THE CHEMISTRY OF LIGHT PRODUCTION 129 



izing that in bacterial cultures in test tubes, anaerobic 

 conditions soon appear, and also the strong reducing 

 action of bacteria upon many substances (for instance, 

 nitrates or methylene blue) under anaerobic conditions, 

 it struck me that the bacteria might be reducing the oxida- 

 tion product of luciferin to luciferin again. We must 

 remember that since crude luciferase solution is a cold- 

 water extract of a luminous animal allowed to stand until 

 all the luciferin has been oxidized, it must contain oxy- 

 luciferin as well as luciferase and will give light if 

 the oxyluciferin is again reduced and oxygen ad- 

 mitted. This appears to be the correct explanation of 

 the above experiments. 



Oxyluciferin may also be readily reduced by the use 

 of the blood of the horse-shoe crab (Limulus) allowed to 

 stand until bacteria develop. This experiment is of special 

 interest because the blood contains haemocyanin, which is 

 colorless in the reduced condition and blue in the oxy- 

 condition. The color change thus serves as an indicator 

 of the oxygen concentration in the blood. A sample of 

 foul-smelling Limulus blood full of bacteria will become 

 colorless on standing in a test tube for 10 to 15 minutes, but 

 the blue color quickly returns if shaken with air. Such 

 a blood has the power of reducing oxyluciferin through the 

 activity of the bacteria which it contains. Fresh blood 

 has very little if any reducing action. 



Not only bacteria but also tissue extracts have a strong 

 reducing action in absence of oxygen. Thus, muscle tissue 

 stained in methylene blue will very quickly decolorize 

 (reduce) the methylene blue if oxygen (air) is kept away, 

 but the blue color immediately returns if air is admitted. 

 Oxyluciferin {i.e., a solution of luciferin which has been 



