Synthesis and characterization of biologically active new Schiff bases containing 3-functionalized 1,2,4-triazoles and their zinc(II) complexes: Crystal structure of 4-bromo-2-[(E)-(1H-1,2,4-triazol-3-ylimino)- methyl]phenol

Article abstract of DOI:10.1002/aoc.1833

Biologically active triazole Schiff bases (L¹-L³) derived from the reaction of 3-amino-1,2,4-triazole with chloro-, bromo- and nitro- substituted salicylaldehydes and their Zn(II) complexes (1-3) have been synthesized and characteriz

Full text of DOI:10.1002/aoc.1833

Full Paper
Received: 16 May 2011
Revised: 2 July 2011
Accepted: 12 July 2011
Published online in Wiley Online Library: 6 September 2011
(wileyonlinelibrary.com) DOI 10.1002/aoc.1833
Synthesis and characterization of biologically
active new Schiff bases containing
3-functionalized 1,2,4-triazoles and their
zinc(II) complexes: crystal structure of
4-bromo-2-[(E)-(1H-1,2,4-triazol-3-ylimino)-
methyl]phenol
Zahid H. Chohan∗ and M. Hanif
Biologically active triazole Schiff bases (L1 –L3) derived from the reaction of 3-amino-1,2,4-triazole with chloro-, bromo- and
nitro- substituted salicylaldehydes and their Zn(II) complexes (1–3) have been synthesized and characterized by their physical,
spectral and analytical data. Triazole Schiff bases potentially act as tridentate ligands and coordinate with the Zn(II) metal atom
through salicylidene-O, azomethine-N and triazole-N. The complexes have the general formula [M(L-H)2], where M = zinc(II) and
L = (L1 –L3), and observe an octahedral geometry. The Schiff bases and their Zn(II) complexes have been screened for in-vitro
antibacterial, antifungal and brine shrimp bioassay. The biological activity data show the Zn(II) complexes to be more potent
c
antibacterial and antifungal than the parent simple Schiff bases. Copyright ꢀ 2011 John Wiley & Sons, Ltd.
Supporting information may be found in the online version of this article.
Keywords: triazole Schiff bases; zinc(II) complexes; antibacterial; antifungal activity
Introduction
Experimental
Materials and Methods
Compounds containing triazole moiety have been revealed to
exhibit a wide variety of interesting biological properties such
as anticancer,[1,2] antifungal,[3–6] antimicrobial,[7–9] analgesic,[10]
anti-inflammatory,[11–14] antibacterial,[15–19] anticonvulsant[20–22]
and antitubercular.[23–27] Furthermore, they are used as a versatile
reagent in the synthesis of heterocyclic compounds and/or as a
raw material in drug synthesis[28a–e] (Fig. 1 shows some examples
of clinical drugs in practice containing 1,2,4-triazole ring system).
To expand and further explore the clinical significance of triazoles,
we have undertaken to synthesize new Schiff base derivatives
of triazole (L1 –L3; Scheme 1). These triazole-derived Schiff bases
havepotentialsitesformetalcomplexation,especiallywiththezinc
atom toform theirZn(II)complexes(1–3;Scheme 1). Wehopethat
these newly synthesized Zn(II) complexes will have a tendency to
reduce the resistivity of bacterial strains.[29] The newly synthesized
Schiff bases and their metal complexes were characterized by
their IR, NMR, molar conductance, magnetic moments, electronic
and elemental analyses data. The synthesized ligands and
their Zn(II) chelates were evaluated for in vitro antibacterial
activity against Escherichia coli, Salmonella sonnei, Pseudomonas
aeruginosa,Salmonellaentericaserovartyphi,Staphylococcusaureus
andBacillussubtilis, andforantifungalactivityagainstTrichophyton
longifusus, Candida albican, Aspergillus flavus, Microsporum canis,
Fusarium solani and Candida glabrata strains. In vitro brine shrimp
bioassayhasalsobeencarriedouttostudythecytotoxicproperties
of these compounds.
All chemicals used were of reagent grade. All metal salts were used
as chlorides. Melting points were recorded on Fisher Johns melt-
ing point apparatus. Infrared spectra were recorded on Shimadzu
FT-IR spectrometer. The C, H and N analyses was carried out using
a Perkin–Elmer, USA model. The H and 13C-NMR spectra were
1
recorded in DMSO-d6 using TMS as internal standard on a Bruker
Spectrospin Avance DPX-500 spectrometer. Electron impact mass
spectra (EIMS) were recorded on a Jeol MS Route Instrument.
In vitro antibacterial, antifungal and cytotoxic properties were
studiedatHEJResearchInstituteofChemistry,InternationalCentre
for Chemical Sciences, University of Karachi, Pakistan and Depart-
ment of Chemistry, The Islamia University of Bahawalpur, Pakistan.
