Methyl calcein blue and other analogues of calcein blue

4-Methylumbelliferone-8-methylenesarcosine (Methyl Calcein Blue) and four related metallofluorochromic indicators derived from umbelliferone, 4-methylumbelliferone, and 4-methylesculetin by condensation with formaldehyde and iminodiacetic acid or glycine have been synthesized, the structures established, the absorbance and fluorescence measured as functions of pH, and the reactions with copper(II) and calcium studied with attention to the effects on fluorescence. All of the compounds display a maximum fluorescence at ph about 9. The fluorescence of each is quenched by copper(II). The calcium derivatives of the compounds derived from the umbelliferones and iminodiacetic acid fluoresce at high ph but those from the umbelliferones and glycine or sarcosine do not. At high pH, 4-methylesculetin and the amino-acids derived from it do not fluoresce either alone or in the prescence of calcium.