Synthesis, characterization, antimicrobial activity and ultrastructure of the affected bacteria of new quinoline compounds

Article abstract of DOI:10.1007/s11164-016-2474-2

The reaction on 8-hydroxy quinoline-7-aldehyde azo compounds (HL_n) (where n?=?1–5) with 4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one to obtain HL_n (where n?=?6–10) have been characterized by means of TLC, melting point and spectral data, such as IR, ¹H NMR, mass spectra and thermal studies. The X-ray diffraction patterns of two starting materials 8-hydroxy quinoline-7-aldehyde (start 1), 4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one (start 2) and the ligands (HL_5,10) are investigated in powder form. All the ligands have been screened for their antimicrobial activity against four local bacterial species, two Gram-positive bacteria (Bacillus cereus and Staphylococcus aureus) and two Gram-negative bacteria (Escherichia coli and Klebsiella pneumoniae) as well as against four local fungi; Aspergillus niger, Alternaria alternata, Penicillium italicum and Fusarium oxysporium. The results show that the azo ligands (HL_n) (where n?=?1–5) have no antimicrobial activity against bacteria and fungi while most azomethine ligands (HL_n) (where n?=?6–10) are good antibacterial agents against E. coli and K. pneumoniae as well as antifungal agents against P. italicum and A. alternata. The results were compared to standard substances (start 1) and (start 2). Among the azomethine ligands, HL₁₀ was the most effective against the most microorganisms tested. The size of clear zone was ordered as p-(OCH₃?3?2) as expected from Hammett’s constant (σ^R). Also, the ultrastructure study of the affected bacteria confirmed that HL₈ is good antibacterial agent against E. coli and S. aureus.

Full text of DOI:10.1007/s11164-016-2474-2

Res Chem Intermed
DOI 10.1007/s11164-016-2474-2
Synthesis, characterization, antimicrobial activity
and ultrastructure of the affected bacteria of new
quinoline compounds
A. Z. El-Sonbati2 M. I. Abou-Dobara1
•
•
M. A. Diab2 A. A. El-Bindary2 S. G. Nozha2
•
•
Received: 21 October 2015 / Accepted: 1 February 2016
Ó Springer Science+Business Media Dordrecht 2016
Abstract The reaction on 8-hydroxy quinoline-7-aldehyde azo compounds (HLn)
(where n = 1–5) with 4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one to
obtain HLn (where n = 6–10) have been characterized by means of TLC, melting
point and spectral data, such as IR, 1H NMR, mass spectra and thermal studies. The
X-ray diffraction patterns of two starting materials 8-hydroxy quinoline-7-aldehyde
(start 1), 4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one (start 2) and the
ligands (HL5,10) are investigated in powder form. All the ligands have been screened
for their antimicrobial activity against four local bacterial species, two Gram-pos-
itive bacteria (Bacillus cereus and Staphylococcus aureus) and two Gram-negative
bacteria (Escherichia coli and Klebsiella pneumoniae) as well as against four local
fungi; Aspergillus niger, Alternaria alternata, Penicillium italicum and Fusarium
oxysporium. The results show that the azo ligands (HLn) (where n = 1–5) have no
antimicrobial activity against bacteria and fungi while most azomethine ligands
(HLn) (where n = 6–10) are good antibacterial agents against E. coli and K.
pneumoniae as well as antifungal agents against P. italicum and A. alternata. The
results were compared to standard substances (start 1) and (start 2). Among the
azomethine ligands, HL10 was the most effective against the most microorganisms
tested. The size of clear zone was ordered as p-(OCH3 \ CH3 \ H \ Cl \ NO2) as
expected from Hammett’s constant (rR). Also, the ultrastructure study of the
affected bacteria confirmed that HL8 is good antibacterial agent against E. coli and
S. aureus.
S. G. Nozha: Abstracted from her Ph.D. Thesis.
&
A. Z. El-Sonbati
elsonbatisch@yahoo.com
1
2
Botany Department, Faculty of Science, Damietta University, Damietta, Egypt
Chemistry Department, Faculty of Science, Damietta University, Damietta, Egypt
123

A. Z. El-Sonbati et al.
Keywords Azo Schiff bases Á Azo quinoline Á Azomethine Á Antimicrobial
activities Á Ultrastructure
Introduction
Azo dyes constitute one of the largest and most varied groups of synthetic organic
dyes in use today [1]. The major classes of dyes have anthroquinoid, indigoid and azo
aromatic compounds. The chemical structure of azo dyes is comprised of a
conjugated system of double bonds and aromatic rings. All these structures allow
strong p–p* transitions in the UV–visible (UV–Vis) area, with high extinction
coefficients. The drawback of these dyes is that they are not easily degraded by
aerobic bacteria, and with the action of anaerobic or microaerobic reductive bacteria,
they can form toxic or mutagenic compounds such as aromatic amines [2–4].
In addition, Schiff bases are some of the most widely used organic compounds.
They are used as pigments and dyes, catalysts, intermediates in organic synthesis
and as polymer stabilizers [5]. Schiff bases have also been shown to exhibit a broad
range of biological activities, including antifungal, antibacterial, antimalarial,
antiproliferative, anti-inflammatory, antiviral and antipyretic properties [5–8]. It has
been suggested that azomethine linkage (C=N) might be responsible for the
biological activities of Schiff bases [9].
Because of excellent yield and fast reaction, many researchers have developed
azo Schiff base compounds and studied their properties and different applications
[10, 11].
The quinoline nucleus has gathered immense attention among chemists as well as
biologists as it is one of the key building elements for many naturally occurring
compounds [12], while 8-hydroxyquinolines have been explored as a viable drug
discovery platform in many instances. They can exert their physiological properties
through bidentate chelation of metal ions [13].
Different quinoline compounds have been tested against fungi, a range of Gram-
positive and Gram-negative bacteria, including Staphylococcus aureus and
Escherichia coli [14–16]. Lam et al. [14] synthesized and characterized various
derivatives of 8-hydroxyquinoline and studied their potential antibacterial activity
against Gram-positive S. aureus and Gram-negative E. coli. They found that these
quinoline compounds had no effect on E. coli, while showing antibacterial effect
against S. aureus. They found that the synthetic 5,7-dibromo-2-methylquinolin-8-ol
derivative shows a similar minimum inhibitory concentration (MIC) of 6.25 lg/mL
as compared to that of methicillin (3.125 lg/mL) against Staphylococcus aureus.
El-Behery et al. [15] synthesized a Schiff base hydrazone derived from 7-chloro-4-
hydrazinoquinoline and its complexes with Cu(II), Ni(II), Co(II), Fe(III), and
UO2(II) ions. Also, they tested the Schiff base ligand and its metal complexes
against one stain Gram-positive bacteria (S. aureus), Gram-negative bacteria
(E. coli), and yeast (Candida albicans) and they found that, their compounds
exhibited higher antibacterial acivities. Abou-Dobara et al. [16] studied the
antimicrobial activity of azodye ligands) and their complexes [Cu(II)/Ni(II)] against
four local bacterial and fungal species. They reported that these azodye compounds
123