General Procedure for the Synthesis of Schiff Bases (L1 –L3)
To a hot magnetically stirred solution of 3-amino-1,2,4-triazole
(0.84 g, 10 mmol) in methanol (15 ml) was added a solution of 5-
chlorosalicylaldehydes (1.56 g, 10 mmol) in methanol (25 ml).The
∗
Correspondenceto:Zahid H. Chohan,DepartmentofChemistry,BahauddinZa-
kariyaUniversity,Multan,60800,Pakistan.E-mail: dr.zahidchohan@gmail.com
Department of Chemistry, Bahauddin Zakariya University, Multan, 60800,
Pakistan
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Appl. Organometal. Chem. 2011, 25, 753–760
Copyright ꢀ 2011 John Wiley & Sons, Ltd.

Z. H. Chohan and M. Hanif
Figure 1. Examples of some bioactive compounds containing 1,2,4-triazole moiety.
Scheme 1. Preparation of triazole Schiff bases L1 –L3 and their Zn(II) complexes 1–3.
resultant mixture was refluxed for 3 h. During refluxing the solid
product was precipitated. The solid product was separated by
filtration, washed with hot methanol, then with ether and dried. It
was recrystallized in a hot solution of ethanol–methanol (1 : 1) to
obtain thin layer chromatography (TLC)-checked well-crystallized
product. The same method was applied for the preparation of all
other ligands.
4-Chloro-2-[(E)-(1H-1,2,4-triazol-3-ylimino)methyl]phenol (L1)
Yield: 81% (1.80 g); dark yellow; m.p. 222 ◦C; IR (KBr, cm−1): 3185
(NH), 2775 (H-bonded –OH), 1631 (HC N), 1608 (C N, triazole),
1036 (N–N), 815 (C–Cl); 1H NMR (DMSO-d6): δ 7.00 (d, J = 8.8 Hz,
C3 –H), 7.45 (dd, J = 8.8, 2.6 Hz, C4 –H), 7.88 (d, J = 2.6 Hz, C6 –H),
8.44 (s, C7 –H), 9.40 (s, C9 –H), 12.35 (s, 1H, OH), 14.10 (s, 1H, NH);
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Appl. Organometal. Chem. 2011, 25, 753–760

Biologically active new Schiff bases
13C NMR (DMSO-d6): δ 118.09 (C3), 120.45 (C1), 129.27 (C5), 132.72
(C6), 134.13 (C4), 152.43 (C9), 157.87 (C8), 159.93 (C2), 162.65 (C7);
EIMS (70 eV) m/z (%): 224 ([M]+, 32), 222 (94), 207 (72), 205 (100),
187 (9), 154 (13), 153 (17), 75 (17), 70 (26) 69 (32); Anal. calcd for
C9H7ClN4O (222.63): C, 48.55; H, 3.17; N, 25.17; Found: C, 48.32; H,
3.21; N, 25.08%.
[Zn(L1-H)2] (2)
Yield: 61% (0.365 g); dirty yellow; m.p. 285–287 ◦C (decomp);
IR (KBr, cm−1): 3175 (NH); 1616 (HC N); 1610/1595 (C N);
1384 (C–O); 1040 (N–N); 532 (Zn–N); 445 (Zn–O). Conductivity,
22.7 ꢀ−1 cm2 mol−1. Diamagnetic. λmax 28670 cm−1 1H NMR
.
(DMSO-d6): δ 7.13 (d, J = 8.2 Hz, C3 –H), 7.71 (dd, C4 –H), 8.13
(d, C6 –H), 8.79 (s, C7 –H), 9.68 (s, C9 –H), 14.18 (s, 1H, NH); 13C NMR
(DMSO-d6): δ 116.7 (C5), 119.35 (C3), 121.46 (C1), 133.95 (C6), 136.2
(C4), 152.05 (C9), 158.4 (C8), 160.23 (C2), 163.45 (C7). Anal. calcd for
C18H12N8O2Br2 Zn (597.56): C, 36.18; H, 2.02; N, 18.75; Found: C,
36.71; H, 1.91; N, 18.98%.
4-Bromo-2-[(E)-(1H-1,2,4-triazol-3-ylimino)methyl]phenol(L2)
Yield: 80% (2.13 g); dark yellow; m.p. 239 ◦C; IR (KBr, cm−1):
3175 (NH), 3268 (H-bonded –OH), 1632 (HC N), 1610 (C N,
triazole),1038 (N–N); 1H NMR (DMSO-d6): δ 6.96 (d, J =
8.7 Hz,C3 –H), 7.56 (dd, J = 8.7, 2.5 Hz, C4 –H), 8.00 (d, J = 2.5 Hz,
C6 –H), 8.45 (s, C7 –H), 9.39 (s, C9 –H), 12.3 (s, 1H, OH), 14.05 (s, 1H,
NH); 13C NMR (DMSO-d6): δ 116.57 (C5),119.24 (C3), 121.35 (C1),
133.86 (C6), 136.12 (C4), 151.89 (C9), 158.26 (C8), 160.09 (C2), 163.15
(C7); EIMS (70 eV) m/z (%): 268 ([M]+, 44), 266 (50), 251 (100), 249
(95), 199 (9), 76 (10), 69 (12); Anal. calcd for C9H7BrN4O (267.08): C,
40.47; H, 2.64; N, 20.98; Found: C, 40.29; H, 2.56; N, 21.07%.
