The NMR Study and Antimicrobial Activity of Some Schiff Bases Derived From Sulphonamide Drug

Some Schiff base compounds derived from sulfonamide drug were synthesized by reaction of 4aminobenzenesulfonamide with aromatic aldehydes (2-hydroxy-1-naphthaldehyde, 3,4-dihydroxybenzaldehyde and 2hydroxy benzaldehyde) in good yields. Characterization of synthesized compound was carried by elemental analysis, IR, 1 H, 13 C, HSQC and HMBCNMR spectroscopy. The synthesized compounds were screened for their antibacterial activity against Staphylococcus aureus, Streptococcus sp., Bacillus subtillus, Escherichia coli and Klebsiella pneumonia. Additionally, the compounds were tested for antifungicidal activity against Candida krusei, Candida tropicalis, Aspergillus fumigates and Aspergillus niger.


INTRODUCTION
Sulphonamides were the first drugs found to act selectively and could be used systematically as preventive and the therapeutic agents against various diseases [1]. In medicine, the term "sulfonamide" is sometimes used as a synonym for sulfa drug, a derivative or variation of sulfanilamide. The first sulfonamide was discovered in 1932 [2]. The condensation products of sulpha drugs with aldehydes and ketones are biologically active [3,4].
Schiff bases are used as pigments and dyes, catalysts, intermediates inorganic synthesis and as polymer stabilizers. A number of Schiff's base molecules show biological activities including antibacterial, antifungal, antidiabetic, antitumour, antiproliferative, anticancer, anti-corrosion and anti-inflammatory activities [5][6][7][8]. Sulfa Schiff bases have been subject to thorough studies where a wide diversity of these derivatives have been prepared and used in various biological and pharmacological fields [9][10][11]. The aim of present work is to synthesis of some Schiff base derived from sulphonamide (Scheme1) and study the biological activity theoretically (Molecular modiling and in vitro antimicrobial activity.

EXPERIMANTAL a-Physical mesurments
Infrared spectra (IR) were recorded as KBr discs in the range of 4000-400 cm -1 using FT-IR spectrophotometer Shimadzu model IR. Affinity-1 at the department of Chemistry, College of Education for pure sciences, University of Basrah, Iraq. 1 H, 13 C, Roesy, HSQC and HMBC NMR spectra were measured on a Brucker at 600 MHz, with TMS as internal reference at Konstanz university, Germany. Microanalysis for carbon, hydrogen and nitrogen were carried out by a Perkin-Elmer 240B Elemental Analyzer. Melting points were measured by a Philip Harris melting point apparatus and uncorrected.

b-Synthesis
General Synthesis of Schiff-bases5-7 4-aminobenzenesulfonamide 1 (1.37, 2.00 mmol) and aromatic aldehydes (2-4) (2.1 mmol) were dissolved in absolute ethanol followed by addition of catalytic amount of glacial acetic acid dropwise and the mixture was heated under reflux for 4h. The reaction mixture was then cooled in an ice bath and the crude product thus obtained was collected by filtration, further purified by recrystallization from ethanol.

c-Antimicrobial activity
The synthesized compounds were screened in vitro for their antibacterial activity against:Staphylococcus aureus, Streptococcus sp., Bacillus subtillus, Escherichia coli and Klebsiella pneumonia. Additionally, the compounds were tested also for antifungal activity against Candida albicans, Candida tropicalis, Aspergillus fumigates and Aspergillus niger using the paper disc-agar diffusion technique on Muller Hinton agar and Sabouraud dextrose agaras a culture media for antibacterial activity and antifungal activity, respectively [12]. The test compounds were dissolved in DMSO solvent and recommended concentrations (50, 100 and 200 μg/mL) were used in the disc-agar diffusion technique. Antibiotic drug Ampicillin and Nystatin were used as control for bacteria and fungi, respectively. Petri plates containing 20 mL of Mueller Hinton Agar were used for all the bacteria tested. Fungistrains were cultivated in Sabouraud dextrose agar. Sterile Whatman no. 1 filter paper disks (6mm in diameter) impregnated with the solution in DMSO of the test were placed on the Petri plates. A paper disk impregnated with dimethyl sulfoxide (DMSO) was used as negative control. The plates were incubated for 24 h at 37 o C in the case of bacteria and 72 h that 27 o C for fungi. The inhibition zone diameters were measured in millimeters. The bacteria and fungi were supplied from department of Microbiology, College of Veterinary Medicine, University of Basrah.

