28 (s, 3H, CH3), 2.32 (s, 3H, CH3), 6.08 (s, 2H, OCH2O), 6.97–7.00 (d, 1H, ArH), 7.15 (s, 1H, Pyrrolic ArH), 7.28–7.60 (m, 5H, ArH), 7.80–8.00 (d, 2H, ArH), 8.18 (S, 1H, Pyrrolic ArH), 11.56 (s, 2H, Pyrrolic NH, CONH); 13C NMR (75 MHz, DMSO-d6) δ (ppm): 8.4,10.8, 101.5, Antiinfection Compound Library 116.3, 121.6, 123.2, 126.2, 128.6, 129.3, 144.2, 146.3, 147.9, 152.7, 157.0; MS (ESI) m/z: 390.17 [M + H]+. In vitro antimicrobial activities of the

formazan derivatives (2a–j) were determined using different microorganisms by micro broth dilution assay. 18 and 19 The microbial strains Escherichia coli NCIM 2065, Pseudomonas aeruginosa NCIM 5031, Salmonella typhi NCIM 2501, Klebsiella pneumoniae NCIM 2957, Bacillus subtilis NCIM 2699,

Aspergillus flavus NCIM 549, Aspergillus fumigatus NCIM 902, Aspergillus niger NCIM 620 were obtained from the National Chemical Laboratory, Pune, India. The bacteria were maintained on nutrient broth (NB) and fungal strains were maintained on Sabouraud dextrose broth at 37 °C. For bacteria – the bacterial strains used as inoculums were grown at 37 °C to get optical density 0.6 at 600 nm. Colony forming units (CFU) were counted by using serial plate Hydroxychloroquine in vitro dilution method and bacterial counts were adjusted to 1 × 105 to 1 × 106 CFU/ml for susceptibility testing. For fungus – the fungal inoculums were prepared from 10 days old culture grown on potato dextrose agar medium. The Petri dishes were flooded with 8–10 ml of distilled water and the conidia were scraped using sterile spatula. The spore density of each fungus was adjusted with spectrophotometer (A595 nm) to obtain a final concentration approximately 105 spores/ml. Minimum Inhibitory Concentration (MIC) determination was carried out using micro broth dilution method20 as per NCCLS guidelines. The test was performed in 96-well culture plates (Hi-media). The compounds were dissolved in DMSO to make eight different concentrations viz. 30, 15, 7.5, 3.75, 1.875, 0.9375, 0.46875, 0.2344 mg/ml

in the wells by twofold dilution method. Further dilutions 1, 0.5, 0.25, 0.125, 0.0625, 0.03125, 0.0156, 0.0078 mg/ml were prepared for the compounds 2c mafosfamide & 2h for A. flavus, 2g for all strains of bacteria and fungi, 2i for P. aeruginosa, E. coli, K. pneumonia, B. subtilis & A. flavus, 2j for K. pneumonia & A. flavus which did not shows activities in the concentration range 30–0.23 mg/ml. Negative control was prepared using Dimethyl sulphoxide, and concentrations of Tetracycline for bacteria and Amphotericin B for fungus were used as positive control. The 96 well plates were incubated for 24 h and 48 h at 37 °C for bacteria and fungus respectively. The lowest concentration of each compound which inhibited any visual growth was considered to be the MIC of that respective compound. All such experiments were repeated thrice.