Dioleamide as a novel chemiluminescence reagent: Simple synthesis and highly sensitive bioenzyme detection

In this work, a novel chemiluminescence (CL) reagent dioleamide (DOA) was synthesized, which contains a long chain organic compound with an amide as the core functional group. It exhibited stable and strong CL emission when sodium hypochlorite was used as the oxidant. A mechanistic study indicates that CL is mainly due to the process in which the reactive oxygen species (ROS) produced by the system during the chemical reaction oxidize the DOA to the excited state and then return to the ground state to release energy in the form of light radiation. Alkaline phosphatase (ALP) catalyzes the hydrolysis reaction of p-nitrophenyl phosphate (PNPP) to generate p-nitrophenol (PNP). The maximum absorption wavelength of PNP significantly overlaps with the CL emission wavelength of the DOA, enabling energy transfer from DOA to PNP and thus the CL intensity is weakened. Consequently, a simple, efficient, and sensitive ALP sensor based on chemiluminescence resonance energy transfer (CRET) system was developed with a detection limit of 0.020 U/L. The NaClO-DOA-PNP sensing system, which demonstrated a highly sensitive reaction to ALP in the range of 0.1–30 U/L, was successfully applied to the detection of ALP in fetal bovine serum.