METHODS OF MEASURING ANTIBIOTIC ACTIVITY 71 



1 6 to 24 hours, and readings are made. The highest dilution at which 

 the test organism fails to grow is taken as the end point. Activity is ex- 

 pressed in units, using the ratio between the volume of the medium and 

 the end point of growth or the dilution at which growth is inhibited 



(948). 



The bacteriostatic spectra of a group of antibiotic substances com- 

 pared with certain chemical agents are shown in Table 8. Different bac- 

 teria show different degrees of sensitivity to the different substances. A 

 comparison with the action of phenol can result in what may become 

 known as the "bacteriostatic phenol coefficient" for each active sub- 

 stance. 



Serial Dilution Method 



Once a substance is characterized as regards its selective action upon 

 specific bacteria, its activity or concentration can be measured more ac- 

 curately by the liquid dilution or titration method. One test organism is 

 selected, usually a strain of S. aureus. Different strains may vary in their 

 action. In some cases, Streftococcus hemolyticusy B. subtilis, and others 

 have been used for measuring the activity of a substance against gram- 

 positive bacteria, and E. coli for gram-negative bacteria. Definite vol- 

 umes of the test medium are placed in test tubes and sterilized (sterility 

 is essential in this method), and various dilutions of the active sub- 

 stance are added. The dilutions can range in order of 3 : i , 2 : i, or even 

 narrower, namely in series of i .2 : i , i .5 : i , etc. The tubes are inoculated 

 with the test organism and incubated for 16 to 24 hours. In some cases 

 the medium is inoculated before it is distributed into the tubes. The 

 highest dilution of the antibiotic substance giving complete inhibition of 

 growth, as expressed by a lack of turbidity of medium, is taken as the 

 end point. Activity is expressed in units as above (804). 



The dilution method has two disadvantages (276) : first, every assay 

 takes much time j second, during chemical fractionation, the substance 

 may become contaminated with bacteria not sensitive to the active sub- 

 stances. 



One modification of the method has been adapted for measuring the 

 actjivity of penicillin. Several dilutions of the active agent are prepared 

 and 0.5 ml. portions added to 4.5 cm. quantities of liquid medium in 