[Zn(L3-H)2] (3)
Yield:67%(0.355 g);lghtyellow;m.p.258–260 ◦C(decomp);IR(KBr,
cm−1): 3196 (NH); 1619 (HC N); 1612/1596 (C N); 1384 (C–O);
1030 (N–N); 1370 (NO2); 528 (Zn–N); 455 (Zn–O). Conductivity,
24.9 ꢀ−1 cm2 mol−1. Diamagnetic. λmax 28 738 cm−1 1H NMR
.
(DMSO-d6): δ 7.30 (d, J = 8.3 Hz, C3 –H), 8.40 (dd, C4 –H), 8.88 (s,
C7 –H), 8.90 (d, C6 –H), 9.84 (s, C9 –H), 14.20 (s, 1H, NH); 13C NMR
(DMSO-d6): δ 117.23 (C3), 120.2 (C1), 125.25 (C4), 127.85 (C6),142.35
(C5), 152.55 (C9), 158.3 (C8), 161.9 (C2),164.15 (C7). Anal. calcd for
C18H12N10O6Zn (527.79): C, 41.81; H, 2.28; N, 26.44; Found: C, 42.09;
H, 2.44; N, 26.69%.
4-Nitro-2-[(E)-(1H-1,2,4-triazol-3-ylimino)methyl]phenol (L3)
Yield: 78% (1.82 g); yellow; m.p. 210 ◦C; IR (KBr, cm−1): 3196 (NH),
3285 (H-bonded –OH), 1635 (HC N), 1612 (C N, triazole), 1370
(NO2), 1029 (N–N); 1H NMR (DMSO-d6): δ 7.17 (d, J = 9.0 Hz,
C3 –H), 8.28 (dd, J = 9.0, 2.3 Hz, C4 –H), 8.6 (s, C7 –H), 8.81 (d,
J = 2.3 Hz, C6 –H), 9.53 (s, C9 –H), 13.05 (s, 1H, OH), 14.10 (s, 1H,
NH); 13C NMR (DMSO-d6): δ 117.09 (C3), 120.05 (C1), 125.13 (C4),
127.72 (C6),142.21 (C5), 152.38 (C9), 158.17 (C8), 161.73 (C2), 163.85
(C7); EIMS (70 eV) m/z (%): 397 ([M]+, 48), 380 (100), 335 (36), 259
(18), 245 (26), 191 (11), 164 (13), 150 (10), 138 (9), 121 (21), 76
(14); Anal. calcd for C9H7N5O3 (233.18): C, 46.36; H, 3.03; N, 30.03;
Found: C, 46.28; H, 2.96; N, 30.15%.
Pharmacology
In vitro antibacterial bioassay
All the newly synthesized compounds (L1 –L3) and their respective
metal (II) chelates (1–3) were tested against four Gram-negative
(E. coli, S. sonnei, P. aeruginosa and S. typhi) and two Gram-positive
(S. aureus and B. subtilis) bacterial strains by the disk diffusion
method.[30,31] The tested compounds (ligand and complex) were
dissolved in DMSO to get 10 mg ml−1 solution. A known volume
(10 µl) of the solution was applied with the help of a micropipette
onto the sterilized filter paper disks. The disks were dried at room
temperature overnight and stored in sterile dry containers. Disks
soaked with 10 µl of DMSO and dried in air at room temperature
were used as the negative control. The standard antibiotic disks
used as positive control were either purchased from manufacturer
or prepared as above in the laboratory by applying a known
concentrationofthestandardantibioticsolution.Bacterialcultures
were grown in nutrient broth medium at 37 ◦C overnight and
spread on to solidified nutrient agar medium in Petri plates
using sterilized cotton swabs in standard microbiological working
environment. Test and control disks were then applied to the
solidified medium surface with the help of sterilized forceps. The
plates were incubated at 37 ◦C for 12–15 h. The results were
recorded by measuring the zone of inhibition in mm against each
compound. As reference compound, ampicillin was used and the
experiments were carried out in triplicate and the values obtained
were statistically analyzed.
Procedure for Preparation of Zinc(II) Complexes (1–3)
Zn(II) complex with 4-chloro-2-[(E)-(1H-1,2,4-triazol-3-ylimino)
methyl]phenol (L1)
To a hot magnetically stirred solution of (L1; 0.445 g, 2 mmol) in
·
ethanol (20 ml) was added dropwise a solution of Zn(II) Cl2 6H2O
(0.238 g, 1 mmol) in ethanol (15 ml). The resultant mixture was
refluxed for 2 h. The precipitated product formed during refluxing
was collected by filtration, thorough washing with hot ethanol
followed by ether and dried. It was then recrystallized in dioxane.
