CULTURAL CHARACTERS OF BACTERIA 37 



calcium carbonate. The flask is then inoculated with the 

 culture, and allowed to incubate at the optimum temperature 

 for 10 days. The culture is then filtered to remove undissolved 

 calcium carbonate. The filtrate is then made slightly acid 

 with HC1, and then slightly alkaline with Na 2 C0 3 . The pre- 

 cipitated CaC0 3 is filtered off. The filtrate is then introduced 

 into a retort of the proper size, and distilled. The distillate, 

 50 cc. of which should be collected, may contain alcohol, acetone, 

 or aldehyde. To test the presence of these bodies, add to 10 cc. 

 of the distillate 5 or 6 drops of a 10 per cent solution of caustic 

 potash, and warm the liquid to about 50 C. A solution of 

 iodide of potassium, saturated with free iodine, is added drop by 

 drop, until the liquid becomes a permanent yellowish brown 

 color. It is then carefully decolorized by adding, drop by drop, 

 the caustic potash solution. If any of the preceding bodies are 

 present, iodoform is gradually deposited at the bottom of the 

 tube as yellow crystals. 



For the separate detection of alcohol, aceton, and aldehyde, 

 consult Allen's " Commercial Organic Analysis," Vol. I. 



The residue in the retort will contain the volatile and fixed 

 organic acids. To separate the volatile acids, the contents of the 

 retort is made strongly acid with sulphuric acid, and as much 

 distilled over as is consistent with the safety of the retort. The 

 distillate may contain acetic, formic, propionic, and butyric acids. 



The quantity of the volatile acids is determined by titration. 

 The preceding distillate is made alkaline with baryta water, 

 and the solution evaporated to dryness. Then add 20 cc. of 

 absolute alcohol, and let stand for 1-2 hours, with frequent 

 stirring. Filter, and wash with alcohol. The residue will 

 contain barium formate and acetate, and the filtrate mainly 

 barium propionate and butyrate, and a little formate and 

 acetate. Evaporate the filtrate to dryness ; dissolve in 1 50 cc. 

 of water ; saturate with calcium chloride, and distil off the pure 

 butyric acid. Dissolve the residue on the filter in hot water ; 



