57 



solution of amino acids become acid on the addition of neutral formalde- 

 hyde. Sugars may be considered as polymers of formaldehyde, and, as some 

 sugars have aldehyde groups capable of reacting, it is suggested that the 

 increased acidity in sterilized sugar media may be due to the interaction 

 of the amino acids in the medium with the aldehyde group of the sugar. 

 This reaction takes place very slowly and does not become evident before 

 30 minutes autoclaving. In Table XXVI. this contention is very clearly 

 demonstrated. Neither asparagine, maltose, lactose, nor rafinose become 

 acid on autoclaving, even for an hour,, but if maltose or lactose are 

 heated in the presence of asparagine a distinct acidity develops in 30 min- 

 utes, and a much more marked acidity after one hour. With raifinose, on 

 the other hand, there is no increase in acidity. The disaccharids (maltose 

 and lactose) contain reactable aldehyde groups whereas the trisaccrid, 

 raffinose, does not. The contention of Mudge that it is the presence of 

 an unstable sugar molecule together with an ammo acid which gives rise 

 to the acidity on sterilization appears very plausible. 



TABLE XXVI. EFFECT OF AN AMINO ACID (ASPARAGINE) ON PRODUCTION 

 OF ACID FROM SUGARS, BY AUTOCLAVING. 



(After Mudge 1917) 



In the revised Standard Methods for 1920, two important modifications 

 in the preparation of lactose broth were recognized: 



(1) The method of sterilization recommended for autoclaving is 15 

 pounds for 15 minutes provided the total time of exposure to heat is not 

 more than a half-hour. "Otherwise a 10 percent solution of the required 

 carbohydrate shall be made in distilled water and sterilized at 100 (for 

 one and a half hours), and this solution shall be added to sterile nutri- 

 ment broth in amounts sufficient to make a 0.5 percent solution of the 

 carbohydrates and the mixture shall then be tubed and sterilized at 100 C. 

 for 30 minutes, or it is permissable to add by means of a sterile pipette 

 directly to a tube of sterile neutral broth enough of the carbohydrates to 

 make the required 0.5 percent. The tubes so made shall be incubated at 

 37 C. for 24 hours, as a test for sterility." 



(2) The concentration of lactose is reduced to 0.5 percent. This 

 seems to be desirable as it tends to eliminate the danger of loss of the 

 colon group through destruction by acid formed in fermentation. 



