Microwave assisted synthesis, spectroscopic characterization and biological aspects of some new chromium(iii) complexes derived from NO donor Schiff bases

Article abstract of DOI:10.1039/c3nj40907d

The present study is structured to address the development of green microwave processes in the manufacture of a new series of biologically potent (NO) donor Schiff bases and their Cr(iii) complexes. These have been characterized by elemental analysis, melting point and molecular weight determinations, IR, UV-vis and EPR spectral analysis, and X-ray powder diffraction studies. The bioefficient compounds have been synthesized by thermal as well as microwave assisted methods. The azomethine ligands 1-(2-pyridyl) ethanone isonicotinoyl hydrazone (L¹H) and 1-(2-naphthyl) ethanone isonicotinoyl hydrazone (L²H) have been synthesized by the condensation of 1-(2-pyridyl) ethanone and 1-(2-naphthyl) ethanone with isonicotinic acid hydrazide in a 1 : 1 molar ratio, using ethanol as a solvent. The Cr(iii) complexes have been prepared by mixing CrCl₃· 6H₂O in 1 : 1 and 1 : 2 molar ratios with ligands L¹H and L²H in methanol. The synthesized ligands and their new metal complexes have been screened in vitro for antimicrobial activity and in vivo for antifertility activity on male albino rats. The spectral data suggested a hexa-coordinated environment around the central metal ion. Physicochemical studies of the ligands and their Cr(iii) complexes aided the construction of their proposed structures.

Full text of DOI:10.1039/c3nj40907d

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Microwave assisted synthesis, spectroscopic
characterization and biological aspects of some
new chromium(III) complexes derived from
N^O donor Schiff bases
Cite this: NewJ.Chem., 2013,
37, 1445
Nighat Fahmi,*a Sumit Shrivastava,a Ramhari Meena,a S. C. Joshib and R. V. Singha
The present study is structured to address the development of green microwave processes in the
manufacture of a new series of biologically potent (N^O) donor Schiff bases and their Cr(III) complexes.
These have been characterized by elemental analysis, melting point and molecular weight
determinations, IR, UV-vis and EPR spectral analysis, and X-ray powder diffraction studies. The
bioefficient compounds have been synthesized by thermal as well as microwave assisted methods. The
azomethine ligands 1-(2-pyridyl) ethanone isonicotinoyl hydrazone (L1H) and 1-(2-naphthyl) ethanone
isonicotinoyl hydrazone (L2H) have been synthesized by the condensation of 1-(2-pyridyl) ethanone and
1-(2-naphthyl) ethanone with isonicotinic acid hydrazide in a 1 : 1 molar ratio, using ethanol as a
solvent. The Cr(III) complexes have been prepared by mixing CrCl3ꢀ6H2O in 1 : 1 and 1 : 2 molar ratios
with ligands L1H and L2H in methanol. The synthesized ligands and their new metal complexes have
been screened in vitro for antimicrobial activity and in vivo for antifertility activity on male albino rats.
Received (in Montpellier, France)
9th October 2012,
Accepted 6th February 2013
The spectral data suggested
a hexa-coordinated environment around the central metal ion.
DOI: 10.1039/c3nj40907d
Physicochemical studies of the ligands and their Cr(III) complexes aided the construction of their
proposed structures.
www.rsc.org/njc
rise in temperature. Since the process is not limited by the
thermal conductivity of the vessel, the result is an instantaneous
1. Introduction
The use of microwave irradiation in chemical synthesis has
become increasingly popular within the pharmaceutical and
academic arenas, because it is a new enabling technology for
drug discovery and development1 and due to certain advan-
tages, particularly shorter reaction times and rapid optimiza-
tion of chemical reactions.2 Presently, thermally driven
chemical synthesis takes place by one of two ways: conventional
heating or microwave accelerated heating. For the conventional
heating method, reactants are slowly activated by a conven-
tional external heat source.3 Heat is driven into the substance,
passing first through the walls of the vessel in order to reach
the solvent and reactants. This is a slow and inefficient method
for transferring energy into the reacting system. For the micro-
wave heating method, microwaves couple directly with the
molecules of the entire reaction mixture, leading to a rapid
localized superheating of any substance that will respond to
either dipole rotation or ionic conduction.4 Many of the metal
complexes of Schiff base ligands containing N^O donor atoms,
such as semicarbazones5,6 and thiosemicarbazones,7,8 iso-
nicotinoyl hydrazones and their metal chelates, have been
found to exhibit fungicidal, bactericidal, antiviral, and anti-
tubercular activities. Thus the metal complexes of such type of
Schiff base ligands are continuing to attract the interest of
scientists and the coordination of these ligands with transition
9–11
metals has been explored more thoroughly
than their
coordination with non-transition metals. Isonicotinic acid
hydrazide (INH) is a drug of proven therapeutic importance
and is used against a wide spectrum of bacterial ailments.12
In the past Agarwal and Sarin13 have investigated the coordi-
nating ability of INH-derivatives with metal ions. Hydrazones
derived from the condensation of isonicotinic acid hydrazide
with heterocyclic aldehydes have been found to show better
antitubercular activity than INH. Because of the wide range
of medicinal applications of isonicotinic acid hydrazides14,15
and their abilities to coordinate with the transition metal ions,
a Department of Chemistry, University of Rajasthan, Jaipur-302004, India.