The same method was used for the preparation of all other
complexes. The physical and analytical data for each metal
complex are given below.
[Zn(L1-H)2] (1)
Yield: 64% (0.325 g); dirty yellow; m.p. 282–284 ◦C (decomp); IR
(KBr, cm−1): 3185 (NH); 1618 (HC N); 1608/1592 (C N); 1388
(C–O); 1035 (N–N); 815 (C–Cl); 522 (Zn–N); 450 (Zn–O). Conduc-
In vitro antifungal bioassay
tivity, 25.6 ꢀ−1 cm2 mol−1. Diamagnetic. λmax 28 856 cm−1 1H
.
Antifungal activities of all compounds were studied[31] against six
fungalstrains(T.longifusus,C.albican,A.flavus,M.canis,F.solaniand
C. glabrata) using the disk diffusion method. Sabouraud dextrose
agar (Oxoid, Hampshire, UK) was seeded with 105 (cfu) ml−1 fungal
spore suspensions and transferred to Petri plates. Disks soaked in
20 ml (200 µg ml−1 in DMSO) of the compounds were placed at
NMR (DMSO-d6): δ 7.15 (d, J = 8.2 Hz, C3 –H), 7.60 (dd, C4 –H), 8.02
(d, C6 –H), 8.78 (s, C7 –H), 9.65 (s, C9 –H), 14.20 (s, 1H, NH); 13C NMR
(DMSO-d6): δ 118.2 (C3), 120.58 (C1), 129.4 (C5), 132.8 (C6), 134.25
(C4), 152.65 (C9), 158.0 (C8), 160.0 (C2), 162.89 (C7). Anal. calcd for
C18H12N8O2 Cl2Zn (508.63): C, 42.50; H, 2.38; N, 13.94; Found: C,
42.83; H, 2.24; N, 13.69%.
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Z. H. Chohan and M. Hanif
different positions on the agar surface. The plates were incubated
at 32 ◦C for 7 days. The results were recorded as percentage
inhibition and compared with standard drugs miconazole and
amphotericin B.
in the spectra of Schiff bases exhibited the intramolecular H-
bonded –OH. The medium intensity peak in the spectra of ligands
at 3175–3196 cm−1 was attributed to the v(NH) of triazole. The
appearance of a strong new band at 1631–1635 cm−1 in the
spectra of ligands was assigned[35] to azomethine (HC N) linkage,
whichresultedfromcondensationofamino(NH2)groupoftriazole
with carbonyl v(C O) group of salicylaldehydes. The azomethine
(HC N) linkage is confirmed by the disappearance of bands
at 1722 and 3330 cm−1 owing to carbonyl v(C O) and amine
v(NH2) stretching, respectively. The other bands appearing in
the spectra of ligands (newly synthesized) at 1608–1612 and
1029–1038 cm−1 were assigned to the vibrations of C N and
N–N of triazole. The medium intensity band at 3175–3196 cm−1
present in all the Zn(II) complexes was attributed[36] to the v(NH)
vibration. The strong new band appearing at 1631–1635 cm−1
owing to azomethine v(HC N), exhibited a lower shift of
10–20 cm−1 (1613–1621 cm−1) in all the metal complexes. This
shift indicates that the azomethine v(HC N) is coordinated with
the zinc metal ion via its nitrogen atom. Similarly, the band at
1608–1612 cm−1 already assigned to triazole v (C N) is also
Minimum Inhibitory Concentration
Compoundscontainingsignificantantibacterialactivity(over80%)
were selected for minimum inhibitory concentration (MIC) studies.
The minimum inhibitory concentration was determined using the
disk diffusion technique by preparing disks containing 10, 25, 50
and 100 µg ml−1 of the compounds and applying the literature
protocol.[32]
In Vitro Cytotoxic Bioassay
Brine shrimp (Artemiasalina leach) eggs were hatched in a shallow
rectangular plastic dish (22 × 32 cm), filled with artificial seawater,
which was prepared with commercial salt mixture and double-
distilled water. An unequal partition was made in the plastic dish
with the help of a perforated device. Approximately 50 mg of eggs
were sprinkled into the large compartment, which was darkened
while the matter compartment was opened to ordinary light. After
2 days nauplii were collected by a pipette from the lighted side. A
sample of the test compound was prepared by dissolving 20 mg
of each compound in 2 ml of DMSO. From this stock solution 500,
50 and 5 µg ml−1 were transferred to nine vials (three for each
dilutions were used for each test sample and LD50 is the mean
of three values) and one vial was kept as control having 2 ml of
DMSO only. The solvent was allowed to evaporate overnight. After
2 days, when shrimp larvae were ready, 1 ml of sea water and 10
shrimps were added to each vial (30 shrimps per dilution) and the
volume was adjusted with sea water to 5 ml per vial. After 24 h the
number of survivors was counted. Data were analyzed by using
the Finney computer program to determine the LD50 values.[33,34]
shifted to a lower frequency at 1592–1599 cm−1 by 10–20 cm−1
,
indicating the coordination of v(C N) of triazole with the Zn(II)
metal atom. Moreover, the disappearance of the broad band at
3268–3290 cm−1 in the spectra of ligands and the appearance of
new band at 1384–1388 cm−1 owing to v(C–O) was evidence[37]
of deprotonation and coordination of the zinc metal atom with the
ligand through oxygen of salicylidene. All the above evidence was
further supported by the emergence of new bands at 520–532
and 445–460 cm−1 owing to v(Zn–N) and v(Zn–O) vibrations.[38]
These new bands were only observed in the spectra of the zinc
complexes and not in its ligands. As a conclusion, comparison of
the spectra of the ligands (L1 –L3) and their Zn(II) complexes
(1–3) confirmed the coordination of ligands with the Zn(II)
metal tridentately through oxygen of salicylidene, nitrogen of
azomethine (HC N) and nitrogen of triazole (C N).
