LEAFLET II 

 PREPARATION OF MEDL/^ 



STERILIZATION 



General directions for preparation of media are hardly called for 

 here as they can be found in all bacteriological laboratory guides. In 

 the matter of sterilization, however, a few specific instructions seem 

 advisable. 



Ordinary bacteriological media are sterilized for 20 to 30 minutes 

 in an autoclave under steam pressure at 121°C (15 pounds pressure 

 after driving out all air) . In determining this temperature dependence 

 should not be laid upon a pressure gauge; the autoclave should be 

 equipped wuth a thermometer. In general, the smaller the container, 

 and the smaller the number of flasks or tubes sterilized at one time, 

 the shorter the sterilizing time can be. In the case of small batches 

 of media, 15 minutes at 15 pounds are ordinarily sufficient, a fact 

 which is worth taking into account when the media contain sub- 

 stances likely to be decomposed by heat. 



Oils are difficult to sterilize, and when they are added to media it is 

 well to sterilize them separately by dry heat (165-75° for 1 hour) 

 or by autoclaving in small quantities at 121°C. 



Fractional sterilization in flowing steam at 100° for 30-60 minutes 

 on three successive days was formerly recommended to avoid this 

 decomposition in the case of carbohydrates. Recent investigation, 

 however, tends to show that this procedure can be more harmful 

 than the higher temperature for 15 minutes; fractional sterilization, 

 therefore, is used much less than formerly. Instead it is recommended 

 that those sugars especially susceptible to the effects of heat (e. g., 

 xylose, arabinose, fructose, maltose, and under some conditions 

 sucrose and lactose) be dissolved separately and sterilized by filtra- 

 tion before adding to the rest of the medium after it has been auto- 

 claved. The Seitz filter or sintered glass filters prove suitable for 

 this purpose. Where facilities for such filtration are lacking, these 

 sugars can ordinarily be autoclaved successfully if sterilized separately 

 from the rest of the medium and in concentrated solution, employing 

 as brief heating as possible — e. g., 10 minutes at 10 pounds pressure 

 (115°C) if serological tubes are used. 



MEDIA USED IN PURE CULTURE STUDY 



It is a matter of some difficulty to decide just what media should 

 be included here. It would obviously be beyond the scope of this 



n„-3 



