)f6 R. H. A. PLIMMER AND J. L. ROSEDALE 



small size of the crop, proventriculus and pancreas, the organs from four to 

 eight birds were collected and examined together. A single small intestine 

 provided sufficient material, but in most experiments several were combined 

 as the whole series of sucro- or proteo-clastic enzymes were tested for simul- 

 taneously. Separate tests were made for lactase. At least two experiments 

 were made with each part, except the caeca. 



Preparation of enzyme solutions. 



The pancreas, on removal, was cut up into small pieces and ground with 

 sand in a mortar; the ground mass was put into glycerol in which it was kept 

 for several days in the presence of a few cc. of toluene. The solution was then 

 prepared by diluting with rather more than an equal volume of water and 

 filtering from sand, etc. 



The other parts of the alimentary canal were cut open and washed with 

 running water to remove the contents. The mucous membrane was scraped 

 off, ground up with sand and water and extracted for 24-48 hours with water 

 in the presence of a little toluene to prevent putrefaction. The aqueous 

 portion was strained off through cloth to remove sand and larger pieces and 

 used for testing for enzymes. 



It was not possible to scrape off mucous membrane from the inside of the 

 proventriculus. The organ is glandular, covered with numerous small teats, 

 which, on pressing with a scalpel, emit a yellowish, viscous, distinctly acid 

 secretion. This secretion was the material actually used after grinding with 

 sand and mixing with water. Nothing could be scraped off the gizzard, the 

 interior surface of which resembled parchment. 



Detection of enzymes. 



(a) Diastase and invertase. As substrates 100 cc. of 1 % starch solution 

 and 50 cc. of 3 % cane sugar solution were used. Two portions were measured 

 out with a pipette in separate flasks; a know!Q volume of enzyme solution was 

 added to one, and the same volume of boiled enzyme solution, after cooling, 

 to the other; 2 or 3 cc. of toluene were added to each, the flasks corked and 

 put into an incubator at 37° for one or more days. A test for starch by the 

 :odine reaction was made from time to time with a drop removed from the 

 mixture. At the end of the reaction time, the mixtures were washed into a 

 250 cc. measuring flask, a slight excess of colloidal ferric hydroxide added, 

 any excess of the latter removed by a few crystals of magnesium sulphate, 

 the volumes made up to the mark, the solutions filtered and reducing sugar 

 tested for by the complete reduction of 10 cc. of Fehling's solution. The 

 control solutions containing boiled enzyme did not reduce, or only gave a 

 slight reduction due to sugar present in the extract. 



(6) Lactase. The detection of lactase was carried out in a similar way to 

 that of diastase and invertase, using 50 cc. of 4 % lactose solution as substrate. 

 The enzyme and control mixtures were put directly into 250 cc. measuring flasks 