1H NMR spectra
Results and Discussion
1H NMR spectral data of the triazole-derived Schiff bases (L1 –L3)
and their diamagnetic Zn(II) complexes (1–3) are provided in the
Experimentalal section. The displayed signals of all the protons
of free ligands owing to heteroaromatic/aromatic groups were
Chemistry
The triazole-derived Schiff bases (L1 –L3) were prepared by the
reaction of 3-amino-1,2,4 triazole with a series of chloro-, bromo-
and nitro- substituted salicylaldehydes under reflux as shown in
Scheme 1. All the Schiff base derivatives (L1 –L3) were soluble in
ethanol, dioxane, dimethylformamide and dimethyl sulfoxide. The
composition of which is consistent with their micro-analytical and
mass spectral data. The Zn(II) complexes (1–3) were obtained
by stoichiometric reaction of the corresponding ligand with the
zinc metal as its chloride salt in a molar ratio M : L of 1 : 2. All
the complexes were stable against air and moisture at room
temperature. They were insoluble in common organic solvents
and only soluble in dimethylformamide (DMF) and DMSO. Physical
measurements and analytical data of the Zn(II) complexes (1–3)
are given in the Experimental section.
1
found to be in their expected regions.[39] The H NMR spectrum
of Schiff bases (L1) and (L2) displayed the OH proton of the
phenol moiety at δ12.3–12.35 and of (L3) at δ13.0 as singlets. The
compounds (L1 –L3) showed characteristic azomethine C7 –H and
triazole C9 –H protons at δ 8.44–8.60 and δ 9.39–9.53, respectively,
1
as a singlet. The H NMR spectra of L1 and L2 exhibited phenyl
C3 –H and C6 –H as doublet at 6.96–7.00 and 7.88–8.00 ppm,
respectively. However, the phenyl C4 –H of (L1) and (L2) appeared
as a double doublet at 7.45–7.56 ppm. The 1H NMR spectrum of
L3 showed the phenyl C3 –H and C6 –H protons as multiplets at
7.17 and 8.81 ppm, respectively. The proton, C4 –H appearing at
8.28 ppm was assigned to the phenyl as double doublet. A broad
singlet at 14.05–14.10 ppm displayed the NH proton of triazole in
all the ligands that disappeared on exchangement with D2O. The
coordination of the azomethine (CH N) nitrogen and nitrogen
of triazole (C N) ring were assigned by the downfield shifting
of the azomethine C7 –H proton and triazole C9 –H proton signal
from 8.44–8.60 and 9.39–9.53 ppm in free ligand to 8.78–8.88 and
9.65–9.84 ppm in its Zn(II) complexes, respectively. The downfield
shifting of azomethine proton in Zn(II) complexes was attributed
Spectroscopic Characterization of Ligands (L1 –L3) and their
Zn(II) Complexes (1–3)
IR spectra
The characteristic infrared spectral assignment of ligands (L1 –L3)
and their Zn(II) complexes are reported in experimental section.
The presence of a broad band in the region at 2768–2790 cm−1
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Appl. Organometal. Chem. 2011, 25, 753–760

Biologically active new Schiff bases
to the discharging of electronic cloud towards the Zn(II) ion. The
hydroxyl (OH) proton at 12.30–13.0 ppm in the ligands L1 –L3
disappeared in the spectra of its Zn(II) complexes, indicating
deprotonation and coordination of the oxygen with the metal ion.
All other protons underwent downfield shift by 0.15–0.30 ppm
owing to the increased conjugation[40] on complexation with the
zinc metal atom.
