Synthesis of Novel Benzimidazolyl-substituted Acrylonitriles and Amidino-substituted Benzimidazo[1,2-a]Quinolines

A series of novel benzimidazole derivatives 3-10 were synthesized. Benzimidazolyl-substituted acrylonitriles 3 and 4 underwent a photochemical dehydrocyclization reaction to give the corresponding mono- and dicyano-substituted benzimidazo[1,2-a] quinolines 5 and 6. Pinner reaction of these compounds did not give the expected mono- and diamidines, but rather only compounds 7-10, with amido groups at 6-position were isolated. A mechanism for the reaction is proposed. Acyclic compounds 3 and 4, as well as cyclic benzimidazo[1,2-a]quinolines 5-8, exhibit interesting spectroscopic properties and are potential biologically active compounds.


Introduction
Over the past years substituted benzimidazoles and their azino fused derivatives have been one of the most extensively studied classes of heterocyclic compounds, receiving much attention from synthetic organic chemists because of their applications in several areas such as optical laser and polymer dyes, fluorescent tags in DNA sequencing, cosmetic ingredients, etc. [1 -2].Benzimidazoles also posses broad spectrum of biological properties such as antiviral (anti-HIV), anticancer, antibacterial, antifungal and many others [3][4][5][6][7][8][9][10][11][12][13].Benzoannulated benzimidazole analogues such as benzimidazo[1,2-a]quinolines or benzimidazo[1, 2-a]quinazolines, due to their planar chromophore, have the ability to undergo an intercalation process and thus become inserted between adjacent base pairs of a DNA molecule [14][15], or have potential applications as fluorescent probes in homogeneous assays of biological molecules, as well as in optoelectronics [16].A positive charge is generally needed for activity and it can be provided by quaternization of nitrogen atom or by introduction of substituents containing such a positive charge, such as an amidino group.Amidines, the nitrogen analogues of carboxylic acids, are structural components of numerous compounds of biological interest, including important medical and biochemical agents [17][18][19].Herein we present the synthesis and preliminary spectroscopic properties of some novel potentially biologically active acyclic and cyclic benzimidazole derivatives.

Scheme 1.
In the photochemical dehydrocyclization reaction of 3 and 4 (ethanolic solution, c = 1.3 × 10 -3 mol dm -3 ) cyclic bezimidazo[1,2-a]quinolines 5 and 6 were prepared [23].Irradiation was performed for about 12-15 h with a 400-W high-pressure mercury lamp using a Pyrex filter under oxidative conditions, with a small amount of iodine as a catalyst.The photochemical dehydrocyclization reaction was followed by UV/Vis spectroscopy.Cyclic derivatives were obtained in good yields (40-50%).In the 1 H-NMR spectra of both cyclic compounds it was noted that the cyclization reactions lead to a downfield shift of the signal of the ethylenic proton and most other protons.The proton of the NH group of the benzimidazole ring, as well as one proton of the phenyl ring disappeared, which also confirmed the cyclic nature of the products.In the 1 H-NMR spectra of acyclic compound 4 two doublets at 8.11 and 8.04 ppm, corresponding to four phenyl protons, were observed, while in the spectra of compound 6, due to the cyclization reaction, two doublets and one singlet for three phenyl protons were observed.
Compound 5 had been previously prepared by classic thermal cyclization [24].In the Pinner reaction [25][26], carried out under acidic conditions and using absolute EtOH as well as carbitol as solvents, compounds 7-10 were prepared.The formation of the imino-ester was confirmed by the disappearance of a nitrile band at ~2200 cm -1 in the IR spectrum.Attempts to prepare monoamidino-(amidino group at position 6) or diamidino (amidino groups at positions 2 and 6)-substituted benzimidazo[1,2-a]quinolines were not successful.Instead of the desired 6-amidino-benzimidazo[1,2a]quinolines, the benzimidazo[1,2-a]quinoline-6-carboxamide 7 was unexpectedly isolated [27].After isolation and identification of the crude imino-ester by NMR and MS, we confirmed that the amido group at the 6-position is formed immediately in the first phase of Pinner reaction.In 1 H-NMR spectra of compound 7 two novel singlets can be observed for the amide protons.From the Pinner reaction of dicyano-substituted benzimidazo[1,2-a]quinoline 6 only 2-amidino-substituted-benzimidazo[1,2-a]quinoline-6-carboxamides 8-10 were isolated.Their structures were confirmed by their NMR and MS spectra.In the 1 H-NMR spectrum, singlets for amidine protons can be observed in the 11.63 -9.60 ppm region.The presence of the corresponding amidine group in compounds 8, 9 and 10 lead to downfield shifts of most proton signals.In the IR spectrum of amidino compounds 8, 9 and 10, bands in the ~3500 cm -1 region can be observed.
It appears that the cyano groups at position 2 and 6 reacted quite differently under the "Pinner" conditions used.The cyano group at the 2-position of the cyclic benzimidazo[1,2-a]quinoline 6 gave in the first phase an imino-ester, while in the second phase, after reaction with corresponding amine, an amidino-substituted group is obtained.The cyano groups at the 6-position of the cyclic benzimidazo[1,2-a]quinolines 5 and 6 gave, in the second phase of the Pinner reaction, an unsubstituted amido group instead of corresponding amidine.The amido group at the 6-position is also formed immediately in the first phase of Pinner reaction.The proposed mechanism of the abovementioned reaction is presented in Scheme 2. The intermediate protonated imino-ester, formed at the 6-position, underwent an S N -transformation by ethanol giving an amide.
In order to study the spectroscopic properties of substitued benzimidazo[1,2-a]quinolines 5-8 and their acyclic precursors 3-4, a preliminary investigation of their UV/Vis, fluorescence emission and excitation spectra, recorded in EtOH, was undertaken.The results are shown in Figures 1-2  Acyclic compound 3 showed an absorption maximum at 355 nm, while dicyano-substituted acyclic compound 4 showed it at 371 nm (batochromic effect).At 280 and 278 nm both compounds showed another absorption band.Cyclic compounds 5-8, as it can be visualized in Figure 1, showed several absorption bands.Di-substituted benzimidazo[1,2-a]quinolines 6 and 8 showed one absorption band in 370 and 374 nm, while mono-substituted benzimidazo[1,2-a]quinolines 5 and 7 showed two absorption bands from 344 nm to 387 nm.Fluorescence intensity (ar.un.)

