The objective of this proposal is the elucidation of the process known as the chemical production of electronically excited states. In some cases the excited states emit light, and the overall process is termed chemiluminescence (or bioluminescence if the process is catalyzed by an enzyme). In other cases the excited states (singlet and triplet) enter into chemical and biochemical reactions. Because of the high sensitivity of the measurement of light emission from """"""""black bodies"""""""" at room temperature chemiluminescence shows promise as a safe replacement for many techniques utilizing radioactivity. For example, the cyclic hydrazides we investigated a decade ago are currently being used in antibody binding assays. Chemiluminescence is also used widely to assay foretals, ATP, enzymatically generated peroxides, etc. The present proposal is aimed at an understanding of: (1) the generation of excited states from cyclic hydrazides such as luminol. We propose to (a) synthesize the monohydrazide of 5-methyldehydroluciferin (firefly) and test it for light emission on oxidation, (b) synthesize efficient chemiluminescent phthalazinediones derived from N,N-dibutylisoluminol, (c) attempt a low temperature synthesis and study of the bicyclic peroxides proposed to be crucial intermediates in cyclic hydrazide chemiluminescence, and (d) examine the basic chemistry of acyldiazenes at low temperatures and in the absence of oxygen. (2) the generation of excited states from dioxetanes and dioxetaneones based on the highly fluorescent acridine ring systems. Emission from the weakly acidic acridone in both chemiluminescence and fluorescece will be compared; the data may permit a decision as to whether the CIEEL mechanism is operating in these intramolecular cases of excitation. The synthesis, structure, and chemistry of photochemically produced peroxides based on 9-substituted acridine ring systems will be examined. (3) the generation of excited states from firefly luciferin. Emphasis will be placed on fast reactions of the initially formed excited state to yield (possibly?) a yellow-green light-emitting molecule-in an effort to perfect a model system for the yellow-green light emission of the firefly. Analytically pure LH2AMP will be prepared for this study. Physical-organic approaches will be used with emphasis on low temperature spectroscopic techniques.
