Surface and biocidal activity of some synthesized metallo azobenzene isothiouronium salts

Article abstract of DOI:10.1016/j.bmc.2006.08.021

A novel series of azobenzene isothiouronium salts of different alkyl chains (propyl, hexyl and dodecyl) were synthesized by reaction of 4-((4-methylphenyl)azo)phenol with 1,3-dibromopropane, 1,6-dibromohexane and 1,12-dibromododecane, respectively. These

Full text of DOI:10.1016/j.bmc.2006.08.021

Bioorganic & Medicinal Chemistry 14 (2006) 8661–8665
Surface and biocidal activity of some synthesized metallo
azobenzene isothiouronium salts
A. M. Badawi, E. M. S. Azzam* and S. M. I. Morsy
Egyptian Petroleum Research Institute, Applied Surfactants Laboratory, Petrochemicals Department, Cairo, Egypt
Received 18 June 2006; revised 10 August 2006; accepted 15 August 2006
Available online 14 September 2006
Abstract—A novel series of azobenzene isothiouronium salts of different alkyl chains (propyl, hexyl and dodecyl) were synthesized
by reaction of 4-((4-methylphenyl)azo)phenol with 1,3-dibromopropane, 1,6-dibromohexane and 1,12-dibromododecane, respec-
tively. These salts were reacted with copper (II) halide to give their corresponding metallo complexes. The surface tension measure-
ments for the synthesized compounds show that the metallo complexes have adsorption and micellization better than that of the
parent azobenzene isothiouronium salts. The pathogenic Gram-negative bacteria, Gram-positive bacteria, fungi and yeast were used
to determine the biocidal activity of these compounds using gradient plate technique. The results indicate that the copper complexes
of the synthesized azobenzene isothiouronium salts have a relatively better biocidal activity than the parent salts.
Ó 2006 Elsevier Ltd. All rights reserved.
1. Introduction
lower binding constant to Ac-D-Ala-D-Ala, a bacterial
cell-wall model, than the parent antibiotic and isothio-
uronium salt derivatives.3 Sjouke et al. described the
properties of a novel family of aromatic isothiouronium
derivatives.6 These isothiouronium derivatives appear to
be promising candidates for further development as
affinity labels of cation-binding domains, for kinetic
analysis of isoforms or mutated Na/K pumps, or as
probes of other cation transport proteins.6 The struc-
tures of the new gamma-lactam derivatives (N15-isothio-
Isothiouronium salts show a variety of biocidal activi-
ties.1–3 In the quest for biologically active sulfur com-
pounds, studies included isolation, characterization
and synthesis of novel compounds that can be used in
medicine. A wide variety of reactions to explore the syn-
thetic utility of these compounds were carried out to
synthesize more potent biocidal derivatives.4 Many of
the publications describe the synthesis, structural activi-
ty relationship, mode of action and pharmacological ef-
fects such as germicidal, antibacterial, antifungal and
anticancer activities of these compounds. Thiadiazole
isothiouronium derivatives have been synthesized and
their antifungal and antibacterial activities were tested.5
A series of thiourea and isothiouronium salt derivatives
of the aglycone of teicoplanin was prepared by reaction
of the terminal amino group with isothiocyanates, fol-
lowed by S-alkylation of the thiourea compounds.
Unexpectedly, the two classes of derivatives showed a
similar in vitro antibacterial activity against Gram-posi-
tive bacteria. Thiourea compounds, due to the lack of
apositively charged N-terminus group, have a 10-fold
1
uronium) were determined by using H NMR, IR and
fast atom-bombardment mass spectra. The cyclization
mechanism was interpreted on the basis of the identifica-
tion of the intermediate structure. The poor in vitro
antibacterial activity of the new cyclic compounds and
the negligible affinity for the synthetic peptidoglycan
model Ac2-L-Lys-D-Ala-D-Ala are probably due to the
lack of the N-17 amidic proton and to the lack of the
basic character of the nitrogen in position (15).7 The
antimicrobial and antifungal activities of the isothiouro-
nium derivatives, S-sec-butyl isothiouronium 2-pyrimid-
inyl sulfonamide; S-sec-butyl isothiouronium 2-(4,
6-dimethyl pyrimidinyl) sulfonamide and S-sec-butyl
isothiouronium 3-(6-methoxy) pyrydazinyl sulfonamide,
were investigated.8 Several publications have investigat-
ed the biocidal activity of copper (II) complexes and
show that the copper complexes have a better biocidal
activity than the uncomplexed compounds.9–12 In this
paper, we investigate the surface properties and the
Keywords: Azobenzene isothiouronium salts; Metallo complexes;
Adsorption; Micellization and biocidal activity.
*
Corresponding author. Tel.: +210 71 69 665; fax: +202 274 7433;
e-mail: eazzamep@yahoo.com
0968-0896/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2006.08.021