E-mail: nighat.fahmi@gmail.com, rvsjpr@hotmail.com; Fax: +91 141 2708621;
Tel: +91 141 2704677 (NF), +91 141 2742835 (RS)
b Department of Zoology, University of Rajasthan, Jaipur-302004, India
c
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it is highly desirable to synthesize and characterize transition The solution was then concentrated under reduced pressure,
metal complexes with such Schiff base ligands. Keeping which upon cooling gave dark yellow crystalline precipitates.
all these facts under consideration, during the present investi- The products were washed in alcohol and recrystallized in the
gations we have synthesized, characterized and screened same solvent. The structures of ligands (L1H and L2H) are
the biologically potent ligands and their chromium com- shown in (Fig. 1).
plexes against a variety of pathogenic fungal and bacterial
A comparison between the thermal method and microwave
strains. Further, the complexes were also tested for their method is given in (Table 1).
antifertility activity in male albino rats and the results were
2.2. Preparation of the complexes
indeed positive.
For comparison purposes, two different routes were employed
for the synthesis of the metal complexes with each of the
ligands (L1H and L2H). A comparison between the thermal
method and microwave method has been given in Table 1.
(A) Microwave method: The complexes were prepared by
irradiating a reaction mixture of CrCl3ꢀ6H2O and the respective
ligand (L1H and L2H) in an appropriate stoichiometric pro-
portion in 3–5 mL dry methanol using NaOH. The dry products
were recovered from the microwave oven and dissolved in
B5 mL of dry methanol, where the sodium chloride precipitate
(formed during the course of the reaction) was removed by
filtration and the filtrate was then concentrated under reduced
pressure. The resulting compounds were washed with cyclohexane
and recrystallized with methanol.
2. Experimental
All chemicals and reagents used were of AR grade and dried
and distilled before use. The CrCl3ꢀ6H2O was purchased from
Alfa Aesar. All the reagents were dried and distilled before use.
1-(2-pyridyl) ethanone and 1-(2-naphthyl) ethanone were pur-
chased and used as such. Apparatus fitted with Quickfit inter-
changeable joints were used to carry out the reactions.
2.1. Preparation of the ligands
2.1.1. Preparation of the azomethine ligands (L1H and L2H).
Two different routes were used for the synthesis of (L1H and L2H)
ligands (Scheme 1).
(B) Thermal method: The complexes were also synthesized
by the thermal method, where instead of 10–14 min, reactions
were completed in 14–16 hours and the yield of the products
was also less than that obtained by the microwave assisted
synthesis. In this method a methanolic solution of CrCl3ꢀ6H2O
was added to a methanolic solution of ligand (L1H and L2H) in
1 : 1 and 1 : 2 molar ratios using NaOH in appropriate stoichio-
metric proportions. The resulting mixture was heated under
reflux for 14–16 hours, filtered to remove NaCl and the solvent
was concentrated under reduced pressure. The product was
dried in a vacuum. The resulting compounds were washed with
cyclohexane and recrystallized with methanol.
(A) Microwave method: In the microwave assisted method,
1-(2-pyridyl) ethanone isonicotinoyl hydrazone (L1H) and 1-(2-
naphthyl) ethanone isonicotinoyl hydrazone (L2H) were formed
by the condensation of 1-(2-pyridyl) ethanone (0.05 mol)
and 1-(2-naphthyl) ethanone (0.05 mol) with isonicotinic acid
hydrazide (0.05 mol) in a 1 : 1 molar ratio using a microwave
oven, with 5 mL ethanol as solvent. The reactions were com-
plete in a short period of 10–12 min. The solution was then
concentrated under reduced pressure, which upon cooling gave
dark yellow crystalline precipitates. The products were washed
in alcohol and recrystallized in the same solvent.
(B) Thermal method: For comparison purposes, the above
ligands were also synthesized by the thermal method, where
instead of a few drops of alcohol, a hot ethanolic solution
2.3. Physical measurements and analytical methods
(25 mL) of 1-(2-pyridyl) ethanone (0.01 mol) and 1-(2-naphthyl) The molecular weights were determined by the Rast Camphor
ethanone (0.01 mol) was mixed with a hot ethanolic solution method.16 The metal contents were analysed gravimetrically.
(30 mL) of isonicotinic acid hydrazide (0.01 mol) in a 1 : 1 molar Sulfur and nitrogen were determined by Messenger’s17 and
ratio. The contents were refluxed for about 3–4 hours in a water bath. Kjeldahl’s methods,18 respectively. Carbon and hydrogen analyses
Scheme 1 Synthesis of ligand L2H and its tautomeric form.
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Fig. 1 Structures of the azomethine ligands (L1H and L2H).
