Simple Nanochannel-Modified Electrode for Sensitive Detection of Alkaline Phosphatase Through Electrochemiluminescence Signal Quenching by Enzymatic Reaction

Development of sensitive and convenient alkaline phosphatase (ALP) detection methods is of great significance for food analysis, biomedical applications, and clinical tests. In this work, a sensitive detection method for ALP was established based on nanochannel-modified electrodes, where the electrochemical luminescence (ECL) signal was quenched by the enzymatic reaction product. Vertically ordered mesoporous silica film (VMSF) was rapidly grown on low-cost ITO via the electrochemically assisted self-assembly (EASA) method. The resulting modified electrode (VMSF/ITO) exhibited a uniform and ordered nanochannel structure with nanochannel diameter of 2-3 nm and charge-selective permeability, enabling the enrichment of cationic ECL emitter tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺). Compared to the ITO electrode, VMSF/ITO increased the ECL signal by 60 times. In the presence of ALP, it catalyzes the hydrolysis of its substrate, disodium phenyl phosphate hydrate (DPP), generating phenol (Phe), which quenched the ECL signal of Ru(bpy)₃²⁺ and the co-reactant N,N-Dipropyl-1-propanamine (TPA). Based on this principle, ECL detection of ALP can be achieved. The linear detection range for ALP was 0.01 U/L to 30 U/L, with a limit of detection (LOD) of 0.008 U/L. The sensor exhibited high selectivity. Combined with the anti-contamination and anti-interference capabilities of VMSF, the constructed sensor enabled the detection of ALP levels in milk samples.