PubMed · 12979147
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D FILIPO. 1952. [Studies of tonsillar vascularization with the benzidine method].. https://pubmed.ncbi.nlm.nih.gov/12979147/
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3,3',5,5'-Tetramethylbenzidine (TMB) is the most commonly used chromogen for horseradish peroxidase (HRP) and so its performance as an electrochemical substrate was evaluated. Measurements of HRP activity in solution were carried out by using an amperometric detector coupled to a flow injection analysis (FIA) system. The enzymatic product was easily detected at a potential of +0.1 V (vs. Ag-pseudoreference electrode) at a bare screen-printed electrode placed in a homemade electrochemical flow cell. A high flow rate (4.3 mL min(-1)) of 0.5 M H2SO4 was used to obtain repeatable signals and a short analysis time. The detection limit achieved after 15 min of incubation was 2x10(-14) M of HRP. The applicability of the amperometric detector to ELISAs was demonstrated by using a commercially available kit for the quantification of interleukin-6 (IL-6) without modifying the kit manufacturer's protocol or the reagents for this test.
The benzidine photodegradation process was studied using UV/Vis spectroscopy and electrospray ionization mass spectrometry (ESI-MS) combined with collision-induced dissociation (CID) and tandem mass spectrometry (MS/MS). Mass spectrometry was used to characterize benzidine and to identify and characterize possible degradation products and intermediates. The MS data showed that benzidine is quite persistent in aqueous medium. Moreover, the MS analysis enabled us to propose the following three degradation products/intermediates: 4'-nitro-4-biphenylamine, tetrahydroxybiphenyl and 4,4'-dinitrobiphenyl. For the benzidine molecular ion and protonated molecule and for the protonated molecules of the degradation products/intermediates detected, fragmentation patterns are proposed based on CID and MS/MS data. For the photodegradation process different catalysts were used, namely the commercial TiO2 Degussa P25, and the laboratory-prepared ZnO, TiO2 anatase and a titanium-zinc oxide with a perovskite type structure. Comparison of the different catalysts showed that degradation was favoured with the commercial TiO2. Nevertheless, the other catalysts appear to be promising and economic alternatives for potential future remediation studies.