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A. M. Badawi et al. / Bioorg. Med. Chem. 14 (2006) 8661–8665
Table 3. Critical micelle concentration (CMC) and surface parameters
for the synthesized azobenzene isothiouronium salts and their copper
complexes
biocidal activity of a series of azobenzene isothiouro-
nium salts and their complexes with copper metal.
Compound
CMC
(mol/l)
Surface tension
Effectiveness
(ECMC) dyne/cm
at CMC (cCMC
)
2. Results and discussion
2.1. Surface properties
dyne/cm
C3
C6
0.05
55
51
41
54
49
38
17
21
31
18
23
34
0.025
0.005
0.05
C12
C3m
C6m
C12m
The surface tension (c) of the synthesized azobenzene
isothiouronium salts and their corresponding copper
complexes against different concentrations at 25 °C is
plotted in Figure 1. The results show that the copper
complexes have lower surface tension than the parent
azobenzene isothiouronium salts. The critical micelle
concentration (CMC) values were determined from the
plot of surface tension and concentration as shown in
Table 3. It is observed from Table 3 that the CMC
values of copper complexes with the exception of C3m
are lower than those of the parent azobenzene isothio-
uronium salts which reflect greater surface properties.
These results explain that the complexes retain its unity
of structure in their solutions, which increased their
0.01
0.0025
volume in the aqueous media with repulsion between
the hydrophobic chains and water molecules. This repul-
sion facilitates two processes at the same time. The
adsorption process of the molecules of metal complex
at the air/water interface of extremely lower concentra-
tions (below CMC) and the micellization process of
these molecules at concentrations lower than their par-
ent azobenzene isothiouronium salts.13 Table 3 reveals
that the CMC values decrease as the alkyl chain moiety
increases. This is due to the decrease in the solubility of
the prepared azobenzene isothiouronium salts and their
complexes as the alkyl chain in the hydrophobic moiety
increases. Effectiveness (PCMC) is calculated according
to the following equation as shown in the previous
publication.14
75
C3
C3m
C6m
C12m
70
65
60
55
50
45
40
35
30
25
C6
C12
PCMC ¼ co ꢀ c
where co measures the surface tension of pure water at
the appropriate temperature and c is the surface tension
at critical micelle concentration (CMC).
The data of effectiveness are summarized in Table 3. The
effectiveness of the prepared compounds increases as the
alkyl chain length increases. This indicates that the abil-
ity of the synthesized compounds to adsorb at air/water
interface and decrease the surface tension. The decreas-
0.0005 0.001 0.0025 0.005 0.01 0.025 0.05
Conc. (mol/l)
0.1
Figure 1. The relation between the surface tension and concentration
of the prepared isothiouronium salts and their complexes at 25 °C.
Table 1. The elemental analysis of the synthesized azobenzene isothiouronium salts and their copper complexes
Compound
MW
% C
Found
% H
Found
% N
Found
% S
Found
% Br
Found
% Cl
Found
Calcd
Calcd
Calcd
Calcd
Calcd
Calcd
C3
C6
409
451
49.88
53.21
58.31
42.83
46.30
51.80
49.86
53.19
58.30
42.80
46.32
51.82
5.17
6.03
7.34
4.40
5.20
6.47
5.15
6.01
7.31
4.37
5.17
6.45
13.69
12.41
10.46
11.75
10.80
9.29
13.67
12.40
10.44
11.77
10.81
9.30
7.83
7.09
5.98
6.71
6.17
5.31
7.81
7.10
5.99
6.72
6.20
5.29
19.55
17.73
14.95
16.79
15.43
13.28
19.53
17.70
14.97
16.77
15.41
13.29
—
—
—
—
C12
C3m
C6m
C12m
535
—
—
952.5
1036.5
1204.5
7.45
6.84
5.89
7.42
6.87
5.88
Table 2. The FTIR spectroscopy of the synthesized azobenzene isothiouronium salts and their copper complexes
Compound
CH3
CH2
C–S
C–O–C
Para substituted
benzene ring
C3
C6
2913 cmꢀ1
2920 cmꢀ1
2935 cmꢀ1
2947 cmꢀ1
2960 cmꢀ1
2955 cmꢀ1
2848 cmꢀ1
2840 cmꢀ1
2835 cmꢀ1
2860 cmꢀ1
2872 cmꢀ1
2865 cmꢀ1
710 cmꢀ1
716 cmꢀ1
734 cmꢀ1
722 cmꢀ1
711 cmꢀ1
720 cmꢀ1
1026 cmꢀ1
1027 cmꢀ1
1028 cmꢀ1
1025 cmꢀ1
1026 cmꢀ1
1027 cmꢀ1
1005–825 cmꢀ1
1005–822 cmꢀ1
1005–820 cmꢀ1
1005–840 cmꢀ1
1005–835 cmꢀ1
1005–825 cmꢀ1
C12
C3m
C6m
C12m