Table 1 Comparison between the microwave and thermal method
Yield %
Solvent (mL)
Thermal
Time
Thermal
Microwave
Microwave
Thermal (h)
Microwave (m)
Compounds
L1H
65
73
73
66
70
77
82
90
83
87
80
84
100
100
50
45
40
5
5
3
5
3
3.5
3
4
15
14
16
14
10
12
10
10
12
14
L2H
[CrCl2(L1)(H2O)2]
[CrCl(L1)2(H2O)]
[CrCl2(L2)(H2O)2]
[CrCl(L2)2(H2O)]
50
were performed at the CDRI, Lucknow. Infrared spectra were inoculum needle. These Petri plates were wrapped in polythene
recorded on a Nicolet Megna FTIR-550 spectrophotometer bags containing a few drops of alcohol and placed in an
using KBr pellets. The electronic spectra were recorded on a incubator at 25 ꢁ 2 1C. The activity was determined after
Varian–Cary/5E spectrophotometer at SAIF, IIT, Madras, Chennai. 96 h of incubation at room temperature (25 1C). The controls
EPR spectra of the complexes were monitored on Varian E-4X were also run and three replicates were used in each case. The
band spectrometer at SAIF, IIT, Madras, Chennai.
linear growth of the fungus was obtained by measuring the
diameter of the fungal colony after four days and the per-
centage inhibition was calculated as 100 ꢂ (C ꢃ T)/C, where
2.4. Antimicrobial studies
2.4.1. Antifungal studies. The bioefficiencies of the ligands C = diameter of the fungus colony in the control plate after 96 h
and their metal complexes, synthesized by thermal as well as and T = diameter of the fungal colony in the test plates after the
microwave methods, were checked in vitro. The antifungal same period. The antifungal screening data of compounds were
activities of the ligands and their complexes have been compared with the standard (Flucanazone).
evaluated against two pathogenic fungi, Candida albicans and
2.4.2. Antibacterial studies. The antibacterial activity of the
Aspergillus niger, by the agar plate technique. The potato ligands and their chromium complexes were evaluated against
dextrose agar (PDA) medium was prepared in the laboratory two bacteria including Gram-positive (Bacillus subtilis), and
to maintain the fungal growth. For PDA preparation, 20 g Gram-negative (Escherichia coli). The nutrient agar medium
potato was extracted with distilled water (100 mL) at 100 1C [composition: peptone (5 g), beef extract (5 g), NaCl (5 g),
for 1 h and was filtered off using a cotton filter. The potato juice agar–agar (20 g) and distilled water (1000 mL)] was pipetted
was then mixed with 2 g dextrose and 1.5 g agar and finally the into the Petri dish. When it solidified, 5 mL of warm seeded
pH of the prepared PDA media was adjusted to 7. Solutions of agar was applied. The seeded agar was prepared by cooling the
the test compounds in methanol at 100 and 200 ppm concen- molten agar to 40 1C and then adding 10 mL of the bacterial
trations were prepared and mixed with the medium. The suspension. The compounds were dissolved in methanol in
medium was then poured into Petri plates and the spores of 500 and 1000 ppm concentrations. Paper discs of Whatman
the fungi were placed on the medium with the help of an No. 1 filter paper, with diameters of 5 mm, were soaked in the
c
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solutions of varied concentrations. The discs were dried and
Biochemical analysis. Testicular cholesterol, testicular glycogen,
placed on the medium previously seeded with organisms in protein and sialic acid in the testes, epididymis, ventral prostate
Petri plates at a suitable distance. The Petri plates were stored and seminal vesicle were estimated by standard laboratory
in an incubator at 28 ꢁ 2 1C for 24 hours. The diameters of the methods.
zone of inhibition produced by the compounds were compared
Serum testosterone. Testosterone in the serum was measured
with the standard antibiotic (streptomycin). The zone of inhibi-
by the radioimmuno assay.19 Differences between the control
tion thus formed around each disc containing the test com-
and treated animals were evaluated statistically using the
pounds was measured accurately in mm.
Student’s ‘‘t’’ test. A t-test is any statistical hypothesis test in
which the test statistic follows a Student’s t distribution if the
null hypothesis is supported. It is most commonly applied
when the test statistic would follow a normal distribution if
2.5. Antifertility activity
Material and methods. Fertile healthy adult male rats of the
Wistar Strain, with an average body weight between 200–230 g,
the value of a scaling term in the test statistic was known. When
were used for the present study. The animals were housed in
the scaling term is unknown and is replaced by an estimate
plastic cages and were maintained at a room temperature
based on the data, the test statistic (under certain conditions)
(221 ꢁ 2 1C) and uniform light (14 : 10 : L : D). The rats had a
follows a Student’s t distribution.
diet of standard commercial pelleted feed (Ashirwad Food
Industries Ltd., Chandigarh, India) and fresh water ad libitum
throughout the study.
