Antimicrobial efficacy of metal-barbiturate conjugates against pathogenic strains of escherichia coli and staphylococcus aureus

Article abstract of DOI:10.2174/157018011797655232

Resistance to commercially available antimicrobials is increasing at alarming rate therefore there is an urgent need of new antimicrobial compounds and materials with novel modes of action to tackle the problem of resistance in pathogenic gram positive and gram negative bacteria. The current study involves the design and synthesis of new molecular structures which should not resemble the basic molecular structures of the existing classes of antimicrobial agents in order to avoid the risks of cross resistance development. A total of 14 metal-barbiturate complexes (newly synthesized compounds) were evaluated for their antimicrobial properties against multiple drug resistance clinical isolates of S. aureus and E. coli. Broth dilution method was used to determine minimum inhibitory and bactericidal activity of the newly synthesized compounds. The results showed that the metal-barbiturate complexes with cobalt ions were more effective against E. coli and S. aureus as compared to other compounds. Moreover the activity of these compounds was higher against S. aureus as compared to E. coli which indicates species specific variable antimicrobial effects of these compounds.

Full text of DOI:10.2174/157018011797655232

980
Letters in Drug Design & Discovery, 2011, 8, 980-987
Antimicrobial Efficacy of Metal-Barbiturate Conjugates Against Patho-
genic Strains of Escherichia coli and Staphylococcus aureus
Ashfaq Mahmood Qureshi*,1, Mehmood Qadir2, Abdul Rauf4, Muhammad Idrees2, Siara Mumtaz1,
Muhammad Najam-ul-Haq1, Aziz-ur-rehman4, Muhammad Ismail1, Muhammad Athar1,
Rashid Khushal3, Sara Riaz1 and Habib Bokhari*,2
1Department of Chemistry, Baha-uddin Zakriya University, Multan, Pakistan
2Department of Biosciences, COMSATS Institute of Information Technology, Islamabad, Pakistan
3Astra Zeneca, Leicester, UK
4Department of Chemistry, The Islamia University, Bahawalpur, Pakistan
Received April 11, 2011: Revised July 27, 2011: Accepted August 12, 2011
Abstract: Resistance to commercially available antimicrobials is increasing at alarming rate therefore there is an urgent
need of new antimicrobial compounds and materials with novel modes of action to tackle the problem of resistance in
pathogenic gram positive and gram negative bacteria. The current study involves the design and synthesis of new
molecular structures which should not resemble the basic molecular structures of the existing classes of antimicrobial
agents in order to avoid the risks of cross resistance development. A total of 14 metal-barbiturate complexes (newly
synthesized compounds) were evaluated for their antimicrobial properties against multiple drug resistance clinical isolates
of S. aureus and E. coli. Broth dilution method was used to determine minimum inhibitory and bactericidal activity of the
newly synthesized compounds. The results showed that the metal-barbiturate complexes with cobalt ions were more
effective against E. coli and S. aureus as compared to other compounds. Moreover the activity of these compounds was
higher against S. aureus as compared to E. coli which indicates species specific variable antimicrobial effects of these
compounds.
Keywords: Antimicrobial, Barbiturate Complexes, Drug resistance, E. coli, S. aureus.
INTRODUCTION
Barbiturates are a class of drugs that are used as
anesthetics and sleep inducing agents [11]. However, some
previous studies have already shown that barbiturate such as
methohexitone (1%) and thiopentone (2.5%) are bactericidal
against coagulase-negative staphylococci [12-13]. Moreover,
thiopentone (2.5%) has already shown bactericidal activity
against E. coli and P. aeruginosa [14].
Antimicrobial resistance is one of the major threats to the
life quality of human beings as well as animals. Since the
discovery of first antibiotic, the new antibiotics have been
discovered but the incidence of antimicrobial resistance in
the bacteria has also been reported continuously [1-2].