Synthesis, characterization, antimicrobial activity and…
have good antibacterial activity against B. cereus, E. coli and K. pneumoniae, the
complexes were being more toxic than ligands against fungi and Cu2? com-
plexes were being more active than Ni2? complexes against B. cereus, E. coli and
K. pneumoniae.
In view of the facts mentioned above, a new series of azo Schiff bases of
8-hydroxyquinoline-7-aldehyde azodye ligands with 4-amino-1,2-dihydro-2,3-
dimethyl-1-phenylpyrazol-5-one were synthesized and have been evaluated for
their antimicrobial activity. Azodye ligands of 8-hydroxyquinoline-7-aldehyde and
1
its derivatives have been prepared and characterized by elemental analyses, H-
NMR and IR spectra [17].
The aim of the present work is to synthesize 4-{[5-(4-substituted-phenylazo)-8-
hydroxy-quinolin-7-ylmethylene]-amino}-1,5-dimethyl-2-phenyl-1,2-dihydropyra-
zol-3-one derivatives (HLn) (where n = 6–10), and then test the antimicrobial
activity of the HL(1–10) ligands against two Gram-positive and two Gram-negative
bacteria in addition to four different types of fungi. The effect of HL8 on the
ultrastructure of two affected bacteria is also investigated.
Experimental
Materials
4-Amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one and various aniline or p-
substituted anilines were purchased from Aldrich, and the other chemicals were of
analytical grade quality.
Preparation of (5-(40-derivatives phenyldiazo)-8-hydroxy-7-
quinolinecarboxaldehyde) (HL1–5
)
8-Hydroxy-7-quinolinecarboxaldehyde was prepared according to El-Sonbati and
El-Bindary [18]. (5-(40-derivatives phenylazo)-8-hydroxy-7-quinolinecarboxalde-
hyde) HLn where (n = 1–5) prepared according to El-Sonbati et al. [17, 19]
(Table 1). Azo dye ligands (HLn) were prepared from aniline or its p-substituted
derivatives (10 mmol) dissolved in hydrochloric acid (20 mmol/25 ml distilled
Table 1 Physical properties and elemental analysis data of quinoline azodyes ligands HLn (n = 1–5)
Compounda
Yield %
Color
Exp. (calc) %
C
H
N
HL1
HL2
HL3
HL4
HL5
a
55.22
58.38
69.4
Dirty brown
Reddish brown
Dark brown
Light brown
Dark red
66.6 (66.5)
70.0 (70.1)
69.4 (69.3)
61.5 (61.6)
59.7 (59.6)
4.2 (4.2)
4.6 (4.5)
3.9 (4.0)
3.2 (3.2)
3.0 (3.1)
13.5 (13.7)
14.6 (14.4)
15.0 (15.2)
13.3 (13.5)
17.5 (17.4)
76.12
90.67
The analytical data agree satisfactorily with the expected formulae as given in Fig. 1
123

A. Z. El-Sonbati et al.
H2O). The hydrochloric compound was diazotized below 0–5 °C with a solution of
sodium nitrite (0.8 g, 10 mmol, 30 ml distilled H2O). The diazonium chloride was
coupled with an alkaline solution of oxine (1.7 g, 10 mmol) in 20 ml of pyridine.
The crude dye was collected by filtration and was crystallized from DMF, then dried
in a vacuum desiccator over P2O5. The preparation of (HL1–5) is summarized in
Fig. 1.
Preparation of 4-{[5-(4-methoxy-phenylazo)-8-hydroxy-quinolin-7-
ylmethylene]-amino}-1,5-dimethyl-2-phenyl-1,2-dihydro-pyrazol-3-one
(HL6–10
)
Ethanolic solutions of 4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one
(0.1 mol) and [5-(40-derivatives phenyldiazo)-8-hydroxy-7-quinolinecarboxaldehyde)]
(HL1–HL5) (0.1 mol) were refluxed for 4 h over a steam bath. The excess solvent was
removed by evaporation and the concentrated solution was cooled in an ice bath with
stirring. The azo Schiff base (HLn) separated out as a colored powder and was
recrystallized from ethanol, then dried in a vacuum desiccator over P2O5. The purity of
the ligands was checked by TLC. Our synthetic route ofazo Schiff base ligands is shown
in Scheme 1. The color, yield % and analytical data are given in Table 2.
The resulting formed ligands are:
(5-(40-methoxyphenylazo)-8-hydroxy-7-quinolinecarboxaldehyde) (HL1)
(5-(40-methyl phenylazo)-8-hydroxy-7-quinolinecarboxaldehyde) (HL2)
(5-(40-phenylazo)-8-hydroxy-7-quinolinecarboxaldehyde) (HL3)
(5-(40-chlorophenylazo)-8-hydroxy-7-quinolinecarboxaldehyde) (HL4)
(5-(40-nitrophenylazo)-8-hydroxy-7-quinolinecarboxaldehyde) (HL5)
4-{[5-(4-methoxy-phenylazo)-8-hydroxy-quinolin-7-ylmethylene]-amino}-1,5-
dimethyl-2-phenyl-1,2-dihydro-pyrazol-3-one (HL6)
N
O
O
O
N
O
H
O
R
R
H
H
O
H
R
O
O
R
N
N
H
O
O
N
R
O
R
O
H
(C)
(B)
(D)
N
N
H
N
2
0-5 o
C
2
O
R
O
R
1-Diazonium
2-Coupling
P-alkylaniline
N
H
O
O
O
OH
(A)
R=
N=N C6H4X
X= OCH3, CH3, H, Cl , NO2
Fig. 1 Structure for the dimmer and tautomer forms of 8-hydroxy-7-quinoline-carboxaldehyde azodyes
(HL1–5
)
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Synthesis, characterization, antimicrobial activity and…
N
N
R
N
N
N
R
CH3
OH
N
0-5 oC
(Start 2)
4-Amino-1-phenyl-2,3-
N
N
O
N
CH3
1-Diazonium
2-Coupling
P-alkylaniline
N
OH
O
dimethylpyrazolin-5-one
OH
O
Start 1
HL1-5
R= C6H5X
HL6-10
X= OCH3, CH3, H, Cl, NO2
Scheme 1 Synthetic route of azo Schiff base ligands HLn (where n = 6–10)
Table 2 Physical properties and elemental analysis data of quinoline azo Schiff base ligands HLn
(n = 6–10)
Compounda
Yield %
Color
Exp. (calc) %
C
H
N
HL6
HL7
HL8
HL9
HL10
a
39
Pale yellow
Dark yellow
Bright yellow
Dark orange
Dark brown
68.13 (68.29)
70.47 (70.59)
70.01 (70.13)
65.14 (65.26)
63.84 (63.91)
4.64 (4.88)
4.90 (5.04)
4.54 (4.76)
4.12 (4.23)
4.03 (4.14)
16.82 (17.07)
17.37 (17.65)
17.79 (18.18)
16.68 (16.92)
19.08 (19.33)
40.7
43.3
46.6
53
The analytical data agree satisfactory with the expected formulae represented as given in Scheme 1
4-{[5-(4-methyl-phenylazo)-8-hydroxy-quinolin-7-ylmethylene]-amino}-1,5-
dimethyl-2-phenyl-1,2-dihydro-pyrazol-3-one (HL7)
4-{[5-(4-phenylazo)-8-hydroxy-quinolin-7-ylmethylene]-amino}-1,5-dimethyl-2-
phenyl-1,2-dihydro-pyrazol-3-one (HL8)
4-{[5-(4-chlorophenylazo)-8-hydroxy-quinolin-7-ylmethylene]-amino}-1,5-
dimethyl-2-phenyl-1,2-dihydro-pyrazol-3-one (HL9)
4-{[5-(4-nitrophenylazo)-8-hydroxy-quinolin-7-ylmethylene]-amino}-1,5-
dimethyl-2-phenyl-1,2-dihydro-pyrazol-3-one (HL10).
Microbiological investigation
For this investigation, the agar well diffusion method was applied [20]. The
antibacterial activities of the investigated compounds were tested against two local
Gram-positive bacteria (B. cereus and S. aureus) and two local Gram-negative
bacteria (E. coli and K. pneumoniae) on nutrient agar medium. Also, the antifungal
effect was tested against four local fungal species (Aspergillus niger, Alternaria
alternata, Penicillium italicum and Fusarium oxysporium) on DOX agar medium.
The concentrations of each solution were 50, 100 and 150 lg/mL. By using a sterile
123

