesis and production of molecules having value azolothiadiazines have been synthesized by the modifications of
as human therapeutic agents remain one of the main objectives of the parent skeletons by the introduction of new substituents on 3
organic and medicinal chemistry. During the past decade, combi- and 6 positions. This review summarizes the developments in this
natorial chemistry has provided access to chemical libraries based area to date with a focus on how the biological utility scope of these
on privileged structures [1], with heterocyclic motifs receiving conjugated heterocycles has been expanded with the appropriate
special attention as they belong to a class of compounds with inclusion of various substituents on the basic frameworks.
proven utility in medicinal chemistry [2].
Synthesis of nitrogen containing heterocyclic systems has been
2. Chemistry
attracting increasing interest over the past decade because of their
utility in various applications. 1,2,4-Triazoles coupled to another
2.1. Synthetic strategies toward triazolothiadiazoles and
heterocyclic ring exhibited wide spread applications as antibacte-
triazolothiadiazines
rial, antiviral, antihypertensive, antidepressant, anti-inflammatory,
anticonvulsant and antitumorial agents, pesticides, herbicides, lu-
Several protocols for the synthesis of triazolothiadiazoles and
bricants, dyes and analytical reagents [3,4]. Among these, the most
triazolothiadiazines are available in the literature but 4-amino-
common systems are triazoles combined to thiadiazoles or thia-
1,2,4-triazol-3-thiones are considered as useful tools in combining
diazines, incorporated into a wide variety of therapeutically
to triazolothiadiazoles and triazolothiadiazines. This can be attrib-
important compounds possessing a broad spectrum of biological
uted to the amino and mercapto groups which are ready-made
activities such as antiviral, antifungal, antihelmintic, antitumor,
nucleophilic centers for the synthesis of condensed heterocyclic
antibacterial, anti-inflammatory, antitubercular, analgesic,
rings [15,16] as demonstrated by Demirbas and co-workers [17]
(Scheme 1).
The most widely used approach for the synthesis of tri-
azolothiadiazines is illustrated in Scheme 2. 1,2,4-Triazole is
* Corresponding author. Tel.: þ92 51 9064 2216; fax: þ92 51 9064 2241. refluxed with a-haloacetophenone to afford the title compound as
E-mail address: [email protected] (I. Khan). described by Turan-Zitouni et al. [18].

0223-5234/$ e see front matter Ó 2013 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2013.01.060
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 855

Raval and co-workers [23] achieved the synthesis of coupled

nucleus by the condensation of 4-amino triazole with aromatic
aldehydes in the presence of catalytic amount of p-TsOH in DMF in
borosil beaker under microwave irradiation as well as by conven-
tional method (Scheme 7).
Recently, Badr and Barwa [24] accomplished the synthesis of
triazolothiadiazoles by using 4-amino-1,2,4-triazole as a key in-
termediate, obtained by the coupling of aromatic acid with thio-
carbohydrazide as sketched in Scheme 8.
Fascinated by the ongoing pioneering work in heterocyclic
chemistry, Devi et al. [25] synthesized various triazolothiadiazoles
and triazolothiadiazines encompassing 3-nitronaphtho[2,1-b]furan
(Scheme 9).
Almajan et al. [26] disclosed the synthesis of triazolothiadiazine
analogs by intermolecular condensation of 2-chloro-N-phenyl-
acetamide, 2-chloroacetic acid, oxalylchloride and bromo-
Scheme 1. Synthetic pathway of triazolothiadiazoles and triazolothiadiazines.
diethylmalonate with 4-amino-1,2,4-triazole-3-thiol as presented
in Scheme 10.
Farghaly et al. [19] reported the synthesis of triazolothiadiazoles Recently, intramolecular base catalyzed CeC bond formation
and triazolothiadiazines by using oxadiazole thione as the key leading to exclusive stereoselective syntheses of trans 6,7-dihydro-
starting material and by the interaction of the triazolethione with 5H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazines has been carried out by
chloro acetonitrile and sodium acetate fused in refluxing ethanol, Al-Etaibi et al. [27] as sketched in Scheme 11. The formation of the
respectively (Scheme 3). thiadiazine ring involves condensation through the amino group of
An efficient conventional heating and microwave assisted pro- the starting compound and the carbonyl group of acetophenone,
tocol for the synthesis of fused heterocycles has been developed by abstraction of the methylene proton, and a nucleophilic attack of
El-Ashry and co-workers [20] as sketched in Scheme 4. 3-Chloro-2- the resulting carbanion on the azomethine C atom with closure of
chlorocarbonylbenzo[b]thiophene yielded 4-amino-1,2,4-triazole the CeC bond of the thiadiazine ring. The cyclization is diaster-
which was used as a versatile intermediate for condensed hetero- eospecific, probably because of steric hindrances: during the attack
cycles. Better yields and shorter reaction times were achieved using of the carbanion on the plane of the C]N bond with the formation
MW. of the CeC bond, the bulky aryl fragments are trans with respect to
Taha [21] accomplished the synthesis of triazolothiadiazoles by the thiadiazine ring. During stereoselective reactions, one of the
using 4-amino-4H-3-methylthio{7H-1,2,4-triazolo[1,5-d]tetrazol- two possible stereoisomers is formed in high yield as compared to
6-yl}-1,2,4-triazole-5-thiol as a key intermediate. On the other other, so in this case trans-stereoisomer is formed in high yield that
hand, one-pot four component synthesis of triazolothiadiazines is converted into its cis-stereoisomer in polar solvents.
was done with two-carbon cyclizing reagents, namely chlor- Another improved synthesis of dihydroindeno- and indeno-
oacetone, pyruvic acid, benzoylformic acid and benzoin (Scheme 5). triazolothiadiazines have been introduced by the cyclo-
Microwave irradiation assisted synthesis of few novel tri- condensation of a-bromoindanones and a,a-dibromoindanones
azolothiadiazole and triazolothiadiazine derivatives has been with 4-amino-5-mercapto-1,2,4-triazoles by Prakash and co-
developed by Al-Soud et al. [22] as presented in Scheme 6. workers [28] as shown in Scheme 12.
The introduction of different substituents on the parent skeleton
of triazolothiadiazoles and triazolothiadiazines by the aforemen-
tioned synthetic methodologies alter biological activities. This re-
view article focuses on the biological potential of recently
synthesized (during the last decade) compounds differing in
chemical structures.

3. Biological applications of triazolothiadiazoles and

triazolothiadiazines

Nitrogen containing heterocycles particularly tri-

azolothiadiazoles and triazolothiadiazines are the central structural
motifs of various compounds with activities of relevance to
different areas such as pharmaceutical and agrochemicals. The
most common applications are highlighted here.

