INTRODUCTION:

The discipline of medicinal chemistry is devoted to the discovery and development of new agents for treating diseases. Most of this activity is directed to new natural or synthetic organic compounds. Paralleling the development of medicinal agents has come a better understanding of the chemistry of the receptors. The latter has been greatly facilitated by low-cost computers running software that calculates molecular properties and structure and pictures it using high-resolution graphics. Development of organic compounds has grown beyond traditional synthetic methods. It now includes the exciting field of biotechnology using the cell’s biochemistry to synthesize new compounds. Techniques ranging from recombinant DNA and site-directed mutagenesis to fusion of cell lines have greatly broadened the possibilities for new entities that treat disease. It has become possible to develop and dispense modified human insulin that provide more convenient dosing schedules, cell-stimulating factors that have changed the dosing regimens for chemotherapy, humanized monoclonal antibodies that target specific tissues, and fused receptors that intercept immune cell–generated cytokines ¹.

Infectious diseases seemed to be as old as life itself. They have certainly played a major part in shaping human history, not only because of the decimating effects of the various plagues through the centuries, but also because of the intense efforts made to find cures for them, thus advancing medical science. In the past 200 years, empirical science and serendipity have combined to bring us to our current state of knowledge. Infectious diseases caused by bacteria and fungi affect millions of people worldwide and in the United States alone cause a disease burden of more than $20 billion annually. Concerted and systematic programs to discover and develop new antibiotics and anti-fungals have been driven to a considerable extent by the development of resistance by these organisms to the drugs commonly used against them.

The inhibition of bacterial DNA gyrase has been the target of a worldwide research effort which began with the discovery of nalidixic acid in the early 1960s. Quinolines are a class of organic compound of the hetero aromatic series characterized by a double-ring structure composed of benzene and a pyridine ring fused at two adjacent carbon atoms. Structure activity relationships (SAR) of compounds based on nalidixic acid have led to a large group of synthetic antibacterial agents known collectively as the quinolones. The compounds prepared early in this effort such as oxolinic acid, rosoxacin and pipemidic acids were most active against gram negative bacteria. Introduction of a fluorine atom at the 6- position of the quinoline ring system led to norfloxacin, which had a broad spectrum antibacterial activity. Relatively newer members of this family include ciprofloxacin, ofloxacin, enoxacin, lomefloxacin, fluroxacin and temafloxacin. Different quinoline derivatives were found to be reported as antimicrobial², antimalarial^3,4, cytotoxic agents^5,6, antitumors⁷ etc.

The benzimidazoles are known also as benziminazoles or benzoglyoxalines. They have been named also as derivatives of o-phenylenediamine, especially in the early literature⁸. Benzimidazole and their derivatives were reported to have wide biological activities like antifungal⁹, antibacterial¹⁰, antianxiety¹¹, inhibitors of gastric acid secretion¹², anti-inflammatory^13,14 activities etc. Benzimidazoles are very useful intermediates for the development of Molecules of biological interest.

Since benzimidazole and quinoline derivatives both are reported to possess antimicrobial and antifungal activities, it is speculated that the presence of both these two skeletons in a single molecular framework should also have appreciable antimicrobial property. Therefore in the perspective of development of some novel antimicrobial agents, reliancing the literatures, in our present study we propose to synthesize a few quinolino substituted benzimidazole derivatives as possible anti bacterial agents.

MATERIALS AND METHODS:

All the chemicals used to synthesize the title compounds were of analytical grade. Melting points were determined by open capillary tube method and are quoted as uncorrected values. The FT-IR spectra of the synthesized compounds were recorded on a Shimadzu 8400 S FTIR spectrophotometer using KBr discs in the region of 4000-400 cm^-1. NMR spectra were recorded in CDCl₃ at 60 MHz on a Brukar Spectrospin 200 spectrometer using TMS as internal standard. Mass spectral studies were carried out using JEOL GC mate instrument. Thin Layer Chromatographic studies were performed using precoated Silica gel G TLC plates to check the purity of the compounds (Benzene: Chloroform: Ethyl acetate= 1:1:1 as mobile phase), and detections were either done in UV chamber or by using iodine vapour.

Scheme of synthesis:

Synthesis of quinoline-4-carboxylic acid (1a)¹⁵.

Purified acetaldehyde (0.059 mol), freshly distilled Pyruvic acid (0.0625 mol) and 50ml of absolute ethanol were placed in a round bottomed flask. The mixture was then heated to the boiling point on a water bath and added slowly, a solution of p-nitro aniline (0.62 mol) in 25 ml of absolute ethanol. The mixture was then refluxed for 3hrs and allowed to stand over-night. The product was filtered off and washed with a little cold ether. Then it was recrystallized from ethanol. Yield 53%. Compounds 1b-h were synthesized in a similar manner.

