156 NOTE LPOX THE EXTRACTION OF ALIUS FROM CLLTURES. 



ex(-ess of baryta, and evaporated to small volume, with the idea of hydrolysmg 

 any aeid esters. The li(niid wa.s treated with sulphuric acid and extracted for 12 

 hours with ether. The residual acids were neutralised with 9.15 c.c. of N/10 

 baryta water and were treated with sufBcient alcohol to make a 75 % solution. 

 The precipitated barium succinate was filtered off and dried at 130° when it 

 gave 0.041 G gi-am which contained 54.7G % Ba. Barium succinate contains 5-4.16 

 %. The filtrate, when evaporated and dried at 130°, gave 0.1038 gram contain- 

 ing 33.33 % Ba. The barium succinate corresponded to 3.29 c.c. N/10, which 

 left 5.80 c.c. for the acid in the alcohol-soluble salt. Deducting the Ba. and 

 allowing for the H2 equivalent, the acid was calculated as weighing 0.0697 gram. 

 This gave a neutralisation e(|uivalent of 119 and, as we have seen, the Ba. content 

 was 33.33 %. Isovaleric or levulinic acid is indicated, but the ((uantity worked 

 upon was vei'y small for absolutely definite identiticatiun. 



As a matter of fact, the substance was probably lactic acid, for in working 

 through another culture, the succinic acid was pieci]>itated as the barium salt in 

 75 % alcohol and the filtrate, after evaporation, acidification and extraction with 

 ether, yielded a syrupy residue which, when boiled wth zinc oxide, furnished 

 crystals of zinc lactate. 



One ha-s to be very (careful in deducing the nature of the acid from small 

 iiuantities of material as, even with comparatively large (juantities, one may lie 

 led into error. For example, a culture of the same yeast was made with dextrose, 

 ammonium nitrate, and sodium phosphate in the presence of chalk. In working 

 up the products, the ethereal solution was evaporated in the air and then dis- 

 tilled in a current of steam. On concenti-ating the solution on the water bath 

 a disengagement of nitric oxide occurred. Tiie nitric acid had formed nitrous 

 ether during a part of the process, and this had not lieen removed by the steam 

 and liad decomposed on warming in the open. The residual acids were neutra- 

 lised with baryta water in the presence of phenolphthalciii to a permanent pink 

 colour, when a precipitate of barium phosphate and oxalate wa-s thrown dow'n. 

 The filtrate on concentration threw down successive crops of a crystalline pre- 

 cipitate containing 47.8 % and 48.9 % of Barium. A portion of the precipitate 

 acidified and extracted with ether yielded crystals of succinic acid melting at 

 183°. The mother liciuor now required a considerable (|uantity of baryta water 

 to give it a permanent i)ink tint. Thus the original addition of baryta to the 

 first solutiim sufficient to give a permanent pink colour was not 

 sufficient to jiroduce normal barium salts, for if so the amount of 

 Barium in the precipitated succinate would have been neaier the theoretical .54.1 (i 

 % and tlio furtlier addition of baryta to tlie mother liquor would have been un- 

 necessary. 



Summari/. — The extraction of the fixed acids from bacterifil or from yeast 

 cultures is, as has been shown by other writers, of the nature of a mononioleeular 

 reacti(m ami should be continued until no more acid is extracted. 



The preparation of salts, such as those of barium, by neutralising the cx- 

 tract«d acids until a jiink colour is obtained in the presence of plicnolphtliidcin. 

 may be faulty as the reaction is much slowei- than is generally supposed. 