3.1. Anticancer activity

Holla and co-workers [3] designed the synthesis of a series of

novel 1,4-bis-[4-amino-5-mercapto-1,2,4-triazol-3-yl-methoxy]
phenylenes and their triazolothiadiazole derivatives and studied
their anticancer activity against the three cell lines like NCI-H 460
(lung), MCF 7 (breast) and SF 268 (CNS). Interestingly, 3,30 -(2,5-
dichloro-1,4-phenylene)bis(oxy)bis(methylene)di-[1,2,4]triazolo
[3,4-b][1,3,4]thiadiazole-6-thiol (1) was found to be the most active
Scheme 2. Synthesis of triazolothiadiazines. which was further tested against a panel of 60 cell lines derived
856 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868

Scheme 3. Synthesis of the title compounds.

from seven cancer types namely, lung, colon, melanoma, renal, to exhibit better anticancer activities than those of benzimidazoles
ovarian, CNS and leukemia. clubbed with triazolothiadiazine (6).
In another study, Holla et al. [29] reported several structurally
modified triazolothiadiazines and evaluated for their anticancer 3.2. Antitumor activity
activity against full panel of 60 cell lines derived from seven cancer
types. The screening results showed that compound (2) incorpo- A notable advance was achieved by Poojary and co-workers [30]
rating 2-chloroaryloxy methyl at C-3 and 4-chloro benzylidene at by screening a series of triazolothiadiazole derivatives for their
C-7 positions exhibited highest activity with GI50 value < 10 mM antitumor activity. They were first evaluated in a 3-cell line, one-
against all the tested cell lines. A sharp decrease in activity was dose preliminary anticancer assay followed by a full panel of 60
observed when the substituent at C-3 was changed to 4-chloro-3- human cell lines which showed variable antitumor activities.
methyl-aryloxy methyl. Compounds (7) and (8) were found to be highly active against
Subsequently, Poojary and co-workers [30] also reported several leukemia MOLT-4, ovarian cancer OVCAR-3 and prostate cancer PC-
triazolothiadiazines and screened for their anticancer activity. 3. This increase in activity may be attributed to the chloro groups
Three of the newly synthesized compounds were screened for their present on the phenyl ring at position 6 of the title compounds.
anticancer activities under NCI screening programme [31,32] using New 3,6-disubstituted [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole
3-cell line one dose assay. The 3-cell lines used for investigation are and triazolothiadiazine analogs have been explored by Al-Soud et
NCI-H 460 (Lung), MCF 7 (Breast) and SF 268 (CNS). The structuree al. [22] for in vitro antitumor activity against a panel of tumor cell
activity relationship studies indicated that the compounds con- lines, using microculture tetrazolium assay. The synthesized com-
taining chlorine atoms at various positions are more active as pounds (9) and (10) exhibited activity against CD4þ human acute T-
compared to compounds with other substituents and among them, lymphoblastic leukemia. The SAR analysis indicated that the syn-
6-(4-chlorophenyl)-3-(2,4-dichloro-5-fluorophenyl)-7H-[1,2,4]tri- thesis of new analogs of the triazolothiadiazine series may lead to
azolo[3,4-b][1,3,4]thiadiazine (3) was found to be the most potent. the discovery of more potent and selective inhibitors.
Fascinated by the previous studies, Isloor and co-workers [33] Following the developments in this area, Ibrahim [35] turned his
carried out the synthesis of 6-[3-(4-fluorophenyl)-1H-pyrazol-4-yl]- attention toward the synthesis of 4-(3-substituted[1,2,4]triazolo
3[(2-naphthyloxy)methyl][1,2,4]triazolo[3,4-b][1,3,4]thiadiazole. The [3,4-b][1,3,4]thiadiazol-6-yl) aniline, 2-amino-5-substituted-[1,2,4]
synthesized compound was evaluated for anticancer activity by using triazolo[3,4-b][1,3,4]thiadiazol-6-yl methyl-1,3-thiazol-4-ol and
MTT assay which revealed the FPNT (4) as a potent compound. This triazolo[3,4-b]thiadiazole derivatives to optimize the biological
activity may be due to the presence of fluorine on the aryl group. response of the new lead triazolothiadiazole derivatives. The syn-
Moreover, recently, in pioneering studies, Husain et al. [34] re- thesized compounds demonstrated inhibitory effects on the
ported a library of triazolothiadiazole and triazolothiadiazine de- growth of a wide range of cancer cell lines generally at 105e107 M
rivatives incorporating benzimidazole scaffold. These derivatives concentrations. Their antitumor activity appears to be related to
have been screened for their in vitro anticancer activity at the Na- some structural requirements and to the presence of particular
tional Cancer Institute (NCI) against full NCI 60 human cell lines substituents, as a matter of fact, 4-chlorophenoxymethylene moi-
panel. The compounds possessed good results and among them, ety plays an important role for the activity. Compounds (11) and
compound (5) turned out to be a lead candidate and may serve as a (12) maintained the highest growth inhibition [36].
novel template for development of potential and selective agents in Recently, Hu and co-workers [37] explored a series of C-3/C-3
the field of cancer chemotherapy. The screened results indicated bis-fluoroquinolone coupled heterocycles cross-linked with a
that benzimidazoles clubbed with triazolothiadiazole were found [1,2,4]-triazolo[3,4-b][1,3,4]-thiadiazole core against murine
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 857

Scheme 4. Conventional and microwave-assisted synthesis of the title compounds.

leukemia cell line (L1210), human leukocytoma cell line (HL60) and inflammatory activity as shown by compound (16). From SAR
Chinese hamster ovary cell line (CHO) using the MTT assay. Com- analysis, it is revealed that the introduction of electron releasing
pounds (13) and (14) exhibited the most potent activity against groups like methyl or methoxy at the 5th position of indole ring
HL60 cells. This preliminary indication of antitumor activity sug- produced a decrease in the anti-inflammatory activity.
gests that di-(1-cyclopropyl)-substituted fluoroquinolones and bis- Furthermore, triazolothiadiazole and triazolothiadiazine nuclei
fluoroquinolone hybrid molecules are promising lead compounds bearing trichlorophenyl moiety have been reported by Karegoudar
for further developments. and co-workers [41]. The prepared compounds were evaluated for
anti-inflammatory activity which was conducted in acute inflam-
3.3. Anti-inflammatory activity matory model. The compound (17) showed good activity among
the tested compounds as it structurally resembles a known anti-
Mathew et al. [38] accomplished a series of fused heterocycles for inflammatory agent Dichlofenac sodium [42]. 6-(4-Chlorophenyl)-
the evaluation of their biological profile. All the synthesized com- 3-(2,3,5-trichlorophenyl)-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadia-
pounds were tested for their anti-inflammatory activity using carra- zine (18) has been found to exhibit less activity. So, in preliminary
geenan induced rat hind paw edema method of Winter et al. [39]. examination, triazolothiadiazoles have found to be more potent
Tested compounds exhibited weak to moderate activity as depicted by anti-inflammatory agents as compared to thiadiazines.
3-[2-(N-methyl amino)phenyl]-6-[2-(2,4-dichlorophenoxy)ethyl]- Aytaç et al. [43] reported several triazolothiadiazine derivatives
1,2,4-triazolo[3,4-b]-1,3,4-thiadiazole (15). and screened for their anti-inflammation profile. Among the
In another study, Mathew and co-workers [40] investigated that condensed derivatives, compound (19) bearing 3,4,5-
triazolothiadiazoles possessing indole ring exhibited good anti- trimethoxyphenylethyl at the 3rd position and 4-fluorophenyl at

Scheme 5. Synthesis of triazolothiadiazoles and thiadiazines.

858 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868

position of triazolothiadiazole skeleton showed maximum anti-

inflammatory activity [47].
More recently, Malladi and co-workers [48] synthesized a series
of 3,6-disubstituted-1,2,4-triazolothiadiazoles bearing pyrazole
moiety and evaluated their anti-inflammation efficacy. The results
disclosed that 3-propyl-6-(3-(4-chlorophenyl)-1H-pyrazol-4-yl)
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (24) possess significant in-
hibition as compared to the standard drug Dichlofenac sodium.

