INDIRECT METHODS 



131 



different coefficient of velocity; the velocity of reduction was a 

 linear function of the temperature (up to 37° C.) and, finally, 

 ceased with exhaustion of the medium. It was formerly sug- 

 gested that the reduction of methylene blue in this '' slow 

 reductase test " as it is usually termed, was due to enzymes 

 present in the intramammary milk, but it is now generally held 

 that such milk does not contain reducing substances and that 

 the reduction is due to intra and extra cellular products of bac- 

 terial origin. 



Fred ^^ in an examination of 200 samples of milk by this 

 method (adding 1 c.cm. of a 0.05 per cent solution of pure 

 methylene blue in 0.4 per cent saline to 10 c.cms. of milk and 

 holding at 40° C.) found that the time required for reduction 

 was proportional to the bacterial count. His figures are given 

 in Table LI, each group representing the average of 20 samples. 



Table LI 



Barthol ^^ found that samples containing more than 10,000,- 

 000 bacteria per c.cm. and 50 per cent of those containing 

 4-10 millions per c.cm. reduced within one hour. He concluded 

 that 10 millions per c.cm. was the lowest limit that could be 

 estimated by this method and that below this limit there is no 



I 



