Benzazolylcoumarin-based ion indicators

A method of measuring the concentration of a polycationic metal ion in a sample, comprising:

• a) adding to said sample, in an amount sufficient to generate a detectable fluorescent response to said metal ion, a compound according to the formula ##STR13## wherein m is either 2 or 3;
• A is either of the formula ##STR14## or A is of the formula ##STR15## wherein R.sup.1 is H, an alkyl group having 1-5 carbons, a benzyl, an alpha-acyloxyalkyl or a pharmaceutically acceptable esterifying group, a t-butyldimethylsilyl ester, or a pharmaceutically acceptable salt;
• R.sup.2, R.sup.3, R.sup.4 and R.sup.5 are independently H or methyl;
• R.sup.6 and R.sup.7 are independently H, CN, CH.sub.3, CF.sub.3 or CONH.sub.2 ;
• X and X' are independently O, S, or C(CH.sub.3).sub.2 ;
• Y and Y' are independently H, an alkyl having 1-18 carbons, --NO.sub.2, --NH.sub.2, --NH(C.dbd.O)(CH.sub.2).sub.n CH.sub.3, --CF.sub.3, F, Cl, Br, I, --OR.sup.8, --CO.sub.2 R.sup.9, or --OCH.sub.2 CO.sub.2 R.sup.9, where n=0-16;
• R.sup.9 is H, an alkyl group having 1-18 carbons, a benzyl (C.sub.6 H.sub.5 CH.sub.2 --), an alpha-acyloxyalkyl, acetate, or a t-butyldimethylsilyl ether;
• and R.sup.9 is H, an alkyl group having 1-17 carbons, a benzyl (C.sub.6 H.sub.5 CH.sub.2 --) an alpha-acyloxyalkyl or other pharmaceutically acceptable esterifying group, a t-butyldimethylsilyl ester, or a pharmaceutically acceptable salt;
• or one of Y and Y' is a covalently attached conjugant;
• and
• Z is one of H, an alkyl having 1-18 carbons, --NH.sub.2, --NH(C.dbd.O)(CH.sub.2).sub.n CH.sub.3, --CF.sub.3, F, Cl, Br, I, --OR.sup.8, --CO.sub.2 R.sup.9, or --OCH.sub.2 CO.sub.2 R.sup.9 ;
• or Z is a covalently attached conjugant;
• b) illuminating said sample to generate an absorbance or fluorescence response;
• and
• c) observing said absorbance or fluorescence response.