A. Z. El-Sonbati et al.
cork borer (10 mm diameter), wells were made in agar medium plates previously
seeded with the test microorganism. Then, 200 ll of each compound was applied in
each well. The agar plates were kept at 4 °C for at least 30 min to allow the
diffusion of the compound, then incubated at 30 °C or 37 °C for bacteria and fungi,
respectively. Penicillin and miconazole were used as antibacterial and antifungal
substrates, respectively. The diameters of inhibition zones were determined after
24 h for bacteria and after 7 days for fungi. For electron microscopic study,
overnight cultures of E. coli and S. aureus were subjected to 150 lg/mL of HL8 for
2 h. Non-treated (control) bacteria and DMF-treated bacteria were also included.
After that, bacterial cells were centrifuged and washed with distilled water. The
bacteria were fixed with formalin-glutaraldehyde fixative (4F1G) in 0.1 M phosphate
buffer pH 7.4. After rinsing in the buffer, samples were post-fixed in 2 % OsO4 for
2 h at 4 °C in the same buffer. The cells were washed and dehydrated at 4 °C
through a graded series of ethanol. Cells were then treated with propylene oxide
solution and embedded in a mixture of 1:1 of Epon-Araldite for 1 h. Polymerization
was done in the oven at 65 °C for 24 h. Ultrathin sections (50 lm) were cut on an
ultratome (Model LKB), then mounted on copper grids, double-stained with uranyl
acetate and lead citrate and investigated on a JEOL 100CX (TEM).
Measurements
Elemental microanalyses of the separated ligands chelates for C, H and N were
performed in the Microanalytical Center at Cairo University, Egypt. The analyses
were repeated twice to check the accuracy of the analyzed data. Melting or
1
decomposition points were carried out on a melting point apparatus. The H-NMR
spectrum was obtained with a JEOL FX90 Fourier transform spectrometer with
DMSO-d6 as the solvent and TMS as an internal reference. Infrared spectra were
recorded as KBr pellets using a Pye Unicam SP 2000 spectrophotometer.
Ultraviolet–visible (UV–Vis) spectra of the compounds were recorded in Nujol
mulls using a Unicom SP 8800 spectrophotometer. X-ray diffraction patterns of the
powder forms were recorded on X-ray diffractometer (Philips x’pert) with Ni-
˚
filtered CuKa-radiation (k = 1.5418 A) in the range of diffraction angle (2h° = 4°–
70°). The applied voltage and the tube current are 30 kV and 30 mA, respectively.
Mass spectra were recorded by the EI technique at 70 eV using MS-5988 GS-MS
Hewlett-Packard. Thermal studies were computed on Simultaneous Thermal
Analyzer (STA) 6000 system using thermogravimetric analysis (TGA) method.
Thermal properties of the samples were analyzed in the temperature range from 30
to 800 °C at the heating rate of 10 °C/min under dynamic nitrogen atmosphere.
Morphometric analysis
Morphometric analyses were achieved by using UTHSCSA Image tool software for
Wndows v.3.
123

Synthesis, characterization, antimicrobial activity and…
Statistical analysis
All the data of morphometric analysis are expressed as mean SE. The statistical
significance was evaluated by ANOVA using SPSS v.10 and the individual
comparisons were obtained by the LSD method at 0.05 significance levels.
Results and discussion
Characterization of azo dye ligands HLn (n 5 1–5)
The structure of [5-(40-derivatives phenylazo)-8-hydroxy-7-quinolinecarboxalde-
hyde] HLn where (n = 1–5) using various physico-chemical properties like color,
yield and elemental analysis which are given in Table 1. The elemental analytical
data of ligands agreed well with the calculated values as shown in Fig. 1.
The 1H NMR spectra of quinolone and its alkyl derivatives found two signals in
the *9.10–12.43 and *8.89–9.28 ppm regions assigned to d(OH) and d(CN),
1
respectively. H NMR studies for the free ligands under investigation in DMSO-d6
also show two signals at d 9.3–9.7 and 8.95–9.3 ppm which are similar to those
observed previously [21–24]. Those signals are also assigned to the protons of OH
and CN, respectively, in the spectra of the free ligands under investigation. The
–OH proton signals exhibit fast exchange in the presence of D2O, i.e., it disappeared
in the presence of D2O and this is attributed to the strongly hydrogen bonding as in
Fig. 1. The shifts are in the sequence: p-NO2 [ p-Cl [ H [ p-CH3 [ p-OCH3. In
the 13C NMR spectra, there is no detectable signal for the carbon of carbonyl group
(C=O) at the quinoline ring for the hydrazone form. Also, two broad bands (OH
1
signals) were determined from the azo form of ligands in the H NMR spectrum.
The signal for the N–H proton of ligands for the hydrazone form was not observed
in DMSO-d6. The singlet peaks at 56.32 ppm are due to the methoxy (OCH3)
Fig. 2 The relationship
between Hammett’s substitution
coefficients (rR) versus yield %
of azo Schiff base ligands (HLn,
n = 6–10)
123

