User:Carly M. Montanero/Notebook/CHEM-571/2013/09/17: Difference between revisions
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== | ==Objective== | ||
Today we are going to determine the amount of reagent that is required to fully oxidize or fully reduce horseradish peroxidase. For HRP oxidation, we will be using [http://en.wikipedia.org/wiki/Potassium_ferricyanide potassium ferricyanide], K<sub>3</sub>[Fe(CN)<sub>6</sub>]. K<sub>3</sub>[Fe(CN)<sub>6</sub>] has a standard reduction potential of 424mV ([http://pubs.acs.org/doi/abs/10.1021/ja00902a014 ref1] and [http://pubs.acs.org/doi/abs/10.1021/ja01018a013 ref2]) vs [http://en.wikipedia.org/wiki/NHE NHE]. For HRP reduction, we will be using [http://en.wikipedia.org/wiki/Sodium_dithionite sodium dithionite], which has a reduction potential of -460mV vs NHE. We will be monitoring oxidation and reduction through changes in the UV-Vis spectrum of HRP. In order to do this we will also have to account for the absorbance of the K<sub>3</sub>[Fe(CN)<sub>6</sub>, which has an absorption feature at 420nm (for Fe<sup>2+</sup>, ε = 4.7 M<sup>-1</sup>cm<sup>-1</sup>). This is being done in preparation for our experiments tomorrow where we will be determining the redox potential of HRP. | |||
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Revision as of 06:27, 24 September 2013
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ObjectiveToday we are going to determine the amount of reagent that is required to fully oxidize or fully reduce horseradish peroxidase. For HRP oxidation, we will be using potassium ferricyanide, K3[Fe(CN)6]. K3[Fe(CN)6] has a standard reduction potential of 424mV (ref1 and ref2) vs NHE. For HRP reduction, we will be using sodium dithionite, which has a reduction potential of -460mV vs NHE. We will be monitoring oxidation and reduction through changes in the UV-Vis spectrum of HRP. In order to do this we will also have to account for the absorbance of the K3[Fe(CN)6, which has an absorption feature at 420nm (for Fe2+, ε = 4.7 M-1cm-1). This is being done in preparation for our experiments tomorrow where we will be determining the redox potential of HRP. From
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