184 



Prof. H. E. Armstrong and Mr. H. W. Gosney. 



In so far as our results can be brought into comparison with those of 

 previous workers, they appear to be in harmony with their observations: 

 but the activity of the enzyme we have had at our disposal, thanks to 

 Tanaka's important discovery, appears to have been in excess of that used by 

 others. 



As it was obvious that if the limit reached in our synthetic experiments 

 (about 40 per cent, when equivalents are used) were to be exceeded, the 

 water produced in the interaction must be removed as it is formed, we 

 endeavoured to secure this end by carrying out the synthesis in vacuo in a 

 flask connected with drying apparatus. The results obtained have been 

 uniformly unsatisfactory, inferior, in fact, to those obtained under ordinary 

 conditions. Apparently, as pointed out by us previously, the intervention of 

 a film of water is necessary at the interface of the system, where interaction 

 takes place ; if this be removed, action comes to an end. 



In working with the Tanaka preparation, it is noticeable that the activity 

 varies considerably, in a manner which is difficult to understand at first. On 

 more than one occasion we have found that an enzyme which was quite active 

 hydrolytically was inert when used as a synthetic agent with a mixture of 

 acid and glycerol free from water : ultimately, this behaviour was traced to 

 " overdrying," as on the addition of a very small amount of water the enzyme 

 became active. 



Enzyme which has been used and then recovered, by washing it free from 

 oily matter by means of light petroleum, is found, as a rule, to be still 

 active but usually less active than it was originally : the variable behaviour 

 of such preparations is not surprising, however, in view of the colloid nature 

 of the material and the effect which alterations in the state of aggregation 

 and of surface conditions must have. It is noteworthy that the hydrolytic 

 activity — in presence of a relatively large excess of water — of the enzyme is 

 much more reduced by such treatment than the synthetic activity. 



Nature of the Products of Change. — In order to ascertain whether the product 

 of the synthetic action of Lipase is a nearly pure triglyceride like the natural 

 fats and oils, the amount of glycerol uncombined in each experiment of the 

 first series was determined, following the directions given by Lewkowitsch. 



After titration, the contents of each flask was washed into an evaporating 

 basin, boiled to expel most of the alcohol and then just acidified by sulphuric 

 acid. After heating the liquid to the boiling point, the solution of glycerol 

 was filtered off and the fatty acids and enzyme on the filter were then well 

 washed with hot water : the filtrate was purified by addition of a solution of 

 basic lead acetate and the glycerol estimated in the clear nitrate by Hehner's 

 method (oxidation by an acid solution of potassium bichromate). 



