The effect of the media properties on the horseradish peroxidase-catalyzed fluorogenic reaction is investigated. The dimer product exhibited acid-base equilibrium For p-hydroxyphenylpropionic acid, the pKa of the product is 8.3. The fluorescence of the product increased with higher pH; however, a longer time was required to reach reaction equilibrium due to the mismatch of this pH value with that associated with enzyme activity. The cationic micelles cetyltrimethylammonium bromide and cetyltrimethylammonium chloride displayed catalytic and enhancing effects on the reaction at neutral pH. A more sensitive method for the determination of horseradish peroxidase is thus suggested.

Key words: HYDROGEN PEROXIDE, FLUOROSPECTROPHOTOMETRY, EXCITATION SPECTRUM