Fluorescent and colorimetric dual-readout ratiometric assay for highly selective ascorbic acid detection based on gold(I) triggered in situ reaction

A dual-ratiometric assay combining fluorescence and colorimetry is constructed based on a gold(I)-triggered in situ reaction between o-phenylenediamine (OPD) and ascorbic acid (AA). OPD is oxidized by gold(I) to form fluorescent 2,3-diaminophenazine (OPDox), which has an absorption peak centered at 445 nm and a fluorescence emission at 555 nm. When AA is introduced, AA can be oxidized to dehydroascorbic acid (DHAA) by gold(I), inhibiting the production of OPDox. Simultaneously, DHAA reacts with OPD to form a fluorescent quinoxaline derivative (DFQ), which exhibits an absorption peak around 340 nm and a fluorescence emission at 425 nm. Based on this redox-modulated strategy, a highly selective dual-mode sensing assay is established with ratiometric colorimetric (A₃₄₀/A₄₄₅) and ratiometric fluorescence (F₄₂₅/F₅₅₅) signals. This method enables the quantitative analysis of AA, with limits of detection (LOD) of 0.09 μmol/L and 0.04 μmol/L, respectively. Owing to its facile analytical procedure, this assay exhibits many merits including simplicity, excellent selectivity and good biocompatibility, and has been successfully applied to detect AA in human serum samples. Such an approach based on the in situ formation of fluorophores provides a valuable tool for AA detection in clinical diagnosis and drug screening.