Fluorometric Determination of Alkaline Phosphatase Activity

4-methylumpholone phosphate (MUF-P) was used as the substrate to analyze APA. MUF-P was converted to PO₄^3- and 4-methylumpholone (MUF) by the alkaline phosphatase, in which MUF showed fluorescence when pH is above 10^{81 (link)}. For 2 L bottle experiments in 2019 and 20 L bottle experiments in 2018 in summer and in 2019, 45 mL sample with 200–1000 nM MUF-P was incubated in the microcosm tank in the dark after gently mixing^{11 (link)}. The fluorescence and therefore the APA (nM P·h⁻¹) were calculated estimated by time-course measurements (0, 1.5, and 3 h) using the 7200-000 Trilogy fluorometer (Turner Designs) with the long wavelength UV module (P/N 7000-967) after adding borate buffer (pH = 10.8–11.2) to the samples^{82 (link)}. The concentration of MUF corresponding to the fluorescence was calculated using the standard curve (0–1 μM) configured with MUF-2Na^{5 (link),82 (link)}.