Synthesis of Firefly Luciferin Analogues and Evaluation of the Luminescent Properties

Five new firefly luciferin (1) analogues were synthesized and their light emission properties were examined. Modifications of the thiazoline moiety in 1 were employed to produce analogues containing acyclic amino acid side chains (2–4) and heterocyclic rings derived from amino acids (5 and 6) linked to the benzothiazole moiety. Although methyl esters of all of the synthetic derivatives exhibited chemiluminescence activity, only carboluciferin (6), possessing a pyrroline‐substituted benzothiazole structure, had bioluminescence (BL) activity (λmax=547 nm). Results of bioluminescence studies with AMP‐carboluciferin (AMP=adenosine monophosphate) and AMP‐firefly luciferin showed that the nature of the thiazoline mimicking moiety affected the adenylation step of the luciferin–luciferase reaction required for production of potent BL. In addition, BL of 6 in living mice differed from that of 1 in that its luminescence decay rate was slower. All aglow: Luciferin derivatives possessing acyclic amino acid and heterocyclic substituents were synthesized. All synthetic derivatives displayed chemiluminescence, but only d‐carboluciferin showed 0.5 % bioluminescence (BL) activity compared with that of firefly luciferin. d‐Carboluciferin differed from firefly luciferin in that decay of its BL in live mice was slower than that of the natural luciferin (see figure).