A. M. Badawi et al. / Bioorg. Med. Chem. 14 (2006) 8661–8665
Table 4. Biocidal activity for the prepared azobenzene isothiouronium salts
8663
Sample
Inhibition zone diameter (mm/mg sample)
Pseudomonae aeruginosa (Gꢀ)
Staphylococcus aureus (G+)
Aspergillus flavus (fungus)
Candida albicans (yeast)
0.0
14
Control: DMSO
C3
0.0
13
0.0
13
0.0
0.0
0.0
0.0
C6
C12
12
13
13
14
13
13
G, Gram reaction.
Solvent: DMSO.
Table 5. Biocidal activity for the prepared azobenzene isothiouronium copper complexes
Sample
Inhibition zone diameter (mm/mg sample)
Pseudomonae aeruginosa (Gꢀ)
Staphylococcus aureus (G+)
Aspergillus flavus (fungus)
0.0
14
Candida albicans (yeast)
Control: DMSO
C3m
C6m
0.0
15
0.0
14
0.0
16
13
14
14
13
12
13
14
13
C12m
G, Gram reaction.
Solvent: DMSO.
ing in the surface tension is increase as the alkyl chain
length increase. The synthesized copper complexes
(C3m, C6m and C12m) show higher PCMC than their
parent azobenzene isothiouronium salts (C3, C6 and
C12) indicating that the copper complexes have more
efficiency towards adsorption at air/water interface than
their parent azobenzene isothiouronium salts.
and C. albicans yeast. This behaviour could be explained
through their ability towards adsorption at the cell
membrane of the microorganisms under investigation.
The increase in biocidal activity of the copper complexes
is attributed to the lower electronegativity and large vol-
ume of copper ions, which increase their molecular area.
Hence, the effective area of the complex molecules on
the cell membrane increases resulting in a higher biocid-
al action.13
2.2. Biocidal activity
The biological activity of the synthesized azobenzene
isothiouronium salts and their copper complexes against
Gram-negative bacteria (Pseudomonae aeruginosa),
Gram-positive bacteria (Staphylococcus aureus), Asper-
gillus flavus fungus and Candida albicans yeast are repre-
sented in Tables 4 and 5. The biocidal activity of the
compounds under investigation (C3, C6, C12, C3m,
C6m and C12m) agrees with their ability for adsorption
at the water/cell membrane interface. This adsorption
increases solubility through the cell membrane increas-
ing its permeability towards the media ingredients. Thus
biological reactions disturb within the cell cytoplasm.13
Tables 4 and 5 show that, the prepared azobenzene iso-
thiouronium salts and their copper complexes have a
good biocidal activity against bacteria, fungi and yeast
used in this investigation. Table 4 shows that, com-
pounds C3 and C12 give relatively better inhibition
zones against bacteria and fungi than compound C6.
It is noticed that, compounds C3, C6 and C12 show neg-
ative biological activity against A. flavus fungus. The
comparison of the data in Tables 4 and 5 indicates that,
the copper complexes have a better biological activity
towards bacteria, fungi and yeast under study than their
parent azobenzene isothiouronium salts. Also the results
show that, the synthesized 4-methyl-40-propyloxy-azo-
benzene isothiouronium dibromo-dichlorocuparate
complex (C3m) compound gives the highest inhibition
zones against Gram-negative bacteria (P. aeruginosa),
Gram-positive bacteria (S. aureus), A. flavus fungus
3. Conclusions
1. The copper complexes have more effectiveness
towards adsorption at air/water interface than their
parent azobenzene isothiouronium salts.
2. The azobenzene isothiouronium salts and their
copper complexes have a good biological activity
against Gram-positive and Gram-negative bacteria,
fungi and yeast.
3. The biocidal activity of the copper complexes is great-
er than that of their parent azobenzene isothiouroni-
um salts.
4. Experimental technique
4.1. Materials
4.1.1. Synthesis of azobenzene isothiouronium salts. A
series of azobenzene isothiouronium salts (C3, C6
and C12) of different alkyl chains (propyl, hexyl and
dodecyl) was synthesized by refluxing 4-((4-methylphe-
nyl)azo)phenol with 1,3-dibromopropane, 1,6-dibromo-
hexane or 1,12-dibromododecane, respectively, in
acetone for 6 h in presence of potassium carbonate.
The refluxing product was crystallized from 2-propanol
to afford the corresponding bromoazobenzene

