Method to determine guaiacol oxidase activity in soil

A precise and sensitive method to assay guaiacol oxidase activity in soil is needed. The objective of this study was to set up this method by determining residual guaiacol spectrophotometrically after its extraction from soil with 0.5 M NaOH and 1 M CaCl2 and its separation from reaction products and humic molecules of the soil extract by high-performance liquid chromatography (HPLC). A soil sample sterilized by autoclaving was used as a control. The rate of guaiacol oxidation reached the highest value at pH 4.5 to 6.0. The reaction rate of the enzyme approached zero-order kinetics when the substrate concentration was 50 mM. Disappearance of guaiacol was linear with time for at least 1 h. The optimal temperature for guaiacol oxidation activity occurred at 60°C. Guaiacol oxidase activity was significantly correlated with soil total N.