3. Results and discussion
Arrangement of animals for the study. The animals were
The elemental analysis and spectral data are consistent with
divided into seven groups of six animals. The animals in group
the formulation of compounds [CrCl2(L)(H2O)2] and [(CrCl(L)2-
A served as a control group and received the vehicle (0.5 mL
(H2O)]. The resulting chromium complexes are green solids,
olive oil) through oral insterbation at the same time every day.
soluble in MeOH, DMF and DMSO. Molecular weight determi-
The animals in groups B and C received ligands (L1H and L2H),
nations indicate their monomeric nature. The reactions of
whereas the animals in groups D, E, F and G received
CrCl3ꢀ6H2O with the ligands (L1H and L2H) were carried out
chromium(III), complexes of ligands (L1H and L2H), respec-
in unimolar and bimolar ratios in methanol solution with the
tively. The animals were maintained under perfect supervision
and in accordance to the guidelines of committee for the
purpose of control and supervision of experiment on animals
formation of M’N and M–O bonds yielding the substitution
products. The reactions proceed as shown in Scheme 1 and 2.
CrCl3 ꢀ 6H2O þ LH þ NaOH
(CPCSEA) for the regulations of scientific experiments on
(1)
(2)
animals. The experimental protocol has approval of the institu-
tional ethical committee Dept of zoology UOR Jaipur.
Mode of administration of the compound. In groups B
and C, the ligands (20 mg kgꢃ1 b.wt) were dissolved in
0.5 mL olive oil and given orally by a pearl point needle for a
period of 60 days. The animals in groups D, E, F and G, received
same doses of the respective compounds for the same period.
No rat mortality occurred during the study period. The mating
exposure test was done on day 55 of the experiment. The rats
were sacrificed after 24 h of the last dose (61st day) to perform
various tests.
Fertility test. The mating exposure test of all the animals was
performed. They were cohabited with pro-estrous females in
the ratio of 1 : 3. The vaginal plug and presence of sperms in the
vaginal smear was checked for positive mating. Females were
separated and the resulting pregnancies were noted, when
dams gave birth. Fertility was calculated in the control as well
as in the treated groups. At the end of the experimental period,
the animals were weighed and autopsied under light ether
anesthesia.
Â
Ã
1 : 1
ꢃ! CrCl2ðLÞðH2OÞ2 þ NaCl þ 5H2O
MeOH
CrCl3 ꢀ 6H2O þ 2LH þ 2NaOH
Â
Ã
1 : 2
ꢃ! CrClðLÞ2ðH2OÞ þ 2NaCl þ 7H2O
MeOH
Metal complexes were prepared by the microwave method, in
addition to the conventional method. The reason for synthesizing
the metal complexes by the microwave method is due to its
ecofriendly nature. The reaction time is brought from hours to
seconds and a small amount of solvent is required for com-
pletion of the reaction. An enhancement of the yield of the
resulting products is observed.
The physical properties and analytical data of the ligands
and their metal complexes, synthesized by a green chemical
approach as well as conventional method are detailed in
Tables 2 and 3.
3.1. Infrared spectral data
The significant IR bands of the ligands and their metal complexes,
along with their tentative assignments, are reported in Table 4,
which were used for the establishment of the mode of coordination
of the bidentate ligands towards the metal ion. The IR spectra of
Parameters studied
Body and organ weights. The initial and final weights of the
testes, epididymis, seminal vesicle and ventral prostate were
recorded and processed for biochemical estimations.
the ligands (L1H and L2H) show a strong band at 1615–1610 cmꢃ1
,
Sperm dynamics. Sperm density and sperm motility were due to the 4CQN group, which shifts to a lower wave number
assessed by standard methods.
(B15–10 cmꢃ1) in the chromium(III) complexes, indicating the
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Table 2 Analytical data and physical properties of the ligands and their complexes – synthesized by conventional heating
Mol. wt. found
(calculated)
Found (calculated) (%)
C
H
N
Cl
M
Compounds
Colour
Melting point (1C)
L1H
L2H
Dark yellow 140–150
Dark yellow 130–142
63.97 (64.98)
73.70 (74.72) 13.82 (14.52)
38.18 (39.21)
52.46 (53.47)
47.30 (48.32)
62.35 (63.38)
4.53 (5.33)
22.61 (23.31)
4.74 (5.22)
—
—
—
—
238.30 (240.26)
287.23 (289.33)
[CrCl2(L1)(H2O)2] Green
[CrCl(L1)2(H2O)] Green
[CrCl2(L2)(H2O)2] Green
[CrCl(L2)2(H2O)] Green
260–265 (da)
220 (da)
3.19 (3.79)
3.64 (4.14)
3.98 (4.05)
4.19 (4.43)
13.87 (14.13) 16.21 (17.80) 12.28 (13.05) 397.06 (398.18)
18.94 (19.18)
8.78 (9.39)
5.46 (6.07)
8.20 (8.90)
581.42 (583.97)
266–270 (da)
223–225 (da)
15.12 (15.85) 10.94 (11.62) 445.57 (447.30)
4.33 (5.19)
11.91 (12.32)
6.87 (7.62)
679.32 (682.11)
a
d = decomposition.