Hence, continuous burden of infectious diseases and
emergence of drug resistance is a global issue [3-4]. The
resistance to commercially available antimicrobials is
increasing day by day by various bacteria [5]. Therefore,
there is an urgent need of new compounds with new modes
of action and combination therapies to tackle the problem of
resistance in gram positive and gram negative bacteria [6].
Pathogenic bacterial organisms on the other hand under
constant exposure to antibacterials are evolving rapidly and
emerging as more resistant entities to existing antimicrobial
agents [5, 15]. Therefore it is envisaged that new compounds
with new modes of action and combination therapies are
required to tackle the problem of resistance in gram positive
and gram negative bacteria [6].
Staphylococcus aureus is a gram-positive bacterium
which colonizes the skin and is present in nares of 25-30%
of healthy population [7] and is one of the most common
causative agents of nosocomial infections worldwide.
Staphylococcal infections are of particular concern because
of the causative bacteria offering resistance to a wide range
of commonly used antibiotics. On the other hand E. coli have
certain well known resistance determinants therefore they are
used as indicator bacteria internationally for antimicrobial
resistance [8-10].
Structurally similar antimicrobials are likely to face the
similar fate i.e. resistance to one may result in resistance to
others. In order to combat the development of cross
resistance, we intended to explore the basic ring structures
different from existing classes of antimicrobial compounds.
So we used barbiturate as the basic ring structure with metal-
barbiturate complex formation at various positions of the
basic ring in order to explore the best suitable metal ligand
or position on the ring to show maximum antimicrobial
activity. In this way we can counteract the phenomenon of
cross resistance by exploring the chemical formulas and ring
structures dissimilar to the existing antimicrobial compound
*Address correspondence to these authors at the Department of Biosciences,
COMSATS Institute of Information Technology, Islamabad, Pakistan; Tel:
0092-3005127684; Fax: 0092-514442805; E-mail: habib@comsats.edu.pk
structures. For instance, PNU-286607,
a
barbiturate
Department of Chemistry, Baha-uddin Zakriya University, Multan,
Pakistan; Tel: 0092-3064782968; Fax: 0092619210138;
E-mail: shamashfaq@yahoo.com
derivative displays little cross-resistance with marketed
antibacterial agents and is active against methicillin-resistant
1570-1808/11 $58.00+.00
© 2011 Bentham Science Publishers

Antimicrobial Properties of Metal Barbiturate Complexes
Letters in Drug Design & Discovery, 2011, Vol. 8, No. 10 981
staphylococcus aureus (MRSA) and fluoroquinoline-resis-
tant bacterial strains [16]. Some other examples of related
compounds are thioxanthene flupenthixo Prochlorperazine,
and fluphenazine [17-18]. These are different antipsychotic
compounds which have already been found to be effective as
antimicrobial agents by their in vitro studies.
127.8 (68), 119 (33), 105 (39), 92 (34), 77 (20), 63 (50), 51
(13).
5-(4-methoxybenzylidene)pyrimidine-2,4,6(1H,3H,5H)-
trione (2)
Orange yellow powder, 78%; m.p 255 oC; max(ꢀ KBr)/cm-
11212 (C-N str), 1507 (C=C aromatic str), 1600, 1674, 1730
(C=O str), 2840 (C-H aliphatic str), 3071 (=C-H str), 3208
(N-H str); ꢀH(300 MHz; DMSO) 3.86 (3H, s, OCH3), 7.05
(2H, d, J 9, ArH), 8.24 (1H, s, C=CH), 8.36 (2H, d, J 9,
ArH). 11.16 (1H, s, NH), 11.28 (1H, s, NH); 66.2 (CH3),
118.0 (C), 118.6 (CH), 118.8 (CH), 123.5 (CH), 123.8 (CH),
125.0 (C), 148.7 (CH), 151.7 (C), 153.1 (C=O), 164.0
(C=O), 164.1 (C=O); m/z (EI) 246 (100), 245 (87), 231 (10),
215 (20), 202 (65), 172 (13), 159 (19), 132 (17), 117 (18), 89
(25), 61.9 (11).
