THE PREPARATION OF BOUILLON 9 



up to that amount. To this meat infusion add 1% peptone 

 (Witte's) and ^% sodium chloride. Add enough of a nor- 

 mal solution of sodium hydrate to give the liquid a faintly 

 alkaline reaction. In this work the alkalinity can be deter- 

 mined by the "use of sensitive litmus paper. (For neutralizing 

 culture media for special or research work, see Appendix I., 

 p. 126.) The infusion is then boiled in a water bath for three- 

 quarters of an hour, and allowed to cool. When cool, filter it 

 through ordinary filter paper. The filtrate should be perfectly 

 clear. The color will vary according to the amount of blood pig- 

 ment in the meat used, and according to the length of time it is 

 steamed or boiled, i.e. on the amount of material precipitated 

 out. After filtering, distribute the bouillon in tubes and flasks 

 (see above) . Stand the tubes containing the bouillon in a wire 

 basket for sterilization. SteriHze them by boiling in a closed 

 water bath or steaming in the Arnold's steam sterilizer for 30 

 minutes,^ the time to be computed from the time the water 

 boils or the temperature in the steamer reaches 99°. The flasks 

 of bouillon should be boiled or steamed for 20 minutes on each 

 of the two succeeding days (certain anaerobic bacteria may not 

 be destroyed by this treatment). When they have cooled, the 

 outside of the tubes should be carefully wiped with a moist 



1 The customary method of sterilizing culture media is to steam or boil 

 it for about 10 minutes on each of 3 consecutive days. This was 

 found very troublesome by the students, and feeling that it vfas not neces- 

 sary, a long series of test experiments was made by Mr. R. C. Reed, who 

 found that I boiling or steaming for 30 minutes gave just as good results 

 as the customary 3 boilings. As the media is not used for 2 or 3 days after 

 its steriUzation, during which time it is kept in an incubator, the method 

 is well suited to student laboratories, not for the reason that it saves time 

 in preparing the media, but it relieves the congestion in the sterilizer and 

 appreciably aids the student. When sterilized by this method the media 

 must not he inoculated for several days after its preparation. 



Media can be quickly sterilized by means of the autoclave when the 

 temperature is raised from no to 115° C. While this method is quick and 

 convenient, the high temperature seems to be detrimental to media for 

 certain pathogenic bacteria. The autoclave, however, is quite extensively 

 used. 