Chemistry
Treatment of 4-aminobenzenesulfonamide1 with three aldehyde derivatives (2-hydroxy-naphthaldehyde 2, 3,4dihydroxybenzaldehyde 3 and 2-hydroxybenzaldehyde 4 in ethanol and catalytic by 3-4 drops of glacial acetic acid under reflux afforded the desired imine derivatives 5-7 in 82, 78 and 72 % yields, respectively (Scheme 1).The structures of the synthesized compounds were assigned by the elemental analysis (CHN),IR and 1 H, 13 C and 2D NMR. The IR spectra confirm the presence of the azomethine group (-CH=N) stretching with a sharp region for compounds 5-7 at 1602,1598 and 1600 cm -1 , respectively. In addition, the bands at the region 1622-1610 cm -1 were assigned to the C=C aromatic group. In the 1 H NMR spectra of compounds 5-7, the singlets at δ9.69, 9.71 and 8.97 ppm respectively, were assigned for the imine protons (CH=N). The multiplets at the regions δ 8.52-6.59 ppm were attributed to the aromatic protons.
In the 13       The gradient-selected 1 H, 13 C, HMBC NMR spectrum of compound 5 revealed two 1,3 JC,H . Thus, the imino proton (CH=N) at δ9.70 ppm showed two 1,3 JC,H correlations: first one with C-1 of the naphthyl ring at δ160.3 ppm, the second correlation with the aromatic carbon atom C-6' at δ 109.3 ppm and the last one with the aromatic carbon atom C-1' at δ147.2 ppm.
Othercorrelations between protons and carbon atoms can be assigned in Figure 5.
The HSQC and HMBC-NMR Spectra of compounds 6 and 7 were supported the structures of synthesized compounds, Figures 7,8,10 and11. D e c e m b e r 2 3 , 2 0 1 5

Antimicrobial activity
The studied compounds have been evaluated in vitro for their antibacterial and antifungal activities, using the paper discagar diffusion technique [12] by measuring the inhibition zone in mm. Antibiotic drug ampicillin and Nystatin were used as control for bacteria and fungi, respectively. The antibacterial activity of the synthesized compounds were tested against three Gram positive bacteria (Staphylococcus aureus,Streptococcus sp., Bacillus subtillus) and two Gram negative bacteria (Escherichia coli and Klebsiella pneumonia) at a concentration of 50, 100 and 200μg/ml using DMSO as a D e c e m b e r 2 3 , 2 0 1 5 solvent, which not effected in the growth of microbes. Mueller Hinton agar and Sabouraud dextrose agar were used as culture media for antibacterial and antifungal activity respectively. The results of the antimicrobial activity are shown in Table (1).
The screening results indicate that the activity of compounds increases with an increase in the concentration of the solutions. The synthesized compound 1 show high activity against E.coli and Streptococcus spp additionally, the compound 1 also exhibit high biological activity against all tested fungi. Whereas the compound 2 show relatively a good activity against E.coli and all fungi. On the other hand, the compound 3 also show a good biological activity against E.coli and high antifungal activity against the Candida tropicalis, and Aspergillus niger, Table 1.
Concerning to these findings, the possible explanations of our results attribute to the fact that different antibiotics, chemical compounds and drugs have different modes of action, owing to the nature of their structure and degree of attraction to certain objective sites within bacterial and fungi cells walls and membranes.