3.4. Analgesic activity

Turan-Zitouni et al. [18] reported several triazolothiadiazine

derivatives which have been tolerated for their analgesic activity by
acetic acid test (Modified Koster Test) [49] in male albino mice (25e
30 g). The results clearly demonstrated that the synthesized com-
pounds exhibited an interesting profile of analgesic activity
particularly compound (25) as compared to aspirin used as a
standard. The presence of hydroxy and methoxy groups on the
phenyl ring at position 6 may be responsible for the promising
activity.
Scheme 6. MWI assisted synthesis of triazolothiadiazoles and triazolothiadiazines.
Mathew et al. [38] reported a series of 3,6-disubstituted-1,2,4-
triazolo[3,4-b][1,3,4]thiadiazoles. The synthesized compounds
have been screened for their analgesic activity by using Eddy’s hot
the 6th position of the fused ring possessed the most prominent plate technique. 3-[4-(N,N-Dimethyl amino)phenyl]-6-[2-(2,4-
and consistent activity. dichlorophenoxy)ethyl]-1,2,4-triazolo[3,4-b]-1,3,4-thiadiazole (26)
Husain and Naseer [44] synthesized a variety of 3,6- was found to be the most active among the tested series.
disubstituted-1,2,4-triazolo-[3,4-b]-1,3,4-thiadiazoles and in vivo A noteworthy contribution was made by the Karthikeyan and
anti-inflammatory activity was evaluated. The compounds exhibi- co-workers [50] who reported numerous triazolothiadiazole de-
ted good anti-inflammation profile, specifically, 3-(phenox- rivatives. The synthesized compounds were evaluated for analgesic
ymethyl)-6-(3-oxo-3-phenylpropyl)-[1,2,4]triazolo[3,4-b][1,3,4] activity by hot plate test by Eddy and Leimbach. The analgesic
thiadiazole (20) presented the highest anti-inflammatory activity screening results revealed that the compounds (27) and (28) were
better than the standard drug indomethacin. It was observed that found to be the most potent. The structureeactivity relationship
the replacement of 3-oxo-3-phenylpropyl group at C-6 position by investigations indicated that the presence of halo groups like chloro
other groups resulted a decrease in anti-inflammatory activity. and fluoro on both aryl rings at position 3 and 6 may be responsible
From the view point of structureeactivity relationship, it is clear for good results.
that the triazolothiadiazole derivatives of phenoxy acetic acid were Moreover, Mathew and co-workers [40,51] found that the tri-
found to be more active than phenyl acetic acid and salicylic acid azolothiadiazoles possessing indole ring at the 6th position
derivatives. exhibited good analgesic activity as shown by compound (29).
Moreover, El-Shehry et al. [45] prepared a series of tri- Aytaç et al. [43] reported a series of triazolothiadiazine de-
azolothiadiazoles and triazolothiadiazines and assessed their anti- rivatives and screened for their analgesic activity. Among the syn-
inflammatory activity. These condensed derivatives having 2,4- thesized compounds, 3-[2-(3,4,5-trimethoxyphenyl)ethyl]-6-(4-
dichlorophenoxy group at C-6 position like (21) possess highest chlorophenyl)-7H-1,2,4-triazolo[3,4-b]-1,3,4-thiadiazine (30)
activity compared with indomethacin by protecting rats by 36e56% showed prominent activity. The methoxy groups are contributing
from inflammation. On the other hand, 3-((2,4-dichlorophenoxy) significantly toward pronounced biological activity.
methyl)-6-phenyl-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (22) Husain and Naseer [44] prepared a series of triazolothiadiazole
exhibited less activity as compared to triazolothiadiazole series. derivatives which were screened for the analgesic effects using
Hussein et al. [46] reported a series of triazolothiadiazoles acetic acid-induced writhing method. The results of analgesic ac-
bearing pyridine moiety and screened for their in vitro anti- tivity indicated that compounds (20) incorporating 3-oxo-3-
inflammatory effect by the carrageenan induced paw edema phenylpropyl group at C-3 of triazolothiadiazole ring showed
bioassay in rats using indomethacin as a reference drug. The better activity than that of standard drug acetylsalicylic acid
compound (23) possessing pyridine moiety both at 3rd and 6th (aspirin).
Recently, experiments conducted by Devi et al. [25] for the
synthesis of triazolothiadiazoles and triazolothiadiazines encom-
passing 3-nitronaphtho[2,1-b]furan made notable contribution to
the library of heterocyclic compounds showing analgesic effects.
The activity of the synthesized compounds was compared with that
of standard drug tremadol. The results revealed that compound
(31) was found to be less active than that of (32).

3.5. Antitubercular activity

Interesting observations on the antitubercular activity of tri-

azolothiadiazole derivatives were made by Karthikeyan and co-
workers [50]. Tuberculosis Activity Antimicrobial Acquisition and
Scheme 7. Synthesis of triazolothiadiazoles by conventional and microwave methods. Coordinating Facility (TAACF) of Southern Research evaluated some
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 859

Scheme 8. Synthetic pathway of the title compounds.

of the selected compounds for in vitro antitubercular activity 3.6. Anti-HIV activity
against Mycobacterium tuberculosis HRv (ATCC 27294) in BACTEC
12B medium using a broth microdilution assay, the Microplate A series of 1,2,4-triazolo[3,4-b][1,3,4]thiadiazole analogs has
Alamar Blue Assay (MABA) [51]. The antitubercular screening re- been synthesized under microwave irradiation (MWI) by Al-Soud
sults revealed that 3,6-bis(2,4-dichloro-5-fluorophenyl)-1,2,4- et al. [22]. These compounds were tested for their in vitro anti-
triazolo[3,4-b]-1,3,4-thiadiazole (33) showed an excellent inhibi- HIV-1 (strain IIIB) and HIV-2 (strain ROD) activity in human T-
tion of 97%. The presence of chloro and fluoro substituents on aryl lymphocyte (MT-4) cells using the MT-4/MTT assay [53]. Efavirenz
ring may be responsible for inhibitory activity. [54] and capravirine [55] were used as standard drugs. Compound
Kumar et al. [52] reported a novel series of 3-(4- (36) possessing pyrazine moiety was found to be equipotent
isopropylthiazol-2-yl)-6-methyl-[1,2,4]triazolo[3,4-b][1,3,4]thia- against HIV-1 and HIV-2 replication in vitro as compared to (37)
diazoles and triazolothiadiazines and screened for antitubercular which incorporate an unsubstituted phenyl ring.
activity against H37Rv strain. The in vitro antituberculosis screening
results showed that all compounds were active, in particular, 3.7. Growth promoters
compound (34) exhibited excellent antitubercular activity when
compared with first line drug such as Isoniazid. 6-(2,4- Zhang et al. [56] reported a series of fused heterocycles con-
Difluorophenyl)-3-(4-isopropylthiazol-2-yl)-5,6-dihydro-[1,2,4]tri- taining furan ring to investigate the effect of these compounds on
azolo[3,4-b][1,3,4]thiadiazole (35) possess low activity as compared sprouting of mung bean seeds. The results obtained revealed that
to the thiadiazine (34). compounds (38) and (39) increase mung bean sprout growth by

Scheme 9. Protocol for synthesis of the title compounds.

860 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868

Scheme 10. Synthesis of triazolothiadiazine derivatives.

40%. This increase in growth may be attributed to the chloro and triazolothiadiazole derivatives have been found more active as
methoxy substituents present on the aryl ring at position 3 of tri- compared to the thiadiazine series (43).
azolothiadiazole nucleus.

