Boosting the oxidase-like activity of platinum nanozyme in MBTH-TOOS chromogenic system for detection of trypsin and its inhibitor

Nanozymes, as a new type of artificial enzyme, have recently become a research hotspot in the field of catalysis and biomedicine. However, the application of nanozyme is limited by catalytic activity changes of different substrates and low specificity. This work shows that citrate-capped platinum nanoparticles (Cit-PtNPs) exhibit stronger oxidase-like activity than other platinum nanozymes at different pH when 3-methyl-2-benzothiazolinonehydrazone hydrochloride (MBTH) and n-ethyl-n- (2-hydroxy-3-sulfopropyl)-m-toluidine sodium salt (TOOS) were used as chromogenic substrates. This phenomenon has important reference value for different nanozymes to choose chromogenic substrates in catalysis. In MBTH-TOOS chromogenic system, MBTH (-NH) radical is first produced during the reaction through catalytic oxidation of Cit-PtNPs, which reacts with TOOS to produce a colorless compound. The blue-purple quinoid dye was produced through the dismutation of the colorless compound. The catalytic mechanism of the oxidase-like activity of Cit-PtNPs is that two-electron reduction process and four-electron reduction process are simultaneously carried out in the catalytic process. Furthermore, to solve the problem of low specificity of metal nanozymes, protamine is designed as aggregation promoter of Cit-PtNPs and the specifichydrolysis substrate of trypsin. In this work, it can achieve one-step detection of trypsin by the boosting oxidase activity of Cit-PtNPs at pH8. The catalytic activity of Cit-PtNPs is proportional to the concentration of trypsin. The linear range for trypsin is 1.0–70.0 ngmL−1 and the limit of detection is measured to be 0.6 ngmL−1. This novel method has also been successfully applied to the detection of inhibitors and trypsin in urine samples.