1. A method is described for the extraction, purification and separation of acid-soluble phosphorus compounds from mammalian semen. [8-14C]ATP and [8-14C]AMP were used as internal recovery standards to measure the breakdown and loss of these nucleotides in the procedure. 2. Bull, ram, boar and stallion semen was separated into seminal plasma and spermatozoa and the two fractions were examined separately. The overall composition of the mixture of the phosphorus compounds extracted from the two fractions was similar for the four species. 3. Glycerylphosphorylcholine and glycerylphosphorylinositol were the two phosphorus compounds identified in extracts of seminal plasma. ATP, ADP, AMP, GTP, GDP, NAD, fructose 1,6-diphosphate and glucose 6-phosphate were identified in extracts prepared from spermatozoa.

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