13C NMR spectra
The 13C NMR spectra of the free ligands and their Zn(II) chelates
were determined in DMSO-d6. The 13C NMR spectral data are
reportedalongwiththeirpossibleassignmentsintheExperimental
section and all the carbons were found in the expected regions.[39]
The conclusion obtained from these studies provides further
support to the mode of bonding explained in their IR and 1H NMR
spectral data. The azomethine carbon (CH N) of the ligands
appeared in the region δ 162.65–163.85 ppm. The aromatic
carbons of all the free ligands were found in the region δ
116.57–161.73 ppm. All the Schiff bases exhibited triazole carbons
in the region δ 151.89–158.26 ppm. Downfield shifting of the
azomethine carbon from δ 162.65–163.85 ppm in the free ligands
toδ 162.89–164.15 ppminitszinc(II)complexeswasduetoshifting
of electronic density towards the Zn(II) ion. The downfield shifting
also confirmed the coordination of the azomethine to the zinc
metal atom. Furthermore, the presence of the number of carbons
agreed well with their expected values.[40]
Figure 2. Crystal structure of ligand L2.
a nonelectrolytic nature.[43] The room temperature magnetic
moment values of Zn(II) complexes were found to be diamagnetic,
as expected. The electronic spectral values of Zn(II) complexes
1–3 are recorded in the Experimental section. The diamagnetic
Zn(II) chelates did not show any d–d transition and their
spectra were dominated only by the charge transfer band at
28 670–28 856 cm−1
.
Crystallographic structure of ligand L2
The molecular structure of ligand L2 (Fig. 2) shows the expected
bond lengths and angles. The triazol ring and the benzene
ring of the triazole-imino-phenol unit are not perfectly planar,
making a dihedral angle of 9.16◦(3). The functionalized arm at C(1)
Mass spectra
· · ·
atom adopts a cisoidal-conformation, with O–H N(1) distance
The electron impact mass spectral (EIMS) data of the ligands
was found to show[41,42] the compositions as: L1, C9H7N4ClO,
222.2 (calcd 222.63); L2, C9H7N4BrO, 267.0 (calcd 267.08); and
L3, C9H7N5O3 233.08 (calcd 233.18). The L1 showed a base peak
at 205.0 for the fragment [C9H6N4Cl]+. Similarly, L2 showed its
base peak for the fragment [C9H6N4Br]+ at 249 and L3 at 216.05
for fragment [C9H6N5O2]+. These were the most expected stable
fragments. The most fragmented pattern was followed by the
cleavage of C N (exocyclic as well as endocyclic), C C, C–C,
C–N, C–X (X = Cl, Br) and C–O bonds.
of 1.837(6) Å and C1–C2 and N1–C7 bonds in a common plane of
central system and atoms N1–C8 bond deviating from this plane
by 3.03◦(2).
The molecular structure data with the atom-numbering scheme
and 30% probability displacement ellipsoids are shown in
· · ·
Tables 1–3. H atoms have been saved for O–H N interaction
elucidation. Intermolecular interactions in the crystal structure,
C9H7BrN4O, has spatial group: P 43 21 2; a = 5.8778(3) Å;
b = 5.8778(3) Å; c = 55.458(6) Å; α = 90◦; β = 90◦; γ = 90◦;
V = 1915.99(6) Å3; Z = 8.
The hydroxyl-C2 group makes a dihedral angle of 4.48◦(3) with
the plane of the 1,2,4-triazole ring system. The hydroxyl group
adopts a cisoidal conformation with N3 atom. Within the
Conductance, magnetic susceptibility and electronic spectra of Zn(II)
complexes
· · ·
molecule, there are O–H N intramolecular contacts, as detailed
in Table 3.
The molar conductance values of the Zn(II) complexes 1–3
in DMF fell in the range 21.2–29.5 ꢀ−1 cm2 mol−1, showing
Table 1. Selected geometric parameters (Å)
O1–H1
0.839(4)
1.348(6)
1.413(7)
1.406(7)
C1–C6
1.406(7)
1.895(5)
1.457(7)
1.295(7)
117.5(5)
101.2(4)
101.2(4)
109.9(4)
101.6(4)
110.9(5)
116.3(5)
120.1(5)
119.9(5)
N1–C8
1.395(7)
1.355(7)
1.371(7)
1.333(7)
O1–C2
Br1–C5
N2–C8
C1–C2
C1–C7
N3–N4
C1–C6
N1–C7
N4–C9
O1–C2–C1
O1–C2–C3
C1–C2–C3
Br1–C5–C4
Br1–C5–C6
C4–C5–C6
C1–C6–C5
C1–C7–N1
C7–N1–C8
121.6(4)
N1–C8–N3
C8–N3–N4
C8–N3–N4
N3–N4–C9
C8–N2–C9
N2–C9–N4
N2–C8–N3
C2–C3–C4
C3–C4–C5
C2–C1–C6
C2–O1–H1
C2–C3–H3
C1–C6–H6
C1–C7–H7
N1–C7–H7
N3–N4–H4A
N4–C9–C9
N2–C9–H9
119.2(5)
118.6(5)
119.8(5)
118.6(4)
120.7(4)
120.7(5)
120.2(5)
120.0(5)
119.2(4)
109.4(4)
120.0(5)
119.9(5)
120.0(5)
120.0(5)
125.1(5)
124.5(5)
124.6(5)
c
Appl. Organometal. Chem. 2011, 25, 753–760
Copyright ꢀ 2011 John Wiley & Sons, Ltd.