IR data of 1a (KBr, cm^-1): 3609 (O-H str), 3035 (C-H str aromatic), 2918 (C-H str aliphatic), 1720 (C=O str), 1656 (C=N str), 1687 (N-O str), 1499 (C=C str aromatic), 1335 (C-H bending aliphatic), 1237 (C-N str), 787 (C-H bending aromatic). ¹HNMR data of 1a (CDCl₃, δ): 2.5 3H s CH₃, 6.7-6.8 2H Ar H, 7.0-7.2 2H m Ar H, 10.2-10.3 1H s COOH. Mass (M+1): 233

Synthesis of substituted quinolino benzimidazole compounds (2a)¹⁶.

o-Phenylenediamine (0.08 mol) was taken in a round bottomed flask and added quinoline-4- carboxylic acid (1a) (0.08 mol). The mixture was then heated at 100⁰C for 3 hrs. The resulting solution was treated with sodium hydroxide it was just alkaline to litmus. The product was filtered and washed with ice-cold water. The crude product was dissolved in boiling water and treated with de-colorizing carbon. The benzimidazole derivative (2a) appeared to precipitate after cooling. It was washed with cold water and dried at 100⁰C. Yield 70%. In a similar manner compounds 2 b-h were synthesized.

IR data of 2a (KBr, cm^-1): 3612 (O-H str), 3486 (N-H str), 3041 (C-H str aromatic), 2912 (C-H str aliphatic), 1721 (C=O str), 1659 (C=N str), 1691 (N-O str), 1527 (N-H bending), 1489 (C=C str aromatic), 1336 (C-H bending aliphatic), 1237 (C-N str), 794 (C-H bending aromatic). ¹HNMR data of 2a (CDCl₃, δ): 2.5 3H s CH₃, 3.01 1H NH, 6.6-6.9 4H m Ar H, 7.1-7.4 4H m Ar H, Mass (M+1): 305

Reaction of quinolino benzimidazole compounds with DMF-POCl₃(3a) ¹⁷

The substituted quinolino benzimidazole compound (2a) (0.01mole) was added to the Vilsmier Haac reagent at 0⁰C. The reaction mixture was stirred at room temperature for 2 hr. Then it was treated with cold aqueous sodium carbonate solution and heated to 90⁰C. After cooling down to room temperature the solution was extracted with chloroform. The chloroform layer was dried over anhydrous sodium sulfate. The combined extracts were evaporated to dryness and purified by recrystallization from rectified spirit. Yield 50%. Compounds 3b-h were prepared following this method.

IR data of 3a (KBr, cm^-1): 3028 (C-H str aromatic), 2902 (C-H str aliphatic), 1731 (C=O str), 1689 (N-O str), 1657 (C=N str), 1494 (C=C str aromatic), 1339 (C-H bending aliphatic), 1190 (C-N str), 814 (C-H bending aromatic). ¹HNMR data of 3a (CDCl₃, δ): 2.5 3H s CH₃, 6.65-6.92 4H m Ar H, 7.21-7.38 4H m Ar H, 9.98 1H s CHO. Mass (M+1): 333

General procedure for the synthesis of title compounds (4a-l):

A mixture of 3a (2.35mmol) and methyl amine (2.35mmol) was stirred in about 5 ml of water at room temperature for 5 hrs and kept for 48 hours at cold condition. The crystalline solid obtained was collected and washed with ice-cold water and dried to give amino substituted quinolino benzimidazole derivatives (4a). Then it was recrystallized from ethanol. Compounds 4b-l were prepared similar to this method. The physical data are reported in table 2.

IR data of 4a (KBr, cm^-1): 3031 (C-H str aromatic), 2921 (C-H str aliphatic), 1689 (N-O str), 1675 (C=N str), 1494 (C=C str aromatic), 1334 (C-H bending aliphatic), 1194 (C-N str), 824 (C-H bending aromatic). ¹HNMR data of 4a (CDCl₃, δ): 2.43 3H s CH₃, 3.59 3H s N-CH₃, 5.87 1H s N=CH, 6.35-6.82 4H m Ar H, 7.31-7.54 4H m Ar H. Mass (M+1): 346

Antibacterial screening^18-20:

All the synthesized title compounds were subjected to screen in DMF solution for their antibacterial activity by cup plate Agar diffusion method against Staphylococcus aureus, Pseudomonus aeruginosa, Bacillus subtilis and Escherichia coli. Nutrient agar plates were prepared using sterilized petridishes by pouring melted agar media onto the petridishes and allowed to solidify. Then it was inoculated over the surface of agar media with sterile cotton. The cups are then made with the help of borer and filled with solution of suitable concentration of sample and standard and incubated at 37⁰C for 24 hours. The antimicrobial agents diffuse through the agar media around the cups made and produce a characteristic zone of inhibition of the microbial growth which was then measured and represented (Table 3) depending upon their activity and the diameter of the inhibited zone in microbial growth. The control (CHCl₃) with solvent (DMF) in identical condition showed no activity. Norfloxacin was used as a standard.

