A RAPID AND SIMPLE INDOL TEST

It is well known that tryptophane in an inorganic solution fur-nishes an excellent medium for the demonstration of indol pro-duction by bacteria. Zipfel (1) in 1912 found that indol could be demonstrated by this method at the end of twenty-four hours, giving the best reaction with p-dimethyl-amido-benzaldehyde. The test is made by adding to the culture to be tested one cubic centimeter of a solution consisting of p-dimethyl-amido-benzal-dehyde, 4 parts; absolute alcohol, 380 parts; and concentrated hydrochloric acid, 80 parts. The expense of preparing tryptophane and the great difficulty in obtaining it at all in the United States at the present time are drawbacks to the introduction of the tryptophane method. As a substitute, I have found that hydrolyzed casein can be used. Cow casein contains about 1.5 per cent tryptophane, which may be obtained in its amino-acid form by hydrolyzing the casein. The method is as follows: 10 grams of casein are hydrolyzed by 200 cc. of 10 per cent sulphuric acid, the mixture being kept on the water bath for twenty-four hours. At the end of this time, the casein is completely dissolved and the solution is a dark brown. Next, the solution is neutralized by the addition of saturated barium hydrate, thus precipitating out the sulphate. The resulting solution is then evaporated until the amino-acids crystallize. Half of the crystalline mass is dissolved in 500 cc. of Zipfel's inorganic solution consisting of Asparagin and Ammo-nium lactate, 5 grams each; Potassium acid phosphate, 2 grams; Magnesium sulphate, 0.2 grams; and distilled water, 1000 grams. The medium is tubed and sterilized. Assuming that there are 0.15 grams of tryptophane in the cow casein, tryptophane should be present in the mixture to the extent of about 0.03 per cent