3.10. Cytotoxic activity

3.8. Anticonvulsant activity
Sumangala et al. [58] reported a series of triazolothiadiazine
Husain et al. [57] disclosed the synthesis of 3,6-disubstituted- derivatives containing methylsulfonyl group and evaluated for their
1,2,4-triazolo-[3,4-b]1,3,4-thiadiazoles and screened for their anti- cytotoxicity. The cytotoxicity study revealed that some of the 6-
convulsant potential. The results indicated that halo-substituted substituted-3-[2-(methylsulfonyl)benzyl]-7H-[1,2,4]triazolo[3,4-b]
aryl (bromophenyl) at position 6 of the triazolothiadiazole motif [1,3,4]thiadiazines, (44) and (45) containing phenyl and biphenyl
was essential for the activity. Few compounds like (40) and (41) groups, respectively at C-6 position of thiadiazine nucleus showed
showed more lipophilic character and were more active. significant activity compared to the standard Doxorubicin. The
cytotoxicity of thiadiazine containing biphenyl derivative is much
3.9. Molluscicidal agents better than the phenyl derivative.
A novel combinatorial library of 6-arylsubstituted-3-[2-(4-
El-Shehry et al. [45] synthesized a series of 3-((2,4-dichloro- substitutedphenyl)propan-2-yl]-7H-[1,2,4]triazolo[3,4-b][1,3,4]
phenoxy)methyl)-1,2,4-triazolothiadiazoles and thiadiazines and thiadiazines were designed, synthesized and screened for their
screened for their toxicity toward Biomphalaria alexandrina snails. cytotoxic activities using trypan blue exclusion and MTT assay by
An insight inspection of the results showed that 6-((2,4- Puthiyapurayil et al. [59]. The results of the short term in vitro
dichlorophenoxy)methyl)-3-(4-methoxyphenyl)-[1,2,4]triazolo cytotoxic activity against Ehrlich Ascites Carcinoma (EAC) cell lines
[3,4-b][1,3,4]thiadiazole (42) has high effect on the snails. The by trypan blue exclusion method showed poor to moderate activity.

Scheme 11. Stereoselective synthesis of 6,7-dihydro-5H-[1,2,4]triazolo[3,4-b]1,3,4]

thiadiazines. Scheme 12. Synthesis of triazolothiadiazines.
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 861

In vitro cytotoxicity by MTT assay showed that, in EAC cells few eNH and 2,3-dichloro groups attached to phenyl ring at position 6 of
compounds were found to be active. The enhanced activity of the the thiadiazole nucleus as depicted by compounds (54) and (55).
compounds was mainly attributed to the presence of (2-
phenylpropan-2-yl) and [2-(4-chlorophenyl)propan-2-yl] substit-
uent at C-3 position of the triazolothiadiazine ring system. Com-
pound (46) carrying 4-trifluoromethoxyphenyl group at C-6
position and [2-(4-chlorophenyl)propan-2-yl] substituent at C-3
position was the most lead molecule.

3.11. Anthelmintic activity

3,3'-(2,5-Dichloro-1,4- (Z)-7-(4-Chlorobenzylidene)-3-((2-
phenylene)bis(oxy)bis(methylene)di- chlorophenoxy)methyl)-6-(2,4-dichloro-5-
Puthiyapurayil et al. [59] synthesized numerous triazolothiadiazine [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole-6-thiol fluorophenyl)-7H-[1,2,4]triazolo[3,4-
derivatives and screened for their anthelmintic activity. The results (1) b][1,3,4]thiadiazine (2)
revealed that compounds (47) and (48) showed good anthlemintic
activity almost comparable with standard drug. The excellent activity
of these compounds was attributed to the presence of 2,4-dichloro-5-
fluorophenyl and 4-chloro-3-nitrophenyl groups on the C-6 phenyl
ring and halogen groups (4-chloro and 4-bromo) on the C-3 position of
the condensed triazolothiadiazine ring system.

3.12. Antimicrobial agents

Holla et al. [4] reported several nitrophenylfurfurylidene-1,2,4- 6-(4-Chlorophenyl)-3-(2,4-dichloro-5- 6-[3-(4-Fluorophenyl)-1H-pyrazol-4-yl]-3[(2-

fluorophenyl)-7H-[1,2,4]triazolo[3,4- naphthyloxy)methyl][1,2,4]triazolo[3,4-
triazolo[3,4-b]-1,3,4-thiadiazines and tested for antibacterial ac- b][1,3,4]thiadiazine (3) b][1,3,4]thiadiazole (4)
tivity against Escherichia coli, Staphylococcus aureus, Pseudomonas
aeruginosa and Bacillus subtilis. Among the tested compounds, (49)
carrying p-nitrophenyl and a methyl group showed excellent
antibacterial activities against all the bacterial strains.
Demirbas and co-workers [17] described a series of new [1,2,4]
triazolo[3,4-b][1,3,4]thiadiazoles and triazolothiadiazines. The
possible antimicrobial activities of the synthesized compounds
were investigated to ten standard organisms including bacterial
and fungal strains. All newly synthesized compounds exhibited
promising activities against Enterococcus faecalis (Ef), S. aureus (Sa) 1-(1H-Benzo[d]imidazol-2-yl)-3-(6-(2,4- 1-(1H-Benzo[d]imidazol-2-yl)-3-(6-chloro-7H-
dichlorophenyl)-[1,2,4]triazolo[3,4- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-3-
and B. subtilis (Bs). The highest activity was observed for 4-amino- b][1,3,4]thiadiazol-3-yl)propan-1-one (5) yl)propan-1-one (6)
3-methyl-1-((6-methyl-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-3-yl)
methyl)-1H-1,2,4-triazol-5(4H)-one (50).
Holla et al. [29] reported a series of fluorine containing 7-
arylidene-6-(2,4-dichlorophenyl)-3-aryloxymethyl/anilinomethyl-
1,2,4-triazolo[3,4-b]-1,3,4-thiadiazines and screened for their anti-
bacterial activity against E. coli, S. aureus, P. aeruginosa and B. subtilis
bacterial strains by serial plate dilution method. Almost all the
screened compounds exhibited comparable activity with standard
nitrofurazone. Z-3-((4-Chloroaryloxy)methyl)-6-(2,4-dichloro-5- 6-(4-Chlorobenzyloxy)-3-(4-fluoro-3- 6-(2,4-Dichlorobenzyloxy)-3-(4-fluoro-3-
phenoxyphenyl)-[1,2,4]triazolo[3,4- phenoxyphenyl)-[1,2,4]triazolo[3,4-
fluorophenyl)-7-(3,4-dimethoxybenzylidene)-7H-[1,2,4]triazolo b][1,3,4]thiadiazole (7) b][1,3,4]thiadiazole (8)
[3,4-b][1,3,4]thiadiazine (51) showed highest activity against all
the tested bacteria.
In another study, Holla and co-workers [60] reported a series of
triazolothiadiazoles and triazolothiadiazines containing 6-
chloropyridin-3-yl moiety and initially screened for their in vitro
antibacterial activities against different bacterial strains by serial
plate dilution method. Furacin was used as a standard drug. The
synthesized derivatives exhibited mild activities as represented by
compounds (52) and (53). 4-(3-(3,4-Dichlorophenyl)-[1,2,4]triazolo[3,4- 3-(3,4-Dichlorophenyl)-5H-[1,2,4]triazolo[3,4-
1,2,4-Triazolo[3,4-b]-1,3,4-thiadiazoles and 1,2,4-triazolo[3,4-b]- b][1,3,4]thiadiazol-6-yl)aniline (9) b][1,3,4]thiadiazin-6(7H)-one (10)