RESULTS AND DISCUSSION:

Structural modification of quinolino substituted benzimidazole derivatives was carried out by substituting the amino group of benzimidazole with different aldehydes followed by the reaction with two amines. The derivatives were synthesized successfully in moderate to good yields. The structures of the title compounds (4a-l) were confirmed by their IR and ¹H NMR and Mass spectral analysis. The IR spectra of the synthesized title compounds (4a-l) shown the characteristic absorption band for the vibrations like C-H str aromatic around 3030 cm^-1, C-H str aliphatic around 2920 cm^-1, N-O str around 1680 cm^-1, C=N str around 1670 cm^-1, C=C str aromatic around 1490 cm^-1, C-H bending aliphatic around 1340 cm^-1, C-N str around 1190 cm^-1, and C-H bending aromatic around 825 cm^-1. The ¹H NMR spectrum of 1a revealed the presence of a singlet peak at 10.2-10.3 which was assigned as the acidic proton at the 4^th position of the quinoline ring system. Progressing to the next step, the proton signal for acidic hydrogen of 1a disappeared in the ¹H NMR spectrum of 2a with the appearance of a NH proton of benzimidazole part, thus confirming the cyclisation of the carboxylic acidic group with o-phenylene diamine to give quinoline benzimidazole moiety (2a). A close observation of the proton signals of 3a revealed the presence of an aldehydic proton which was noticed further to appear at far downfield. This behavior of the aldehydic proton may be attributed to the fact that presence of electronegative oxygen with a pair of π electron framework made the proton to be acidic enough to appear at downfield. The ¹H NMR spectrum of 4a exhibited a signal for three methyl protons (N-CH₃) appeared at 3.59 which is generally a bit higher downfield value than the normal alkyl protons; deshielding due to the direct attachment of the -CH₃ group with electronegative nitrogen resulted the peak for 3 protons to appear so. Considering the relatively less electronegativity of nitrogen than oxygen it was ascertained that the N=CH proton (4a) appeared at somewhat up field when compared to the aldehydic proton of 3a. All the other aromatic and aliphatic protons were appeared according to their extent of shielding and deshielding.

It was worthy to note that some of the synthesized compounds like 4a and 4e were appreciably effective against all the four strains of bacteria, although with an extent of variation. The results of the antibacterial activity showed that compounds 4b and 4f were moderately active against P. aeruginosa. Remaining compounds were weakly active or inactive.

CONCLUSION:

It can be concluded that the some of the quinolino substituted benzimidazole derivatives with different amines showed appreciable antibacterial activity against the organisms used. Further in detailed study is required to design these compounds for its optimal structural requirements to derive more effective antibacterial agents.

Table- 1. Structure of various derivatives condensed with different amines:

Compound code	R1	R2	R3
4a
4b
4c
4d
4e	Cl
4f	Cl
4g	Cl
4h	Cl
4i
4j
4k
4l

Table- 2. Data of the synthesized derivatives:

Compound code	Mol. Formula	Mol. Wt	Melting point (⁰C)	R_fvalue	Yield (%)
4a		344	151	0.52	58
4b		358	167	0.62	51
4c		358	180	0.67	54
4d		372	192	0.74	57
4e		333	154	0.56	52
4f		347	178	0.59	59
4g		347	190	0.68	50
4h		361	153	0.53	56
4i		313	181	0.72	59
4j		327	166	0.51	50
4k		327	192	0.67	55
4l		341	157	0.58	58

Table- 3. Antibacterial activity of the synthesized compounds.

Compound code	Zone of inhibition
Compound code	S. aureus	P. aeruginosa	B. subtilis	E. coli
4a	++	+++	+++	++
4b	+	++	+	+
4c	+	+	-	+
4d	+	+	+	-
4e	++	+++	+++	+++
4f	+	++	+	+
4g	-	+	-	+
4h	+	-	+	+
4i	-	-	+	+
4j	-	+	+	-
4k	+	+	-	+
4l	+	+	+	+

= inactive, + = weakly active (10-13 mm), ++ = moderately active (14-17 mm), +++ = highly active (18-23 mm)

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Received on 02.07.2014 Modified on 20.07.2014

Asian J. Research Chem. 7(10): October- 2014; Page 841-845

*Corresponding Author E-mail: sukhen18@rediffmail.com

INTRODUCTION: