LABORATORY AND GREENHOUSE STUDIES, 99 



and lactic acids were present, and succinic in small amount. The 

 coconut organism is so similar to Bacillus coli in its cultural charac- 

 teristics that it would be very surprising if it were not likewise simi- 

 lar in its chemical products. The foregoing single analysis is not 

 sufficient to show that they are not the same in this respect, and it 

 should be repeated. 



Dr. Smith gave flasks of this organism (grown in river water con- 

 taining Witte's peptone, Merck's dextrose, and calcium carbonate) 

 to Dr. Carl L. Alsberg for quantitative chemical analysis, who 

 reported as follows: 



Received from Dr. Erwin F. Smith, February 23, 1910, one flask labeled "4101 

 February 4, 1910, Coconut from Agar, February 2, 5083, fr. 5 January 26." 



The culture flask contained a white deposit, which on close inspection was seen 

 not to be homogeneous, for in addition to the calcium carbonate put into the flasks 

 before inoculation there were a few crystalline crusts, the total bulk of which was 

 small. The precipitate was removed by filtration. The filtrate was acid and on 

 warming some carbonic acid gas was liberated. 



A part of the culture liquid filtered free from the calcium carbonate was acidified 

 with sulphuric acid and exhausted with ether. The ether on evaporation left a mass 

 of white crystals which after repeated recrystallization from hot water had a melting 

 point of 183^° C. uncorr. These crystals gave a very powerful pyrrol reaction (Neu- 

 berg). The aqueous solution was neutralized with ammonia and an excess of silver 

 nitrate added. The resulting white silver salt was filtered off with suction and washed 

 successively with water, alcohol, and ether. After drying in a desiccator 0.6655 

 gram was weighed into a crucible and ignited to constant weight. 0.4305 gram 

 silver remained, or 65.12 per cent. The amount calculated for silver succinate is 

 64.70 per cent. On the basis of the silver content of the silver salt, the melting 

 point of the free acid, and the pyrrol reaction, it is safe to say that this substance is 

 undoubtedly succinic acid. 



The mother liquor from which the succinic acid had been removed was subjected 

 to distillation with steam. The distillate was quite acid. It was neutralized with 

 ammonia, and silver nitrate added. The latter was immediately reduced to metallic 

 silver, so that formic acid was probably present. The black silver precipitate was 

 removed, and the clear filtrate concentrated in a desiccator. A crystalline crust, gray 

 in color, formed in the course of a few days. This was removed, washed and dried, 

 and, though obviously impure, its silver content was determined. 0.2305 gram 

 yielded 0.1365 gram silver, or 59.23 per cent. As this corresponds neither to silver 

 acetate (64.67 per cent Ag) nor to silver propionate (59.67 per cent Ag), and as the 

 preparation was obviously impm-e, the determination was repeated. 



Another culture was taken and after removal of the calcium carbonate distilled 

 with the addition of a little sirupy phosphoric acid. The acids in the distillate were 

 converted into the barium salts by evaporating on the water bath with an excess of 

 barium carbonate. To the solution of the barium salts an amount of silver nitrate 

 was added sufficient to combine with only a portion of the acid. On standing over 

 night beautiful long white needles were formed. These were removed, washed and 

 dried, and the silver content determined. 0.3925 gram yielded 0.2530 gram silver, 

 or 64.46 per cent. Silver acetate contains 64.67 per cent. It is therefore evident 

 that beside formic acid there can be present no other volatile acid but acetic. 



The presence of formic acid was further verified by distilling a fresh portion of the 

 culture liquid after it had been rendered faintly alkaline with sodium carbonate. 

 Under these conditions any aldehyde which may have reduced the silver in the 

 228 



