1909.] 



Latent Life of Resting Seeds. 



425 



enzymes were precipitated from their solutions. The method followed was 

 almost identical with that described by Dean* when dealing with the 

 proteolytic enzymes of Cucurlita pepo. Twenty grammes of the grain were 

 ground and mixed with 100 c.c. of cold, boiled water, and shaken for 

 two hours. The mixture was then filtered and the filtrate precipitated by 

 a volume of saturated ammonium sulphate equal to that of the original 

 filtrate. A white, more or less flocculent precipitate was thrown down, 

 which was filtered off and dried overnight in an exhausted desiccator over 

 strong H 2 S0 4 . 



The precipitate was scraped off by means of a sterilised knife as before, 

 and dissolved in a small amount of cold, boiled water. 



In testing for fibrin-digesting enzymes, 1 c.c. of the aqueous solution 

 produced above was put into each of three sterilised test-tubes A, B, and C. 

 The contents of the test-tube C were boiled well and allowed to cool. Into 

 B and C were put - 2 gramme of well- washed fibrin, and drops of 

 chloroform were added to each of the three test-tubes. The three test-tubes 

 provided with corks were placed in the oven at 35° C. and left for an 

 interval of time varying from two to four days. • 



Examination of the contents of all these test-tubes for the detection of 

 peptone showed that B and C both gave faint biuret reaction, the boiled as 

 well as the unboiled specimens. 



A little of the liquid from each test-tube was examined under the micro- 

 scope, with the result that it was found to be swarming with bacilli, by which 

 the conversion of proteids into peptones had been wholly or partially effected. 



This difficulty regarding the bacteria was due to the prolonged time 

 necessary for the action of these ferments to become manifest ; and it was 

 evident that the addition of 5 drops of chloroform, and the subsequent 

 plugging of the test-tubes, was inadequate to maintain antisepsis, so that 

 it was necessary to adopt more stringent measures to destroy the bacteria, 

 but still not to impede in any way the action of any digestive ferments 

 which might be extracted from the resting seeds. 



After many unsuccessful attempts to find some means of effectively 

 fulfilling these conditions, the required end was attained by soaking the 

 grain in strong chloroform for about five minutes after weighing it, and 

 grinding it in the mill while still wet. Care was taken that all the bottles, 

 apparatus, and water employed throughout the experiments were thoroughly 

 sterilised beforehand. Prior to putting the test-tubes into the oven 5 drops 

 of chloroform were added, and the tubes were stopped with plugs of cotton 

 wool. In this way complete sterility was produced. 



* ' Botanical Gazette,' vol. 39, May, 1905, " Proteolytic Enzymes," p. 331. 



