Histidine-copper microenvironment-engineered high-entropy LDH nanozyme enables a neutral-pH colorimetric sensor array for antibiotic discrimination.

Nanozyme-based sensor arrays are attractive for multianalyte analysis, but the low catalytic activity of most high-entropy nanozymes near neutral pH limits their application in complex samples. Here, a histidine-copper coordination microenvironment was engineered on a CoZnCuFeAl layered double hydroxide (LDH) through sequential histidine functionalization and Cu(I) incorporation, yielding LDH-His-Cu. Spectroscopic characterization indicated the formation of Cu-N coordination, the coexistence of Cu(I)/Cu(II) species, and an increased abundance of oxygen vacancies. The resulting nanozyme exhibited markedly enhanced peroxidase-like and laccase-like activities at pH 7. Its peroxidase-like specific activity reached 12.9 × 10-2 U·mg-1, representing 58.6- and 107.5-fold enhancements over LDH-His and pristine LDH, respectively. The enhanced catalytic performance arises from a synergistic catalytic microenvironment constructed by three indispensable components: the LDH framework, histidine-mediated interfacial regulation, and Cu(I) incorporation. Histidine functionalization modulates the local catalytic environment and facilitates neutral-pH catalysis, while oxygen vacancies provide reactive sites for oxygen activation and Cu(I)/Cu(II) redox cycling accelerates electron transfer processes, collectively promoting the generation of multiple reactive oxygen species, including •OH, •O2-, and 1O2. Further, by recording the two enzyme-mimicking reactions at two reaction times, a four-channel colorimetric fingerprint was generated from a single nanozyme. Combined with principal component analysis, the array enabled clear discrimination of eight antibiotics at concentrations down to 1 μM, as well as binary and ternary antibiotic mixtures. The discrimination capability was further demonstrated in spiked milk, egg, and honey samples. This work establishes catalytic microenvironment engineering as an effective strategy for neutral-pH nanozyme catalysis and illustrates the integration of reaction-type and time-domain information for multianalyte pattern recognition.