The objective of the current study was to evaluate the
efficacy of newly synthesized metal-barbiturate complexes
against resistant bacterial isolates. Antimicrobial effects of
some of the newly synthesized barbiturate compounds
(metal-barbiturate conjugates) against common pathogens
including gram negative bacteria (multiple drug resistant E.
coli causing Urinary Tract Infections) and gram positive
bacteria (multiple drug resistant S. aureus causing skin
infections) were identified.
5-(3-bromobenzylidene)pyrimidine - 2, 4, 6 (1H, 3H, 5H)-
trione (3)
MATERIAL AND METHODS
Synthesis of Compounds (1-3)
Bright yellow powder, 20%; m.p 247 oC; max(ꢀ KBr)/cm-1
516 (C-Br str), 1274 (C-N str), 1577 (C=C str), 1678, 1704,
1752 (C=O str), 3098 (=C-H str), 3372 (N-H str); ꢀH (300
MHz; DMSO) 7.41 (1H, t, J 7.9, ArH), 7.69 (1H, d, J 8.8,
ArH), 7.88 (1H,d, J 7.9, ArH), 8.21 (1H, s, ArH), 8.28 (1H,
s, C=CH), 11.26 (1H, s, NH), 11.41 (1H, s, NH); 116.4 (C),
117.6 (CH), 125.3 (C), 128.0 (CH), 128.2 (CH), 128.5 (CH),
146.4 (CH), 154.8 (C), 155.0 (C=O), 162.0 (C=O), 162.4
(C=O); m/z (EI) 294.9 (83), 251.9 (40), 224.9 (7.4), 215
(100), 172 (75), 155 (4), 128 (18.5), 116 (16), 101 (43), 89
(21), 78 (64), 75 (32), 61.9 (85).
All chemicals used in this study are purchased from
Sigma-Aldrich (Czech Republic). Synthesis of all ligands
was done according to Scheme 1. In a round bottom flask,
fitted with reflux condenser, a mixture of equimolar
barbituric acid and aromatic aldehyde was refluxed in
absolute ethanol for about 4 hrs to overnight. After the
formation of precipitates, the reaction mixture was cooled
and filtered off. The precipitates were washed several time
with hot ethanol, dried in an oven and subjected for further
characterization i.e., m.p (John Fisher melting point
apparatus), I.R, 1HNMR, 13C NMR (Jan19 NMR
1
spectrometer, operating at 300 MHz for H and 75 MHz for
Synthesis of Compounds (4-14)
13C) and mass spectrometry (on JEOL JMS600 instrument).
Barbituric acid can act as an alterdentate ligand as it has
more than one equivalent coordination sites available to a
metal ion. In this regard rearrangement is possible in which
metal is transferred from one site to another. Three
barbiturates used in this study for preparation of metal-
barbiturate complexes are compounds 1-3. All the complexes
of barbiturate were prepared in two different molar ratios of
metal: ligand i.e., 1:1 and 1:2 by dissolving ligand and metal
salt separately in minimum amount of ethanol. Then the two
solutions were mixed followed by addition of small amount
of ammonia in a round bottom flask and refluxed for about
45 minutes. The precipitates formed were filtered, washed
5-(5-hydroxy-2-nitrobenzylidene)pyrimidine-2,4,6(1H,3H,
5H)-trione (1)
o
Light grey powder, 61%; m.p 260 C; max(ꢀ KBr)/cm-1
1220 (C-N str), 1425 (C=C str), 1500 (-NO2 antisymmetric
str), 1610, 1710 (C=O str), 3050 (=C-H str), 3069 (-N-H str),
3300 (-OH str); ꢀH(300 MHz; DMSO) 6.78 (1H, d, J 2.1,
ArH), 6.95 (1H,dd, J 9.1, 2.5, ArH), 8.15 (1H, d, J 9.1,
ArH), 8.52 (1H, s, CH=C), 11.12 (1H, s, NH), 11.19 (1H, s,
NH), 11.45 (1H, s, OH); 13CNMR(d6-DMSO, ꢀ, ppm): 116.1
(C), 116.9 (CH), 119.7 (CH), 125.4 (CH), 130.8 (C), 137.3
(C), 146.0 (CH), 149.8 (C), 152.0 (C=O), 161.3 (C=O),
164.2 (C=O); m/z (EI) 277.9 (1), 230 (100), 217.9 (17),
187.8 (100), 174.9 (27), 160 (24), 148.9 (75), 135 (29),
o
with hot ethanol, dried in an oven at 50 C and characterized
by IR and UV and then subjected to biological activity.
