METABOLISM OF ACTINOMYCETES 6 



EXPEEIMENTAL 



Since glycerol proved to be a very favorable source of energy 

 (see paper II of series), it was used in the following experiments. 

 To each liter of medium containing the following ingredients: 

 30 grams glycerol, 1 gram K 2 HP0 4 , 0.5 gram KC1, 0.5 gram 

 MgS0 4 , 0.01 gram FeS0 4 , were added fibrin, casein, powdered 

 egg-albumin, Witte peptone, asparagin, leucin, glycocoll, or urea, 

 5 grams each; NaN0 3 , NaN0 2 , (NH 4 ) 2 S0 4 or (NH 4 ) 2 C0 3 , 2 

 grams each. The casein and egg-albumin were first dissolved in 

 a dilute NaOH(x^) solution, then added to the medium; the 

 fibrin was added in small pieces to the individual tubes. The 

 media were mixed, tubed, 10 to 12 cc. to each tube, and steri- 

 lized at 15 pounds pressure for fifteen minutes. Several tubes 

 from each medium were inoculated with each of a series of rep- 

 resentative Actinomyces and incubated at 25° for a period of 

 fifteen to sixty days. 



Growth is designated by figures as follows: — none, 1 — scant, 

 2 — fair, 3 — good, 4 — very good, 5 — excellent. In studying the 

 figures for growth one should keep in mind that they are only 

 relative and 2 does not designate twice as much growth as 1 or 

 half as much as 4. The scantest growth, even if only a few 

 tiny flakes at the bottom of the tube or a few minute masses 

 floating on the surface or distributed through the medium, was 

 designated as 1. The most abundant growth was designated 

 by 5, while the other figures fall between. All the cultures were 

 compared on the same basis considering the set as a whole and 

 not each organism separately. In describing the aerial my- 

 celium the signs, + and + + were used, when it was present, 

 the first to designate a thin powdery layer and the second a 

 heavy, usually cottony cover. The ammonia was not deter- 

 mined quantitatively in this experiment, but merely qualita- 

 tively, by means of Nessler's reagent. The amino-nitrogen was 

 determined by means of the micro-apparatus of Van Slyke. 

 The hydrogen-ion concentration was obtained by means of the 

 phenol-sulphon-phthalein series of indicators suggested by Clark 

 and Lubs (1917); this was designated by the terminology of 

 Sorensen, using the pH values. 



