82 ROPIXESS IJf WATTLE BARK INFUSIONS, 



potassium citrate 0.3 <^-r with 0.1 St <'t taunic aciil atlded at tlie time of poiiriiig 

 the jilates. After 17 clays at 22°, the tihiis were jiicked off. suspeiuled in water 

 overnight, and coagulated with alcohol. The water and alcohol treatment was 

 repeated. The films suspended ill water were heated in the autoclave fo'" 15 

 minutes at three atmospheres pressure, l)ut tlie treatment did not liquefy them. 

 The water was acidified with two c.c. of normal sulplnnic acid wliich produced an 

 acidity of -)-5°, and the suspension was again autoclaved for an liour. The 

 films liad dissolved. The solution was carefully evaporated to smaller volume, 

 and a ijoi'tion was clarified with alumina cream and the rotation of the fluid 

 observed. The ash-free solids had a specific rotation of [a]D = -|-0.017°. The 

 solution gave a yellow precipitate with Fehling's .solution, and it apjieared that 

 the treatment had partly hydmlysed the gum. It was treated witli alcohol, and 

 the unaltacked gum was filtered off. The ash-free solids in the filtrate had a 

 specific rotation of [,i]ji ^=-|-0.002°. The ditlcrence between these two rota- 

 tions shows that the gum. ]ireci|>itabl(' by alcohol is slightly dextro-rotatory. 



Tlie Aciih fiiriiicd hij the Biirleriii. 



In the routine testing, the bacteria, A and B, were found to produce acid 

 and gas from dextrose and saccharose when these sugars were present in broth. 

 The nature of the acids was further examined. Tlie bacteria were grown in a 

 medium containing 5 % of dextrose, 1 % of meat-extract, and 0.5 % of sodium 

 jihosphate with the addition of chalk from time to time. The bacterium B2 used 

 up the carbonate more quickly than Al or A2, and naturally yielded a greater 

 quantity of acids when tlie cultures were worked up at the end of a month's 

 inculiation. 



Tlie methoil followed in determining the nature of the acids, etc., was es.sen- 

 tially that described in these Proceedings* 



Ethyl alcohol was found in small amount in the cultures from both bacteria. 

 It was proved by giving the iodoform test, by burning with a blue flame and by 

 having aB.P. of 79°. 



A small quantity of insoluble fatly acid was ohlaiiicd fnim the culture of each 

 bacterium. That from A melted at 37°, and from B at 32°. Both were pro- 

 bably mixtures, but the quantities were too small to separate. The softer acids 

 of B were spread on a piece of filter paper and incubated at 28°. when the 

 more fluid portion was absorbed, leaving a resid\ie which melted at 40°, and be- 

 came clear at 42.5°. 



The volatile acids did not contain formic acid. The solutions were neutralised 

 with baryta water, and after evaporation were dried at 140°. The A salts con- 

 tained .52.36 % of barium, the B salts 53.8 %. As barium acetate contains 

 53.73 % of l)ariiuu. it is clear that the volatile acids in both ceases consisted 

 entirely of acetic acid. 



The non-volatile acids contained a small (|uantity of an acid giving a lime 

 salt insoluble in 70 9r alcohol. After aciilification and extraction with ether, 

 monoclinic prisms, melting at 182°, were ol)tained. Succinic acid under the 

 same conditions melted at the same temperatnre, and thus it was proxcil that 

 botli bacteria form a small (|uantity of succinic acid. 



The only other non-volatile acid was lactic. The zinc salt of lactic acid was 

 jircpared from two cultures of the A bacterium originally seeded with Al and 



•/.or. cit., 1901, 606; 190.% 114. 