O
R1
R1
O
R2
R3
HN
OHC
R2
R3
EtOH, reflux
HN
+
4 hrs - overnight
O
N
H
O
O
O
N
H
R 4
R4
Where ( 1 ) = R1 =NO2, R2, R3 =H and R4 =OH
( 2 ) = R1 = R2 =R4 = H and R3 =OCH3
( 3) = R1 = R3 = R4 = H and R2 =Br
Scheme 1. Preparation of Barbiturate Ligands used for complexation.

982 Letters in Drug Design & Discovery, 2011, Vol. 8, No. 10
Qureshi et al.
H
N
O
O
O
Ar
HN
HN
Ar
2+
O
EtOH, reflux
45 minutes
M
O
2
O
O
MX2.xH2O
X2
O
N
H
O
NH
NH
Ar
where M = Zn+2, Ni+2, Co+2 and Mn+2
O
HN
Ar
O
O
N
O
H
HN
Ar
X2
EtOH, reflux
45 minutes
MX2.xH2O
2+
M
O
N
H
O
H2O
OH2
H2O
where M = Zn+2, Ni+2, Co+2 and Mn+2
Scheme 2. Preparation of metal barbiturate complexes.
Antimicrobial Studies
diffusion method for susceptibility pattern. The strain was
resistant to Nalidixic acid, Sulphamethoxazole-Trimetho-
prim, Ampicillin, Trimethoprim. The multidrug resistant
strain of S. aureus was selected for antimicrobial assay.
Bacterial Strains
All clinical isolates used in the present study were
isolated from patients in the hospital. All chemicals were
purchased from Sigma unless otherwise stated.
Antimicrobial Assay
E. coli
Broth dilution method was used to determine minimal
concentration of newly synthesized compounds to inhibit or
kill the microorganism. LB broth was prepared by adding 2g
of media in 100ml distilled water and sterilized using
autoclave at 121ºC for 15 min. Bacterial isolates and quality
control organisms’ suspensions (107cfu/ml) in PBS
(phosphate buffer solution) were prepared by making
dilution using plate count method.10 ꢀl inoculums of these
suspensions (107cfu/ml) were used in the assay.
The strains were isolated from Urine samples collected
from Urinary Tract infection patients. Urine samples were
immediately cultured on CLED (cysteine lactose electrolyte–
deficient) agar. The bacterial growth was subcultured on
MacConkey agar. Lactose- and indole-positive colonies were
presumptively identified as E. coli. The antimicrobial
resistance profiles for E. coli isolates were determined by
standard disk diffusion techniques (Neo-Sensitabs; Oxoid
Ltd), according to the recommendations of CLSI (Clinical
Laboratory Standards Institute) whereas virulence factors
profile was also determined as described in our previous
study [19]. The clinical strain used in this study was resistant
to Nalidixic acid, Ceftazedime, Ciprofloxacin, Sulphametho-
xazole Trimethoprim and Ampicillin.
