Kinetic Determination of Ascorbic Acid

One method for determining the concentration of glucose in fruit juices is to take advantage of its enzymatic oxidation in the presence of glucose oxidase:

β-D-glucose + O2 + H2O → δ-D-gluconolactone + H2O2

To follow the kinetics of this reaction, the hydrogen peroxide is reacted with phenol and 4-aminoantipyrene to form a colored antipyrene dye:

2H2O2 + phenol + 4-aminoantipyrene → antipyrene dye + 4H2O

a reaction that is catalyzed using horseradish peroxidase. The formation of the dye is monitored by measuring the absorbance at 505 nm. Concentrations of the enzymes are chosen such that the second reaction is much faster than the first reaction; thus, any kinetic data is controlled by the first reaction only. One complication is that ascorbic acid, which is often present in fruit juices, also reacts with hydrogen peroxide

2H2O2 + ascorbic acid → dehydroascorbic acid + 2H2O

a reaction that is significantly faster than the enzymatic reaction between H2O2, phenol, and 4-aminoantipyrene.  Shown below are the results of three kinetic runs using a fixed concentration of glucose and variable concentrations of ascorbic acid (Series 1 has the most ascorbic acid and Series 3 has the least).  Clearly explain, in as much detail as you can, the similarities and differences between these three curves.

This entry was posted in Essay Question, Kinetic Method and tagged . Bookmark the permalink.

Leave a Reply

Your email address will not be published. Required fields are marked *