132 



Invertase. 



A slant tube of 10 per cent cane sugar agar, fragments of which gave 

 no precipitate of copper oxide on boiling 2 minutes in Soxhlet's solu- 

 tion, gave after ^6'. hyacinthi had been grown on it for 29 days, a very 

 copious rusty precipitate after boiling 2 minutes in the same solution. 

 Cane-sugar bouillon gave the same result. This indicates that cane 

 sugar is inverted, and to a much greater extent than is needed for the 

 growth of the organism, but we may not therefore assume the existence 

 of an invertase. The fact that cane sugar was not inverted when put 

 into dead or sterile tubes of Ps. hyacinthi cultivated in beef broth and 

 peptonized beef broth, seems to show either that the living organism 

 itself is necessar}^ to bring about the inversion or else that invertase is 

 formed onl}^ when it is required, i. e. , in the presence of cane sugar. 



My first experiments were in non-peptonized alkaline beef broth 

 (stock 382). The contents of tubes 3, 4, 7, 10, 12, and 15 of February 

 7 (cover-glass inoculations 24 days old) were poured together and forced 

 through a Chamberland filter. Two 10 c. c. portions of the sterile fluid 

 were then pipetted into cotton-plugged sterile test tubes, and to each 

 was added 300 milligrams of cane sugar. To one of these tubes chloro- 

 form was added and to the other thymol. They were then set away at 

 18° to 24° C. 



On the fifth da}^ each tube was tested by pipetting 2 c.c. of the clear 

 fluid into boiling Soxhlet's solution, and continuing the boiling 1^ 

 minutes. In neither case was there an}^ reduction. These tests were 

 repeated on the thirty-fifth da}^ with the same negative result. 



A duplicate series from tubes 1, 8, 14, and 18 of Februar}^ 7 (same 

 stock) led to the same result. In neither portion was there an^^ reduc- 

 ing sugar on the fifth or thirt^^-fifth day. 



Thinking that the invertase might possibly have been retained in the 

 walls of the filter, or that the presence of peptone might be essential to 

 the formation of invertase, the experiment was repeated as follows: 



Three old cultures of Ps. hyacinthi — (1) in beef broth with Wittes's peptone (459); 

 (2) in beef broth without peptone (382), and (3) in beef broth with the trace of 

 muscle sugar removed by B. coli (404) — were sterilized by heating them for 10 min- 

 utes at 54° C, viz, at a temperature high enough to kill the organism and low enough 

 to be harmless to invertase. To each tube was then added 500 milligrams of cane 

 sugar and 150 milligrams of thymol. The sugar was transferred from a sterile solu- 

 tion by means of a sterile pipette. Along with these three cultures two other old 

 cultures were tested, viz, one of P.s. cumpestris and one of Ps. stewarti, each in stock 

 382. These tubes were set away for 19 days at 25° to 30° C. 



At the end of this period they were tested as follows for the presence of reducing 

 sugars: 



Twenty-five cubic centimeters of Soxhlet's standard alkaline solution was added to 

 25 c. c. of his standard copper sulphate solution, and after mixing was divided into 

 5 equal parts in 5 clean porcelain capsules and 40 c. c. of distilled water added to 