Stock solutions of barbituric acid and 14 newly
synthesized compounds were prepared by dissolving 5 mg of
each compound in 1ml mixture of DMSO (50% v/v) and
PBS (50% v/v). Two fold serial dilutions for each newly
synthesized compound from its stock solution were prepared
in DMSO/ PBS mixture (50 % / 50 % v/v). For each
compound, test tubes were labeled with different dilution
concentrations. Assay was performed by adding broth media
and compound dilutions in each test tube and the volume of
broth/compound mixture was adjusted to 1 ml i.e. containing
variable amount of broth and newly synthesized compound
dilutions.
S. aureus
The clinical samples from patients suffering from
multiple skin diseases were obtained from the skin lesions
for the isolation of S. aureus with the help of sterile swabs
which were then placed back in the sterile Mannitol Salt
Broth and transported to the laboratory. Samples were then
inoculated on to Mannitol Salt Agar for the selective growth
of S. aureus. S. aureus isolates were then subjected to disk
10 ꢀl of prepared bacterial suspension was added into
each test tube and inoculated tubes were then incubated for

Antimicrobial Properties of Metal Barbiturate Complexes
Letters in Drug Design & Discovery, 2011, Vol. 8, No. 10 983
18 – 24 hr at 37ºC. Positive control was performed by adding
10 ꢀl of S. aureus (ATCC 6538 – Oxoid Basingstoke UK)
and E. coli (ATCC 8739 – Oxoid Basingstoke UK) to
separate test tubes containing broth media only whereas
uninoculated tube containing broth and synthetic compound
serve as negative control for turbidity measurement.
reincubated for additional 4 hours. After 4 hours of
additional incubation, visually clear tubes were again
vortexed and 0.1 ml from each test tube was spread across
the surface of LB agar plates with the help of sterile glass
rod. The plates were incubated at 37ºC for 18-24 hours and
after incubation the concentration in which the petriplate
showed either no growth or reduction of 99.9% from
inoculums was considered as bactericidal concentration of
the specific compound.
After incubation effects of synthetic compounds on
bacterial count in each test tube were recorded by finding out
the turbidity as compared to the control tubes. Minimum
inhibitory concentration was determined by visual inspection
of the tube as MIC is the lowest concentration of compound
that inhibits visible growth. Tubes without growth were
vortexed vigorously for 15 seconds to resuspend any bacteria
that might have adhered to the wall of the tube and were
RESULTS AND DISCUSSION
The spectral data of the synthesized compounds is found
to be supportive. Table 1 shows HRMS and IR data of
synthesized compounds.
Table 1. Spectral Data of Barbiturate/ Barbiturate Metal Complexes
Compound
1
Ligand:Metal HRMS
Selected I.R data (ꢀ cm-1)
L-1
L-2
L-3
C11H7N3O6
277.18978(Cal)
277.18981(Found
1220 (C-N), 1425 (C=C), 1500 (-NO2), 1610, 1710 (C=O), 3050 (=C-H), 3069 (-N-H), 3300 (-
OH)
2
1212 (C-N), 1507 (C=C), 1600, 1674, 1730 (C=O), 2840 (C-H), 3071 (=C-H), 3208 (N-H)
516 (C-Br), 1274 (C-N), 1577 (C=C), 1678, 1704, 1752 (C=O), 3098 (=C-H), 3372 (N-H)
