Concentration upon the Optimum Temperature of a Ferment. 411 



Measurement of the hydrogen ion concentration of the medium, resulting 

 from the addition of various quantities of acid and of alkali per 5 cm. 3 of the 

 reaction mixture, containing 3 mgrm. of dissolved enzyme, were made by the 

 colorimetric method of Sorensen (loc. cit.). The results are contained in 

 Table II, and for convenience of description, in what follows, they have been 

 reproduced on the base line of fig. 1, underneath the respective quantities of 

 M/100 H 2 S0 4 and M/100 Na 2 C0 3 which give rise to them. 



Table II. 



Quantities of acid and of alkali added per 3 mgrm. of 

 enzyme contained in 5 cm. 3 of the reaction mixture. 



Corresponding H + concentrations. 



-60 M/100 H.,S0 4 1 



q !_ H • concentrations greater than 

 q _2Q [ the optimum of fig. 1. 



0-15 „ J 



'07 „ H + concentrations equal to the 



optimum of fig. 1. 



n .nn / j. "i l- \ 1 H + concentrations less than the 

 00 (natural reaction) > ,. r « , 

 0-05 M/100 Na.CO, J optimum of fig. 1. 



10~ 3U ; citrate, methvl orange. 



IO- 30 '; 



10- 34 _ ; 



10 -4 "; phosphate, neutral red. 

 10~ 5 ' 6 ; „ 



10 -6,2 ; 



10 -6 ' 8 ; 

 10 -7 ' 2 ; 

 10-'- 5 ; 



The second stage of the enquiry, which consists in determining the optimum 

 temperature of the enzyme for a series of hydrogen ion concentrations 

 corresponding to different points on the optimum reaction curve, figured in 

 fig. 1, was next undertaken. Nine different H + concentrations of the medium 

 were thus studied: lO" 3 ' , 10" 32 , 10" 34 , 10~ 4 ' 7 , lO" 5 - , lO" 6 ' 2 , lO" 6 ' 8 , 10" 7 - 2 , 

 and 10" 7 - 5 . 



The practical details of the first determination in the series may be given as 

 an example, to show how these optimum temperature determinations were 

 carried out. A solution of the enzyme was prepared containing 3 mgrm. of 

 the preparation per cubic centimetre and, after standing for a half to one 

 hour, was introduced in portions of 1 cm. 3 into eight clean test-tubes, already 

 containing 90 mgrm. of maltose, 06 cm. 3 of M/100 H2SO4 and 3 - 4 cm. 3 of 

 redistilled water. Such a mixture, according to Table II (or fig. 1), 

 corresponds to a H + concentration of 10 -3-0 . The tubes were closed with 

 sterile corks, plunged into water thermostats at known temperatures, and 

 incubated for 16 hours, when the enzyme action was stopped and the quantity 

 of maltose hydrolysed in each tube determined as before. The numbers 

 found are set out in Table III, together with the numbers obtained for the 

 other members of the series. 



VOL. LXXXVIII. — B. 



