128 THE NATURE OF ANIMAL LIGHT 



when oxylucif erin is reduced or whether oxygen is actually 

 added as in formation of haemoglobin cannot be definitely 

 stated at present. We may write equations representing 

 these possibilities as follows : 



Ci6 H20 N3 SCI (leuco-methylene blue) + ^ Cie His N3 SCI 

 (methylene blue) + H2O 



Haemoglobin + O ^ oxyhaemoglobin. 



Let us now turn to the methods which may be used in 

 reduction of oxylucif erin. We may then endeavor to write 

 an equation which will represent the fundamental changes 

 in the luminescence reaction. 



My attempts to reduce the oxidation product of lucif- 

 erin started from the observation that if one places a 

 clear solution of luciferase in a tall test tube, although 

 it may give off no light at first when shaken, after standing 

 a day or so a very bright light would appear on shaking. 

 This was especially true when the luciferase had become 

 turbid and ill-smelling from the growth of bacteria. 

 Thinking that the bacteria produced a substance which 

 could be oxidized by the luciferase, I tried growing bac- 

 teria and also yeast on appropriate culture media, and 

 after some days of growth mixing the culture media con- 

 taining the products of bacterial or yeast growth with 

 luciferase, expecting to obtain light ; but no light appeared. 

 However, if a little crude luciferase solution was added 

 to the bacterial or yeast cultures and then allowed to stand 

 for some hours, light appeared whenever they were 

 shaken. Indeed such cultures behaved much as a suspen- 

 sion of luminous bacteria which has used up all the oxygen 

 in the culture fluid and will only luminesce when, by shak- 

 ing, more oxygen dissolves in the culture medium. Real- 



