CULTURE MEDIA FOR CULTIVATION OF MICROORGANISMS 



307 



phthalein is turned red (hot titra- 

 tion) to the clear light greenish 

 yellow filtrate. 



(6) Boil for one hour. 



(7) Filter off the albumin. 



(8) Readjust the reaction (pH = 7.8). 



(9) Distribute the crystal clear fluid into 

 flasks. 



Sterilization: Method not given. 



Use : Cultivation of diphtheria bacillus and 



toxin production. 

 Reference: Groer and Srnka (1918-19 



p. 334). 



995. Boyd's Hydrolyzed Meat Solution 



Constituents: 



1. Water, tap 1000.0 cc, 



2. H,S04 (strong) 100.0 cc. 



3. NaCl 5.0 g. 



4. KCl 2.0 g. 



5. Lean meat 1.0 lb. 



Preparation : 



(1) To 300.0 cc. of tap water add 100.0 cc. 

 strong H2SO4 gradually with con- 

 stant stirring. 



(2) Cut up 1 pound lean meat into small 

 pieces and add to (1) after the reac- 

 tion has subsided somewhat. 



(3) Place (2) in a suitable water bath 

 and bring quickly to a boil. Boil 

 for 3 to 4 hours with frequent 

 stirring. 



(4) After 3 hours withdraw about 10.0 cc. 

 and test for absence of proteoses 

 with nitric acid. If negative with- 

 draw another 5 or 10.0 cc. and carry 

 out Biuret reaction. Whether hy- 

 drolysis is to be continued or not 

 depends upon the color obtained. 



(5) When the color is satisfactory with- 

 draw the flask from the boiling water 

 and add calcium hydrate (quicklime, 

 CaO, that has been e.xposed to the 

 free air for some time is best) until 

 a large precipitate of CaC04 is 

 formed. Continue adding lime until 

 the precipitate occupies about 1 of 

 the fluid in the flask. 



(6) Filter thru lint and wash thru the 

 lint with a little tap water. Con- 

 tinue the neutralization of the fil- 

 trate obtained with more lime, best 

 carried out in 3 stages so as to avoid 

 an excess of precipitate at any one 

 time. 



(7) Neutralize the acid completely and 

 make the medium faintly alkaline to 

 phenolphthalein. 



(8) Boil (7) for 15 minutes. 



(9) Filter hot thru filter paper. 



(10) Dilute to 1000.0 cc. by the addition 

 of water. 



(11) Dissolve 3 and 4 in (10). 



(12) Make the desired final adjustment of 

 the reaction. 



Sterilization: Sterilize in the autoclave. 

 Use: General culture medium. 

 Variants : 



(a) The author suggested the steriliza- 

 tion of solution (9) above by auto- 

 claving and storing until ready for 

 use. When desired continue as in 

 steps (10), (11) and (12) as indicated. 



(b) Norris prepared the medium as 

 follows : 



(1) Add 500.0 g. lean meat to 40.0 cc. of 

 strong H2SO4. 



(2) Pound together in a mortar. 



(3) Mix thoroly and digest in a water 

 bath for 4 days at 70°C. 



(4) Test for proteose with nitric acid 

 and then the Biuret test. 



The remainder of the preparation is 

 identical with Boyd's medium, step 

 (5) thru (12). 

 References: Boyd (1917-18 p. 410), Norris 

 (1918-19 p. 176). 



996. Harvey's Hydrolyzed Meat Solution 



Constituents: 



1. Water. 



2. Meat, goat, sheep or ox. 

 Preparation : 



(1) Mince finely fat-free goat, sheep or 

 ox flesh. 



(2) Add to it 80.0 cc. strong commercial 

 hydrochloric acid of specific gravity 

 1.16, per kilogramme. 



(3) Mix thoroly. 



(4) Keep in water bath 4 days at 70°C. 



(5) Bring the volume up to 1000.0 cc. 

 for each kilogramme of minced meat 

 used. 



(6) Mix 1 part (5) with 2 parts boiling 

 water. 



(7) Add 75.0 cc. 40.0% sodium hydroxide 

 for each kilogramme of minced meat 

 used. 



(8) Make the reaction faintly alkaline to 

 litmus. 