Table 3 Analytical data and physical properties of the ligands and their complexes – synthesized by microwave heating
Found (calculated) (%)
Mol. Wt. found
(calculated)
C
H
N
Cl
M
Compounds
Colour
Melting point (1C)
L1H
L2H
Dark yellow 140–150
Dark yellow 130–142
64.20 (64.98)
74.15 (74.72) 14.20 (14.52)
38.86 (39.21)
53.09 (53.47)
47.91 (48.32)
62.96 (63.38)
5.01 (5.33)
22.82 (23.31)
4.98 (5.22)
—
—
—
—
239.06 (240.26)
288.13 (289.33)
[CrCl2(L1)(H2O)2] Green
[CrCl(L1)2(H2O)] Green
[CrCl2(L2)(H2O)2] Green
[CrCl(L2)2(H2O)] Green
260–265 (da)
220 (da)
3.54 (3.79)
3.78 (4.14)
3.98 (4.05)
4.23 (4.43)
13.92 (14.13) 16.90 (17.80) 12.88 (13.05) 397.32 (398.18)
18.95 (19.18)
8.98 (9.39)
5.65 (6.07)
15.32 (15.85) 11.06 (11.62) 446.18 (447.30)
4.84 (5.19) 7.02 (7.62) 680.41 (682.11)
8.34 (8.90)
582.12 (583.97)
266–270 (da)
223–225 (da)
11.96 (12.32)
a
d = decomposition.
Table 4 IR (cmꢃ1) spectral data of the ligands and their metal complexes
IR spectral data (cmꢃ1
the electronic spectra of the chromium(III) complexes that
were recorded in DMSO. In the case of the chromium(III)
complexes, the three d–d transitions are expected and are
also observed experimentally. Three bands at 16556–17513,
22780–24038 and 30211–33000 cmꢃ1 were observed due to
the 4A2g - 4T2g (n1), 4A2g - 4T1g (n2) and 4A2g - 4T1g (P)
(n3) transitions, respectively, suggesting an octahedral geometry
around the Cr3+ ion.20 Various ligand field parameters like Dq,
B and b have been calculated and given in Table 5. The
)
v(NH)
n(OH)
n(CQN)
n(M–O)
n(M’N)
Compound
L1H
3244
3250
—
—
—
—
—
3465
3452
3467
3460
1615
1610
1602
1605
1600
1595
—
—
588
585
600
598
—
—
435
423
440
428
L2H
[CrCl2(L1)(H2O)2]
[CrCl(L1)2(H2O)]
[CrCl2(L2)(H2O)2]
[CrCl(L2)2(H2O)]
—
4
energy of the first spin-allowed transition [4A2g (F) - T2g (F)]
directly gives the value of 10Dq. The electronic repulsion
parameter is expressed in terms of Racah parameter and ‘B’
has been evaluated during these studies. The nephelauxetic
ratio b indicates that the complexes have appreciable covalent
character.
coordination of the azomethine nitrogen to the metal atom. The IR
spectra of the ligands show bands at 3244–3250 cmꢃ1, due to n(NH)
group, which has disappeared from the spectra of the complexes.
The medium intensity bands in the region 1675–1690 cmꢃ1, due to
n(CQO) vibrations in the ligands (L1H and L2H), shifted to a lower
frequency in the spectra of their complexes, which indicates
amide–imidol tautomerism and their subsequent coordination
through the imidol oxygen. In the spectra of all the chromium(III)
complexes, a band is observed in the range 880–876 cmꢃ1, which
may be attributed to the coordinated water molecule. Further, a
broad band at around 3467–3452 cmꢃ1 may be due to the n(O–H) of
the water molecule. The single band observed at ca. 320–315 cmꢃ1
is due to n(Cr–Cl), suggesting that all the complexes have a trans
structure. The presence of non-ligand bands, due to n(Cr’N) and
n(Cr–O), in the spectra of the chromium(III) complexes around
440–421 cmꢃ1 and 600–585 cmꢃ1, further confirm the com-
plexation of the ligands to the metal ion.
Table 5 Electronic spectral data (cmꢃ1) of the chromium(III) complexesa
Spectral
bands
Compounds
Transitions
(cmꢃ1
)
Dq B1 b = B/B1 n2/n1
4
[CrCl2(L1)(H2O)2] A2g (F) - 4T2g(F) (16 863) 1684 636 0.69
1.38
1.36
1.37
1.36
4A2g(F) - T1g(F) (23 310)
4
4A2g(F) - T1g(P) (30 959)
4
4
[CrCl(L1)2(H2O)] A2g (F) - 4T2g(F) (17 241) 1721 607 0.66
4A2g(F) - T1g(F) (23 474)
4
4A2g(F) - T1g(P) (32 786)
4
4
[CrCl2(L2)(H2O)2] A2g (F) - 4T2g(F) (16 556) 1654 605 0.65
4A2g(F) - T1g(F) (22 780)
4
4A2g(F) - T1g(P) (32 362)
4
4
[CrCl(L2)2(H2O)] A2g (F) - 4T2g(F) (17 391) 1738 618 0.67
4A2g(F) - T1g(F) (23 752)
4
3.2. Electronic spectral analysis
4A2g(F) - T1g(P) (33 000)
4
The nature of the ligand field around the metal ion and the
geometry of the metal complexes have been deduced from
a
B = complex, B1 = free ion.
c
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3.3. 1H-NMR Spectra
reported on the basis of X-ray powder diffraction studies.22
The results show that the compound belongs to an ‘ortho-
rhombic’ crystal system, having unit cell parameters of a = 9.57,
b = 17.21 c = 21.15 and a maximum deviation of 2y =
0.039–0.0451 and a = 901, b = 901, g = 901.