A. Z. El-Sonbati et al.
carbons. The aromatic carbons (C=C), hydroxyl rings and aldehydic group are
observed in the extensive 111.5–153.2 ppm range.
The electronic spectra of HLn ligands exhibited bands at 32,500–32,150 cm-1 (CN)
(p–p*), 33,450–33,340 cm-1 (H-bonding and association), 40,038–39,460 cm-1
(phenyl) (Ph–Ph*, p–p*) [20] and 29,340–29,230 cm-1 transition of phenyl rings
overlapped by composite broad (p–p*) of the azo structure. The band due to the
n ? p* transition obtained in the visible region is associated mainly with the color of
the respective compound [23]. The analytical data and the spectral properties of the
ligands are consistent with their structure in Fig. 1.
Fig. 3 X-ray diffraction pattern for two starts: a start 1 and b start 2
Fig. 4 X-ray diffraction pattern for (HL5 and HL10) ligand derivatives: a HL5 and b HL10
123

Synthesis, characterization, antimicrobial activity and…
Characterization of azo Schiff base ligands HLn (n 5 6–10)
The ligands (HL6–10) were prepared by stirring an appropriate amount of reactive p-
derivatives 8-hydroxy quinoline-7-aldehyde (azo compounds) with the correspond-
ing 4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one in ethanol. The
formed Schiff base azo ligands (HL6–10) were characterized with respect to their
composition by elemental and spectral analysis. All five ligands HL6–10, gave
Fig. 5 Mass spectrum of HL2 ligand
Scheme 2 Fragmentation patterns of HL2 ligand
123

A. Z. El-Sonbati et al.
Fig. 6 Mass spectrum of a HL6 ligand, b HL8 ligand, c HL9 ligand
123

Synthesis, characterization, antimicrobial activity and…
493
(a)
(b)
(c)
Scheme 3 Fragmentation patterns of a HL6 ligand, b HL8 ligand, c HL9 ligand
123

A. Z. El-Sonbati et al.
(a) 100
80
60
40
20
0
HL1
HL2
HL3
HL4
HL5
100
200
300
400
500
600
Temperature °C
(b) 100
80
60
40
20
0
HL6
HL7
HL8
HL9
HL10
100 200 300 400 500 600 700 800
Temperature °C
Fig. 7 Thermogravimetric analysis (TGA) for ligands. a HL1–5 and b HL6–10
satisfactory elemental analyses (Table 2). Detailed solution and solid state studies
of these ligands were carried out to establish their geometry.
As shown in Table 2, the values of yield % is related to the nature of the
p-substituent as they increase according to the following order p-(NO2 [
Cl[ H [ CH3 [ OCH3). This can be attributed to the fact that the effective charge
experienced by the d-electrons increased due to the electron-withdrawing p-
substituent (HL9 and HL10) while it decreased with the electron-donating character
of (HL6 and HL7). This is in accordance with that expected from Hammett’s
substitution coefficients (rR), as in Fig. 2 the yield % values correlate with (rR), and
it is clear that all these values increase with increasing rR [22–26].
123

Synthesis, characterization, antimicrobial activity and…
Table 3 Thermal decomposition data for azo quinoline ligands (HL1–5
)
Compound
Temp.
range (°C)
Found mass
loss (calc) (%)
Assignment
HL1
35–291
291–389
389–620
[620
27.35 (24.72)
31.95 (33.80)
40.99 (41.0)
–
Evaluation of CO ? loss of OH & OCH3
Decomposition of C6H4N2
Decomposition of C9H4N
Ash
HL2
HL3
120–400
400–620
[620
24 (23.36)
Decomposition of C4H6N
Decomposition of C4H7N2O2
Loss of 8C
40.89 (39.52)
34.33 (32.98)
10.64 (10.09)
56.8 (56.62)
OH (6.13)
134–305
305–618
Evaluation of CO
Loss of OH ? decomposition of C9H4N2
C9H4N2 (50.49)
32.56 (32.45)
3.6 (4.17)
[620
Decomposition of C6H4N
Decomposition of CH
HL4
HL5
55–202
202–423
423–620
[620
23.8 (22.8)
48 (49.9)
Decomposition of C3H3O2
Decomposition of C6H6N3Cl
Loss of 6C
24.64 (23.12)
28.43 (28.22)
CO (8.68)
170–299
Evaluation of CO ? Loss of OH & NO2
OH (5.27)
NO2 (14.27)
39.30 (39.09)
32.27 (32.27)
299–620
Decomposition of C9H4N
Decomposition of C6H4N2
[620
X-ray diffraction analysis
The X-ray diffraction patterns (XRD) of 8-hydroxy quinoline-7-aldehyde (start 1),
4-amino-1,2-dihydro-2,3-dimethyl-1-phenylpyrazol-5-one (start 2) and the ligands
(HL5 and HL10) in powder form are shown in Figs. 3 and 4, respectively. Broad
peaks for start 1 at 2h = 16° and 23° indicating an amorphous structure (Fig. 3a).
Many peaks are observed which indicate the polycrystalline nature of start 2
(Fig. 3b). According to Fig. 4, the XRD patterns of (HL5 and HL10) show that the
peak appeared at 2h = 16° for HL5 is a mixture of amorphous and polycrystalline
structures (Fig. 4a), while HL10 is completely amorphous with two peaks appearing
at 2h = 24° and 45° (Fig. 4b).
Mass spectra
The electron impact mass spectrum of HL2, HL6, HL8 and HL9 ligands were
recorded and investigated at 70 eV of electron energy. For HL2, It is obvious that
the molecular ion peaks are in good agreement with their suggested empirical
formulae as indicated from the elemental analysis (Table 1). The mass spectrum
123