1,3,4-thiadiazines carrying thioalkyl and sulfonyl phenoxy moieties

were reported by Karabasanagouda et al. [61]. The newly synthesized
compounds were screened for their antibacterial activity against E.
coli (ATTC-25922), S. aureus (ATTC-25923), P. aeruginosa (ATCC-
27853) and Klebsiella pneumoniae (recultured) bacterial strains by
serial plate dilution method. The investigation of antibacterial and
antifungal screening data revealed that all the tested compounds
showed moderate to good inhibition and they deserve more 4-(3-(4-Chlorobenzyloxy)-[1,2,4]triazolo[3,4- 2-Amino-5-((3-(4-chlorobenzyloxy)-
consideration as potential antimicrobials. The good activity is b][1,3,4]thiadiazol-6-yl)aniline (11) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-6-
attributed to the presence of pharmacologically active eCH3, eOCH3, yl)methyl)-thiazol-4-ol (12)
862 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868

1-Cyclopropyl-3-(3-(1-cyclopropyl-6-fluoro-4-oxo-7-(piperazin-1-yl)-1,4-dihydroquinolin-3-yl)-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-6-yl)-6-fluoro-7-(4-methylpiperazin-1-yl)quinolin-4(1H)-
one (13)
3,6-Di(pyridin-4-yl)-[1,2,4]triazolo[3,4- 3-Propyl-6-(3-(4-chlorophenyl)-1H-pyrazol-4-
b][1,3,4]thiadiazole (23) yl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (24)

6-(3-(1-Cyclopropyl-6-fluoro-7-(4-methylpiperazin-1-yl)-4-oxo-1,4-dihydroquinolin-3-yl)-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-6-yl)-9-fluoro-3-methyl-10-(4-methylpiperazin-1-yl)-2H-
[1,4]oxazino[2,3,4-ij]quinolin-7(3H)-one (14)

5-Methoxy-2-(3-((5,6,7,8- 3-[4-(N,N-Dimethyl amino)phenyl]-6-[2-(2,4-

tetrahydronaphthalen-2-yloxy)methyl)-7H- dichlorophenoxy)ethyl]-1,2,4-triazolo[3,4-b]-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-6- 1,3,4-thiadiazole (26)
yl)phenol (25)

A series of dichlorofluorophenyl containing triazolothiadiazoles

were reported by Karthikeyan and co-workers [50] and tested their
antimicrobial efficacy using disc diffusion method. The antibacterial
3-[2-(N-Methyl amino)phenyl]-6-[2-(2,4- 3-(2-Chloro-5-methoxyphenyl)-6-((5-methoxy- screening data revealed that few compounds were active against
dichlorophenoxy)ethyl]-1,2,4-triazolo[3,4-b]- 1H-indol-3-yl)methyl)-[1,2,4]triazolo[3,4-
1,3,4-thiadiazole (15) b][1,3,4]thiadiazole (16) S. aureus and E. coli especially 3-(2,4-dichloro-5-fluorophenyl)-6-(4-
chlorophenyl)-1,2,4-triazolo[3,4-b]-1,3,4-thiadiazole (56) exhibited
good antibacterial activity against all tested bacterial strains almost
equivalent to that of the standard drug Ciprofloxacin. The newly
prepared compounds were also evaluated for their antifungal ac-
tivity against Aspergillus niger, Candida albicans, Aspergillus fumiga-
tus, Penicillium marneffei and Trichophyton mentagrophytes in DMSO
by agar diffusion method. The antifungal screening data showed that
several compounds exhibited good activity against C. albicans and
A. fumigatus especially compounds (56) exhibited good antifungal
activity against all tested fungal strains almost equivalent to that of
6-(4-Methoxyphenyl)-3-(2,3,5- 6-(4-Chlorophenyl)-3-(2,3,5-trichlorophenyl)-
trichlorophenyl)-[1,2,4]triazolo[3,4- 7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (18) the standard drug Griseofulvin. This enhanced activity may be
b][1,3,4]thiadiazole (17) attributed to the presence of chloro and fluoro groups on both the
aryl rings at 3rd and 6th position of condensed nucleus.
The research group of Karegoudar [41] reported a series of new
triazolothiadiazole and triazolothiadiazine derivatives and screened
for their antibacterial activity against E. coli, S. aureus, P. aeruginosa
and K. pneumoniae bacterial strains by serial plate dilution method.
The newly prepared compounds were also screened for their anti-
fungal activity against Aspergillus flavus, A. fumigatus, P. marneffei
and T. mentagrophytes in DMSO by serial plate dilution method. The
investigation of antimicrobial screening data revealed that all the
3-[2-(3,4,5-Trimethoxyphenyl)ethyl]-6-(4- 3-(Phenoxymethyl)-6-(3-oxo-3-phenylpropyl)- tested compounds showed moderate to good inhibition. The good
fluorophenyl)-7H-1,2,4-triazolo[3,4-b]-1,3,4- [1,2,4]-triazolo[3,4-b][1,3,4]-thiadiazole (20)
thiadiazine (19)
activity is attributed to the presence of pharmacologically active e
OCH3, 2,3-dichloro, 4-hydoxy-3-amide, 4-chloro, eSCH3 groups
attached to phenyl ring, pyridyl, bromopyridyl and aryloxy groups of
the thiadiazole as shown by compounds (57) and (58).
Furthermore, in a continuation of their previous studies, Prasad and
co-workers [62] reported some new triazolothiadiazoles bearing 4-
methylthiobenzyl moiety and evaluated for antimicrobial activities.
The investigation of the antibacterial and antifungal screening studies
revealed that all the tested compounds showed moderate to good
inhibition in DMSO. The good activity can be attributed to the presence
of pharmacologically active groups like 2,3,4-trichloro, 4-methylthio,
3,6-Bis((2,4-dichlorophenoxy)methyl)- 3-((2,4-Dichlorophenoxy)methyl)-6-phenyl- 2,4-dichloro-5-fluoro, which are directly attached to the phenyl ring of
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (21) 7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (22) the triazole system, and the groups like 4-nitro, 4-fluoro, 2-nitro-4-
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 863

