Microwave Assisted Synthesis, Characterization and Evaluation for Antimicrobial Activity of Novel 1, 5- Benzothiazepines

Aims: 1,5-benzothiazepine ring is privileged aromatic heterocycles of interest to organic and medicinal chemists because of its ease of synthesis and biological activities. This study aims to synthesize new series of 1, 5-benzothiazepine by direct and efficient microwave assistance and to evaluate for antimicrobial activity by MIC method. Place and Duration of Study: The study was conducted at Department of Pharmaceutical Chemistry, Chebrolu Hanumaiah Institute of Pharmaceutical Sciences, Guntur-19, A. P. from January, 2019 to October, 2021. Methodology: 1, 5-benzothiazepines (BT-21 to BT-40) were synthesized by microwave irradiation. The structures of the products were established by elemental analysis, FTIR, 1 H-NMR, 13 C-NMR and mass spectroscopic studies. The synthesized compounds were also evaluated for their Antimicrobial activity by MIC method. Results: The microwave assisted synthetic procedure adopted yielded the 1,5-benzothiazepine derivatives BT-21 to BT-40 in good amounts and at a lesser time span. The synthesized 1, 5benzothiazepine derivatives showed good to moderate antibacterial and antifungal activities.BT-25 having a dihydroxy-methyl-phenyl moiety proved to be more potent against all selected bacterial Original Research Article Kumar et al.; JPRI, 33(62A): 39-53, 2021; Article no.JPRI.81212 40 strains, B. subtilis, S. aureus, E. coli and P. aeruginosa with a MIC value of 64 μg/ml. BT-33 having fluorophenyl moiety, BT-35 having hydroxyl-nitrophenyl moiety and BT-40having dibromophenyl moiety proved to be more potent against all selected fungal strains, A. niger and C. tropicalis with a MIC value of 16 μg/ml. Conclusion: These results showed that the synthesized 1, 5-benzothiazepine derivatives have better scope for further development as antimicrobial agents.


INTRODUCTION
The design and synthesis of heterocyclic hybrids have received greater attention due to their ease of synthesis and improved biological properties [1]. The structures that evolve from such conjugation are usually rigid frameworks that can show the appended rings in a well-defined fashion that is necessary for molecular recognition of the biological target. Usually, the variable nature of these functionalities defines the selectivity on a privileged core for a particular target.1, 5-Benzothiazepines (1 and 2) are sevenmembered heterocyclic compounds containing nitrogen and sulphur as hetero atoms with diverse bioactivities [2][3][4][5][6]. 1,5-Benzothiazepines are the most well-known representatives of benzologs of 1,4-thiazepine (3) and one of the three possible benzo-condensed derivatives, viz. 1,4-(4), 4,1-(5) and 1,5-benzothiazepines [6][7][8][9][10]. 1,5-benzothiazepine ring is privileged aromatic heterocycles of interest to organic and medicinal chemists because of its ease of synthesis and biological activities.

MATERIALS AND METHODS
All the reactions were carried out under specified laboratory conditions. All the synthetic work was done by procuring laboratory grade reagents and analytical grade solvents. Pre-coated silica gel 60 F 254 plates were used for thin-layer chromatography (TLC) and the spots on the TLC plates were visualized by UV lamp (254 nm). The products were purified by recrystallization using suitable solvents. Melting points were determined by Digital melting point apparatus and were uncorrected.FT-IR spectra were recorded on Bruker Vertex 80v spectrometer using potassium bromide discs and the absorption band values are given in cm

General Procedure for Synthesis of 1, 3-substituted-prop-2-ene-1-ones [C-21 to C-40]
In this method, acetylated α-naphthol (0.01 mol), aromatic aldehyde (0.011mol) were taken in 5ml of ethanol and poured in 100ml Erlenmeyer borosil flask. To this reaction mixture, 4ml basic alumina was added. The reaction mixture was thoroughly mixed and irradiated inside a microwave for 2-3 minutes at medium level 600W. After completion of reaction, mixture was cooled and the product was extracted with ethanol ( Fig. 1).

General Procedure for Synthesis of 2,3-dihydro-2-substituted-4(naphthalene-2-ol)-yl--1,5-Benzothiazepines (BT-21 to BT-40)
A mixture of (0.01 mol) 1,3-substitutedprop-2-en-1-one and (0.01 mol, 1.25ml) 2-aminothiophenol and pinch of potassium acetate as catalyst were thoroughly mixed and taken in a clean borosil beaker. The solvent-free reaction mixture was then subjected to microwave irradiation for 2-3 minutes at 80-85 o C. The reaction mixture was then allowed to cool to room temperature and then poured cold water in the mixture and stirred vigorously. Products were washed with water to remove the catalyst, filtered, dried and recrystallized by ethanol (Fig.  2).

