Abstract
We described 1 a substance found in cultures of veal brain broth medium during the growth of H. pertussis. Subsequently, 2 further experiments with this material were reported. The present experiments were done (1) to ascertain the specific element in the filtrate, and (2) to obtain the sticky material that is produced in these cultures in more concentrated form.
After varied experimentation, I found that 4 fractions could be obtained in the following manner: Fraction I, termed the acid precipitation fraction. The filtrate obtained as previously described was brought up to pH 4.0 with acetic acid and then left in the icebox overnight. The bulky dark brown precipitate which formed was centrifuged off and was found to be slightly soluble in water or physiological saline. The remaining filtrate was adjusted to pH 8.0 with 25% NaOH and then back to pH 4.0 with acetic acid and put in the icebox overnight. The small precipitate formed was added to the previous one mentioned. Usually all the acid fraction came down after 2 such procedures, but occasionally a small amount could be obtained by a third precipitation. The combined precipitates formed the acid fraction of the filtrate.
Fraction II, termed the alkaline alcohol precipitate. The filtrate remaining after the previous fractionation was brought up to pH 8.0 with 25% NaOH; 2 volumes of 95% alcohol were added and the solution was put in the icebox over night. A heavy precipitate was obtained which was soapy white in appearance and easily soluble in water or physiological saline. The same procedure was repeated once or twice, each time the pH of the remaining filtrate being brought up to 8.0, and 2 volumes of alcohol added.
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