1
The H-NMR spectra of the ligands were recorded in DMSO-d6
using TMS as the internal standard. The 1H-NMR spectra of the
ligands show a signal at d 10.08–11.03 ppm, which is due to the
NH proton. A singlet observed at 8.84–8.90 ppm may be
assigned to the a-protons of the pyridine ring. The multiplet
observed at 6.97–8.28 ppm may be assigned to the b-protons of
the pyridine ring and the protons of the aromatic ring. The
On the basis of above studies an octahedral environment
around the metal atoms has been proposed (Fig. 2).
spectra also exhibit signals due to the methyl protons attached 3.6. Antimicrobial assay
to the azomethine group at d 2.33–2.50 ppm.
The ligands and their chromium complexes, synthesized through
the conventional as well as the microwave heating method, were
evaluated for their antimicrobial activity against two bacteria
(Bacillus subtilis and Escherichia coli) and two fungi (Candida
albicans and Aspergillus niger). The results are summarized in
Chart 1 and 2. The results were compared with those of the
standard drug streptomycin for bacteria and fluconazole for
fungi. All the ligands and their respective Cr(III) complexes were
found to be sensitive against all the fungal and bacterial strains
tested. The results reveal that there is a considerable increase in
the toxicity of the complexes as compared to the free ligands. The
biological activity of the ligands exhibited a marked enhance-
ment on the coordination with the metal ions against all the
tested bacterial/fungal strains, which shows that metal chelates
are more active than the ligands. This may be explained by
Tweedy’s chelation theory,23 according to which, chelation
reduces the polarity of the central metal atom because of partial
sharing of its positive charge with the ligand, due to which the
lipophilic character of the metal chelate increases and favours its
permeation through the lipid layer of the cell membrane. It has
also been proposed that the ultimate action of the compounds is
the denaturation of one or more proteins of the cell as a result
of which normal cellular processes are impaired,24 and the
deactivation of various cellular enzymes that play vital roles in
the different metabolic pathways of these microorganisms.
3.4. ESR spectral analysis and magnetic moments
The ESR spectra of the 1 : 1 and 1 : 2 chromium(III) complexes,
synthesized by different routes, were recorded at room tem-
perature. These consist of a single broad peak in each case and
from which the Lande splitting factor (‘g’ values) has been
calculated and given in Table 6. For the present complexes, the
g values lie in the range 1.9350–1.9980, with giso (2.0), which
are characteristic of an octahedral geometry.21 The room tem-
perature magnetic moment for the chromium(III) complexes is
slightly less than required. The observed magnetic moment
values of 3.63–3.78 BM and the electronic spectra of the
complexes support the octahedral structure of the complexes.
3.5. X-ray powder diffraction study
The possible lattice dynamics of the finely powdered [CrCl2(L)-
(H2O)2] and [(CrCl(L)2(H2O)] type of products have been
Table 6 ESR spectral data of the chromium(III) complexes
Compound
H0
g value Temp./1C Magnetic moment (m)
[CrCl2(L1)(H2O)2] 3371.12 1.9879 25
[CrCl(L1)2(H2O)] 3354.07 1.9980 25
[CrCl2(L2)(H2O)2] 3463.28 1.9350 25
[CrCl(L2)2(H2O)] 3417.36 1.9610 25
3.70
3.75
3.66
3.74
Fig. 2 Suggested structures for the chromium complexes.
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Chart 1 Antifungal screening data for the ligands (L1H and L2H) and their chromium(III) complexes.
Chart 2 Antibacterial screening data for the ligands (L1H and L2H) and their chromium(III) complexes.
Antifertility test
Sperm dynamics. The sperm motility of the spermatozoa and
Results. Treatment with the ligands (L1H and L2H) and their sperm density in testes and cauda epididymis were decreased
chromium(III) complexes at the dose level of 20 mg kgꢃ1 b.wt, significantly (p r 0.1 to 0.001) after treatment with the ligands
for a period of 60 days, showed the following variation in the (L1H and L2H) and their chromium(III) complexes in all the
different end points.
treated groups Table 8.
Body and organ weights. No significant change in body
weight over the experimental period among the treated groups
Tissue biochemistry. Oral administration of the ligands
was noticed. However, a significant reduction in the weight of and their metal complexes caused a significant reduction in
the testes, epididymis, ventral prostate and seminal vesicle glycogen and sialic acid, whereas the testicular protein and
were noticed after treatment with the ligands (L1H and L2H) cholesterol contents of the reproductive sex organs were
and their chromium(III) complexes Table 7.
increased significantly (p r 0.1 to 0.001) Table 9.
