Properties of Dinoflagellate Luciferin

The solution of purified dinoflagellate luciferin is yellow, showing absorption maxima at 245 and 390nm in an aqueous solution and at 241 and 388 nm in 40% acetonitrile containing 85 mM NaCl and 3 mM NaHCOs (Fig. 8.4). The compound is strongly fluorescent in blue (excitation maximum at 390 nm, emission maximum at 474 nm Fig. 8.5). The properties of this luciferin are nearly identical with those of the compound F of euphausiid shrimps (Section 3.2). The luciferin is rapidly oxidized in the presence of a trace of oxygen, and also inactivated by a weak acid, even by an acidity of pH 4 or the acidity 

6 Chemical Structures of Dinoflagellate Luciferin and its Oxidation Products (Nakamura et al., 1989) 

Mild chromic acid oxidation of luciferin (CrOs/KHSC /HiO, room temperature) yielded 3-methyl-4-vinylmaleimide (1, Fig. 8.7), 3-methyl-4-ethylmaleimide (2), and an aldehyde (3), whereas vigorous chromic acid oxidation (CrOs/2N H2SO4, 90°C) gave hema-tinic acid (4) (Dunlap et al., 1981). These results closely resemble the results of the chromic acid oxidation of the fluorescent compound F of euphausiid (p. 76), indicating a structural similarity between dinoflagellate luciferin and the compound F. 

When a methanolic solution of luciferin was left at -20° C in the presence of air, most of the luciferin was oxidized in three days, based on its 1H-NMR spectrum. The air oxidation product was purified by HPLC on a TSK DEAE-5PW column using 35% acetonitrile containing 85mM NaCl and 3mM NaHCC 3. The purified product in the HPLC eluent showed absorption maxima at 237nm 

The luciferin produces a blue oxidation product during its purification process. In the DEAE chromatography of luciferin, this blue compound is eluted before the fractions of luciferin. The fractions of the blue compound were combined and purified by HPLC on a column of Hamilton PRP-1 (7 x 300 mm) using methanol-water (8 2) containing 0.1% ammonium acetate. The purified blue compound showed absorption peaks at 234, 254, 315, 370, 410, 590 (shoulder) and 633 nm. High-resolution FAB mass spectrometry of this compound indicated a molecular formula of C l C Nai m/z 609.2672 (M - Na + 2H)+, and mlz 631.2524 (M + H)+]. These data, together with the HNMR spectral data, indicated the structure of the blue compound to be 8. 

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