1696 (C=O), 1594 (C=C), 3137 (=CH), 3444 (-NH), 851 (C-N), 1347 (NO2), 549 (M-N)
1674 (C=O), 1622 (C=C), 3050 (=CH), 3420 (-NH), 850 (C-N), 1348 (NO2), 525 (M-N)
1691 (C=O), 1594 (C=C), 3010 (=CH), 3384(-NH), 851 (C-N), 1346 (NO2), 520 (M-N)
1692 (C=O), 1586 (C=C), 3015 (=CH), 3418 (-NH), 859 (C-N), 1350 (NO2), 519 (M-N)
1694 (C=O), 1585 (C=C), 3195 (=CH), 3384 (-NH), 851 (C-N), 1346 (NO2), 541(M-N)
1692 (C=O), 1594 (C=C), 3040 (=CH), 3376 (-NH), 851 (C-N), 532(M-N)
1682 (C=O), 3195 (-NH), 3398 (-OH), 518 (M-N)
C12H10N2O4
246.2188(Cal)
246.2195(Found)
3
C11H7BrN2O3
295.08888(Cal)
295.08919(Found)
4
L-1: Zn
(1:1)
C11H13Cl2ZnN3O9
467.55062(Cal)
467.55078(Found)
5
L-1: Mn
(1:1)
C11H13Cl2MnN3O9
457.079665(Cal)
457.079698(Found)
6
L-1: Ni
(1:1)
C11H13Cl2NiN3O9
460.83502(Cal)
460.83499(Found)
7
L-1: Co
(1:1)
C11H13Cl2CoN3O9
461.074815(Cal)
461.074845(Found)
8
L-1: Co
(1:2)
C22H14Cl2CoN6O12
684.218755(Cal)
684.218777(Found)
9
L-2: Mn
(1:1)
C12H16Cl2MnN2O7
426.108685(Cal)
426.108697(Found)
10
11
12
13
14
L-2: Co
(1:1)
C12H16Cl2CoN2O7
430.103835(Cal)
430.103877(Found)
L-2: Co
(1:2)
1703 (C=O), 3202 (-NH), 3413 (-OH), 517 (M-N)
C24H20Cl2CoN4O8
622.276795(Cal)
622.276799(Found)
L-3: Mn
(1:1)
1694 (C=O), 1589 (C=C), 3010 (=CH), 3405 (-NH), 850, 1348 (NO2), 540(M-N)
1694 (C=O), 1583 (C=C), 3055 (=CH), 3400 (-NH), 851 (C-N), 1346 (NO2), 517 (M-N)
1682 (C=O), 3144 (-NH), 3400 (-OH), 518 (M-N)
C11H13BrCl2MnN2O6
474.978765(Cal)
474.978789(Found)
L-3: Ni
(1:1)
C11H13BrCl2NiN2O6
478.73412(Cal)
478.73455(Found l)
L-3: Co
(1:2)
C22H14Br2Cl2CoN4O6
720.016955(Cal)
720.016972(Found)

984 Letters in Drug Design & Discovery, 2011, Vol. 8, No. 10
Qureshi et al.
H
O
NO2
OH
HN
O
N
H
O
m/z 278
-HNO2
-OH
HN
17
47
O
H
O
NO2
HN
O
N
H
O
O
N
H
O
OH
m/z 260.9
m/z 230.8 (100%)
43
43
-HNC=O
O
-HNC=O
H
NO2
O
HN
HN
O
OH
m/z 187.8 (100%)
O
m/z 217.9
28
-CO
-HNC=O
43
m/z 160
m/z 174.9
47
-HNO2
m/z 127.9
Fig. (1). Mass fragmentation pattern of Compound 1.
The 13CNMR spectrum showed resonance of three
carbonyl groups at each instance ranging from 152-164.2
ppm. The carbon of alkene moiety resonated at 117.6-119.7
ppm and its adjacent carbon resonated highfield in the range
149.8-154.8 ppm due to two carbonyl groups in its vicinity.
MIC of DMSO for E. coli was 17 % (v/v) (IC50 = 12 %),
and MBC at 29 % (v/v) whereas the MIC for S. aureus was 8
% (v/v) (IC50 = 6 %) and MBC was 20 % (v/v). Barbituric
acid was the parent compound for the syntheses of derived
metal-barbiturate complexes. Antimicrobial properties of
Barbituric acid were studied to see the antimicrobial effects
of the parent compound. Barbituric acid did not show any
bacteriostatic or bactericidal effects against E. coli and S.
aureus at concentrations upto 2000 ꢀg/ml. The antimicrobial
effects of compounds (1-3) were also studied. (1) did not
show any bacteriostatic or bactericidal effects against E. coli
and S. aureus till 2000 ꢀg/ml. IC 50 values of (2) (1350
ꢀg/ml for S. aureus; 1300 ꢀg/ml for E. coli) and (3) (1150
ꢀg/ml of S. aureus; 1100 ꢀg/ml for E. coli) were higher as
compared to metal-barbiturate complexes (10) and (11)
which indicate that metal-barbiturate complexes are more
effective as compared to parent compounds.
