A ratiometric fluorescent strategy for alkaline phosphatase activity assay based on g-C3N4/CoOOH nanohybrid via target-triggered competitive redox reaction

The development of ratiometric fluorescent strategy is very significant and challenging in bioanalysis. Herein, graphitic carbon nitride (g-C3N4) nanosheets and cobalt oxyhydroxide (CoOOH) nanoflakes are exploited for ratiometric fluorescence assay of alkaline phosphatase (ALP) activity. In the g-C3N4/CoOOH nanohybrid, g-C3N4 nanosheets serve as a signal unit and the CoOOH nanoflakes function as a recognition element, and initially, the fluorescence of g-C3N4 is quenched by the CoOOH nanoflakes. In the absence of target, the CoOOH nanoflakes of the nanohybrid system are able to oxidize o-phenylenediamine (OPD), and the resultant oxidation product (OxOPD) quenches the blue emission of g-C3N4 and meanwhile emits orange fluorescence which acts as another signal element. However, an efficient redox reaction between ascorbic acid (AA) and CoOOH can cause decomposition of the CoOOH nanoflakes, and additionally, ALP can catalytically hydrolyze L-ascorbic acid-2-phosphate (AAP) to generate AA. Thus, in the presence of target, the CoOOH nanoflakes are destroyed by AA preferentially and the OPD is rarely oxidized to OxOPD, accompanied with strong blue emission of g-C3N4 and weak orange fluorescence from OxOPD. Target-dependent dual-signal change made the ratiometric assay possible, and also AA-induced signal variation was investigated and attributed to the stronger reducing capacity of AA than OPD. The ratiometric sensing platform for ALP activity assay provides a new perspective for the applications of two-dimensional nanomaterials to develop novel and sensitive biosensors.