The standard reaction for culture media for water analysis shall be +1.0 per cent., as determined by tests of the sterilized medium. As ordinarily prepared, broth and agar will be found to have a reaction between +0.5 and +1.0. For such media no adjustment shall be made. The reaction of media containing sugar shall be neutral to phenolphthalein. Whenever reactions other than the standard are used, it shall be so stated.
b. Sterilization.
All media and dilution water shall be sterilized in the autoclav at 15 lbs. (120° C.) for 15 minutes after the pressure reaches 15 lbs. All air must be forced out of the autoclav before the pressure is allowed to rise. As soon as possible after sterilization the media shall be removed from the autoclav and cooled rapidly. Rapid and immediate cooling of gelatin is imperative.
Media shall be sterilized in small containers, and these must not be closely packed together. No part of the medium shall be more than 2.5 cm. from the outside surface of the glass. All glassware shall be sterilized in the dry oven at 170° C. for at least 1½ hours.
c. Nutrient Broth. To Make One Liter.
1. Add 3 grams of beef extract and 5 grams of peptone to 1,000 cc. of distilled water.
2. Heat slowly on a steam bath to at least 65° C.
3. Make up lost weight and adjust the reaction to a faint pink with phenol red, or if the phenolphthalein titration is used, and the reaction is not already between +0.5 and +1, adjust to +1.
4. Cool to 25° C. and filter through filter paper until clear.
5. Distribute in test-tubes, 10 cc. to each tube.