A. Z. El-Sonbati et al.
Table 4 Thermal decomposition data for azo Schiff base quinoline ligands (HL6–10
)
Compound
Temp.
range (°C)
Found mass
loss (calc) (%)
Assignment
HL6
35–46
2 (2.64)
92.357 (92.48)
4.685 (4.878)
9.22 (8.82)
Loss of CH
46–570
Decomposition of C25H23N6O3
Loss of 2C
570–800
100–240
240–590
590–800
50–170
HL7
HL8
HL9
HL10
Decomposition of C3H6
Decomposition of C23H14N6O2
Decomposition of C2H4
Loss of CH
85.12 (85.29)
6.28 (5.88)
2.97 (2.81)
170–550
550–800
105–170
170–580
580–800
120–200
200–580
580–800
83.26 (83.11)
13.2 (14.06)
4.82 (5.24)
Decomposition of C21H16N6O2
Decomposition of C5H5
Decomposition of C2H2
Decomposition of C20H14N6O2Cl
Decomposition of C5H5
Decomposition of C3H3
Decomposition of C17H4N7O4
Decomposition of C7H14
82.3 (81.66)
12.3 (13.09)
7.5 (7.69)
72.79 (72.97)
19.47 (19.33)
Table 5 The values of the Ed,
of HLn (where n = 1–10)
according to the Arrhenius
relationship
Compound
Ed (kJ mol-1
)
HL1
HL2
HL3
HL4
HL5
HL6
HL7
HL8
HL9
HL10
22.91583
40.12523
142.1689
29.44023
143.2887
50.80982
51.81592
18.60734
44.23005
40.58675
fragmentation mode of ligand (HL2) shows the exact mass of 291 corresponding to
the formula C17H13N3O2 (Fig. 5). The ion of m/z = 291 undergoes fragmentation to
a stable peak at m/z = 263 by losing C=O atoms (structure I) as shown in
Scheme 2. The loss of CHO leads to the fragmentation with m/z = 234 (structure
II). The loss of C5H5 atoms leads to the fragmentation with m/z = 169 (structure
III). The loss of C2H2 atoms leads to the fragmentation with m/z = 143 (structure
IV). A breakdown of the backbone of HL2 ligand gives the fragments (V) and (VI).
It is obvious that the molecular ion peaks for ligands (HL6, HL8 and HL9) are in
good agreement with their suggested empirical formula as indicated from the
123

Synthesis, characterization, antimicrobial activity and…
HL1
HL
2
-12
-13
-14
-15
-16
-12
-14
220 - 295 oC
105 - 217 oC
-16
2.1x10-3 2.2x10-3 2.3x10-3 2.4x10-3 2.5x10-3 2.6x10-3
1.8x10-3 1.8x10-3 1.8x10-3 1.9x10-3 1.9x10-3 2.0x10-3
1/T (K-1)
1/T (K-1)
-12
-13
-14
-15
-12
HL
3
HL
4
-13
-14
-15
100 - 180 oC
120 - 190 oC
-16
2.3x10-3
2.4x10-3
2.5x10-3
2.6x10-3
2.2x10-3
2.3x10-3
2.4x10-3
2.5x10-3
1/T (K-1)
1/T (K-1)
HL
5
-13
-14
-15
-16
170 - 290 oC
1.9x10-3
2.0x10-3
2.1x10-3
2.2x10-3
1/T (K-1)
(a)
Fig. 8 Coats–Redfern (CR) of azo ligands a (HLn, n = 1–5), b (HLn, n = 6–10)
123

A. Z. El-Sonbati et al.
-13.0
-13.5
-14.0
-14.5
-13.0
-13.5
-14.0
-14.5
HL7
HL6
107-489oC
140-490oC
0.0015
0.0018
1/T (K-1)
0.0021
0.0024
0.0014 0.0016 0.0018 0.0020 0.0022
1/T (K-1)
-13
-14
-15
-16
-13.2
-13.5
-13.8
-14.1
-14.4
-14.7
HL9
HL8
120-480oC
122-450oC
0.0014 0.0016 0.0018 0.0020 0.0022
1/T (K-1)
0.0015
0.0018
0.0021
0.0024
1/T (K-1)
HL10
-13.0
-13.5
-14.0
-14.5
132-480oC
0.0014
0.0016
0.0018
1/T (K-1)
(b)
0.0020
0.0022
Fig. 8 continued
elemental analysis (Table 2). The mass spectrum fragmentation mode of ligand
(HL6) shows the exact mass of 493 corresponding to the formula C28H25N6O3
(Fig. 6a). The ion of m/z = 493 undergoes fragmentation to a stable peak at
m/z = 279 by losing C12H12N3O atoms (structure I) as shown in Scheme 3a. The
loss of C6H7O leads to the fragmentation with m/z = 185 (structure II). The loss of
123

Synthesis, characterization, antimicrobial activity and…
HL2
HL1
0
-1
-2
-3
0.0
-0.5
-1.0
-1.5
220-295 °C
105-217 °C
-2.0
-30
-15
0
15
30
-60
-40
-20
0
20
40
60
θ (K)
θ (K)
0.0
-0.5
-1.0
-1.5
-2.0
-2.5
0.0
-0.5
-1.0
-1.5
-2.0
HL4
HL3
100-180 °C
15 30
120-190 °C
-30
-15
0
-30
-15
0
15
30
θ (K)
θ (K)
0.0
-0.5
-1.0
-1.5
-2.0
-2.5
HL5
170-290 °C
-60
-40
-20
0
20
40
θ (K)
(a)
Fig. 9 a Horowitz–Metzger (HM) of azo ligands (HLn, n = 1–5). b Horowitz–Metzger (HM) of azo
Schiff base ligands (HLn, n = 6–10)
123

A. Z. El-Sonbati et al.
HL7
HL6
-0.3
-0.6
-0.9
-1.2
-1.5
-0.3
-0.6
-0.9
-1.2
-1.5
140-490 °C
100 150
107-489 °C
-150 -100 -50
0
50
-150 -100 -50
0
50 100 150
θ (K)
θ (K)
-0.3
-0.6
-0.9
-1.2
-1.5
-1.8
-2.1
HL8
HL9
-0.3
-0.6
-0.9
-1.2
-1.5
122-450 °C
100 150
120-480 °C
-150 -100 -50
0
50
-150 -100 -50
0
50 100 150
θ (K)
θ (K)
HL10
-0.3
-0.6
-0.9
-1.2
-1.5
132-480 °C
-150
-100
-50
0
50
100
150
θ (K)
(b)
Fig. 9 continued
C3H5 atoms leads to the fragmentation with m/z = 142 (structure III). A breakdown
of the backbone of HL6 ligand gives the fragments (IV) and (V).
The mass spectrum fragmentation mode of ligand (HL8) shows the exact mass of
462 corresponding to the formula C27H22N6O2 (Fig. 6b). The ion of m/z = 462
undergoes fragmentation to a stable peak at m/z = 249 by losing C12H12N3O atoms
(structure I) as shown in Scheme 3b. The loss of C6H5N2 leads to the fragmentation
123