Conclusions
We have prepared benzimidazolyl substituted acrylonitriles 3-4, which were converted into corresponding benzimidazo[1,2-a]quinolines 5-6 by a photochemical dehydrocyclization reaction.From the Pinner reaction of compounds 5 and 6 we did not obtain the desired mono-and diamidinosubstituted benzimidazo[1,2-a]quinolines.The cyano group at the 6-position of both cyclic compounds 5 and 6 gave an unsubstituted amido group, instead of an amidine group.An amido group was formed in the first phase of the Pinner reactions.All the prepared compounds, especially the cyclic benzimidazo[1,2-a]quinolines 5-8, are potential biologically active compounds and may be expected to be DNA intercalators, which will be investigated during the course of our ongoing work in this area.

General
Melting points were determinated on a Koffler hot stage microscope and are uncorrected.IR spectra were recorded on Perkin-Elmer 297 spectrophotometer with KBr disks. 1 H-and 13 C-NMR spectra were recorded in DMSO-d 6 on a Varian Gemini 300 or Bruker Avance DPX 300 spectrometers using TMS as an internal standard.Mass spectra were recorded on an Agilent 1100 series LC/MSD Trap SL.Elemental analysis for carbon, hydrogen and nitrogen were performed on a Perkin-Elmer, Series II, CHNS Analyzer 2400.Where analyses are indicated only as symbols of elements, analytical results obtained are within 0.4% of the theoretical values.All compounds were routinely checked by TLC with Merck silica gel 60F-254 glass plates.The electronic absorption spectra were obtained on Varian Cary 100 spectrometer and fluorescence emission spectra were recorded on Varian Eclipse fluorimeter, in both cases using quartz cuvettes (1 cm).

General method for preparation of 3 and 4.
A solution of equimolar amounts of corresponding benzaldehydes 1a,b and benzimidazole-2acetonitrile 2 in absolute EtOH (15 mL) was refluxed in the presence of piperidine (0.03 mL) for 4 h.The dark yellow product which separated on cooling was filtered off, washed with petroleum ether and EtOH and recrystallized from EtOH to give the products.

General method for preparation of benzimidazo[1,2-a]quinoline-6-carboxamide 7 and 2-amidino-benzimidazo[1,2-a]quinoline-6-carboxamides 8-10
A stirred suspension of compounds 5 and 6 in absolute EtOH was cooled in an ice-salt bath and then saturated with HCl gas.The flask was then tightly stoppered and the mixture was maintained at room temperature for 20 days until the nitrile band disappeared (monitored by IR analysis at 2200 cm -1 ).The reaction mixture was purged with N 2 gas and diluted with diethyl ether (50 mL).The crude imidate was filtered off and was immediately suspended in absolute EtOH (10 mL).The corresponding amine was added and the mixture was stirred for one to three days at room temperature.The crude product was then filtered off and washed with diethyl ether.