wileyonlinelibrary.com/journal/aoc

Z. H. Chohan and M. Hanif
Table 2. Torsion angle (degree) of ligand L2
O1–C2–C1–C7
N1–C7–C1–C2
1.9(8)
3.0(8)
N2–C8–N1–C7
N3–C8–N1–C7
−7.8(8)
N1–C7–C1–C6
N1–C8–N2–C9
−178.0(5)
170.8(5)
178.6(5)
groups. Further, the delocalization of π-electrons over the whole
chelate ring is increased, which in turn increases the lipophilicity
of the complexes. The increased lipophilicity then enhances the
penetrationofthecomplexesintolipidmembranesofthebacterial
strains, thus killing more of them.[47,48]
Table 3. Hydrogen-bond parameters (Å)
· · ·
· · ·
· · ·
D–H
H
A
D
A
D–H
A
O1–H1
1.837
2.190
2.583
2.963
1477.31
146.43
N4–H4A
In Vitro Antifungal Bioassay
The antifungal screening of all the synthesized compounds was
carried out against T. longifusus, C. albican, A. flavus, M. canis,
F. solani and C. glabrata fungal strains (Table 5) according to the
literature protocol.[31] The obtained results were compared with
the standard drugs miconazole and amphotericin B. The activity of
standard drug is considered as 100%. The Schiff base derivatives
of triazole and their Zn(II) complexes exhibited a moderate to
significant degree of inhibitory effects on the growth of testing
fungal strains. However, only a few compounds showed low
degree of inhibitory effect on the growth of testing strains. The
antifungal results illustrated in Table 5 indicate that ligand L1
showedsignificantactivity(57–65%)againstcande,andmoderate
activity (42–45%) against a and d. Schiff base ligand L2 possessed
significant activity (53–66%) against a, d and f and moderate
activity (43%) against d. L3 exhibited significant activity (64–68%)
againstbanddandmoderateactivity(38–51%)againsta,c,eandf.
L1 showed low activity (18–25%) against b and f and L2 possessed
weak activity (27–28%) against c and e. Compound 1 showed
good antifungal activity (54–69%) against a, c and e and moderate
(37–48%) against b, d and f. Compound 2 was found to show a
significantactivity(57–82%)againsta,dandf,moderate(39–51%)
against b and weaker (29–36%) against c and e. Significant
activity (53–81%) was observed by compounds 3 against b–d,
and moderate (45–52%) against a, e and f. It was evident that
the overall potency of the uncoordinated compounds/ligands was
enhanced on coordination with the metal ion.
Pharmacology
In vitro antibacterial bioassay
The antibacterial activity of newly synthesized Schiff bases and
their corresponding Zn(II) complexes was determined against four
Gram-negative (E. coli, S. sonnei, P. aeruginosa and S. typhi) and two
Gram-positive (S. aureus and B. subtilis) bacterial strains (Table 4)
according to the literature protocol.[30,31] The obtained results
of all the compounds were compared with those of the standard
drug ampicillin (Table 4). The percentage of activity was compared
with the activity of the standard drug considering its activity as
100%. All the ligands (L1 –L3) and their Zn(II) complexes (1–3)
showed moderate to significant activity against all the bacterial
strains; however, weak activity was observed for L1 against f
and L2 against b. The activity of ligand L1 was found to be
significant (59–61%) against strains a and e, moderate (46–50%)
against b–d and weaker (31–33%) against f. The compound L2
showed moderate activity (43–51%) against a and c–f and weaker
(33%) against b. Significant activity (57–75%) was observed by
ligand L3 against b–e and moderate (50–51%) against a and f.
Compounds 1–3 showed overall a significant activity (53–85%)
against strains a–f. The results given in Table 4 more precisely
show that compounds 1–3 possessed activity, 61–84% against
a, 54–87% against b, 56–82% against d, 56–85% against c and
e and 55–77% against f. These results evidently show that the
activity of the Schiff base compounds enhanced on coordination
with the zinc metal. Enhancement in activity upon coordination
can be explained on the basis of our previous studies according to
the chelation theory.[44–46] Chelation reduces the polarity of metal
ion to a significant extent owing to overlapping with the donor
MIC for Antibacterial Activity
The preliminary screening results of all the compounds showed
thattheZn(II)complexes1and3werefoundtobethemost(above
Table 4. Antibacterial bioassay (concentration used 1 mg ml−1 of DMSO) of ligands L1 –L3 and Zn(II)-diaquo complexes 1–3; zone of inhibition
(mm)
Gram-negative
Pseudomonas
Gram-positive
Staphylococcus Bacillus
Escherichia
coli
Salmonella
sonnei
Salmonella enterica
serovar typhi
Compound
aeruginosa
aureus
subtilis
SA
L1
L2
L3
1
16
12
13
21
19
20
26
12
08
18
19
13
21
24
15
14
21
26
24
27
32
14
13
16
18
14
22
28
16
14
17
23
20
23
27
09
14
15
17
16
19
29
2.73
2.34
2.73
3.39
4.13
2.83
2.73
2
3
SD
>10, Weak; >10, moderate; >16, Significant. SD, standard drug (ampicillin). SA, statistical analysis.
c
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Copyright ꢀ 2011 John Wiley & Sons, Ltd.
