PROPERTIES OF THE PURIFIED HYPOTHALAMIC-PITUITARY NA K-ATPASE INHIBITOR/

Previously we described the isolation and final purification of an end ogenous sodium-pump inhibitor from the CNS, mainly from bovine hypotha lamus and pituitary. The purification protocol consisted of lipophilic chromatography, followed by lipid extraction, and semi-preparative an d analytical reverse-phase high-pressure liquid chromatography. The bi oassays used were in vitro Na+/K+-ATPase inhibition, and H-3-ouabain d isplacement from its specific binding site in the enzyme structure, as well as inhibition of Rb-86 uptake from human red blood cells. We hav e obtained, from both tissues, a low-molecular-weight, nonpeptidic, no nlipidic substance that elutes as a single peak highly pure according to criteria of coincidence of its spectra properties. When rechromatog raphed in two different chromatographic systems, the same homogeneous peak is obtained suggesting complete purity. This pure substance can b e isolated from other bovine tissues as well as from human plasma and human placenta. It shows a very distinctive fluorescence spectrum and it behaves as a potent inhibitor of the Ca2+ pump of synaptosomal plas ma membrane.