Activation Products of Panal

Activation procedures (Shimomura, 1989). To activate trace amounts of panal, several granules of the crystals of ammonium sulfate are added to 5-50 il of a sample made in 30% methanol, making sure that (NH4)2S04 is in excess to saturate the solution. The mixture is left standing at room temperature for 1 h (85% activation) or one night (complete activation). After the activation, 1 jig of panal emits about 1.5 x 1010 photons using the assay method given below. 

To activate 1-2 mg of panal with ammonium sulfate, 1 ml of 3 M (NH4)2S04 (pH 3.7 with acetic acid) and 2-3 mg of CTAB are added, 

The yield of luminescence activity is increased by including CTAB, Tergitol 7 (a sulfate detergent), or p-toluenesulfonyl chloride to various extents. 

Assay methods. Activity can be measured at pH 5.5 or at pH 8.0. With the same sample, the pH 5.5 method gives a much higher luminescence intensity than the pH 8.0 method (and, shorter reaction time), although the total amounts of light emitted by the two methods are practically equal (Fig. 9.4). The pH 5.5 method is susceptible to inhibition by various salts, whereas the pH 8.0 method is not affected. 

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Properties of activated products. The luminescence of the activation products of panal is elicited by Fe2+ and H2O2, and the light emission is significantly enhanced by the presence of various detergents, particularly cationic detergents, such as CTAB. The influence... 

Properties of the activation product. The two decylamine-activation products (luciferins) showed similar absorption characteristics (A.max 372 nm in water, and 375 nm in ethanol), which clearly differ from the absorption peak of the natural luciferin (320 nm) reported by Kuwabara and Wassink (1966). The fluorescence emission of the activation products varied significantly by solvents, showing a peak at 460 nm in neutral aqueous solution and a broad peak at 485-522 nm in ethanol. They emitted chemiluminescence (A.max 580 nm) in the presence of CTAB, H2O2 and Fe2+ (Fig. 9.13), in resemblance to the (NH4)2S04-activation product of panal (A.max 570 nm). 

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