Indian Journal of Chemistry Vol. 44A, March 2005, pp. 463-468 Horseradish peroxidase catalyzed and electrochemical oxidations of 2-thiouracil-A comparison Rajendra N Goyal *, Sham M Sondhi, Anand M Lahoti & Adil A Abdulla Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee 247667 (Utta,ranchal), In'dia Received 6 September 2004; revised 10 January 2005 Horseradish peroxidase (type VIII) catalyzed and electrochemical oxidations of 2-thiouracil have been studied in phosphate buffer of pH 7.20 (JL==l.O M) at an ambient temperature of 25±2°C. The peroxidase catalyzed oxidation has been initiated by using hydrogen peroxide and electrooxidation carried out at pyrolytic graphite electrode. The UV-absorbing intermediates generated in both the oxidations have been characterized by voltammetry, spectrophotometry and by kinetics of decay and are found identical. Products of enzymic and electrochemical oxidations have been characterized by using GC- MS stuides. A tentative redox scheme has been proposed for the enzymic oxidation of 2-thiouracil and compared with that of electrooxidation and has also been found similar. Thus, it has been concluded that oxidation of 2-thiouracil by enzymic and electrochemical methods are, in a chemical sense, identical. IPC Code: Int.CI. 7 C25B3/02; C07B33/00 Nucleic acid bases with sulfur atom have been detected in natural t-RNA I and therefore have attracted considerable interest during last decade. 2- Thiouracil (I) and its derivatives have been found to induce modifications in thyroid gland and are claimed as an antithyroid drugs 2 .3. Two well known thiouracil antithyroid drugs, 6-propyl-2-thiouracil and 6-methyl- 2-thiouracil, have also been shown to be competitive inhibitors of neuronal nitric oxide synthase 4 . 2- Thiouracil and its derivatives have also been used as antitumor agents 5 • 6 . It has been reported that electrochemical and horseradish peroxidase catalyzed oxidation of uric acid? and N-methylated uric aGids 8 appear to proceed, in a chemical sense, by very similar if not identiCal mechanisms at around physiological pH. As electrochemical investigations of biologically important compounds provide uniquely invaluable insights about the redox mechanisms of the enzymic reactions, it was considered interesting to study the peroxidase catalyzed oxidation of 2-thiouracil and compare the results with that of electrochemical oxidation 9 • In particular, we were interested in knowing if the detailed mechanism elucidated from electrochemical studies at pyrolytic graphite electrode was applicable to the enzyme catalyzed oxidation of 2-thiouracil also. It is expected that results of these studies will provide insights into overall chemical pathways followed by the substrate molecule in the enzymic oxidation reaction. - (I) Materials and Methods 2-Thiouracil was procured from Sigma Chemical Co., USA and was used as received. Horseradish peroxidase (HRP) type VIII (R z >3.0) was obtained from Bangalore Genei Pvt. Ltd., and catalase was supplied by CSIR Centre for Biochemicals, New Delhi, India. Hydrogen peroxide (30% w/v) was a product of Merck. Silylation grade acetonitrile (CH 3 CN) and N,O-bis[trimethylsilyl]trifluoto- acetamide (BSTFA) were supplied by Supe1co, USA. Phosphate buffers of ionic strength 1.0 M were prepared I 0 from analytical grade chemicals (NaH 2 P0 4 and Na2HP04) in double distilled water. Conventional electrochemical equipments ll ,l2 were used to carry out cyclic voltammetric studies. The numbers of electrons (n) involved in electrooxidation reaction were determined by monitoring the exponential decay of the current-time curve as reported by Li ngane I3 . UVNis spectral changes during electrooxidation were monitored by electrolyzing 0.2 mM 2-thiouracil in a H-cell using pyrolytic graphite plate (lx6 cm 2 ) as working, cylindrical platinum gauze as auxiliary and Ag/AgCI