Synthesis, characterization, antimicrobial activity and…
with m/z = 144 (structure II). A breakdown of the backbone of HL8 ligand gives
the fragments (III) and (IV).
The mass spectrum fragmentation mode of ligand (HL9) shows the exact mass of
496.5 corresponding to the formula C27H21N6O2 Cl (Fig. 6c). The ion of m/z = 496.5
undergoes fragmentation to a stable peak at m/z = 445 by losing C4H4 atoms
(structure I) as shown in Scheme 3c. The loss of C7H5N3O leads to the fragmentation
with m/z = 299 (structure II). The loss of CH3 atoms leads to the fragmentation with
m/z = 283 (structure III). The loss of C6H5N2Cl atoms leads to the fragmentation with
m/z = 144 (structure IV). A breakdown of the backbone of HL9 ligand gives the
fragments (V) and (VI).
Thermogravimetric analysis
The TGA curves of azo (HL1–5) and azo Schiff base ligands (HL6–10) are shown in
Fig. 7(a and b), respectively. It is clear from these figures that the change of azo
group to azomethine strongly affects the decomposition behavior. The detailed
analysis of the behavior of decomposition for both azo (HL1–5) and azo Schiff base
Table 6 Thermodynamic activation energy for all ligands (HL1–10
)
Compounda Temp.
range
Method Parameter
Ea
(kJ mol-1
A (s-1
)
DS* (J mol-1 DH*
DG*
(°C)
)
K-1
)
(kJ mol-1
)
(kJ mol-1
)
HL1
HL2
HL3
HL4
HL5
HL6
HL7
HL8
HL9
HL10
a
220–295
CR
155
156
69
1.06E?13 -3.38E-01
3.96E?13 1.06E?01
6.34E?05 -1.37E?02
5.69E?06 -1.19E?02
9.76E?08 -7.55E?01
2.38E?10 -4.90E?01
1.75E?08 -9.01E?01
4.13E?09 -6.38E?01
1.31E?07 -1.13E?02
1.66E?09 -7.27E?01
4.46E-03 -2.96E?02
1.93E-01 -2.64E?02
2.70E-03 -2.99E?02
8.97E-02 -2.70E?02
9.40E-02 -2.70E?02
3.52E?00 -2.40E?02
2.81E-03 -2.99E?02
1.30E-01 -2.67E?02
5.69E-03 -2.93E?02
1.74E-01 -2.65E?02
150
150
146
125
120
119
113
122
117
146
139
181
173
177
170
171
164
178
170
179
172
HM
CR
152
105–217
100–180
120–190
170–290
140–490
107–489
122–450
120–480
132–480
65.4
68.6
87.6
92.7
83.3
90.1
89.5
103
HM
CR
72.3
91
HM
CR
96.1
86.8
93.7
93.7
107
11.8
22.5
10.8
20.3
24.9
34.3
11.3
21.8
13.6
23.2
HM
CR
HM
CR
6.9
HM
CR
17.6
6.01
15.6
20.3
29.7
6.57
17.1
8.76
18.4
HM
CR
HM
CR
HM
CR
HM
The number corresponds to that used in Tables 1 and 2
123

A. Z. El-Sonbati et al.
(HL6–10) ligands are presented in Tables 3 and 4, respectively. For azo ligands
HL1–5, the TGA curves shown in Fig. 7a reveal that the decomposition process
starts at a temperature in the range 150–200 °C depending on the function group,
while in the first stage of heating, HL1 (p-OCH3) losses about 4.873 % of its weight,
this loss can be attributed to the loss of moisture adsorbed in the matrix [22]. TGA
curves of azo Schiff base ligands HL6–10 shows three steps of decomposition as
shown in Fig. 7b. The decomposition process starts at temperature in the range
35–240 °C depending on the function group.
More results about the degradation process can be extracted from the TGA curves
as follow. The rate constant of the thermal degradation (K) is given by the Arrhenius
relationship [22]:
Table 7 Antibacterial activity data of HLn (where n = 6–10)
Compound
Conc.
(lg/mL)
Gram-positive bacteria
Gram-negative bacteria
Bacillus
cereus
Staphylococcus
aureus
Escherichia
coli
Klebsiella
pneumoniae
Start 1
Start 2
HL6
50
100
150
50
-ve
-ve
1
1
2
2
-ve
-ve
1
2
2
2
-ve
1
-ve
-ve
-ve
-ve
-ve
-ve
-ve
1
2
100
150
50
-ve
-ve
-ve
-ve
-ve
-ve
1
2
2
2
-ve
-ve
-ve
1
2
100
150
50
3
2
HL7
2
-ve
2
100
150
50
2
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
1
1
3
1
HL8
1
-ve
3
-ve
-ve
-ve
-ve
-ve
2
100
150
50
1
1
3
HL9
-ve
1
2
100
150
50
2
-ve
-ve
-ve
-ve
2
2
HL10
Penicillin
3
1
100
150
50
3
1
4
1
1
-ve
-ve
-ve
100
150
3
2
3
3
2
3
The results were recorded as the diameter of inhibition zone (mm)
123

Synthesis, characterization, antimicrobial activity and…
lnK ¼ lnA À Ed=RT
where R is a gas constant (=8.314 JK-1 mol-1) and Ed the thermal activation
energy of decomposition. The values of Ed for HL1–5 azo ligands and azo Schiff
base ligands HL6–10 can be obtained by plotting ln K as a function of 1/T. The
values of Ed of HLn (=-R.slope) are found to be in the range (22.92–143.28) for
HL1–5 and in the range (18.6–51.8) for HL6–10, respectively, as shown in Table 5.
The high values of Ed for HL1–5 than HL6–10 indicate that the azo ligands are more
stable than the Schiff base ligands.
Calculation of activation thermodynamic parameters
The thermodynamic activation parameters of decomposition processes of the azo
(HL1–5) and azo Schiff base ligands (HL6–10), namely activation energy (Ea),
enthalpy (DH*), entropy (DS*), and Gibbs free energy change of the decomposition
(DG*), are evaluated graphically by employing the Coast–Redfern [27] and
Horowitz–Metzger [28] methods.
Coast–Redfern equation
The Coast–Redfern equation, which is a typical integral method, can be represented
as:
ꢀ
ꢁ
Z
Z
a
T2
dx
A
Ea
¼
exp À
dt
ð1Þ
n
ð1 À aÞ
/
RT
0
T1
For the convenience of integration, the lower limit T1 is usually taken as zero.
This equation on integration gives:
Fig. 10 Antibacterial activity data of azo Schiff base ligands (HLn, n = 6–10) and its two starting
materials, namely 8-hydroxy-7-formylquinoline (start 1) and 4-amino-1-phenyl-2,3-dimethylpyrazolin-5-
one (start 2), against Escherichia coli. Inhibition zone was recorded as mm
123

A. Z. El-Sonbati et al.
Fig. 11 Effect of HL8 and HL10
on growth of E. coli on nutrient
agar using concentration
I = 100 lg/mL, II = 150 lg/mL
Fig. 12 Effect of HL7 and HL10
on growth of K. pneumoniae on
nutrient agar using
concentration = 100 lg/mL
ꢂ
ꢃ
ꢂ
ꢃ
lnð1 À aÞ
Ea
AR
ln À
¼ À
þ ln
ð2Þ
T2
RT
/Ea
A plot of the left-hand side (LHS) against 1/T was drawn for the azo ligands
(HL1–5) and azo Schiff base ligands (HL6–10) in Fig. 8a and b, respectively. Ea is the
energy of activation in J mol-1 and calculated from the slope and A in (s-1) from
the intercept value. The entropy of activation (DS*) in (J mol-1 K-1) was
calculated by using the equation:
123