Appl. Organometal. Chem. 2011, 25, 753–760

Biologically active new Schiff bases
Table 5. Antifungal bioassay (concentration used, 200 µg ml−1) of ligands L1 –L3 and their Zn(II) complexes 1–3
Antifungal activity (in vitro)
Trichophyton
longifusus
Candida
albican
Aspergillus
flavus
Microsporum
canis
Fusarium
solani
Candida
glabrata
Compound
L1
L2
L3
1
42
58
49
54
70
51
A
18
43
68
37
51
81
B
57
28
51
69
36
53
C
45
66
64
48
80
67
D
65
27
38
68
29
50
E
25
53
41
37
66
45
F
2
3
SD
SD, standard drugs. MIC (µg ml−1); A, Miconazole (70 µg ml−1; 1.6822 × 10−7 M ml−1); B, miconazole (110.8 µg ml−1; 2.663 × 10−7 M ml−1);
C, amphotericin B (20 µg ml−1; 2.164 × 10−8 M ml−1); D, miconazole (98.4 µg ml−1; 2.365 × 10−7 M ml−1); E, miconazole (73.25 µg ml−1
1.760 × 10−7 M ml−1); F, miconazole (110.8 µg ml−1; 2.663 × 10−7 M ml−1).
;
Table 6. Minimum inhibitory concentration (M ml−1) of the selected compounds 1 and 3 against selected bacteria and in vitro cytotoxic bioassay
of ligands, L1 –L3, and their Zn(II) complexes, 1–3
In vitro antibacterial bioassay. Minimum inhibitory concentration (M ml−1
)
Cytotoxic bioassay
LD50 (M ml−1
Escherichia
coli
Salmonella
sonnei
Psudomonas
aeruginosa
Staphylococcus
aureus
No.
)
L1
L2
L3
1
–
–
–
–
>2.88 × 10−4
>2.77 × 10−4
>2.92 × 10−4
>5.76 × 10−4
>1.05 × 10−4
>4.23 × 10−4
–
–
–
–
–
–
–
–
2.15 × 10−7
–
2.13 × 10−7
1.34 × 10−7
2
–
–
–
–
–
3
0.65 × 10−7
1.29 × 10−7
1.25 × 10−7
80%) active compounds. Therefore, these two compounds were
selected for MIC studies (Table 6). The MIC values of compounds
1 and 3 fell in the range 6.475 × 10−8 to 2.252 × 10−7 M. The
MIC results showed that compound 3 is the most active, showing
a maximum inhibition of 6.475 × 10−8 M against bacterial strain
S. sonnei.
of cell membrane of the microorganism. Further, it has been
suggested that some functional groups such as azomethine
(HC N) or hetero-aromatics present in these compounds also
play an important role in antibacterial and antifungal activity that
may be responsible for the enhancement of the hydrophobic
character and liposolubility of the molecules.
Acknowledgments
In Vitro Cytotoxic Bioassay
One of us (M.H.) is grateful to the Higher Education Commission,
Government of Pakistan for an award to carry out this research.
The synthesized ligands L1 –L3 and their Zn(II) complexes (1–3)
were screened for their cytotoxicity (brine shrimp bioassay) using
the protocol of Meyer et al.[33] The cytotoxic data recorded in
Table 6 revealed that only compound 2 displayed potent cytotoxic
activity (LD50 = 1.676 × 10−4 M) against Artemia salina, while all
other compounds were considered as almost inactive in this assay.
It was interesting to note that the zinc(II) complexes showed
potent cytotoxicity as compared with the ligands. This activity
relationship may serve as a basis for future development of certain
cytotoxic agents in clinical practices.
Supporting Information
Crystallographic data for the structural analysis has been de-
posited with the Cambridge Crystallographic Data Centre, CCDC
no. 774 652 for ligand L2. A copy of this information can be
obtained free of charge from The Director, CCDC, 12 Union
Road, Cambridge, CB2 1EZ, UK (fax: +44-1223–336033; email: de-
posit@ccdc.cam.ac.uk or http://www.ccdc.cam.ac.uk). Supporting
information can be found in the online version of this article.
Conclusions
The improvement of antibacterial and antifungal activity in
ligands L1 –L3 increased upon chelation/coordination. It has been
proposed that the chelation process reduces the polarity of the
metal ion by coordinating with ligands, which in turn increases
the lipophilic nature of the zinc metal. This lipophilic nature of
metal thus enhances its penetration through the lipoid layer
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