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Table 7 Effect of ligands and their corresponding metal complexes on the reproductive organs weight of male ratsa
Body weight (g)
Initial
Organ weight (mg/100 g b. weight)
Final
Testes
Epididymis
Seminal vesicle
Group
Treatment
Ventral prostate
A
B
C
D
E
F
Control
220.0 15.8
228.0 13.7
233.0 14.1
238.0 13.5
225.0 10.7
220.0 13.1
225.0 14.7
237.0 16.5
245.0 17.5
251.0 15.6
247.0 14.7
235.0 13.4
229.0 13.5
220.0 13.8
1100.0 75.8
950.0 45.0b
936.0 17.7b
900.0 17.0b
870.0 15.0c
840.0 14.8c
670.0 18.6c
520.0 14.1
430.8 15.4b
410.0 15.3c
350.0 16.3c
310.0 14.3c
267.0 13.2c
226.0 11.5c
440.0 15.5
380.0 21.7b
366.0 20.3b
300.0 14.4b
310.0 13.2b
288.0 13.8c
245.0 17.4c
390.0 13.2
305.0 15.5b
292.0 15.2b
250.0 14.9b
245.0 13.9c
233.0 16.6c
210.0 14.8c
L1H
L2H
[CrCl2(L1)(H2O)2]
[CrCl(L1)2(H2O)]
[CrCl2(L2)(H2O)2]
[CrCl(L2)2(H2O)]
G
a
b
c
Mean ꢁ 6 SEM of six animals. Groups B, C, D, E, F, G and H compared with Group A. p r 0.01. p r 0.001.
Table 8 Sperm dynamics and fertility after the administration of ligands and
are dependent on androgen for their growth and function. The
reduction in the weights of these sex accessory organs may be
due to the decreased production of androgen.27
their corresponding metal complexesa
Sperm density
(million/mL)
Sperm
motility %
The decreased sperm density in the testes and cauda
epididymis is an indicator of reduced spermatogenesis, and
the reduced sperm motility may be due to the altered enzymatic
activity of the oxidative phosphorylation process.28 Thus,
a decrease in sperm motility and density after oral administra-
tion of the ligands and their corresponding chromium(III)
complexes may be due to an androgen deficiency, which caused
the impairment in testicular function by altering the enzymatic
activities responsible for spermatogenesis, suggesting thereby
an antiandrogenic effect of these compounds. The decrease in
male fertility could be explained by the fact that the ligands
and their metal complexes acted directly on the testes and
influenced the androgen biosynthesis pathway.29 The ligands
and their chromium(III) complexes also induced biochemical
changes in the testes and sex accessory organs.
Cauda
epididymis
Fertility %
(Cauda
epididymis)
Testes
Group Treatment
A
B
C
D
E
F
Control
L1H
75.1 2.0
4.7 0.9 54.0 3.9 100(+ve)
60.1 1.7c 3.9 0.6b 45.0 3.2b 75(ꢃve)
L2H
58.0 1.5c 3.7 0.5b 47.0 3.2b 78(ꢃve)
[CrCl2(L1)(H2O)2] 45.0 1.5c 2.1 0.5c 40.0 1.2c 80(ꢃve)
[CrCl(L1)2(H2O)] 40.0 1.7c 2.0 0.4c 38.0 1.1c 82(ꢃve)
[CrCl2(L2)(H2O)2] 37.0 1.6b 1.8 0.3c 35.0 1.1c 85(ꢃve)
[CrCl(L2)2(H2O)] 36.7 1.4c 1.5 0.7c 34.0 1.2c 88(ꢃve)
G
a
Mean ꢁ 6 SEM of six animals. Groups B, C, D, E, F, G and H compared
b
c
with Group A. p r 0.01. p r 0.001.
Serum testosterone. A significant reduction (p r 0.1 to 0.001)
in the serum testosterone level was observed after treatment
with the ligands and their chromium(III) complexes Table 9.
The results demonstrate a marked decrease in testicular
glycogen, which may be due to interference during glucose
metabolism.30 Inhibition of glycogen synthesis eventually affects
the protein synthesis and thus inhibits spermatogenesis.31
Sialic acids are concerned with changing the membrane
4. Discussion
The administration of ligands (L1H and L2H) and their corre-
sponding chromium(III) complexes brought about a marked surface of maturing spermatozoa as well as with the develop-
reduction in the weight of testes and other accessory sex ment of their fertilizing capacity.32 Thus, decreased levels of
organs. The reduction in testicular weight may be due to the sialic acid in the testes and sex accessory organs may inhibit the
tubule size, alteration of spermatogenesis with marked fertilizing capacity of sperms.
