FLUOROMETRIC DETERMINATION OF PEROXIDES IN AN HPLC SYSTEM

Hemin was used to substitute for horseradish peroxidase (HRP) as catalyst with phydroxyphenyl acetic acid (PHPAA) as substrate for post-column detection of H2O2 and methyl hydroperoxide (CH3OOH, MHP) in an HPLC system. In a hemin-catalyzed system, post-column reaction and fluorescence measurement can both be conducted optimally at a pH of ~10.5. Hydroxymethyl hydroperoxide (HOCH2OOH, HMHP) and bis(hydroxymethyl) peroxide (HOCH2OOCH2OH, BMHP) will rapidly be converted to H2O2 in alkaline solution and can also be detected. The HPLC system is optimized for the analysis of H2O2 and MHP. Reproducibilities are better than 5% for H2O2 and MHP. The detection limits in aqueous solution are 9 nM for H2O2 and 200 nM for MHP.