786 EXPERIMENT STATION RECORD. 



has led Lloyd, working with English Cheddar, and Freudenreich, with the Swiss, 

 both to ascribe to the milk-souring ferments the chief role in these changes. 



"We have been able to confirm these results so far as the bacterial changes are 

 concerned, but early in our work observations were made which did not conform to 

 this theory. 



"From two independent lines of research it became evident that profound changes 

 of a physical and chemical nature occurred in milk in which bacterial fermentations 

 had been excluded. In these experiments the casein of the milk underwent practi- 

 cally the same series of decomposition changes that are to be found in a ripening 

 cheese, viz, the insoluble casein was converted into soluble proteids as indicated 

 above. 



"Parallel experiments with cheese gave the same result qualitatively and quanti- 

 tatively; the products formed could not be distinguished from those of a normally 

 cured cheese. 



"Having eliminated the effect of the organized ferments (bacteria) by means of 

 such chemical agents as ether, chloroform, benzol, etc., which do not affect the action 

 of unorganized ferments, the conclusion was forced upon us that the changes which 

 occurred were of a nonvital character and w«re undoubtedly due to enzyms. Two 

 hypotheses as to the origin of these enzyms suggested themselves — either they were 

 produced by bacteria which developed in the milk before the anaesthetics were 

 applied, or they were inherent in the milk itself. The possibility of bacteria func- 

 tioning in this way was excluded by securing milk drawn from the animal with 

 great care and treated immediately with antiseptics. These freshly secured milks 

 underwent identical changes with the others, showing that the enzyms were natural 

 to the milk." 



It was found that the proteids in separator slime digested more 

 rapidly than those in skim milk, showing that the enzyms were sepa- 

 rated in part in centrifugal creaming. Accordingly the slime was used 

 for isolating the enzyms, the fresh slime being mixed with an equal 

 weight of water containing 40 per cent of alcohol, the paste thus 

 formed strained through cheese cloth, an antiseptic (either thymol or 

 benzol) added to prevent fermentation, and the mass filtered after 24 

 hours. The filtrate was concentrated to about -^ its volume (never 

 heating over 25° C). It was usually quite acid, and when neutralized 

 with sodium carbonate gave a precipitate the filtrate from which was 

 of a clear amber color, decomposed peroxid of hydrogen rapidly (a 

 reaction common to most unorganized ferments), greatly hastened the 

 formation of soluble products in milk, and liquefied gelatin. 



"When added to milk it first coagulated the casein and afterwards redissolved 

 the curd— the time required for this change depending upon the amount of extract 

 added and the temperature. When the acid extract was added to milk the coagu- 

 lation of the casein was hastened and the curd formed was firm and quite similar to 

 that produced by rennet. In acid solution the digestion was much slower than in 

 neutral or slightly alkalin solutions, indicating that the hydrolytic principle which 

 it contains is more nearly related to a tryptic than to a peptic ferment. Attempts 

 to obtain more concentrated extracts by precipitating the enzym from the above 

 extract with absolute alcohol have thus far failed." 



Various experiments are reported showing how the formation of 

 soluble proteids in milk is accelerated by the enzym, that this power 

 of digesting casein is destroyed by heat, and that the change does not 

 occur as readily when even small amounts of hydrochloric acid are 



