The frequency of life threatening infections such as tuberculosis, cancer, AIDS etc caused by pathogenic microorganisms is increasing worldwide and becoming an important cause of morbidity and mortality in immuno-compromised patients. Synthetic compounds constitute an important source of various bioactive compounds such as antibacterial (Zakaria, 2000; Biswas et al.,2002) antifungal and anticancer (Pratt, 1979) compounds. The synthesized compounds which are used for the treatment of infectious diseases are known as chemotherapeutic agents. Every year thousands of compounds are synthesized with an aim to find a potential chemotherapeutic agent to combat pathogenic microorganisms. But very few compounds are withstood as therapeutic agent for various methodological tests. Antimicrobial screening is one of these tests required to perform for primary selection of compounds as the therapeutic agents.
The antimicrobial screening is necessary to find out the suitable candidate of therapeutic agent among the synthesized compounds. Usually many compounds possess antimicrobial properties but have serious toxic effects to the host, therefore in the ideal cases, the drug should be highly toxic to the parasite and completely atoxic for the host. In the continuation of our ongoing efforts aimed to find new compounds for chemotherapy (Islam et al., 2001; Biswas et al., 2002), five new synthesized compounds are selected for antimicrobial screening.
Materials and Methods
Source of Compounds: The compounds used in the present study were synthesized according to the following general procedure:
Preparation of cadmium coordination complexes [Cd(DPH)(LG*)x]
An ethanolic solution of cadmium (II) chloride (1.833g; 1.0 mmol) and deprotonated Phthalic acid (DPH, 1.661g; 1.0 mmol) were mixed in the 1:1 ratio with constant stirring. Then an ethanolic ligand solution [LG*] x (a variable ligand) was added to the resulting mixture and heated gently on a magnetic regulator hot plate with constant stirring. As a result the volume of the solution was reduced to one half and then allowed to cool. The precipitates formed were filtered, washed several times with ethanol and then dried in a vacuum desiccators charged with anhydrous CaCl2.
the compound is [Cd(DPH)(Py)2, C1],
the compound is [Cd(DPH)(8-HQ), C2] and
[LG]x = (Isoquinoline)2
the compound is [Cd(DPH)(IQ), C3].
Preparation of addition compound 1:1 antimony (III) chloride with acetophenone [SbCl3 .C6H5COCH3, C4]
A solution of acetophenone (1.201g, 1.0 mmol) in absolute alcohol was added drop wise with gently stirring to an ethanolic solution o f antimony (III) chloride (2.281g, 1.0 mmol) in warm condition .The resulting mixture was refluxed for seven hours and allowed to cool . Then the volume of this solution was reduced to one half by heating on a hot plate and cooled in a freeze for two days. The precipitate obtained was filtered, washed with ethanol and dried in a vacuum desiccator charged with CaCl2.
Preparation of addition compound 1:1 Arsenic (III) bromide with benzamide, [AsBr3.C6H5CONH2, C5]
The benzamide solution (1.21 g, 1.0 mmol) in absolute alcohol was mixed drop wise with gently stirring to an alcoholic solution of arsenic (III) bromide (3.14g, 1.0 mmol) in warm condition. The resulting mixture was refluxed for seven hours and allowed to cool. Then the volume of this solution was reduced one half by heating on a hot plate and cooled in a freeze for two days. The precipitate obtained was filtered, washed with absolute alcohol and dried in a vacuum desiccator charged with CaCl2.
Antimicrobial screening: Disc diffusion method (Bauer et al., 1966; Barry, 1980; Rios et al., 1988), is a widely accepted procedure for the in-vitro investigation of the susceptibility of microorganisms to the compounds, so this method is adopted in this investigation. The method is essentially a qualitative or semiquantitative test indicating sensitivity or resistance of microorganisms to the test materials as well as bacteriostatic or bactericidal activity of a compound (Reiner, 1982).
The standard test microorganisms were collected from the Department of Microbiology, University of Dhaka and ICDDRB, Dhaka, Bangladesh. The diameters of zones of inhibition produced by the compounds were compared with standard antibiotics (Kanamycin 30 μg disc1 and Nystatin 200 μg disc1). The experiment was performed in duplicate to minimize errors.
