Reduction of the 2,2′-Azinobis(3-Ethylbenzthiazoline-6-Sulfonate) Cation Radical by Physiological Organic Acids in the Absence and Presence of Manganese

Abstract

ABSTRACT Laccase is a copper-containing phenoloxidase, involved in lignin degradation by white rot fungi. The laccase substrate range can be extended to include nonphenolic lignin subunits in the presence of a noncatalytic cooxidant such as 2,2′-azinobis(3-ethylbenzthiazoline-6-sulfonate) (ABTS), with ABTS being oxidized to the stable cation radical, ABTS ·+ , which accumulates. In this report, we demonstrate that the ABTS ·+ can be efficiently reduced back to ABTS by physiologically occurring organic acids such as oxalate, glyoxylate, and malonate. The reduction of the radical by oxalate results in the formation of H 2 O 2 , indicating the formation of O 2 ·− as an intermediate. O 2 ·− itself was shown to act as an ABTS ·+ reductant. ABTS ·+ reduction and H 2 O 2 formation are strongly stimulated by the presence of Mn 2+ , with accumulation of Mn 3+ being observed. Additionally, 4-methyl- O -isoeugenol, an unsaturated lignin monomer model, is capable of directly reducing ABTS ·+ . These data suggest several mechanisms for the reduction of ABTS ·+ which would permit the effective use of ABTS as a laccase cooxidant at catalytic concentrations.