196 Papers from the Department of Marine Biology. 



disconnected and shaken during extraction. To filter the extract it 

 is only necessar}'- to connect one of the arms of d with a desiccator fit- 

 ted with a funnel and filter rack. When the desiccator is exhausted, 

 C and the desiccator are connected, and the pressure of the hydrogen 

 in C drives the extract onto the filter paper. The firefly photogen 

 begins to phosphoresce when the atmospheric pressure reaches 5 to 

 6 mm., which means an oxygen pressure of 1 to 1.2 mm. Consequently 

 it is necessary to use a good vacuum-pump and make connections 

 air-tight. I found small-bore lead tubing sealed with Khotinsky 

 cement the best for the purpose. 



All of my experiments have been carried out in the dark and the 

 material observed at critical stages (as when the oxygen-free water was 

 added) to make sure that no light appeared, and always with negative 

 results. But to make sure that no very slow leakage of oxygen into 

 the filtering chamber occurred, I have carried out the extraction in a 

 special tube provided with a capillary sealed off during the extraction. 

 After extracting in this tube for 1| hours and admitting oxygen, no 

 phosphorescence appeared. Thinking that possibly the photogen dis- 

 solved in the extracting fluid did phosphoresce, but only so faintly as 

 to be invisible because distributed through a relatively large volume of 

 extract fluid, the unfiltered extract was evaporated in vacuo to a small 

 volume. This can be very easily done by placing the rubber tube from 

 the vacuum-pump over the capillary onto the special tube, exhausting, 

 and then breaking the capillary through the walls of the rubber tube to 

 connect with the air-pump. Even when concentrated the extract gave 

 no light on adding oxygen. 



We must conclude that the photogen is destroyed in distilled water, 

 even without oxidation. The search for watery solvent for the photo- 

 gen becomes, then, a search for a solvent in which the photogen is 

 stable. The following solutions were tried in addition to distilled 

 water. Extraction was allowed to proceed for from 1 to 1| hours. 



1. Ringer's solution (as representing fairly accurately the concentration and 



composition of the firefly's blood). 



2. 0.125 m NaCl. 



3. Sea-water (a mixture of chlorides and sulphates of Na, K, Ca, and Mg). 



4. 5 per cent NaCl. 



5. 0.05 m NaOH and 0.1 m NaOH. The dried, powdered firefly organs will 



phosphoresce strongly if sprinkled on the surface of 0.1 m NaOH. 

 G. 0.02 m HCl. Dried firefl}- powder will phosphoresce on 0.0125 m HCl and on 

 0.025 m HCl, but less briUiantly . Only one or two bright dots appear on 0.05 m 

 HCl and no phosphorescence occurs on 0.1 m HCl. If neutraUzed within 

 two minutes after contact with the acid, the hght does not appear in the 0.1 m 

 HCl treated material nor become brighter in the 0.05 m and 0.025 m treated 

 material. 



In each case, after extraction, oxygen was admitted and the solution 

 shaken, yet in no case did light appear either in the undissolved residue 

 or in the solution. The 0.02 m HCl extract was also neutralized, as it 



