Primary product of the horseradish peroxidase-catalyzed oxidation of pentachlorophenol

Peroxidases are a class of enzymes that catalyze the oxidation of various p henolic substrates by hydrogen peroxide. They are common enzymes in soil an d are also available commercially, so that they have been proposed as agent s of phenolic pollutant transformation both in the environment and in engin eered systems. Previous research on the peroxidase-catalyzed oxidation of p entachlorophenol (PCP) has suggested that tetrachloro-p-benzoquinone (chlor anil) is the principal product and that a considerable fraction of the PCP added to reaction mixtures appears to be resistant to oxidation. In experim ents employing alternative methods of product separation and analysis, we f ound that both of these observations a re artifacts of extraction and analy tical methods used in previous studies. The major product of the horseradis h peroxidase-catalyzed oxidation of pentachlorophenol from pH 4-7 was 2,3,4 ,5,6-pentachloro-4-pentachlorophenoxy-2,5-cyclohexadienone (PPCHD), (PPCHD) , which is formed by the coupling of two pentachlorophenoxyl radicals. The yield of chloranil and other soluble products was negligible. PPCHD is inso luble and unreactive in aqueous media but is reactive when dissolved in var ious organic solvents. Substantial amounts of chloranil were formed when PP CHD was dissolved in benzene, ethyl acetate, or methanol; less was formed i n hexane and acetonitrile; and negligible amounts were formed in dimethylfo rmamide. High-pressure liquid chromatography (HPLC) analysis of PPCHD indic ated that it is capable of undergoing dissociation and reduction to pentach lorophenol under typical reversed-phase HPLC conditions. Chlorinated oligom eric products are formed when PPCHD is stored in acetonitrile, either alone or with added pentachlorophenol. Our results demonstrate that the removal of PCP in peroxidase-catalyzed reactions can be much higher than indicated in previous work, as long as the initial product is separated by filtration or other physical means.