Hydrogen peroxide

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Hydrogen peroxide

This information is part of O2k-technical support and open innovation.
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Hydrogen peroxide


Hydrogen peroxide

Hydrogen peroxide, H2O2 or dihydrogen dioxide, is one of several reactive oxygen intermediates generally referred to as reactive oxygen species (ROS). It is formed in various enzyme-catalyzed reactions (e.g. superoxide dismutase) with the potential to damage cellular molecules and structures. H2O2 is dismutated by catalase to water and oxygen. H2O2 is produced as a signaling molecule in aerobic metabolism and passes membranes more easily compared to other ROS.

Abbreviation: H2O2

MitoPedia methods: Respirometry, Fluorometry 

MitoPedia topics: Substrate and metabolite 


Applications in HRR

Measurement of hydrogen peroxide

Two techniques to measure H2O2 concentration and H2O2 production can be combined with the O2k:

  1. O2k-Fluorometry using Amplex red;
  2. Electrochemical measurement with an amperometric H2O2 sensor.


The fluorometric measurement of H2O2 in the O2k is based on the O2k-Fluorescence LED2-Module, using Amplex red (or one of its brand names).[1] Please see Amplex red about the method as well as various application notes: MiPNet18.06_Amplex-Yeast, MiPNet18.05_Amplex-Mouse-heart, MiPNet17.17_Amplex-Mouse-brain.

Electrochemical measurement

For electrochemically (H2O2 electrode) measuring H2O2 (and all other amperometric methods) please see our O2k-NO Amp-Module and its manual MiPNet15.05 NO-Manual. All stated there about measuring NO is technically also correct for using a H2O2 electrode together with the oxygraph. In this manual you will also find which accessories you need to use a H2O2 electrode together with the O2k-MultiSensor System (there are different requirements for different generations of O2ks). The electrode itself has to be bought from a third party.

It is sometimes stated that the available H2O2 electrodes may not have the required sensitivity for most ROS studies. WPI states a lower detection limit of "<100 nM" for their 2 mm "macro" electrode and "<10 nM" for their 0.1 mm "mini sensor". The latter value is quite similar to the stated detection limit for fluorescence based methods, however it is not clear if the "mini sensor", probable developed for direct tissue insertion, works well in macroscopic chambers. A direct comparison would be helpful. Please add your experiences!

Picomolar detection ranges have been reported for a method combining Amplex Red (R) with square wave voltammetry.[2]

  1. Sumbalova Z, Harrison DK, Gradl P, Fasching M, Gnaiger E (2011) Mitochondrial membrane potential, coupling control, H2O2 production, and the upper limit of mitochondrial performance. Abstract Kagoshima.
  2. Lyon J L, Stevenson K J (2006) Picomolar peroxide detection using a chemically activated redox mediator and square wave voltammetry. Anal Chem 78: 8518-8525

Reoxygenation of the respiration medium with H2O2

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