[A novel catalytic spectrophotometric determination for horseradish peroxidase]

A new catalytic-kinetics spectrophotometric method for the determination of horseradish peroxidase was developed. It was based on the catalytic effect of horseradish peroxidase (HRP) on the coloration reaction, in which the reduced rhodamine B was oxidized by H2O2 in pH 6.80 phosphate buffer. It was found that reduced rhodamine B stocking solution could be steady in 0.25% beta-CD solution. The kinetic behavior of the reaction and the effects of some experimental conditions were investigated and discussed in detail. The method has been applied to determine HRP with a satisfactory result. The calibration curve is linear over the range 15-250 pg.10 mL-1 of HRP (r = 0.9989). The limit of detection was 12 pg.10 mL-1 and the relative standard derivative RSD is 4.25% by determining for 20 pg.10 mL-1 HRP (n = 10).