ELECTROCHEMICAL INVESTIGATIONS OF SOME BIOLOGICALLY IMPORTANT ORGANIC COMPOUNDS

Abstract

Electrochemical investigations of the redox chemistry of organic compounds of biological significance or naturally occurring biomolecules present major challenges both from electromechanistic as well as analytical view point. Modern electrochemical techniques provide indispensable, versatile and significant potentialities to extend and deepen the under standing of bio-electrochemical and bio-physicochemical aspects of the biological transformations undergoing in the living systems. Many important biological systems are of redox type and therefore can be studied electrochemically. There are now sufficient amount of evidences which support the view that redox mechanisms deduced for small biomolecules using purely electrochemical and related methodologies do provide rather unique and significant insights into the complex biological redox processes. Therefore, much of the growth that can be expected in the future from electro chemical studies, in general, will be associated with elucidation of mechani sm of electrochemical redox behaviour of such compounds. In view of such an importance of electrochemical investigations, the redox behaviour of different type of biologically important organic compounds such as bisazo dyes, heterocyclic amines, sulpha drug and methylated xanthine (a derivative of purine) is elucidated using various electrochemical techniques in combination with other sophisticated spectro scopic and analytical techniques. Inspite of the importance of this class of organic compounds in medicine and biology, the studies on their reaction mechanism are still obscure and therefore have been investigated in detail. The interesting results of electrochemical studies of some biologically significant organic compounds are presented in following chapters of the thesis. (ii) First chapter of the thesis comprises of general introduction descri bing a brief review on electrochemical investigations of different class of organic compounds selected for present studies and their significant results relevant to the present study. Second chapter of the thesis deals with elucidation of mechanism of electrochemical reduction behaviour of two important bisazo dyes, viz., chrysophenine and Acid Red-150, largely used as cotton dyes which possess better heat resistance capacity than monoazo dyes. The aim behind selecting these two bisazo dyes was to study the effect of various electron releasing and electron withdrawing substituents on the ease of electroreduction and the effect of an azo group on the electroreduction of another azo group. Azo compounds, in general, are electroreduced in 2e, 2H step to give hydrazo derivative as the product. On the other hand, some workers have also reported 4e, 4H reduction of azo group with the cleavage of N-N bond. Literature survey reveals that electrochemical reduction behaviour of monoazo compounds have been studied in great detail in aqueous as well as non-aqueous media. However, very little attention has been paid so far to bisazo compounds i.e., azo compounds substituted with another azo group. Polarographic studies on these two dyes clearly indicate that first 4e, 4H+ reduction causes the cleavage of an azo group, which is followed by further 4e, 4H+ reduction giving amino compounds as the final products of electroreduction in the entire pH range studied. Therefore, the electro reduction of two azo groups at d.m.e. takes place in an overall 8e, 8H process in the entire pH range (2.0 - 10.3). It was very interesting to observe that in the pH range 5.0-6.0, chrysophenine gave rise to a third polarographic 2e, 2H+ wave which was ascribed to the reduction of extra (iii) nucleic^ C = C"^present in the molecule. This wave could not be observed at other pH probably because of high reduction potential of )• C = C< at d.m.e. due to which the reduction waves merged with the background. Furthermore, first 4e, 4H+ reduction of Acid Red-150 caused the cleavage of -N = N - bond attached in the vicinity of bulkier naphthol moiety. The diffusion controlled nature of the electrode process was established by the linearity of i, versus Jh plot. The concentration higher than 0.1 mM of chrysophenine caused the precipitation, of the dye in the solution indicating that this dye can be estimated below 0.1 mM concentration by .polarographic method. However, in case of Acid Red-150, id versus concen tration (0.05- 1.0 mM), was linear passing through the origin indicating that this dye can safely be estimated in this concentration range at d.m.e. In cyclic sweep voltammetry at sweep rate of 150 mVs , chrysophenine exhibited one well defined 8e, 8H+ reduction peak. However, Acid Red- 150, in cyclic sweep voltammetry, exhibited two well defined 4e, 4H reduction peaks in the entire pH range studied. The nature of electrode process at different pH was established. The value of n, number of elec trons involved in the electroreduction process were determined by graphical integration of current-time curve. On the basis of spectral studies and product characterisation by TLC, m.p., I.R. spectra, it has been concluded that unlike bisazo benzene where first 4e, 4H+ reduction gives bishydrazo benzene, the cleavage of -N = N- group occurs in both the dyes indicating the formation of amino compounds as the final product of electroreduction. The larger reduction potentials of the dyes at PGE in comparison to d.m.e. are explained on the basis of adsorption complications associated with the solid electrodes. The third chapter of the thesis is