Minimum inhibitory concentration (MIC) of a compound is defined as the lowest concentration of that compound in a medium without visible growth of the test organisms. The basic principle is the dilution tests which comprises the serial dilution of the antimicrobial agent inoculated with the organism. For the test, standard serial dilution technique (Reiner 1982) was employed. The media used in this respect were nutrient agar & nutrient broth (DIFCO).
Results & Discussion
Antibacterial activity: The inhibitory activity of the compounds C1, C2, C3, C4 & C5 was determined at a concentration of 100μg/disc against a series of Gram positive and Gram negative pathogenic bacteria. The results are shown in Table 1. Results revealed that all the tested compounds have strong activity against most of the Gram positive & Gram negative bacteria. Among the tested materials, the compound C4 (i.e.SbCl3.C6H5COCH3) was found to be most active than others. When compared with standard antibiotic Kanamycin at the same concentration of 30 μg disc1, the compound C4 also has shown substantial antimicrobial activity.
In Table 1, the antibacterial activity of the three synthetic cadmium coordination complexes against 14 pathogenic bacteria is presented. Among these three complexes the antibacterial activity of compound C2 is greater than compound C1 and compound C3. So the structural activity correlation with antibacterial screening data reveal that the cadmium complexation with the ligand 8- hydroxy quinoline rather than pyridine and isoquinoline is necessary for greater antibacterial activity (activity of the compounds can be represented sequentially, C2 > C1> C3). On the other hand, among the two addition compounds C4 and C5, the antibacterial activity is approximately same in both compounds but in case of compound C5 the antibacterial activity against Sarcina lutea and Shigella dysenteriae is so much lower than the compoundC4. So we can select the compound C4 for further tests such as subacute toxicity as for suitable therapeutic antibacterial agent.
Antifungal activity: The compounds C1, C2, C3, C4 & C5 were tested against the pathogenic fungi at a concentration of 200 μg disc1 for each and the result was compared with standard antibiotic Nystatin 200 μg disc1. The antifungal activity was determined after 72 hours of incubation at room temperature (30°C) and the obtained results are cited in Table 2. It was observed that three compounds i.e. C1, C2, & C4 showed more antifungal active than other two compounds C3 and C5.
Minimum Inhibitory Concentration (MIC): The minimum inhibitory concentration (MIC) of the compounds C1, C2, C3, C4 and C5 was determined against Shigella dysenteriae, Salmonella typhi, Bacillus subtilis and Streptococcus β-haemolyticus by serial dilution technique and the values were shown in the Table 3. The MIC values of the compound C1 against Shigella dysenteriae, Salmonella typhi, Bacillus subtilis and Streptococcus β-haemolyticus were 64, 32, 32 and 64 μg ml1 respectively, for compound C2, 64, 32, 16 and 32 μg ml1 respectively, for compound C3, 128, 64, 32 and 32 μg ml1 respectively, for compound C4, 16, 8, 8 and 4 gμ ml1 respectively and that for compound C5, 64, 16, 64 and 32 μg/ ml respectively. From the MIC values, it was found that the compounds C2, C4 and C5 were more potent against Bacillus subtilis and Salmonella typhi.
It was concluded that among the tested compounds, the compound C4 (i.e. SbCl3.C6H5COCH3) possesses substantial antimicrobial activity with a minimum inhibitory concentration. By comparing the results with previously published results (Biswas et al., 2002) of antibacterial activity of benzene sulfonic acid derivative complex compounds, we can say that our tested compounds are more superior for selection of a suitable chemotherapeutic agent.
|Table 2:||In vitro antifungal activity of the compounds C1, C2, C3, C4, C5 and Nystatin
|Table 3:||MIC values of the compounds C1, C2, C3, C4 and C5, against two Gram positive and two Gram negative bacteria
Further, acute toxicity and other pharmacological tests are necessary to utilize the compound C4 as a potential therapeutic agent.