chloro, 2-trifluoromethyl, 4-chloro, 4-methyl which are directly zone of inhibition >25.0 mm as compared with standard drug
attached to the arylfuryl ring as shown by 3-(4-(methylthio)benzyl)-6- ciprofloxacin which showed the zone of inhibition of 26.6 mm
(2,3,4-trichlorophenyl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (59). against S. aureus. It should be noted that out of these compounds, 3-
Mathew et al. [51] reported a new series of conjugated heterocy- propyl-10,10a-dihydroindeno[1,2-e][1,2,4]triazolo[3,4-b][1,3,4]
cles and a filter-paper disc-method was employed for the in vitro study thiadiazine (64) was found to be most potent member showing
of antibacterial and antifungal effects against E. coli, P. aeruginosa, zone of inhibition even greater than the standard drug.
B. subtilis, S. aureus, A. niger and C. albicans, respectively. The screening All the compounds were also tested for their in vitro antifungal
results evidenced that the electron-withdrawing group substituted on activity against three fungal strains, namely, A. niger, A. flavus and
hetero-aromatic ring at the 6th position of triazolothiadiazole system Penicillium species through poisoned food method. Standard drug
enhanced the antimicrobial action of the tested compounds. Pyridine fluconazole was used for comparison with antifungal activity. From
derivatives with 2-fluoro substitution (60) showed maximum inhib- the careful comparison of the results, it has been revealed that
itory action against all the tested bacterial strains. mainly the 9-methylindeno[1,2-e][1,2,4]triazolo[3,4-b][1,3,4]thia-
(4-X-Phenylsulfonyl)phenyl containing 6-amino-[1,2,4]triazolo diazine (65) showed excellent antifungal activity with >65% inhi-
[3,4-b][1,3,4]thiadiazines and [1,2,4]triazolo[3,4-b][1,3,4]thiadiazin- bition of mycelial growth against all the fungal strains.
6-ones were synthesized by Almajan et al. [26]. The synthesized
compounds were tested for their in vitro antibacterial activity against
the Gram-positive (E. faecalis ATCC 29212; S. aureus ATCC 25923;
Staphylococcus epidermidis ATCC 14990; Bacillus cereus ATCC 13061)
and Gram-negative (Acinetobacter baumanii ATCC 19606; Citrobacter
freundii ATCC 27853; Enterobacter clocae ATCC 49141; E. coli ATCC
25922; P. aeruginosa ATCC 9027) bacteria by using the broth dilution 6-(3-Chloro-4-fluorophenyl)-3-(2,4-dichloro-5- 3-(2,4-Dichloro-5-fluorophenyl)-6-[(4-
fluorophenyl)1,2,4-triazolo[3,4-b]-1,3,4- fluorophenoxy)methyl]1,2,4-triazolo[3,4-b]-
method for determination of MIC. Tetracycline and ampicillin were thiadiazole (27) 1,3,4-thiadiazole (28)
used as control drugs. The data generated from this study showed
that compounds displayed low to moderate activity. The obtained
results can be attributed to quite bulky structure of the tested com-
pounds but they may be associated with the nature of tested bacterial
species. The most active compound with triazolothiadiazine nucleus
of this series is 3-[4-(phenylsulfonyl)phenyl]-5H-[1,2,4]triazolo[3,4-
b][1,3,4]thiadiazin-6(7H)-one (61) against S. epidermidis.
In a similar study, Kumar et al. [52] synthesized a novel series of
clubbed isopropylthiazole derived triazolothiadiazoles and tri-
azolothiadiazines and tested for antimicrobial activities. The anti- 6-((1H-Indol-2-yl)methyl)-3-(3,4- 3-[2-(3,4,5-Trimethoxyphenyl)ethyl]-6-(4-
dimethoxybenzyl)-[1,2,4]triazolo[3,4- chlorophenyl)-7H-1,2,4-triazolo[3,4-b]-1,3,4-
bacterial activity of the synthesized compounds was performed by b][1,3,4]thiadiazole (29) thiadiazine (30)
broth dilution method [20,21] against the following standard bac-
terial strains; S. aureus (ATCC 11632), Streptococcus faecalis (ATCC
14506), B. subtilis (ATCC 60511), K. pneumoniae (ATCC 10031), E. coli
and P. aeruginosa (ATCC 10145) and antifungal activity against
Yeasts: Saccharomyces cerevisiae (ATCC 9763, Sc) and Candida tro-
picalis (ATCC 1369, CT), mould: A. niger (ATCC 6275). The prepared
compounds exhibited interesting trends in structureeactivity rela-
tionship (SAR) studies. The linkage between eNH2 and eSH groups
depicted moderate antimicrobial activity whereas the aforemen- 3-(1-Nitronaphtho[2,1-b]furan-2-yl)-6-phenyl- 3-(1-Nitronaphtho[2,1-b]furan-2-yl)-5H-
tioned moieties linked through methylene spacer exhibited signifi- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (31) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-6(7H)-
one (32)
cant activity against both Gram-positive and Gram-negative strains.
Interestingly, 3-(4-isopropylthiazol-2-yl)-6-(4-methoxyphenyl)-
7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (62) demonstrated sig-
nificant activity against tested bacterial and fungal species.
Aggarwal et al. [63] reported a series of novel nalidixic acid based
triazolothiadiazole and triazolothiadiazine derivatives and screened
for their antibacterial efficacy using broth dilution technique.
Streptomycin drug was used as a standard. Compound (63) bearing
chloro substituent at 2-position on aromatic ring showed maximum
antimicrobial potency against all tested microorganisms which was 3,6-Bis(2,4-dichloro-5-fluorophenyl)-1,2,4- 3-(4-Isopropylthiazol-2-yl)-6-(4-
comparable to the standard drug. This could be due to the inductive triazolo[3,4-b]1,3,4-thiadiazole (33) methoxyphenyl)-7H-[1,2,4]triazolo[3,4-
b][1,3,4]thiadiazine (34)
and mesomeric effect. The triazolothiadiazine derivatives were
found to be less active as compared to the triazolothiadiazole series.
Prakash et al. [28] reported a series of dihydroindeno- and
indeno[1,2-e][1,2,4]triazolo[3,4-b] [1,3,4]thiadiazines and explored
their effect on in vitro growth of microorganism causing microbial
infection. Antimicrobial results revealed that all the tested com-
pounds possessed moderate to excellent antibacterial activity
against both Gram-positive (S. aureus, B. subtilis) and Gram-
negative bacteria (E. coli, P. aeruginosa). On the basis of zone of 6-(2,4-Difluorophenyl)-3-(4-isopropylthiazol- 3-(3,4-Dichlorophenyl)-6-(pyrazin-2-yl)-
inhibition against test bacteria, most of the synthesized compounds 2-yl)-5,6-dihydro-[1,2,4]triazolo[3,4-b][1,3,4]- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (36)
thiadiazole (35)
showed excellent activity against S. aureus showing the maximum
864 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868

3-(3,4-Dichlorophenyl)-6-phenyl-7H- 3-(2-Furanyl)-6-(2-chlorophenyl)-1,2,4-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (37) triazolo[3,4-b]-1,3,4-thiadiazole (38)
(Z)-3-((4-Chlorophenoxy)methyl)-6-(2,4- 6-(4-Chlorophenyl)-3-((6-chloropyridin-3-
dichloro-5-fluorophenyl)-7-(3,4- yl)methyl)-[1,2,4]triazolo[3,4-
dimethoxybenzylidene)-7H-[1,2,4]triazolo[3,4- b][1,3,4]thiadiazole (52)
b][1,3,4]thiadiazine (51)

