Functional magnetic nanoparticles with delayed chemiluminescence for tyrosinase detection using delay time as a quantitative parameter.

Chemiluminescence (CL) bioassays traditionally rely on flash-type systems, generating immediate light emission upon liquid mixing. Such systems often suffer from turbulence-induced disruptions during measurements and rely solely on emission intensity as the quantitative parameter, requiring high-sensitivity detectors and resulting in compromised reproducibility. However, the analytical potential of CL emission time as an alternative quantitative parameter remains largely unexplored. To overcome these limitations, 8-amino-5-chloro-2,3-dihydro-7-phenylpyrido-[3,4-d]-pyridazine-1,4-dione (L012) and Mn(II) bifunctional mesoporous silica-coated manganese ferrite core-shell magnetic nanoparticles (MnFe2O4@mSiO2/L012/Mn(Ⅱ) NPs) were synthesized. Using NaHCO3 and H2O2 as coreactants and thiourea as a time-regulating reagent, the as-prepared NPs exhibited intense and delayed CL emission with an ultranarrow full width at half maximum of 0.61 s, attributed to the high loading efficiency of L012 and Mn(II) enabled by the mSiO2 shell. The CL delay time was highly tunable for precise time-resolved measurements, and the NPs possessed robust magnetic separability and excellent long-term CL stability. Leveraging these features, a CL method was developed that used CL delay time as a novel quantitative parameter for tyrosinase (TYR) detection, achieving an impressive detection limit of 26 U/L. The delay time-based approach improved detection accuracy and stability, eliminated reliance on high-sensitivity detectors, and simplified instrumentation requirements. Furthermore, the developed CL method was successfully applied to TYR detection in human serum samples and TYR inhibitor screening, highlighting the broad potential of this time-resolved CL strategy for enzyme activity assays and biomedical applications.