6 



GENERAL BACTERIOLOGY 



SPECIAL DIRECTIONS. All glassware prepared in 1. is to be 

 sterilized for one hour at 150 C. The small pipettes should be 

 placed in brass tubes, provided for the purpose, and also sterilized. 



EXERCISE 4. PREPARATION OF BOUILLON. 



GENERAL DIRECTIONS. Any one of the three methods (A, B 

 or C) may be used. They are arranged in order of preference, but 

 method C is the most convenient, and hence most used. 



A. 



a. From 500 grams (1% Ibs.) of lean 

 beef remove the fat and connective tis- 

 sue and mince (or use Hamburg steak). 



b. Add 1 liter of distilled water and 

 after shaking thoroughly set in ice 

 chest for 12 to 24 hours. 



c. Squeeze through a cloth and add 

 enough distilled water to filtrate to 

 make 1 liter and place in vessel to cook. 



C. 



a. Weigh out 

 three grams of 

 beef extract 

 (such as Lie- 

 big's). 



6. Add 1 liter 

 of distilled wa- 

 ter. 



c. Place in 

 vessel for cook- 

 ing. 



B. 

 a. Ditto. 



b. Add 1 liter of 

 distilled water. 



c. Place in ves- 

 sel for cooking, 

 then cook for y s 

 hour at about 70 

 C., filter through 

 paper and make 

 up to 1 liter. 



d. Add to any of the above solutions: \% (10 gms.) peptone 

 (Witte) and %% (5 gms.) common salt (NaCl), then weigh solu- 

 tion, with vessel, so that the water which is subsequently driven 

 off in cooking can be accurately replaced. 



Cooking may be done either in a flask which is heated in a water- 

 bath or sterilizer, double-walled boiler, or rice-cooker. In case a rice- 

 cooker is used a 50% solution of calcium chloride should be placed 

 in the outer vessel instead of water as by this means the contents 

 of the inner vessel can be brought to a rapid ebullition, something 

 impossible by the use of water alone. 



e. Heat, not above 60 C., until ingredients are in solution, 

 then restore the water lost by evaporation. 



/. Neutralize. This is a very important step and calls for great 

 care. Of the following methods, A is more accurate and should 

 be employed for special or research work. For ordinary routine 

 work B may be employed. 