Badr and Barwa [24] reported a series of triazolothiadiazoles and

triazolothiadiazines derived from 5-nitro furoic acid and tested for
3-(2-Furanyl)-6-(2-methoxyphenyl)-1,2,4- 6-Benzyl-3-(2-bromophenyl)- their in vitro antibacterial activity against S. aureus (Gram-positive
triazolo[3,4-b]-1,3,4-thiadiazole (39) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (40) bacteria) and E. coli (Gram-negative bacteria). For comparison,
ampicillin was used as a reference drug. All the synthesized com-
pounds exhibited considerable activity against S. aureus and most
of them are found to be as effective as reference drug. The obtained
results revealed that antibacterial activity of the newly synthesized
heterocyclic compounds, containing 1,2,4-triazole moiety fused
with 1,3,4-thiadiazole ring depend on the basic skeleton of the
molecule rather the substituents and all were found to be as
6-Benzyl-3-(naphthalen-2-ylmethyl)- 6-((2,4-Dichlorophenoxy)methyl)-3-(4- effective as reference drug ampicillin. The representative example
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (41) methoxyphenyl)-[1,2,4]triazolo[3,4- is 3-(5-nitrofuran-2-yl)-N-phenyl-[1,2,4]triazolo[3,4-b][1,3,4]thia-
b][1,3,4]thiadiazole (42)
diazol-6-amine (66). Meanwhile, among triazolothiadiazine de-
rivatives it was noticed that the activity relies upon the substituents
rather the basic skeleton of the molecule. The compounds, bearing
p-chlorophenyl moiety and p-bromophenyl moiety respectively
at position 6 of the thiadiazine ring were found to be as effective
as reference drug against S. aureus. The representative example is
6-(4-chlorophenyl)-3-(5-nitrofuran-2-yl)-7H-[1,2,4]triazolo[3,4-b]
[1,3,4]thiadiazine (67). However, compounds bearing p-nitro-
3-((2,4-Dichlorophenoxy)methyl)-6-phenyl- 3-(2-(Methylsulfonyl)benzyl)-6-phenyl-7H-
7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (43) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (44)
phenyl moiety and p-methoxyphenyl moiety respectively, at posi-
tion 6 of the thiadiazine ring exhibited moderate activity, while
compound bearing unsubstituted phenyl moiety at position 6 of the
thiadiazine ring showed weak activity.
Hussein et al. [46] reported 3,6-disubstituted-1,2,4-triazolothia-
diazole derivatives and screened for their antibacterial activity. Re-
sults from the tested compounds showed that 6-(pyridin-3-yl)-3-
(pyridin-4-yl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (68) exhibited
higher antibacterial activity than ampicillin, used as a standard drug.
6-(Biphenyl-4-yl)-3-(2- 3-(2-(4-Chlorophenyl)propan-2-yl)-6-(4- Sumangala et al. [58] reported a new series of 6-aryl-3-[4-
(methylsulfonyl)benzyl)-7H- (trifluoromethoxy)phenyl)-7H- (methylsulfonyl)benzyl]-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazines
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (45) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (46)
and evaluated for their antimicrobial activity. Among the screened
samples, compound (69) showed excellent antibacterial activity
against E. coli which may be due to the presence of 2,4-dichlorophenyl
group at C-6 position of thiadiazine. The antifungal activity of the
derivatives has much better results for many of the compounds like
(70). The significant activity of this compound may be due to the
presence of 3-coumarinyl group at C-6 position of thiadiazine ring.
Puthiyapurayil et al. [59] reported a series of triazolothiadiazine
derivatives and screened for their antimicrobial activity. The com-
3-(2-(4-Chlorophenyl)propan-2-yl)-6-(2,4- 6-(4-Chloro-3-nitrophenyl)-3-(2-(4-
dichloro-5-fluorophenyl)-7H- chlorophenyl)propan-2-yl)-7H- pound (71) showed comparatively very good activity against all
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (47) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (48) bacterial strains. The good activity was may be due to the presence
of pharmacologically active eOCF3, fluorine, (highly electro nega-
tive) in the molecules, which increases the lipophilicity and affects
the partitioning of molecules into membranes and facilitates hy-
drophobic interactions of the molecules with specific binding sites
on either receptor or enzyme. Results showed that combination of
triazolothiadiazine containing thioether linkage and presence of
pharmacologically active fluorine, chlorine and eOCF3 groups gave
an enhanced biological activity against all microbial strains.
2-((6-(4-Chlorophenyl)-3-methyl-7H- 4-Amino-3-methyl-1-((6-methyl-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-7- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-3-
Moreover, new triazolothiadiazole derivatives showing consid-
ylidene)methyl)-5-(4-nitrophenyl)-1,3,4- yl)methyl)-1H-1,2,4-triazol-5(4H)-one (50) erable antibacterial efficacy have been reported by Hanif et al. [64].
oxadiazole (49) Ciprofloxacin was used as a standard drug. Compound bearing
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 865

fluoro group on phenyl ring at position 6 exhibited significant ac-

tivity as depicted by 6-(2,5-difluorophenyl)-3-(2,5-
dimethoxybenzyl)-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (72).
Overall, most of the tested compounds in both the series expressed
excellent antibacterial activity as compared to standard drug
ciprofloxacin.
Recently, the research group of Jubie [65] reported some novel 6-(4-Chlorophenyl)-3-(3-methoxybenzyl)- 3-(3-Fluorobenzyl)-6-(furan-3-yl)-
stearic acid analogs such as 6-(heptadecyl)-3-phenyl-[1,2,4]triazolo [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (75) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (76)
[3,4-b][1,3,4]thiadiazole and tested for their antibacterial potential.
The results of preliminary antimicrobial testing of compounds
revealed that (73) showed pronounced antibacterial and antifungal 3.14. Antidepressant activity
activities against B. subtilis, P. aeruginosa and C. albicans, respec-
tively. This good inhibition may be attributed to the presence of an Recently, Jubie et al. [65] synthesized 6-(heptadecyl)-3-phenyl-
alkyl chain at position 6 of the condensed skeleton. [1,2,4]triazolo[3,4-b]-1,3,4-thiadiazole and screened for antide-
Very recently, in a continuation of their previous studies [64], pressant activity in swiss albino mice by forced swim test (FST). The
the research groups of Rama and Lee [66] have reported several compound (73) showed significant antidepressant activity.
pyridyl- and thiophenyl-substituted-1,2,4-triazolo[3,4-b]-1,3,4-
thiadiazole derivatives and screened for their antibacterial activ-
ity. Almost all the tested compounds were potent against four 3.15. Urease inhibitors
different strains of bacteria when compared with that of reference
drug ciprofloxacin. In particular, compound (74) was found to Hanif et al. [64] have reported a series of triazolothiadiazoles
possess the most potent activity (MIC: 0.156 mg/mL) against all the and evaluated for their urease inhibition potential. Almost all the
tested strains. The least activity was shown by compound con- compounds showed remarkable urease inhibition more than the
taining methoxy group on phenyl ring at position 3 and thiophen- standard inhibitor (thiourea) determined through the standard
2-yl methyl group at position 6. urease assay. 3-(2,5-Dimethoxybenzyl)-6-p-tolyl-[1,2,4]triazolo
[3,4-b][1,3,4]thiadiazole (77) was found to be the promising
candidate for urease inhibition.

6-(4-Heptadecylphenyl)-3-phenyl- 3-(2-Chlorophenyl)-6-(3,5-difluoropyridin-2-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (73) yl)-[1,2,4]-triazolo[3,4-b][1,3,4]thiadiazole
(74)

3.13. Antioxidant activity 3-((6-Chloropyridin-3-yl)methyl)-6-(4- 3-((4-(Methylthio)phenoxy)methyl)-6-p-tolyl-

methoxyphenyl)-7H-[1,2,4]triazolo[3,4- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (54)
Sunil et al. [33] reported two triazolothiadiazoles 6-[3-(4- b][1,3,4]thiadiazine (53)
fluorophenyl)-1H-pyrazol-4-yl]-3-[(2-naphthyloxy)methyl][1,2,4]
triazolo[3,4-b][1,3,4]thiadiazole (FPNT) and 6-[3-(4-chlororophenyl)-
1H-pyrazol-4-yl]-3-[(phenyloxy)methyl][1,2,4]triazolo[3,4-b][1,3,4]
thiadiazole (CPPT) and investigated the in vitro antioxidant property
by spectrophotometric DPPH and ABTS radical scavenging methods
as well as by lipid peroxide assay. The screening results proved that
FPNT (4) possess remarkable antioxidant potential.
Recently, Hanif et al. [64] reported various triazolothiadiazole
derivatives possessing methoxybenzyl and methoxyphenethyl
groups and evaluated for their antioxidant potential. The tri-
azolothiadiazole class of compounds was found to be significantly 6-(2,4-Dichlorophenyl)-3-((4- 3-(2,4-Dichloro-5-fluorophenyl)-6-(4-
active against superoxide anion radical. The screening results (methylthio)phenoxy)methyl)-7H- chlorophenyl)-1,2,4-triazolo[3,4-b]-1,3,4-
indicated that 6-(4-chlorophenyl)-3-(3-methoxybenzyl)-[1,2,4]tri- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (55) thiadiazole (56)
azolo[3,4-b][1,3,4]thiadiazole (75) possess significant antioxidant
potential with an IC50 value of 9.04 mM at such a low concentration,
even lower than standard. The activity of (75) is enhanced in this
class due to the presence of methoxy and chloro groups on phenyl
rings at 3rd and 6th positions of condensed skeleton, respectively.
More recently, Rafiq et al. [67] have reported a new series of
triazolothiadiazole derivatives and screened for their antioxidant
potential. In the DPPH free radical scavenging assay, n-propyl gallate
with an IC50 value of 40.8 mM was used as a reference drug. Com-
pound (76) with an IC50 value of 31.54 mM was the most active
compound of the series. Compound (76) has 3-fluorobenzyl group as
substituent attached with triazolothiadiazole ring, showing greater 6-(4-Methoxyphenyl)-3-(2,3,5- 6-(3-Amido-4-hydroxyphenyl)-3-(2,3,5-
electron donating ability. Compound with 3-methoxyphenyl group trichlorophenyl)-[1,2,4]triazolo[3,4- trichlorophenyl)-7H-[1,2,4]triazolo[3,4-
b][1,3,4]thiadiazole (57) b][1,3,4]thiadiazine (58)
as substituent was found to be the least active.
866 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868

