We prepare an oxidase mimic nanomaterial-based enzyme, it could effectively catalyze the conversion of TMB to oxTMB. In this process, we want to know the exact type of ROS in the catalytic process (the current speculation is superoxide anion). So, we used DMPO as a trapper agent and discussed it by using electron paramagnetic resonance spectroscopy. However, six peaks with the same peak intensity are observed (according to the reported results, four peaks with the intensity of 1:1:1:1 should appear). I'm confused about the reason for this result. What details must be paid attention to in the process of testing?
Dear Yuan Zhang thanks for posting this very interesting technical question. Cited below please find a potentially useful reference in which a possible alternative to the reagent DMPO (= 5,5-dimethyl-pyrroline N-oxide) is presented. It is stated in this paper that (citation): "5,5-Dimethyl-pyrroline N-oxide (DMPO) has been used for decades to trap superoxide anion in biological systems. A major disadvantage of DMPO is that the DMPO-superoxide adduct (DMPO–OOH) spontaneously decays to the DMPO–hydroxyl adduct (DMPO–OH) and as a result, superoxide anion was typically detected by monitoring the superoxide dismutase-inhibitable formation of DMPO–OH." Thus it might be worth a try searching for alternative reagents:
Detection of superoxide anion using an isotopically labeled nitrone spin trap: potential biological applications
https://core.ac.uk/download/pdf/82771921.pdf
This paper is freely available as public full text on the internet (please see attached pdf file). Good luck with your work and best wishes, Frank Edelmann
The EPR spectrum of the superoxide-DMPO adduct shows effectively six lines of equal intensity. (different from the spectrum of the HO-DMPO adduct with only four lines).
For further details, please see: VIEIRA, A.J.S.C.; TELO, J.P. "EPR Detection of Oxygen Radical Anion in Aqueous Solution at Room Temperature", Redox Report (1997), 3, 303
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