194 



CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



(4) Filter. 



(5) Dissolve the precipitate in 1000.0 cc. 

 distilled water. 



(6) Use this solution for preparing the 

 medium instead of (6) above. 



(b) This medium should contain little, if 

 any, water soluble B vitamin. 



(1) Take the filtrate from variant (a) 

 step (4) and concentrate to 100.0 cc. 



(2) Add 95.0% alcohol to get a con- 

 centration of 90.0% alcohol. 



(3) Allow to stand over night at 5°C. 



(4) Filter. 



(5) Evaporate the filtrate to dryness. 



(6) Dissolve in 1000.0 cc. distilled 

 water. 



(7) Use this solution for preparing the 

 medium instead of (6) in the origi- 

 nal medium. 



(c) This medium should contain water 

 soluble B vitamin. 



(1) Extract 10.0 g. autolized yeast in a 



Soxhlet apparatus for 8 hours with 

 95.0% alcohol. Fresh alcohol then 

 added and continued another 8 

 hours. Repeat this action for 

 40 hours, breaking up the yeast 

 clumps in thimble several times. 



(2) Evaporate alcohol after extraction 

 at 60-80 °C. 



(3) Add residue to 500.0 cc. distilled 

 water. 



(4) Add (3) to 500.0 cc. of a 2.0% Difco 

 peptone solution. 



(5) Adjust, heat, filter and sterilize as 

 for the original medium. 



(d) This medium should contain no water 

 soluble B vitamin. 



(1) Dissolve the residue from variant 

 (c) step (1) in 500.0 cc. distilled 

 water. 



(2) Add (1) to 500.0 cc. of 2.0% Difco 



peptone solution and prepare the 

 medium as in variant (c) step (4) 

 and (5). 



(e) This medium should contain water 

 soluble B vitamin. 



(1) Add 10.0 g. autolized yeast to 

 500.0 cc. of distilled water. 



(2) Add (2) to 500.0 cc. of a 2.0% Difco 



peptone solution prepare the me- 

 dium as in variant C step (4) 

 and (5). 

 Reference: Ayers and Mudge (1922 pp. 450, 

 451). 



692. Spronck's Yeast Infusion Peptone 

 Solution 

 Constituents: 



1. Water 5000.0 cc. 



2. Yeast 1000.0 g. 



3. NaCl 25.0 g. 



4. Peptone (Witte) 100.0 g. 



Preparation : 



(1) Boil 1 kilogram of commercial yeast 

 in 5 liters of water for 20 minutes, 

 stirring constantly. 



(2) Pour into glass cylinders and allow 

 to stand for 24 hours. 



(3) Decant the clear fluid. 



(4) Carefully acidulate the fluid and add 

 5.0 g. NaCl and 20.0 g. Witte's peptone 

 per liter of fluid. 



(5) Neutralize by the addition of soda 

 and then add 7.0 cc. of a normal soda 

 solution for each liter of medium. 



(6) Heat, and filter on a paper. 



(7) Distribute in flasks. 

 Sterilization: Sterilize at 120°C. (time not 



specified). 

 Use: Production of diphtheria toxin. 

 References: Spronck (1898 p. 702), Tanner 



(1919 p. 57), Besson (1920 p. 35), Harvey 



(1921-22 p. 120). 



693. Cohen and Clark's Yeast Infusion 



Peptone Solution 

 Constituents : 



1. Water 1000.0 cc. 



2. Brewer's yeast 10.0 g. 



3. Peptone (Difco) 10.0 g. 



4. K2HPO4 5.0 g. 



5. Succinic acid 2.0 g. 



6. Glucose 10.0 g. 



Preparation : 



(1) Extract 10.0 g. Brewer's yeast in 

 1000.0 cc. of water containing Difco 

 peptone 1.0%, K2HPO4 0.5%, suc- 

 cinic acid 0.2% and glucose 1.0% 

 (method not given). 



(2) Filter. 



(3) Adjust to desired pH with NaOH 

 or HCl. 



Sterilization: Method not specified. 



Use: To study H-ion influence on growth 



of B. bulgaricus and B. coli. 

 Reference: Cohen and Clark (1919 p. 421). 



694. Ickert's Yeast Extract Peptone Solution 



Constituents: 

 1. Water 1000.0 cc. 