Synthesis, characterization, antimicrobial activity and…
4.0
3.5
3.0
2.5
2.0
1.5
1.0
0.5
0.0
Fig. 13 The relationship
between Hammett’s substitution
coefficients (rR) versus the
inhibition zone (mm) of ligands
(HLn, n = 6–10) (in cases of
E. coli at concentrations 50 lg/mL
(a) and 150 lg/mL (b))
HL10
HL9
HL6
HL7
HL8
0.0
-0.4
-0.2
0.2
0.4
0.6
0.8
1.0
Hammett's substituent coefficient (σR)
(a)
4.5
4.0
3.5
3.0
2.5
2.0
HL10
HL7
HL8
HL6
HL9
-0.4
-0.2
0.0
0.2
0.4
0.6
0.8
1.0
Hammett's substituent coefficients (σR)
(b)
ꢂ
ꢀ
ꢁꢃ
Ah
DSÃ ¼ 2:303 log
R
ð3Þ
kBTs
where kB is the Boltzmann constant, h is the Plank’s constant and Ts is the TG peak
temperature.
Horowitz–Metzger equation
The Horowitz–Metzger equation is illustrative of the approximation methods. These
authors derived the relationship:
123

A. Z. El-Sonbati et al.
Table 8 Antifungal activity data of HLn (where n = 6–10)
Compound
Conc.
(lg/mL)
Aspergillus
niger
Fusarium
oxysporum
Alternaria
alternata
Penicillium
italicum
Start 1
50
100
150
50
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
-ve
1
1
7
2
-ve
-ve
-ve
2
2
2
2
1
Start 2
HL6
5
3
100
150
50
-ve
-ve
-ve
5
2
-ve
-ve
-ve
-ve
-ve
1
2
4
100
150
50
2
-ve
-ve
-ve
7
-ve
5
HL7
100
150
50
1
-ve
-ve
-ve
3
1
HL8
-ve
-ve
-ve
-ve
-ve
7
5
100
150
50
3
-ve
1
HL9
-ve
3
100
150
50
-ve
1
-ve
-ve
1
HL10
-ve
7
3
100
150
50
2
-ve
2
7
1
Miconazole
5
1
100
150
3
3
6
1
4
3
6
2
The results were recorded as the diameter of inhibition zones (mm)
"
#
1Àn
1 À ð1 À aÞ
1 À n
Eah
2:303RTs2
log
¼
;
for n ¼ 1
ð4Þ
when n = 1, the LHS of Eq. (4) would be log[-log(1 - a)] as shown in Fig. 9a and
b, respectively, for (HL1–5) and (HL6–10). For a first-order kinetic process, the
Horowitz–Metzger equation may be written in the form:
ꢂ
ꢀ
ꢁꢃ
Wa
Eah
2:303RTs2
log log
¼
À log 2:303
ð5Þ
Wc
where h = T - Ts, wc = wa - w, wa = mass loss at the completion reaction;
w = mass loss up to time t. The plot of log [log (wa/wc)] versus h was drawn and
123

Synthesis, characterization, antimicrobial activity and…
Fig. 14 Antifungal activity data of azo Schiff base ligands (HLn, n = 6–10) and its two started materials,
namely 8-hydroxy-7-formylquinoline (start 1) and 4-amino-1-phenyl-2,3-dimethylpyrazolin-5-one (start
2) against Alternaria alternata. Inhibition zone was recorded as mm
Table 9 Morphometric analysis of bacterial cells treated with HL8
Treatment E. coli
Cell length
S. aureus
Cell width
Distance between cell
wall and cell membrane
Cell diameter Cell wall
thickness
Control
DMF
HL8
474.7 30.7 243.3 5.5
465.8 11.4 249.3 2.8
17.6 1.9
19.5 0.6
334.9 18.7 14.6
1
274.2 16*
300.8 3.8
19 1.1*
16.8 0.6
433 74.8 199.9 13.1* 22.5 2.4
* Significantly different at 0.05 level
found to be linear from the slope from which Ea was calculated. The pre-exponential
factor, A, calculated from equation:
Ea
RTs2
A
h
ꢄ
ꢅi
¼
ð6Þ
Ea
RTs
/ exp À
The entropy of activation, DS*, is calculated from Eq. (3). The enthalpy
activation, DH*, and Gibbs free energy, DG*, are calculated from:
DHÃ ¼ Ea À RT
ð7Þ
123

A. Z. El-Sonbati et al.
Fig. 15 Transmission electron micrograph of normal S. aureus illustrating different size of cocci cells,
normal thin electron dense cell walls, homogenous electron dense cytoplasm, normal division and
numerous ribosomes (asterisks). (Specimen fixed in 4F1G solution and double-stained with uranyl acetate
and lead citrate; original magnification 97500)
DGÃ ¼ DHÃ À TDSÃ
ð8Þ
The calculated values of Ea, A, DS*, DH* and DG* for the decomposition steps
for azo ligands (HL1–5) and azo Schiff base ligands (HL6–10) are summarized in
Table 6. The entropy of activation energies reflects the thermal stability of the
compounds. The entropy of activation is found to have negative values in the HLn
(where n = 1–10), which indicate that the decomposition reactions proceed
spontaneously [29, 30]. The value of DG* is positive, considered as a favorable
or spontaneous reaction.
From the values of the energy of activation (Ea) of all the ligands (HL1–10), it is
observed that the azo ligands (HL1–5) are more stable than the azo Schiff base
ligands (HL6–10).
Microbiological investigation
The Gram-negative rods have been implicated in infections as diverse as pneumonia to
ear infections. One of the more common Gram-negative rods that cause disease in
humans is E. coli. According to Bailey and Scott’s ‘‘Diagnostic Microbiology,’’ E. coli
123