devoted to the elucidation of mechanism of electrochemical oxidation of a sulpha drug, viz., sulphadiazine (iv) which is an important antibacterial drug used for the control of bacillary dysentery and streptococcal infections. Sulpha drugs, the first drugs in the history to control systemic bacterial diseases, led the way directly or indirectly to a number of the more important therapeutic discoveries of the present time. Despite their long history, very little information is available about the electrochemical oxidation of this class of compounds. Most of the investigations presently reported deal with the determination of these drugs by different techniques in many pharmaceutical preparations. However, in all these investigations, no attempt has been made to identify the products of the reaction and thus mechanistic studies on the electrooxidation behaviour attracted less attention. Linear and cyclic sweep voltammetry, coulometry, spectral studies and product separation and identification by TLC, column chromatography, m.p., I.R. and mass spectra, clearly indicated that aromatic amino group of sulphadiazine underwent electrooxidation in a single well defined 2e , 2H+ peak. The plot of i as a function of concentration clearly indicated that i linearly increases upto about 5 mM concentration of sulphadiazine, P after which the peak current was more or less constant. Thus, sulphadiazine can only be estimated below 5 mM concentration at PGE. The increase of i /ACv1 values with log v indicated adsorption of sulphadiazine at PGE. P z In the acidic pH range, two products, viz., hydroxylamino and azo com pounds, were identified. In the basic pH range only azo product was obtai ned. A tentative mechanism has been proposed to account for the formation of these two products. Anodic voltammetry of aniline and its derivatives has been studied in detail in aqueous and non-aqueous systems and the redox reaction has been found to be complex, giving a mixture of products. However, very (v) little information is available on the electrochemical oxidation of hetero cyclic amines probably because of their resemblance with benzenoid amines. Isomeric aminopyridines i.e., 2-, 3- and 4-aminopyridines are of great importance in the synthesis of various antibacterial compounds, in estimation and extraction of various metal ions and in protecting agricultural crops etc. In view of such an importance of aminopyridines, the electrochemical oxidation of isomeric aminopyridines have been investigated at PGE using different techniques and the interesting results obtained are summarised in the fourth chapter of the thesis. The oxidation of all the three aminopyridines was observed in a single 2e, 2H+ step. The peak current values (i ) increased linearly with increasing concentration of 2-aminopyridine upto a concentration of about 4 mM, and at higher concentrations the peak current had a tendency to become constant. Similarly, the plot of i versus concentration of 3-aminopyridine was linear upto about 10 mM, after which the peak current was practically constant. In case of 4-aminopyridine, the adsorption effect was practically nil due to the higher concentration used otherwise the cyclic voltammetric behaviour of all the three aminopyridines was practi cally identical. UV spectra were recorded at different pH to determine the pKa of 2-, 3- and 4-aminopyridines and were found to be 6.8, 6.2 and 9.3, respectively. These values were also confirmed by Ep versus pH plot in cyclic voltammetry. In UV/Vis spectral changes the intermediate formed in electrooxidation of 3-aminopyridine was more stable in comparison to 2- and 4- aminopyridines. The products of electrooxidation have been separated and characterised by TLC, m.p., I.R. and N.M.R. spectra as azopyridine. . The stereochemistry of the isolated products was established by comparing the NMR data with cis- and trans-isomers of 2,2'-3,3'- and 4,4'-azopyridines and it was inferred that the product in all the cases was trans derivative. (vi) The last chapter of the thesis is concerned with the electrochemical studies on 8-methylxanthine, an important dihydroxy purine, used as diure tics. The electrochemical behaviour of N-methylated xanthines has been studied significantly however, electrochemical oxidation behaviour of Cmethylated xanthine, viz., 8-methylxanthine has not been studied so far. In linear and cyclic voltammetry, 8-methylxanthine was electrooxidised in one well-defined oxidation peak in the pH range 1.35-10.71. The peak potential of this peak was dependent on pH and shifted towards less positive potential with increase in pH. The plot of Ep vs pH showed two breaks at around pH 3.1 and 9.7 indicating the pKa of 8-methylxanthine. These pKa values were also confirmed by recording UV spectrum of 8-methylxan thine at different pH and plotting absorbance as a function of pH where two inflection points at the concomitant pH values further confirmed the above pKa values. The peak current function (i /ACvi) was almost constant at different log (sweep rate) which indicated the diffusion controlled nature of electrode process in the concentration range of 0.1-0.5 mM. The value of n, number of electrons involved in oxidation was determined by exponen tial decay of current with time and was found to be 2.0 ± 0.1 in the pH range 1.35-10.71. The products of controlled potential electrolysis were separated by column chromatography and HPLC and were analysed as alloxan and acetamidine by I.R. spectrum, m.p. and by a comparison of cyclic voltammograms of completely electrolysed solution with that of authentic samples.