F. I. TSUJI, A. M. CHASE AND E. N. HARVEY 139 



spectrum of doubly cycled Cypridina luciferin, which had been 

 washed with benzene to minimize ultraviolet-absorbing impurities. 

 Luciferin itself is insoluble in benzene. The spectra were measured 

 in pH 6.8 phosphate buffer solution, using the Beckman spectrophotom- 

 eter, and are shown in Fig. 2. The heavy line represents the solution 

 immediately after dissolving the luciferin. As shown earher (Chase, 

 1940), an initial absorption maximum in the visible region at about 



o 

 o 



Ul 



o 



0.20 



0.15 - 



0.10 



0.05 



0.00 



250 300 350 400 



WAVELENGTH - mjj 



450 



500 



Fig. 2. Absorption spectra of a solution of doubly cycled and benzene-washed 

 luciferin, measured in pH 6.8 phosphate buffer. The heavy line spectrum was 

 obtained as soon as possible after the solution had been prepared. The solu- 

 tion, standing exposed to air, at room temperature, was then remeasured at 

 the times indicated. Characteristic of a luciferin solution under these condi- 

 tions is the shift of the visible absorption band from a maximum at 435 

 to one at 465 mii. Solutions of pH 1.0 do not show this shift (see Fig. 3). 



435 m/x is rapidly replaced by one at about 465 m^u and the latter 

 then almost entirely disappears during exposure of the solution to air. 

 In the ultraviolet the initial spectrum has a pronounced absorption 

 peak at 265 m/x, and a shoulder at about 310 m^i. On exposure to air 

 the absorptions at 265 and 310 m/j. decrease and, simultaneously, a 

 new band appears at about 365 m/x. Isosbestic points exist at 330 and 



