Does 3,5,dibromo-4-nitrosobenzene sulphonate spin trap superoxide radicals?

Area of Science:

• Biochemistry
• Chemical Kinetics
• Electron Paramagnetic Resonance (EPR) Spectroscopy

Background:

• The spin trap 3,5-dibromo-4-nitrosobenzene sulphonate (I) is used to detect short-lived radicals.
• Superoxide radicals (O2.-) are biologically important reactive oxygen species.
• Previous studies have reported conflicting results regarding radical detection after reacting I with O2.-.

Purpose of the Study:

• To investigate the reaction mechanism between spin trap I and superoxide radicals (O2.-).
• To elucidate the source of the stable radical observed in some studies.
• To determine the role of xanthine oxidase and associated peroxidase activity in radical formation.

Main Methods:

• Pulse radiolysis was employed to study the rapid reaction kinetics between I and O2.-.
• Electron Spin Resonance (ESR) spectroscopy was used to detect radical species.
• The xanthine/xanthine oxidase system was utilized to generate O2.- in the presence and absence of peroxidase activity.

Main Results:

• Pulse radiolysis confirmed a rapid reaction between I and O2.-, yielding an unstable product.
• ESR studies detected no radicals when I reacted with O2.- generated via gamma-radiolysis.
• A stable radical was observed only when peroxidase activity was present in the xanthine oxidase sample, indicating formation via hydrogen peroxide.

Conclusions:

• The reaction of spin trap I with O2.- does not directly produce a stable radical detectable by ESR.
• The previously reported stable radical is a consequence of peroxidatic oxidation by hydrogen peroxide, a byproduct of O2.- dismutation.
• The presence of peroxidase activity in the xanthine oxidase enzyme preparation is critical for the formation of this stable radical.