Antimicrobial Activity
The antimicrobial (antibacterial and antifungal) activities of the novel benzothiazepines was evaluated against selected bacterial and fungal strains using standard experimental procedures as described in the literature [35]. The standard strains were procured from the American Type Culture Collection (ATCC) and Gene Bank, Institute of Microbial Technology, Chandigarh, India. The bacterial strains selected for the study were Bacillus subtilis (ATCC-60511), Staphylococcus aureus (ATCC-11632), Escherichia coli (ATCC-10536), and Pseudomonas aeruginosa (ATCC-10145) whereas the fungal strains include Aspergillus niger  and Candida tropicalis (ATCC-1369). Ampicillin was used as positive control for antibacterial activity and fluconazole for antifungal activity. Antibacterial activity was performed using nutrient agar medium whereas Potato Dextrose-Agar medium was used for antifungal testing. 2.048 mg of each test compound was taken in vials separately. Then 2 ml of methanol was added. Thus, a solution with a concentration of 1.024 mg/ml was obtained. All the experiments were carried out in triplicate and the results are presented as the mean of three independent experiments. The microbial strains were grown at 37 °C in their respective nutrient medium and diluted in sterile nutrient broth medium to get a suspension containing 10 7 cells/ml and this suspension was used as the inoculum. All the test tubes were incubated for 18 h at 37 °C. A similar experiment with inoculum, medium and methanol without compound was furthermore performed to confirm that there is no inhibitory effect of methanol used for the dilutions. The test tube number in which the first sign of the growth of the organism observed was noted using a spectrophotometer. The MIC was determined for all the compounds by taking that concentration used in the test tube number just before the test tube number where the first sign of growth observed [36].

Antibacterial Activity
The antibacterial activity of the novel benzothiazepines was evaluated against selected bacterial strains using Ampicillin as positive control by MIC method. From the results (Table 3 & Fig. 3), it is evident that most of the 1,5-benzothiazepines synthesized showed antibacterial activity with different MIC values against the tested organisms, but not comparable with that of the standard. Among the compounds tested, BT-25 having a dihydroxymethyl-phenyl moiety proved to be more potent against all selected bacterial strains B.subtilis, S.aureus, E.coli and P.aeruginosa with a MIC value of 64 µg/ml. BT-40 having dibromophenyl moiety proved to be more potent against three selected bacterial strains S.aureus, E.coli and P.aeruginosa with a MIC value of 64 µg/ml. These results suggested that, the electron releasing groups present on phenyl moiety of the BT-25 compound affects the charge distribution, which confers significant improvement in biological effect. The enhanced inhibition observed is more likely due to its interaction with some intracellular target [37]. The presence of a strong electron-withdrawing groups in the compound BT-40 alter the nature of the compound in such a way as to promote binding to the target(s) [38].

Antifungal Activity
The antifungal activity of the novel benzothiazepines was evaluated against selected fungal strains using Fluconazole as positive control by MIC method. From the results (Table 4 & Fig. 4), it is evident that most of the 1, 5-benzothiazepines synthesized showed antifungal activity with different MIC values against the tested organisms, but not comparable with that of the standard. Among the compounds tested, BT-33 having fluorophenyl moiety, BT-35 having hydroxyl-nitrophenyl moiety and BT-40having dibromophenyl moiety proved to be more potent against all selected fungal strains, A.niger and C.tropicalis with a MIC value of 16 µg/ml. BT-25 having dihydroxymethylphenyl moiety and BT-26 having dihydroxyphenyl moiety proved to be more potent against all selected fungal strains, A.niger and C.tropicalis with a MIC value of 32 µg/ml. The presence of substituents containing an electron withdrawing group that is surrounded by high electron density might be the reason for the high antifungal activity of the benzyl tested compounds [39].

CONCLUSION
The microwave assisted synthetic procedure adopted was afforded the 1, 5-benzothiazepine derivatives BT-21 to BT-40 in good yield at the cost of shorter reaction time. The synthesized 1, 5-benzothiazepine derivatives showed good to moderate antibacterial and antifungal activities.Out of the synthesized compounds, molecules bearing electron releasing group showed good antibacterial activity and molecules bearing electron withdrawing groups showed good antifungal activity.The compounds BT-25, BT-33 and BT-35 are the promising molecules with antimicrobial properties and have better scope for further development as antimicrobial agents and potency of these compounds is required to be confirmed further by in-vivo screening.

CONSENT
It is not applicable.

ETHICAL APPROVAL
It is not applicable.