Calcein is a fluorescent metal indicator for calcium detection with EDTA titration. It is slightly soluble in water. The proton dissociation constants are reported to be pKa1=2.1, pKa2=2.9, pKa3=4.2, pKa4=5.5, pKa5=10.8 and pKa6=11.7. Calcein solution emits a yellowish-green fluorescence in acidic conditions, and no fluorescence in basic conditions. However, Calcein will emit fluorescence in the presence of metal ions such as Al, Ba, Ca, Cu, Mg, and Zn in basic conditions. Therefore, Calcein is also used for direct fluorometric EDTA titration of these metal ions. Since Calcein has a clear endpoint even if the sample has color or contains phosphate, it is suitable for calcium detection in biological samples or fertilizers. TPC, murexide, or PPC are added to get a clear endpoint. For example, by mixing equal amounts of TPC with Calcein, intense green fluorescence will turn reddish-purple with no fluorescence at the endpoint.
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