New Phenoxyacetic Acid Analogues with Antimicrobial Activity

This paper presents experimental data regarding the synthesis and the antimicrobial activity of new 4phenylazo-phenoxyacetic acids. All compounds were characterized by IR, UV-Vis, mass spectral data and by elemental analysis. These compounds were tested for their antibacterial activity against Staphylococcus aureus, Streptococcus pyogenes, Escherichia coli, Pseudomonas aeruginosa, Proteus vulgaris and for their antifungal activity against Candida albicans by disk diffusion method.

The derivatives of phenoxyacetic acid have been studied intensively due to their pharmacological and phytobiological activity.
Ethacrynic acid (2-[2,3-dichloro-4-(2-ethylprop-2enoyl)phenoxy]ethanoic acid), a monosulfonamyl diuretic, differs form other thiazide diuretics in that a double ring system is incorporated into its structure.Ethacrynic acid inhibits sodium ion transport across the renal tubular epithelium.By increasing the delivery of sodium to the renal tub, ethacrynic acid indirectly increases potassium excretion via the sodium-potassium exchange mechanism [2].
Centrophenoxine (acetic acid, (4-chlorophenoxy)-2-(dimethylamino)ethyl ester), also known as Lucidril® and meclofenoxate, is one of the older nootropic drugs-it was developed in 1959 at the French National Scientific Research Center [3].Beneficial therapeutic effects of centrophenoxine have been observed in various human disorders such as cerebral atrophy, brain injury, post apoplectic status, chronic alcoholism and barbiturate intoxication [4].
We have recently reported the synthesis of 4-phenylazophenoxyacetic acids by the condensation of some 4phenylazo-phenols with chloroacetic acid [5,6].
In this respect, as the most recent results are continuing our concern in the chemistry of azo dyes, and in the present paper we report the synthesis, structure and antimicrobial activity of several new 4-phenylazophenoxyacetic acids.

Techniques
The melting points have been established in capillaries and verified with a Boetius apparatus.Elemental analyses of carbon, hydrogen and nitrogen have been performed using a Carlo-Erba O/EA, 1108, analyser.The electronic spectra have been carried in dioxane, with a UV-Vis Jasco spectrophotometer, within 200-700nm.FTIR spectra were recorded on a Avatar Nicolet spectrophotometer in KBr pellets, within the range 3500-400cm -1 .Mass spectra have been obtained using a HPGC-MS 5890 MD 5971 spectrometer at 70eV, with carrier gas He at 2mL/min.

General procedure
All compounds were tested for antimicrobial activity against 6 microorganisms: Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa, Proteus vulgaris, Escherichia coli and Candida albicans.Test compounds were dissolved in ethanol.Concentrations of 0,2% of the test compounds were obtained.
Tests of different isolates of microorganisms used were carried out by pouring 15mL sterile Mueller Hinton agar in each Petri discs of 9 cm diameter.After solidification, the plates were placed in an incubator at 37 o C for 30 min to remove excessive moisture.
Overnight broth culture was streaked evently onto medium in three directions using a wooden stick cotton swab.Excess suspention was removed from the swab by rotating it firmly against the side of the tube before seeding the plate surface using sterile forceps.The plates were inoculated aerobically at 37 o C within 15 min.After 24 h incubation, the diameters of the inhibition zones were measured (including the 6 mm diameter of the disc) with a rule [7].

Results and discussions
In this paper it is presented the synthesis of four 4phenylazo-phenoxyacetic acids by the condensation of sodium salts of corresponding substituted 4-phenylazophenols with chloroacetic acid in alkaline medium, using Williamson method.
The structure of these compounds has been investigated on the basis of UV-Vis, IR and mass spectra.
The IR spectra show absorption bands specific to -COOH, -N=N-, Ar-O-CH 2 , Ar-CN groups, and also for the aromatic rings.
Carboxylic acids show two characteristic IR absorption that make the COOH group easily identifiable.In our case, the OH bond of the carboxyl group gives rise to a very broad absorption over the range 3473-3453cm -1 and the C=O bond shows an absorption between 1740-1707cm -1 .
Aromatic rings show a characteristic series of peaks in the 1520-1470cm -1 of the infrared spectra.
In the region 1600-1400 cm -1 available the bands due to the vibrations of azo group and those due to the stretching vibrations í C=C for the C-C linkages in aromatic nuclei.The absorptions bands due to the azo group appear in two regions, namely at 1600-1550 cm -1 and 1430-1400 cm -1 .The intensive bands of the first region are a result of conjugation of azo group with aromatic nuclei.The weak bands of region 1430-1400 cm -1 are due to the vibration of azo group and agree with the data presented in literature.[8][9][10] In conformity with our previous papers, the examination of the infrared spectra of compounds 1-4 reveals that the azo group shows a stronger absorption in region 1590-1562cm -1 than in 1417-1406cm -1 [5,6].
Very important and relevant are the bands due to the etheric vibrations; normally, these compounds should have two bands because of antisymmetrical and symmetrical vibrations.Very intense bands were attributed from 1276-1257cm -1 to the antisymmetrical valence vibrations and much weaker from 1078-1014cm -1 to the etheric symmetrical valency vibrations.
The strong absorption band at 1190-1110 cm -1 is characteristic to the vibration of the Ph-N band [9][10].
On the other hand, the moderate absorption band at 870-856cm -1 can be assigned to the vibration of the C-Cl bond.
In the UV-Vis spectra we remark the presence of some absorption bands: from middle intensity at 220-248nm, benzenoid-type E-or B-bands, consequently to the π electrons conjugation from the aromatic rings, intense absorption K-bands 344-362nm, as the result of the conjugated system Ar-N=N-Ar, and low intensity R-bands at 430-452nm, due to the -N=N-chromophore [11][12].
Further evidence for the 4-phenylazo-phenoxy acetic acids structure was obtained from mass spectrum.
For 2-chloro-4-phenylazo-phenoxyacetic acid, scheme 1 show the fragmentation process specific to the phenoxyacetic acids by the cleavage of the bond between the C of the methylene group and C atoms of the carboxyl group followed by CO 2 elimination.
The melting temperatures of 4-phenylazophenoxyacetic acids depend on the nature and position of the existing on the aromatic nuclei.The presence of cyano group in para position explains the increasing of melting points of 4-(4-cyano-phenylazo)-phenoxyacetic acid.
The yields are generally high, depending on the raw materials purity and on solubility of novel acids in the solvent used for recrystallisation.