1912.] B. cloacae {Jordan) on Citric and Malic Acids. 5 



non-production of any notable quantity of alcohol. The distillation of the 

 original liquid was continued with steam, until titration of the distillate 

 showed no further neutralisation of alkali. The combined titrated distillates 

 were evaporated to low bulk, made up to 100 c.c. with distilled water, and 

 examined for volatile acids. Formic acid, when present, was determined by 

 heating 10 c.c. of the solution for two hours on the water-bath with 10 c.c 

 of a 10 per cent, solution of sodium nitrite and 50 c.c. of a saturated solution 

 of mercuric chloride, and weighing the precipitated mercurous chloride. 

 From the weight found, the amount of formic acid was calculated. 



The identity of the acids was established by determination of the molecular 

 weight by means of the barium salt. The residue in the distilling flask was 

 concentrated to 100 c.c, and completely extracted with ether in a continuous 

 extraction apparatus. The ether was distilled off, and the residue dried at 

 100° and weighed. The identity of this residue with succinic acid was 

 established by converting it into its barium salt. A weighed portion was 

 heated for one hour on the water-bath with water and excess of barium 

 carbonate. After filtration, the insoluble portion was well washed with hot 

 water and the filtrate made up to 200 c.c. ; 100 c.c. of this solution were 

 evaporated to dryness, dried, and weighed. Conversion of this residual 

 barium salt into barium sulphate showed it to consist of barium succinate, 

 and proved that lactic acid is not produced by the growth of B. cloacce on 

 either malic or citric acid. Further proof that the residue was succinic acid 

 was afforded by the determination of its melting point and by its reaction 

 with ferric chloride. 



The amount of residual acid, i.e. the acid not used up by the organism, was 

 determined in the residue from which the succinic acid had been extracted 

 with ether. This was made faintly alkaline with caustic potash, and 

 evaporated to low bulk to get rid of ammonia. The residue was then taken 

 up with water, exactly neutralised with acetic acid, and barium acetate 

 added. Four volumes of 95 per cent, alcohol were then added, and the 

 liquid allowed to stand overnight. In the case of the malic acid only a 

 faint precipitation took place, showing that all but traces of the malic acid 

 had been used up. In the case of citric acid, the precipitated barium salt 

 was filtered off, washed with 63 per cent, alcohol, ignited, and finally 

 weighed as barium sulphate, from which the amount of citric acid left 

 was calculated. 



In no case did the fermentation liquids give Vosges and Proskauer's 

 reaction, thus proving that methylacetylcarbinol, which is produced by the 

 action of B. cloacce on mannitol and glucose, is not formed. 