Synthesis, characterization, antimicrobial activity and…
Fig. 16 Transmission electronic micrograph of S. aureus treated with DMF illustrating decreased size of
cocci cells, electron dense boundary cell wall increased in thickness, numerous ribosomes (single
asterisks), electron dense granules (double asterisks) and increased number of cells forming division
septa. (Specimen fixed in F1G solution and double-stained with uranyl acetate and lead citrate; original
4
magnification 97500)
is by far the most common cause of uncomplicated community-acquired urinary tract
infections and is frequently seen in wound infections. Certain strains of E. coli (such as
E. coli O157:H7) can cause serious diarrhea and kidney damage [31, 32].
The antimicrobial activity of HLn (where n = 1–10) ligands were tested against
bacteria and fungi; we used more than one test organism to increase the chance of
detecting their antimicrobial activities. The used organisms in the present
investigations included two Gram-positive bacteria (B. cereus and S. aureus) and
two Gram-negative bacteria (E. coli and K. pneumoniae) in addition to four species
of fungi; (A. niger, P. italicum, A. alternata and F. oxysporium).
None of the used azo ligands (HLn) (where n = 1–5) were found to have any
antibacterial activity. However, the used azo Schiff base ligands (HLn) (where
n = 6–10) were found to have antibacterial activity. The results of the antibacterial
activities of the synthesized azo Schiff base ligands (HL6–10) are recorded in
Table 7.
According to Table 7, HL6–10 have antibacterial activity against E. coli (inhibition
zone of HL6 = 2 and 3 mm at concentrations = 50 and 100 lg/mL, respectively),
(inhibition zone of HL7 = 2 mm at concentrations = 50 and 100 lg/mL
and = 3 mm at concentration = 150 lg/mL), (inhibition zone of HL8 = 3 mm at
123

A. Z. El-Sonbati et al.
Fig. 17 Transmission electronic micrograph of S. aureus treated with ligand (HL8) illustrating cocci
cells, electron lucent or damaged boundary cell wall, numerous ribosomes (single asterisks), electron
dense granules (double asterisks), granular cell contents appear to be exuding from the damaged
membrane (triple asterisks), vacuoles of electron lucent regions are formed in the cytoplasm (four
asterisks). (Specimen fixed in 4F1G solution and double-stained with uranyl acetate and lead citrate;
original magnification 97500)
concentrations = 100 and 150 lg/mL), (inhibition zone of HL9 = 2 mm at concen-
trations = 50, 100 and 150 lg/mL) and (inhibition zone of HL10 = 3 mm at
concentrations = 50 and 100 lg/mL and = 4 mm at concentration = 150 lg/mL).
Addition to that K. pneumoniae was also weekly affected by HL6–10 (inhibition zone
of HL6 = 1 mm and HL9 = 2 mm at concentration = 150 lg/mL), (inhibition zone
of HL7 = 2 mm and = 1 mm at concentrations = 100 and 150 lg/mL, respec-
tively) and (inhibition zone of HL10 = 1 mm at concentrations = 50, 100 and
150 lg/mL, respectively) as shown in Fig. 10. HL6–10 ligands were found to have
antibacterial activity against E. coli than against K. pneumoniae, and HL10 has high
activity more than other ligands as shown in Figs. 11 and 12. HL10 is more active than
penicillin against E. coli and K. pneumoniae, whereas the HLn and their derivatives
have no effect against Gram-positive bacteria.
As shown in Table 7, the values of inhibition zones for ligands (HL6–10) are
related to the nature of the p-substituent as they increase according to the following
order: p-(NO2 [ Cl [ H [ CH3 [ OCH3) [25]. This can be attributed to the fact
that the effective charge experienced by the d-electrons increased due to the
electron-withdrawing p-substituent (HL9 and HL10) while it decreased by the
123

Synthesis, characterization, antimicrobial activity and…
Fig. 18 Transmission electronic micrograph of normal E. coli showing rod shaped cells, normal smooth
continuous cell wall and cell membrane, homogenous electron dense cytoplasm, normal electron lucent
region between cell wall and cell membrane and normal cell division (single asterisks). (Specimen fixed
in 4F1G solution and double stained with uranyl acetate and lead citrate; original magnification 97500)
electron-donating character of (HL6 and HL7). This is in accordance with that
expected from Hammett’s substitution coefficients (rR) as shown in Fig. 13a, b.
Correlating the values of inhibition zone (mm) with rR, it is clear that these values
increase with increasing rR. It is important to note that the existence of a methyl
and/or methoxy group enhances the electron density on the coordination sites and
simultaneously decreases the values of inhibition zone.
The results in Table 7 reveal that the azo Schiff base ligands are moderately toxic
against Gram-positive bacteria. Both ligands (HL6–10) are more active against
E. coli and K. pneumoniae.
Patel [33] found that 5-(4-N,N-diethylaminosulfonly phenylazo)-8-hydroxy
quinoline and their metal chelates have moderate and high toxicity against some
fungi including Aspergillus niger and Penicillium expansum. Our results of the
antifungal activities of the azo Schiff base ligands (HL6–10) are recorded in Table 8.
The results of the examination of antifungal activity of HL6–10 ligands revealed that
the ligands have moderate toxicity against fungi.
The values of inhibition zone for ligands (HL6–10) are shown in Table 8, HL6–10
have antifungal activity against A. alternata (inhibition zone of HL6 = 5 mm at
concentration = 100 lg/mL), (inhibition zone of HL7 and HL9 = 7 mm at
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A. Z. El-Sonbati et al.
Fig. 19 Transmission electronic micrograph of E. coli treated with DMF showing rod shaped cells,
normal smooth continuous cell wall and cell membrane, homogenous electron dense cytoplasm and
normal electron lucent region between cell wall and cell membrane. (Specimen fixed in 4F1G solution and
double-stained with uranyl acetate and lead citrate; original magnification 97500)
concentration = 150 lg/mL) and (inhibition zone of HL10 = 7 mm at concentra-
tions = 100 and 150 lg/mL) as shown in Table 8 and Fig. 14. In addition to that P.
italicum was also affected by HLn (inhibition zone of HL6 = 4 and 2 mm at
concentrations = 50 and 100 lg/mL, respectively), (inhibition zone of HL7 = 5 mm
at concentration = 50 lg/mL and = 1 mm at concentrations = 100 and 150 lg/mL),
(inhibition zone of HL8 = 5 and 3 mm at concentrations = 50 and 100 lg/mL,
respectively), (inhibition zone of HL9 = 1 mm at concentrations = 50 and 150 lg/mL)
and (inhibition zone of HL10 = 3, 2 and 1 mm at concentrations = 50, 100 and
150 lg/mL, respectively). HL6–10 have low antifungal activity against F. oxysporum
(inhibition zone of HL7 and HL10 = 1 mm at concentration = 100 lg/mL) and
(inhibition zone of HL8 and HL9 = 3 mm at concentrations = 150 and 100 lg/mL,
respectively).
The results of the TEM comparative study on the morphology and internal
ultrastuctures; of normal and HL8-treated E. coli and S. aureus provided strong
evidence that HL8 is stressful and toxic for the bacterial cells. In general, treatment
of E. coli with HL8 resulted in significant morphometric changes, whilst similar
exposure of S. aureus produced no significant morphometric changes (Table 9).
However, HL8 prevents S. aureus’ ability to adapt by reducing size, as shown in
Fig. 15, and thickening the cell walls was seen with DMF treatment (Table 9). In
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