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A. M. Badawi et al. / Bioorg. Med. Chem. 14 (2006) 8661–8665
+
NH
2
+
-
CH
N
N
O
(CH )
2 3
S
C
Br
NH
C
3
2
2
-
Br CuCl
NH
CH
N
N
O
(CH )
S
2
2
2
3
3
2
NH
4-methyl -4`- propyloxy-azobenzene isothiouronium bromide (C3)
2
4-methyl -4`- propyloxy-azobenzene isothiouronium dibromo-dichloro
cuparate complex (C3m)
NH
2
2
+
-
O
CH
N
N
(CH )
2 6
S
C
Br
3
+
NH
4-methyl - 4`- hexyloxy-azobenzene isothiouronium bromide (C6)
NH
C
2
2
-
Br CuCl
2
2
CH
N
N
O
(CH )
2 6
S
3
NH
NH
2
+
-
2
O
CH
N
N
(CH )
2 12
S
C
Br
3
4-methyl -4`- hexyloxy-azobenzene isothiouronium dibromo-dichloro
cuparate complex (C6m)
NH
2
4-methyl - 4`- dodecyloxy-azobenzene isothiouronium bromide (C12)
+
Scheme 1. The chemical structure of the prepared azobenzene
isothiouronium salts.
NH
C
2
2
-
Br CuCl
CH
N
N
O
(CH )
2 12
S
2
2
3
NH
compounds which were converted to isothiouronium
salt by refluxing with thiourea in ethanol for 5 h.15
The chemical structure of the synthesized azobenzene
isothiouronium salts (C3, C6 and C12) is shown in
Scheme 1.
2
4-methyl - 4`- dodecyloxy-azobenzene isothiouronium dibromo-dichloro
cuparate complex (C12m)
Scheme 2. The chemical structure of the prepared azobenzene
isothiouronium complexes.
4.1.2. Synthesis of copper azobenzene isothiouronium
salts. Copper complexes of azobenzene isothiouronium
salts (C3m, C6m and C12m) were prepared by refluxing
two moles of azobenzene isothiouronium salts and one
mole CuCl2 in ethyl alcohol for 3 h.16 The resulting crys-
tals were washed twice with petroleum ether, recrystal-
lized from methanol and dried under vacuum at 50 °C
for 4 h (Scheme 2).
the clear zone of inhibition surrounding the sample is
taken as a measure of the inhibitory power of the sample
against the particular test organism.17–20
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isothiouronium salts and their complexes is shown in
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was carried out using ATI Malttson Genesis Series
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for their biocidal activity using diffusion disc method.
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