Cross reactivity between ascorbate peroxidase and phenol (guaiacol) peroxidase

• Ascorbate peroxidase (APX) activity is assessed by a decrease in ascorbate levels.
• However, guaiacol peroxidase (POX) activity results in the same reaction.
• POX activity is also not specific as some APX isozymes use similar phenols as a substrate.
• Therefore, APX activity should always be carefully separated from POX activity.

Adverse conditions often induce an increase in active oxygen species (AOS) such as hydrogen peroxide (H2O2). H2O2 is converted to water, and thus becomes detoxified by enzymes such as ascorbate peroxidase (APX; EC 1.11.1.11). APX activity is estimated by the disappearance rate of ascorbic acid, which becomes oxidized. However, ascorbate is also a substrate of guaiacol peroxidase (POX; EC 1.11.1.7). POX oxidizes phenols (including flavonoids), whereby ascorbate accepts an electron from phenoxyl or flavonoid radicals. Ascorbate becomes thereby converted to the monodehydroascorbate radical, which subsequently can become converted to dehydroascorbate. POX isozymes therefore convert hydrogen peroxide to water and oxidize ascorbate, just as APX does. POX activity is usually estimated by monitoring the formation of tetraguaiacol from guaiacol. This reaction is not specific, as some APX isozymes show rather high activity when using guaiacol or similar phenols as a substrate. It is concluded that APX activity can easily be confounded with POX activity, and vice versa. Proper methods should be used to separate the two enzyme activities.