481 



The quantity of diastase was determined by finding how long it 

 was before the starcii had completely disappeared out of a starch-solution 

 of definite strength, which had been mixed with the enzyme-solution, 

 whence the quantity of enzyme must 1)6 inversely proportional to 

 the time. The pi-esence or absence of starch was investigated by the 

 aid of an iodine-solution of known strength. Theoretically this is 

 not the best method; it would be better to detei-mine the quantity 

 of sugar which is formed in a definite short time from the starch 

 and then to consider the quantities of diastase proportional to the 

 figures thus found. But this is impossible in the case of the diastase 

 of Aspergillus niger, because there is here often such a very small 

 quantity of enzyme, that the sugar could not be deleruiined. For this 

 reason it was thought piefei'able to divide the enzyme-solution to 

 be examined, into a number of equal paits, and to mix each of 

 these with the same small quantities of very dilute starch-solution 

 and successively to test the mixtures after a definite interval for 

 the presence or absence of starch with llie aid of the same verv 

 dilute iodine-solution. 



Preliminary experiments showed that when a solution is used 

 which contains in 100 gr. water 62.5 mg. iodine and 62.5 mg. 

 potassium iodide, 1 ccm. of this is enough to colour a starch-solution 

 distinctly blue, when this contains 1 mg. soluble starch in 10 c.c. 

 of water, whilst when this amount is only 0.2 mg., a definite red- 

 violet colour still results. 



There are formed from starch, before it is completely hydrolysed 

 by the action of the diastatic enzyme of Aspergillus ?iiger, ery thro- 

 dextrin-like intermediate products, which with iodine are coloured 

 red ; so long as such bodies were present it was assumed that the 

 reaction was not ended; it was only when after mixing with the 

 iodine solution, the colour remained yellow, that the starch was 

 considered to have disappeared completely. Naturally all the fluids were 

 under the same condition. For this reason 5 c.c. of the fluid to be 

 investigated was always mixed in test-tubes with 5 c.c. of a solution 

 which contained 0.08 pc. of soluble starch and with some drops 

 of toluene; the latter was for the purpose of preventing any develop- 

 ment of fungi or bacteria in the experiments of longer duration. 

 The tubes were then shaken for a shoi-t time; and then placed in 

 the dark at a constant temperature of 24° C. After a certain time 

 the fluid was filteied off from one of the test-tubes and 1 cc. of 

 the above mentioned solution of iodine was then added. If the colour 

 was blue or red, this process was repeated some time later, until 

 the colour was decidedlv vellow. 