The mas spectra of all the compounds showed molecular
ions of different intensity, which confirmed their molecular
weights. The mass fragmentation pattern of compound (1) is
speculated in Fig. (1). The base peak at m/z 230.8 showed
the most stable fragment formed in this case by the loss of
HNO2.
Antimicrobial Assay
The study has shown that some of the metal-barbiturate
complexes have at least some bacteriostatic activity against
uropathogenic E. coli. The bactericidal effects of some of the
barbiturates were observed against the tested S. aureus. The
variations in the effects of these compounds indicates that
there are some of the metals which when attached to the
barbiturate, the compound become antimicrobial or its
activity is enhanced, while the other metal-barbiturate
complexes are less effective.
We used a total of 14 newly synthesized Barbiturate
derivative compounds to evaluate their antimicrobial
properties against clinical multiple drug resistance
uropathogenic E. coli and S. aureus. Out of the 14
barbiturate derivatives, 9 compounds (1, 4-10 and 14) did
not show any bacteriostatic or bactericidal effects against E.
coli and S. aureus till 2000 ꢀg/ml except 4 which started
inhibition of bacterial growth at 1500 ꢀg/ml for S. aureus
(IC 50 = 1825 ꢀg/ml) with no activity for E. coli till 2000
The solvent system for dissolving compounds was
DMSO, so antimicrobial assay was performed to evaluate
the antimicrobial effect of DMSO on the studied strains.

Antimicrobial Properties of Metal Barbiturate Complexes
Letters in Drug Design & Discovery, 2011, Vol. 8, No. 10 985
Table 2. Melting Temperatures, Solubilities and Antimicrobial Activities (IC 50) of Compounds Against E. coli and S. aureus
Entry
Ligand:Metal
IC 50 (E. coli)
IC 50 (S. aureus)
Solvent
Melting Point °C /
Decomposition Point
1
2
3
4
L-1
L-2
260oC (Sharp)
255 oC (Sharp)
247 oC (Sharp)
255 oC (decomp)
----
----
DMSO
DMSO
DMSO
DMSO
1300 ꢀg/ml
1100 ꢀg/ml
----
1350 ꢀg/ml
1150 ꢀg/ml
1825 ꢀg/ml
L-3
L-1: Zn
(1:1)
5
6
L-1: Mn
(1:1)
140 oC (decomp)
300 oC (decomp)
195 oC (decomp)
220 oC (decomp)
255 oC (decomp)
110 oC (decomp)
155 oC (decomp)
215 oC (decomp)
200 oC (decomp)
170 oC (decomp)
----
----
----
----
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
L-1: Ni
(1:1)
7
L-1: Co
(1:1)
----
----
8
L-1: Co
(1:2)
----
----
9
L-2: Mn
(1:1)
----
----
10
11
12
13
14
L-2: Co
(1:1)
450 ꢀg/ml
425 ꢀg/ml
----
300 ꢀg/ml
325 ꢀg/ml
1400 ꢀg/ml
----
L-2: Co
(1:2)
L-3: Mn
(1:1)
L-3: Ni
(1:1)
----
L-3: Co
(1:2)
----
----
15
16
DMSO
Barbituric acid
----------
----------
12 % (v/v)
----
6% (v/v)
----
-------
-------
ꢀg/ml. This means that either the sites of metal attachment in
these compounds have no role in antimicrobial activity [18,
20] or the type of metal ions ligated at these sites are inert
against bacteria. Many of the studies have reported the rising
level of resistance against other antimicrobial agents as well
in S. aureus [21-22] and uropathogenic E. coli [23-24].
aureus and E. coli [17]. The results indicate that (10) and
(11) were more effective against S. aureus as compared to E.
coli. IC 50 value of (10) for S. aureus was 300 ꢀg/ml and
450 ꢀg/ml for E. coli. On the other hand IC 50 values of (11)
for S. aureus was 325 ꢀg/ml while 425 ꢀg/ml for E. coli.