reduction of gametes production combined with a reduction
The increased testicular cholesterol is attributed to decreased
in the seminiferous diameter in treated animals relative to the concentration of androgen, which results in impaired spermato-
control,25 and inhibition of the steroid biosynthesis of leydig genesis.33 Similarly, the elevation in the testicular protein
cells.26 The testes, epididymis, and other accessory sex organs contents after treatment with the ligands and their metal
Table 9 Biochemical changes in the testes of male rats after treatment with the ligands and their corresponding metal complexesa
Testicular sialic
Testicular protein
(mg gꢃ1
Testicular glycogen
(mg gꢃ1
Testicular cholesterol
(mg gꢃ1
Serum testosterone
Group
Treatment
acid (mg gꢃ1
)
)
)
)
(mg mLꢃ1
2.50 0.50
)
A
B
C
D
E
F
Control
5.08 0.10
4.30 0.08b
4.20 0.08b
3.10 0.07c
3.20 0.30c
2.90 0.10c
2.77 0.08c
250.00 9.80
295.00 10.50b
283.00 10.00b
330.00 8.70c
350.00 7.50c
330.00 10.50c
345.00 9.50c
2.75 0.10
2.30 0.10b
2.12 0.10b
1.90 0.09b
1.80 0.08c
1.60 0.09c
1.55 0.60c
5.38 0.45b
6.80 0.70b
6,70 0.50b
7.50 0.90c
7.70 0.80c
7.40 0.80c
7.20 0.50c
L1H
2.10 0.30b
1.75 0.20b
1.70 0.20b
1.50 0.10c
1.55 0.14c
1.40 0.09c
L2H
[CrCl2(L1)(H2O)2]
[CrCl(L1)2(H2O)]
[CrCl2(L2)(H2O)2]
[CrCl(L2)2(H2O)]
G
a
b
c
Mean ꢁ 6 SEM of six animals. Groups B, C, D, E, F, G and H compared with Group A. p r 0.01. p r 0.001.
c
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9 A. Scozzafava, F. Briganti, M. A. Ilies and C. T. Supuran,
J. Med. Chem., 2000, 43, 292.
10 R. Garg, M. K. Saini, N. Fahmi and R. V. Singh, Transition
Met. Chem., 2006, 31, 362.
complexes may be due to the hepatic detoxification activities
caused by these compounds, which results in an inhibitory
effect on the activities of enzymes involved in the androgen
biotransformation.
11 R. Sharma, S. K. Agarwal, S. Rawat and M. Nagar, Transition
Met. Chem., 2006, 31, 201.
12 H. N. Fox, Science, 1952, 116, 129.
13 R. K. Agarwal and R. K. Sarin, Polyhedron, 1993, 12, 2411.
14 R. K. Agarwal, L. Singh, D. K. Sharmab and R. Singh, Turk. J.
Chem., 2005, 29, 309.
15 R. C. Maurya, P. Patel and D. Sutradhar, Synth. React. Inorg.
Met.-Org. Chem., 2003, 33, 1857.
A marked reduction in testosterone content, in association
with a highly reduced circulating level of this hormone, con-
firmed alterations in the reproductive physiology of the rats.
These results suggested that the ligands (L1H and L2H) and
their chromium(III) complexes exert inhibitory effects on testi-
cular function and lead to the infertility in male rats. Further,
addition of a metal ion to the ligand enhances the activity.
16 A. I. Vogel, A Textbook of Organic Quantitative Analysis,
fifth edn, 2004, p. 243.
4. Conclusions
17 A. I. Vogel, A Textbook of Quantitative Chemical Analysis,
sixth edn, 2006, p. 487.
18 A. I. Vogel, A Textbook of Quantitative Chemical Analysis,
sixth edn, 2000, p. 387.
19 A. Belanger, S. Caron and V. Picard, J. Steroid Biochem. Mol.
Biol., 1980, 13(2), 185.
20 A. C. Fabretti, C. Preti, L. Tassi, G. Tosi and P. Zannini, Aust.
J. Chem., 1986, 39(4), 605.
21 M. M. El-ajaily, A. A. Maihub, S. S. Hudere and S. M.
Ben saber, Asian J. Chem., 2006, 18(4), 2427.
22 S. Shrivastava, N. Fahmi, D. Singh and R. V. Singh, J. Coord.
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23 B. G. Tweedy, Phytopathology, 1964, 55, 910.
24 A. L. Lehninger, Biochemistry, second edn, Worth Publishers,
New York, 1975, p. 513.
Microwave (MW) irradiation is an efficient and environmentally-
benign method to accomplish various inorganic syntheses to
afford products in higher yields in shorter reaction periods.
1-(2-Pyridyl) ethanone isonicotinoyl hydrazone (L1H) and
1-(2-naphthyl) ethanone isonicotinoyl hydrazone (L2H) behave
as monofunctional bidentate ligands with the metal ion under
different reaction conditions. An octahedral geometry for the
Cr(III) complexes have been proposed. The biological screening
data of the ligands and their complexes indicate that the
complexes are more potent than the parent ligands.
Acknowledgements
The authors are thankful to the CSIR, New Delhi, and UGC
(980/5842), New Delhi, for financial assistance.
25 D. Hellengue, K. Rhouma, O. Tebowrbi and M. Sakly, Pol. J.
Environ. Stud., 2003, 12, 557.
26 S. C. Joshi, R. Mathur and N. gulati, Toxicol. Ind. Health,
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