482 



The question was whether in this way, by repeated trials, 

 trustworthy results could be obtained. In order to ascertain this, some 

 preliminary experiments were carried out. From a culture of Aspei'- 

 yiUus niger which was 5 days old, the tluid was filtei-ed off and 

 divided into two equal portions. One half remained unchanged, the 

 other was again divided into two, and one of (he halves was boiled 

 for a short time to ensure destruction of the enzyme, and after 

 cooling it was mixed with the unboiled portion, so that tliere were 

 two fluids of which one must contain double the quantity of the 

 enzyme in the other. 5 cc. of each of (he fluids was mixed in a 

 test-tube with 5 cc. iodine-solution and after certain intervals each 

 tube was tested for the presence of starch. It was found that the 

 undiluted solution after 95 minutes was still coloured some- 

 what orange yellow l)y iodine, whilst aftei' 100 minutes the colour 

 was pure yellow, so that the colour-change had taken place after 

 97.5 minutes. In the twice-diluted solution ihe change had taken 

 place in the interval between 180 and 190 minutes, therefore in 

 about 185 n)inutes. The proportion of 97.5:185 is 1:1.9, whilst 

 the quantities of enzyme were to one another as 2:1, so that the 

 error in deteiinination was not more than 5 " ,. 



In another case the starch was hydrolised hy the undiluted solu- 

 tion in 495 minutes, by the twice-diluted in 907 minutes and by 

 the 10 times diluted in 4365 minutes. These figures are as 1:1.8: 

 8.8 instead of as 1.: 2 : 10. 



Lastly there was a third case in which the enzyme-solution had 

 been diluted 2, 4, 8, and 16 times. The result is shown in the 

 following table. 



Still coloured red ^^ . ' Propor- 



ance of Average 



with I. 3tg^^j^ tion 



Undiluted solution Afterl h. 45 min. After 2hoursl 1 h 52 min. 1.— 



Twice diluted „ „ 3 „ 20 „ « 4 „ 3 „ 40 „ 2.— 



4 times „ „ „7 „ 30 „ „ 8 „ 1 „ 45 „ 4.1 



8 „ „ „ , 13 „ 30 „ „ 16 „ 14 „ 45 „ 7.9 



16 „ „ „ „ 27 . 00 „ „ 28 „ [27 „ 30 „ 14.7 



When it is remembered how tentative these estimations were 

 then the agreement between the determination and reality must be 

 considered very satisfactory; the difference hei-e amounts to much 

 less than 10"/. in most cases. 



