A series of improved Ca2+ chelating agents which can be used for NMR and fluorescence imaging will be synthesized and evaluated for their absorption and emission characteristics as well as their ability to bind Ca2+. The proposed chelators incorporate fluorophores which have desirable high quantum yields and visible wavelength absorption and emission. The fluorophore is attached to an EGTA-like structure which is known to have great specificity for Ca2+ over Mg2+ and other divalent metal ions. The proposed Ca2+ indicators will thus have improved optical characteristics over currently used quin2 and indo-1, which they resemble structurally except for the fluorophore. In addition, the indicator will be fluorine substitute of 19F NMR detection. The indicators will be cell membrane permeable when the acetate groups are esterified, and reconvertible to the acetate form by cytosolic esterases inside the cell. Thus the trapped chelator can be used to follow concentration changes in intracellular calcium concentration, (Ca2+)i, as a function of time in response to various perturbing influences. Intracellular calcium is currently the subject of wide-spread interest as a second messenger that triggers and regulates cell activation. Phase I work involves synthesis of the amino napthopyrone skeleton linked to the F substituted ring through the diether backbone. In Phase II the synthesis will be completed and evaluation of optical constants and Ca2+ binding constants will be carried out. The most promising indicators will be tested for cell membrane permeability and as tomographic imaging agents for 19F NMR.
