6 Variations in Bacteria Caused by Change of Medium 



Loosely Combined Sulphur was estimated as sulphide. The mate- 

 rial was boiled 4 hours in 10 per cent. KOH, neutralized with acetic 

 acid, and precipitated as lead sulphide, care being taken during 

 the process to prevent the formation of lead sulphate or carbonate. 



PHOSPHORUS. The material was first decomposed by boiling nitric 

 acid precipitated by ammonium-phospho-molybdate, redissolved 

 in ammonium hydroxide, reprecipitated and estimated as magne- 

 sium pyrophosphate. * 



CALCIUM. The material was decomposed by boiling sulphuric acid, 

 precipitated by absolute alcohol, washed with 40. per cent alcohol 

 at 34 C., and weighed as sulphate. 



TOTAL NITROGEN. This was estimated by the macro-Kjeldahl method. 

 AMINO-NITROGEN was determined by the Van Slyke method. 



COAGULABLE PROTEIN 



Preliminary. Since a quantitative estimation of the protein content 

 of the bacterial cell would assist in determining biological variations, 

 the ideal method of protein extraction would involve the disintegra- 

 tion of the cell material without injury to the proteins. But the 

 cellulose-like structure of the cell presents interference. To dissipate 

 this, two alternatives are offered: to employ a cellulose enzyme, or 

 to carry on extraction as far as possible by means of chemical reagents. 

 Enzymes, however, are objectionable. Chemical reagents, therefore, 

 must be resorted to, success depending on proper selection. 



In exploring the effect of acids, alkalis, and neutral salts and sol- 

 vents on proteins, it appeared that distilled water and sodium chloride 

 possess greatest efficiency especially at reduced pressure. Of the two 

 solvents distilled water was discarded because it extracted fats together 

 with proteins; sodium chloride was selected as being a more powerful 

 protein solvent. 



The method of determining quantitatively the protein content of 

 bacteria was therefore the following: Weighed amounts of wet bac- 

 teria were subjected to extraction by a definite amount of 6. per cent 

 NaCl for 24 hours at 50 C., under 250 mm. pressure. After sedi- 

 mentation, decantation of the supernatant liquid followed. The sedi- 

 ment was washed, and after the wash waters had been added to the 

 original liquid, the whole was reduced to its original volume by evapo- 