Our observed differences between antimicrobial activities
(IC 50 values) of (2) (1350 ꢀg/ml for S. aureus; 1300 ꢀg/ml
for E. coli), (3) (1150 ꢀg/ml of S. aureus; 1100 ꢀg/ml for E.
coli), (11) (325 ꢀg/ml for S. aureus; 425 ꢀg/ml for E. coli)
and (10) (300 ꢀg/ml for S. aureus; 450 ꢀg/ml for E. coli)
against E. coli and S. aureus may be due to difference of
susceptibility levels between different species due to the wall
structure i.e. Single layer of cell wall of gram positive (S.
aureus) while the other has double layer (E. coli). The wall
structure may result in different outcomes when similar
drugs used on two different bacteria e.g. Penicillin is more
effective against Gram positive bacteria as compared to the
Gram negative bacteria [25].
Metal-barbiturate complex (12) has shown no
bacteriostatic activity against E. coli upto 2000 ꢀg/ml
however it was bacteriostatic against S. aureus (IC 50 =
1400 ꢀg/ml). On the other hand four compounds (2-3, 10-11)
had bacteriostatic effects against uropathogenic E. coli out of
which (10) (IC 50 = 450 ꢀg/ml) and (11) (IC 50 = 425
ꢀg/ml) were bacteriostatic at lower concentrations (Fig. 2,
Table 2). Of these (4) compounds, (2) (IC 50 = 1350 ꢀg/ml)
and (3) (IC 50 = 1150 ꢀg/ml) were bacteriostatic while the
other two i.e., (10) (IC 50 = 300 ꢀg/ml) and (11) (IC 50 =
325 ꢀg/ml) have bactericidal activity (Fig. 3, Table 2)
against S. aureus. Similar antimicrobial effects have already
been reported by different cyclophenamides against S.

986 Letters in Drug Design & Discovery, 2011, Vol. 8, No. 10
Qureshi et al.
It was also noted that the antimicrobial effects of
different compounds (2) (IC 50 = 1350 ꢀg/ml), (3) (IC 50 =
1150 ꢀg/ml), (10) (IC 50 = 300 ꢀg/ml) and (11) (IC 50 = 325
ꢀg/ml)) against the same species, i.e., S. aureus, were not of
the same level; some were bacteriostatic (2-3) and the others
were bactericidal (10-11).
2,4,6(1H,3H,5H)-trione (2) in 1:1 and 1:2 metal: ligand ratio
respectively. These (10-11) were found to have the highest
antimicrobial activity amongst the compounds we studied.
These results suggests that antimicrobial activity of the
compounds may have been regulated by the position at
which metal ion is ligated.
The Barbituric acid derivatives have a possible risk of
side effects due to sedative, hypnotic and anesthetic effects.
The pharmacological effects of barbiturates scale from mild
sedation to anesthesia. These effects can be minimized by
applying the antimicrobial properties of the derived
compounds to treat skin infections with susceptible
pathogens (for example S. aureus) and other conditions
where no oral or systemic administration of the compounds
would be required.
Our study shows that barbiturate compounds when
complexed with cobalt (Co+2) metal at specific position show
antibacterial activity against multiple drug resistant E. coli
and S. aureus.
ACKNOWLEDGEMENT
Fig. (2). Antibacterial effects of (10) and (11) on Multidrug
Resistant Uropathogenic E. coli.
H. Bokhari wants to thank Higher Education
Commission of Pakistan (HEC) for providing research
support to carry out this work. Furthermore, A. M. Qureshi is
extremely grateful to Bahauddin Zakriya University to
provide generous funding and support for carrying out the
work related with chemistry.
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