To provide the basis for clinical diagnosis in an emergencycase, a portable smartphone device-based multi-signal sensing system for on-site determination of alkaline phosphatase (ALP) is introduced. In this system, cobalt hydroxide (CoOOH) nanoflakes can oxidize O-phenylenediamine (OPD) to produce 2,3-diaminophenazine (OxOPD), resulting ina strong fluorescence at 565nm and an absorbance at 420nm, respectively. The ascorbic acid 2-phosphate (AAP) can be hydrolyzed by alkaline phosphatase (ALP) to yield ascorbic acid (AA). Then, AA reduces the CoOOH nanoflakes to produce Co2+, and AA is oxidized to form dehydroascorbic acid (DHAA), thereby inhibiting the formation of OxOPD. The reaction product DHAA further combines with OPD to yield 3-(1,2-dihydroxyethyl)furo[3,4-b]quinoxalin-1(3H)-one (DFQ) accompanied by a strong fluorescence at 430nm. Based on this, the fluorometric assay for ALP hasa wide linear range from 0.8 to 190U/L with a low detection limit of 0.16U/L, and the colorimetric assay from 3 to 130U/L with a detection limit of 1.94U/L. Moreover, a portable smartphone sensing platform integrated with fluorescent and colorimetric signals was established for rapid determination of ALP without spectrometers. Recoveries of 97-104%for spiked samplesand relative standard deviations (RSD) of less than 2% (n= 3) confirmed the feasibility of the developed platform in complicated samples, opening up new horizons for on-site evaluation in the biomedical field.