Luminometric determination of FAD in subpicomole quantities.

Very small quantities of FAD were able to reactivate apo-D-amino acid oxidase. In the presence of D-alanine, luminol, horseradish peroxidase, and an excess of the apoenzyme, a quantitative luminometric determination of FAD was possible. The maximal photon emission measured in a bicarbonate buffer, pH 9.2, at 37 degrees C was proportional to the amount of FAD added. FMN, riboflavin, or 5-deazaflavin produced no chemiluminescence and had no inhibitory effect in the assay when added together with FAD. With this method, FAD could be quantitatively determined with high accuracy in perchloric acid extracts of animal tissue and bacteria.