3-(4-(Methylthio)benzyl)-6-(2,3,4- 3-(4-(N,N-Dimethylamino)phenyl)-6-(2-fluoro- 3-(2-Phenylpropan-2-yl)-6-[4- 6-(2,5-Difluorophenyl)-3-(2,5-

trichlorophenyl)-[1,2,4]triazolo[3,4- pyridin-4-yl)-1,2,4-triazolo[3,4-b]-1,3,4- (trifluoromethoxy)phenyl]-7H- dimethoxybenzyl)-[1,2,4]triazolo[3,4-
b][1,3,4]thiadiazole (59) thiadiazole (60) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (71) b][1,3,4]thiadiazole (72)

Recently, Rafiq et al. [67] have demonstrated the synthesis of a

series of triazolothiadiazole derivatives and screened for their
urease inhibitory activity. Almost all the compounds showed
excellent urease inhibition more than the reference thiourea.
Compound (78) with an IC50 value of 0.764  0.03 and Ki value of
0.450  0.06 mM was the most potent compound of the series. Most
active compound (78) has 4-methoxyphenyl group attached with
rigid triazolothiadiazole moiety. The replacement of methoxy
group with chloro group leads to the decrease in inhibitory activity.
3-[4-(Phenylsulfonyl)phenyl]-5H- 3-(4-Isopropylthiazol-2-yl)-6-(4-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-6(7H)- methoxyphenyl)-7H-[1,2,4]triazolo[3,4-
one (61) b][1,3,4]thiadiazine (62)

3-(2,5-Dimethoxybenzyl)-6-p-tolyl- 6-(Furan-3-yl)-3-(4-methoxyphenyl)-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (77) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (78)

3-{6-(2-Chlorophenyl)-1,2,4-triazolo[3,4-b] 3-Propyl-10,10a-dihydroindeno[1,2- 3.16. Plant growth regulators

[1,3,4]thiadiazol-3-yl}-1-ethyl-7-methyl-1,8- e][1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (64)
naphthyridin-4(1H)-one (63)
Jin et al. [68] synthesized a series of novel 6-aryl-3-(D-gal-
actopentitol-1-yl)-7H-1,2,4-triazolo[3,4-b][1,3,4]thiadiazines and
investigated the effects of these compounds on sprouting of wheat
and radish seeds. From the results, it was interesting to note that 1-
(6-phenyl-7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-3-yl)pentane-
1,2,3,4,5-pentaol (79) showed inhibitory activities toward the
growth of the dicotyledon (radish) at two concentration levels, but
under the same conditions it expressed stimulative activities
9-Methylindeno[1,2-e][1,2,4]triazolo[3,4- 3-(5-Nitrofuran-2-yl)-N-phenyl- toward the growth of the monocotyledon (wheat). It has a good
b][1,3,4]thiadiazine (65) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazol-6-amine level of activity and is worthy of further study to establish a rela-
(66)
tionship between structure and activity.

3.17. PDE4 inhibitors

Thomas and co-workers [14] showcased a series of substituted

7H-[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine and evaluated as PDE4
inhibitors. Several analogs were also explored with varying sub-
stitutions on the phenyl ring attached to the C-3 position of the
1,2,4-triazole ring system. Substitutions included methoxy, fluoro,
chloro and trifluoromethyl groups on the ortho, meta and para po-
6-(4-Chlorophenyl)-3-(5-nitrofuran-2-yl)-7H- 6-(Pyridin-3-yl)-3-(pyridin-4-yl)- sitions of the phenyl ring. Analysis of these analogs confirmed that
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (67) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazole (68)
various substitutions at one of the ortho positions were necessary to
maintain potent PDE4A inhibition. From the screening results,
compound (80) was identified as highly potent PDE4 inhibitor.

6-(2,4-Dichlorophenyl)-3-[4- 3-{3-[4-(Methylsulfonyl)benzyl]-7H- 1-(6-Phenyl-7H-[1,2,4]triazolo[3,4- 3-(2,5-Dimethoxyphenyl)-6-(4-methoxy-3-

(methylsulfonyl)benzyl]-7H- [1,2,4]triazolo[3,4-b][1,3,4]thiadiazin-6-yl}- b][1,3,4]thiadiazin-3-yl)pentane-1,2,3,4,5- (tetrahydrofuran-2-yloxy)phenyl)-7H-
[1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (69) 1H-isochromen-1-one (70) pentaol (79) [1,2,4]triazolo[3,4-b][1,3,4]thiadiazine (80)
 I. Khan et al. / European Journal of Medicinal Chemistry 63 (2013) 854e868 867

3.18. Anticandidal activity revealed that these compounds are the central structural motifs of
biologically interesting molecules which are of high interest to the
Altıntop et al. [69] synthesized several triazolothiadiazine de- pharmaceutical industry. The activity profile of these scaffolds
rivatives and tested in vitro against Candida species and compared relies upon the substitution on the parent skeleton of both het-
with ketoconazole. The compound bearing cyclohexyl moiety and erocycles and the indicated pharmacophores can be synthesized
p-chlorophenyl substituent on triazolothiadiazine ring (81) was based on the SAR analysis for further exploration of biological
found to be the most potent derivative against C. albicans (ATCC efficacy. The potential biological activities such as anticancer,
90028), Candida parapsilosis, C. albicans (NRRL Y-12983) and antitumor, anti-inflammatory, analgesic, anti-HIV, antidepressant,
Candida glabrata. It is apparent that there is a positive correlation anticandidal, anthelmintic, anticonvulsant, antimicrobial, antioxi-
between anticandidal activity and two functional moieties, namely dant, antitubercular, molluscicidal and cytotoxicity have been
cycloaliphatic group and p-chlorophenyl substituent on tri- exhibited by these analogs and comprehensively showcased in this
azolothiadiazine ring. It can be attributed to þp effect of chloro review. These derivatives have also been used as plant growth
substituent and cyclohexyl group. regulators and various enzyme inhibitors such as PDE4, urease,
alkaline phosphatase and acetylcholinesterase and have witnessed
3.19. Alkaline phosphatase inhibition important progress in recent years. In view of the increasing sig-
nificance of triazolothiadiazole and triazolothiadiazine scaffolds,
Rafiq et al. [67] have reported a new series of triazolothiadiazole we speculate that the design and exciting developments of novel
derivatives and screened for their alkaline phosphatase inhibitory synthetic methodologies combined with potential biological pro-
activity. Potassium dihydrogen phosphate was used as a standard file will provide a future insight in this rapidly evolving research
drug. Compound (82) with an IC50 value of 0.061  0.001 mM was area.
the most active compound of the series. It has 4-bromophenyl
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