GEORGE F. REDDISH 



303 



methods also require a greater amount of time and material than the results warrant. 

 This led to a phenol-coefficient test using B. typhosus as a test organism in which the 

 best features of the Rideal- Walker and Hygienic Laboratory tests were retained and 

 their worst features eliminated. In this way a very simple test was obtained which 

 gave consistent results with a minimum amount of time and material.' 



The methods recommended by the writer^ and as approved by the American Pub- 

 lic Health Association are given below: 



B. TYPHOSUS 

 Medium 



Liebig beef extract or Lemco 5 g. 



Peptonum siccum (Armour) 10 g- 



Sodium chloride 5 g. 



Distilled water i ,000 cc. 



Place the ingredients in about 1,000 cc. of distilled water. Boil for 30 minutes and then 

 filter. Add enough sodium hydroxide to bring the broth to a pH of 6.8 with brom thymol 

 blue. Add enough water to make the total volume 1,000 cc. Run into test tubes, placing 

 10 cc. in each tube. Sterilize at 15-pound pressure for 30 minutes. 



Stock culture. — The stock culture is transferred at least once a month on beef-extract 

 agar slants pH 7.0-7.5, and kept at room temperature in tubes plugged with cotton. The 

 agar should be made as follows: 



Percentage 



Liebig beef extract 0.5 



Armour's peptone i . o 



Sodium chloride 0.5 



Agar 1.5 



It is transferred in the foregoing broth, incubating at 37° C, on three consecutive days before 

 using in the test. Fresh broth culture is started from agar-slant stock culture each month. 



Organism. — -The organism used in the test is a 22-to 26-hour culture of B. typhosus 

 grown in the specified medium at 36°-38° C. This culture must be capable of resisting dilu- 

 tions of phenol ranging from 1-90 to i-ioo for at least 5 minutes at 20° C, and must be killed 

 in 15 minutes by dilutions of phenol ranging up to and including 1-90. 



In order to obtain the desired results, it is necessary to make daily transfers; however, 

 the Sunday transfer may be omitted and the Tuesday culture will still be satisfactory. 



Temperature. — The mixture of culture and diluted disinfectant must be held at 20° C. 

 during the test. 



Phenol. — The phenol used must meet all the requirements of the United States Pharma- 

 copoeia and, in addition, must have a congealing point (point of constant temperature on 

 cooling) not below 40° C. If the congealing point of the available phenol is slightly below 

 40° C. the phenol may be distilled and the middle third of the distillate collected. If the con- 

 gealing point of this portion is not below 40° C, it is suitable for use. 



Prepare a 5 per cent (by weight) stock solution of the phenol and standardize it by titra- 



' This method, as well as the procedures which are given below, was approved by the Standard 

 Methods Committee of the Laboratory Section of the American Public Health Association at the 

 fifty-fifth annual meeting, 1926. 



^Reddish, George F.: op. oil., 17, 320. 1927. 



