THE (ESTRUS-PRODUCING HORMONE 85 



being combined. The alcoholic extract is then evaporated to 

 dryness and the residue extracted with ether; the phosphatides 

 are precipitated from the ethereal solution with acetone. The 

 acetone extract contains the active principle. Since 1915, 

 Frank and his collaborators (219-231) have reported a series 

 of observations agreeing roughly with those of Herrmann and 

 Fraenkel. These authors emphasize the presence of the oestrus- 

 producing substance in both placentae and corpora lutea. The 

 extracts were tested upon immature rabbits, and later upon 

 rats. Similar observations have been reported by Aschner 

 (41). Wintz (640), and Seaborn and Champy (553) have used 

 untreated liquor folliculi for injection purposes with positive 

 results. Fellner (193-8) has amplified his original observations 

 from time to time. 



Recent work on the cestrus-producing hormone. The application 

 of the vaginal smear technique by Allen and Doisy to the testing 

 of ovarian extracts made possible a great advance in the 

 whole study of the internal secretions of the ovary. Their extracts 

 were made in much the same way as those of previous workers, 

 but various elaborations were introduced. Allen and Doisy 

 originally used liquor folliculi from cow or pig as their starting 

 material. The liquor folliculi was obtained by aspiration with 

 a hypodermic needle from the larger follicles. The fluid was 

 mixed with twice its volume of 95 °o alcohol and allowed to 

 stand until the precipitated proteins had coagulated. The 

 mixture was then filtered, and the residue again extracted with 

 alcohol to remove the fraction of the hormone adsorbed by the 

 proteins. The two alcoholic filtrates were then mixed and 

 evaporated down to a watery residue, which was then extracted 

 with ether. The ether-soluble fraction was extracted with 

 acetone in order to leave behind the phosphatides. The acetone- 

 soluble fraction was re-extracted with 95*^0 alcohol, by which 

 means much of the inert fat was eliminated. Further puri- 

 fication was obtained by precipitating cholesterol with 

 methyl alcohol at 0° C. A similar process with certain modifica- 

 tions in the early stages was used for the extraction of solid 

 tissue. From their initial investigations, Allen and Doisy con- 

 cluded that an extract could be obtained which would produce 

 all the normal extra-ovarian symptoms of oestrus in the ovariec- 



