OXIDATION CHEMISTRY OF SOME BIOLOGICALLY IMPORTANT N-HETEROCYCLIC COMPOUNDS

Abstract

Electrochemical investigations of biologically important compounds and their precursors present major challenges both from an electro-mechanistic and analytical viewpoint. Electrochemistry, with its unique ability to oxidize or reduce compounds at a well-controlled electrode potential and by just giving (at the anode) or taking (at the cathode) electrons, provides interesting information about the redox reactions of these compounds. The electron transfer between two chemical entities is one of the most fundamental process in chemistry and biochemistry. In biological systems, oxidative processes are well known for their role in energy conversion and substrate metabolism. Typical of these are those involved in catabolism of many biologically important organic compounds. Since redox pathways of such type are usually complex due to enzyme interactions and biologically active site is probably deep within the molecularframe-work, it is very difficult to study in their entirety. The best way of getting details of in vivo redox chemistry of compounds would be by examining the electrochemical behaviour of a compound at the electrode-solution interface. It is well-established that electrochemical studies provide much better picture of the chemical aspects of the enzyme-catalyzed redox reactions of these compounds. In recent years, it has been clearly demonstrated that the electrochemical studies when coupled with other analytical methods such as GC, HPLC, HNMR, mass spectrometry, etc. have the potentialities to probe and elucidate the mechanistic aspects of complex enzyme catalyzed biological transformations undergoing in biosystems. For a number of years, our laboratory has been investigating the redox behaviour of a variety of compounds. Some of these compounds were studied in detail at pyrolytic graphite electrode like Uric acid, Adenine, Guanine etc, whereas others were studied superficially. As purines are the building blocks of two wellknown nucleic acids, viz., RNA and DNA, the enzymic oxidation of these compounds GO was also studied wherever possible. Recently, electrooxidation of Indole and its derivatives was carried out in our laboratory due to their biological and clinical significance in Central nervous systems. The present dissertation deals with the oxidation studies of some Nheterocyclic compounds of biological significance. The compounds selected are 1,3,7,9-tetramethyluric acid, 6-hydroxy-2,4,5-triaminopyrimidine and indole derivatives, viz., indole-3-methanol and indole-3-ethanol. Chemical oxidation by persulfate of indole derivatives and their comparison with electrochemical oxidation has also been studied. The thesis is a small part of the big proposal submitted by Prof. R. N. Goyal in connection with redox chemistry of such types of molecules. For simplicity and clarity the results of the investigation are organized in the dissertation as follows: The first chapter of the thesis is "General Introduction" and presents a brief review and significant results relevant to the present study on the electrochemical investigations of the various compounds selected. The chapter also highlights the salient features of the techniques used in the investigation. Uric acid is the primary end product of purine metabolism and is a constituent of many body fluids. In view of the importance of uric acid in biological process, the second chapter deals with electrochemical investigation of 1,3,7,9-tetramethyluric acid (I) in aqueous and micellar medium. This chapter basically presents the effect of tetra methyl substitution on the oxidation of uric acid. As the studies in aqueous micellar systems have been reported to mimic the microenvironment analogous to that of the active sites of enzymes in a physiological system, the effects of ionic and non-ionic surfactants on the voltammetric behaviour of tetramethyluric acid has also been evaluated. It was observed that the peak potential and peak current significantly alter in the presence of ionic surfactants whereas the non-ionic surfactants did not affect the electrode process. These studies indirectly supported the formation of a cationic free radical in the electrode reaction. The effect of methyl groups on the oxidation of uric acid has also been examined. The methyl groups besides producing an electron releasing effect also restricts the number of resonating structures of uric (iii) acid. It has been suggested that methylation of nitrogen atom at pyrimidine ring of uric acid significantly affects the oxidation mechanism resulting in products different from that obtained in uric acid. "3<K (I) The third chapter of the thesis presents electrochemical oxidation of 6-hydroxy 2,4,5-triaminopyrimidine (II), a well-known precursor of guanine, at pyrolytic graphite electrode. Cyclic sweep voltammetry at a sweep rate of 100 mVs" exhibited one well-defined oxidation peak Ia in the entire pH range studied. In the reverse sweep, two reduction peaks Ic and IIC were observed below pH 4.3. Peak Ic formed quasi-reversible couple with peak Ia in the entire pH range studied. The nature of the electrode reaction was established as EC in which charge transfer is followed by competitive chemical reactions. Double potential step chronoamperometric experiments were also performed to determine the half-life of the intermediate (diimine) generated in the reaction. Controlled potential electrolysis provided the value of 'n', number of electrons transferred in the redox reaction as 2.0 ±0.2. Spectral changes during electrolysis were monitored to detect the formation of UVabsorbing intermediate. The decay of the intermediate generated during the oxidation was found to occur in pseudo-first-order reaction. The products of electrolysis were separated by gel-permeation chromatography and characterized by m.p., TLC, IR and 'HNMR etc. and a plausible mechanism for the formation of products has also been suggested. H2N OH N (ID NH2 NH2 (iv) Fourth chapter of the thesis deals with the oxidation of Indole-3-methanol (III), a phytochemical isolated from cruciferous vegetables and is the most powerful anticancer compound. In view of the importance of indole-3-methanol, an attempt has been made to study the electrochemical as well as the chemical oxidation of this compound in phosphate buffers. A comparison of mechanisms for both the oxidations is also presented. ,CH2OH In electrochemical oxidation, cyclic sweep voltammetry of indole-3-methanol at a sweep rate of 100 mVs"1 exhibited a single anodic peak Ia in the pH range 2.4- 10.7. In the reverse sweep, only one cathodic peak peak IIC was observed at pH > 3.6. Ep vs pH plots for both anodic and cathodic peaks were linear. The nature of the electrode reaction was established as ECE in which two-charge transfer steps are separated by the chemical steps involved. Coulometric studies provided the value of n, number of electrons, as 3.0 ± 0.1. The kinetics of the UV-absorbing intermediate generated was monitored spectrophotometrically. Its decay followed a pseudo-firstorder behaviour. On the other hand, chemical oxidation of indole-3-methanol by persulfate followed pseudo-first-order kinetics having first-order with respect to the substrate. Both electrochemical and chemical oxidation of indole-3-methanol occurred through step-wise electron transfer. It was interesting to note that the electronic spectral changes observed in the initial course of oxidation, both electrochemically and chemically were very similar. These changes exhibited an isosbestic point at 290 nm and depicted increase in absorption in the visible region. A comparison of the electrochemical and chemical oxidation clearly indicates that the final products of oxidation of the two mechanisms are different, in spite of their similar initial le, 1H+oxidation step. (v) The last chapter of the thesis deals with the electrochemical and chemical oxidation of Indole-3-ethanol (IV). Indole-3-ethanol, an important indole derivative, is found both in plants and animals. The nature of the electrode reaction was found as ECE in which two-charge transfer steps are separated by the chemical steps. During chemical oxidation, stoichiometric experiments were carried out by estimating the unused amount of persulfate iodometrically and mole ratio was found to be 1: 5.5 (indole-3-ethanol: S2082"). Products of both oxidations were characterized by GC and GC-mass. A comparison of electrochemical and chemical oxidation indicated that oxidation occurred both in the aromatic as well as pyrrole ring and major products of oxidation were identical. Tentative mechanisms for both the oxidations are also suggested. ^•^ CH2CH2OH N' H