8 Frances Carter, Marion C. Woods and Miriam E. Simpson 



The sheep FSH from DEAE-ccllulosc columns, given at the same total 

 unitage, and under the same time relationships used in determining standard 

 conditions, resulted in ovulation less reliably than did the original preparation 

 containing 10% ICSH. Pilot experiments (Table 7) show the ovulatory 

 response from three FSH preparations of increasing purity, as indicated by 

 lower MED (25 /Mg down to 1.7 ^g) and decreasing percentage of ICSH 

 contamination (5%, 4% and 1.5%, respectively). The results from use of 

 these purified fractions did not equal that obtained from the original less 

 purified preparation; in fact, only about one-third to one-half of the rats 

 ovulated. 



Several possible explanations for the discrepancy between these and earlier 

 results can be offered. In the first place, the rate of absorption and excretion 

 of the more purified preparations may differ from that of cruder ones. Several 

 means for obtaining prolonged action were tried, such as multiple injections 

 and injection of FSH in solutions containing gelatin or serum albumin, but 

 no clear evidence was obtained that this was an important factor. The 

 possibility that pituitary tropic hormones other than gonadotropins may be 

 involved in induction of ovulation also must be considered. This seems 

 improbable, however, as the only important biologically active contaminant 

 of FSH preparations is ICSH. Even the starting material for all preparations, 

 the 40% ethanol extract of whole sheep pituitaries which had an MED for 

 follicle development of 0.250 mg, contained less than 0.1 % growth, thyro- 

 tropic and adrenocorticotropic hormones. It did contain a small amount of 

 lactogenic hormone (0.075 lU/mg) but lactogenic hormone has been tested 

 as an ovulatory supplement in rats (as well as monkeys) and no effect in 

 inducing ovulation has been demonstrated. Purified preparations of FSH 

 with potency of 25 /xg down to 1.7 /Mg contained less than 0.1 % of all the 

 other tropic hormones: growth, thyrotropic, adrenocorticotropic and 

 lactogenic hormones. 



The most obvious explanation of the reduced eflRcacy of more highly 

 purified FSH preparations was that the ICSH content had been depleted too 

 far. Attention was therefore turned to whether the ovulatory stimulus from 

 purified FSH was improved by addition of ICSH. Reconstitution of the 

 fraction of ICSH found in the original preparation, by addition of 10% by 

 unitage of ICSH, was the first approach. 



As the ICSH added was a highly purified product a word should be said 

 regarding its chemical fractionation and assay. These preparations were all 

 standardized in hypophysectomized immature rats by determining the 

 minimal dose (RU) which would repair the deficient interstitial cells (Table 8). 



The ICSH was prepared by methods similar to those used in purifying 

 FSH from a starting material of 40% ethanol extract from whole sheep 

 pituitary. The steps in the procedure, the potency of the preparation, and 

 the multiple of the MED which showed evidence of FSH contamination are 



