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  • Title: Identification of two cardiac glycosides as Na(+)-pump inhibitors in rat urine and diet.
    Author: Tamura M, Harris TM, Phillips D, Blair IA, Wang YF, Hellerqvist CG, Lam SK, Inagami T.
    Journal: J Biol Chem; 1994 Apr 22; 269(16):11972-9. PubMed ID: 8163500.
    Abstract:
    Endogenous Na(+)-pump specific inhibitors are present in the plasma, urine, and tissues of humans and animals. To date, the source of these inhibitors has not been rigorously defined. In the present study, large amounts of several Na(+)-pump specific inhibitors have been demonstrated to exist in the urine of rats raised on a regular chow diet and tap water. All of the inhibitor levels have been found to increase 1.5-8-fold by the surgical preparation of reduced renal mass (RRM) and one-kidney, one-clip (IK, IC) hypertension. These urinary inhibitors, however, except for the ouabain-like inhibitor which eluted from a high performance liquid chromatography C18 column at the same retention time as [3H]ouabain, disappeared within a week after switching the diet from regular diet (number 5001, PMI Feeds, Inc.) to pure synthetic diet (number 5755). The urinary level of the ouabain-like inhibitor decreased to only one-half of the level in the control rat raised on a regular diet. Two of these inhibitors were purified from both urine and diet by a combination of Amberlite XAD-2 adsorption chromatography, reverse phase low pressure liquid chromatography, and several high performance liquid chromatographies. Reverse phase high performance liquid chromatography, liquid secondary ion and gas-liquid mass spectrometries, and proton nuclear magnetic resonance spectroscopy identified these inhibitors as a stereoisomer of convalloside, probably neoconvalloside, and a mono-rhamnoside of periplogenin or its stereoisomer. These cardiac glycosides exhibited inhibitory potencies comparable to ouabain against ouabain-displacement from Na+,K(+)-ATPase and against 86Rb uptake into human erythrocytes, and they also exhibited cross-reactivity to anti-ouabain antibodies and anti-digoxin antibodies. These results clearly demonstrate that the principal source of most of the inhibitors in rat urine is the diet. The results suggest that the ouabain-like inhibitor may be derived from an endogenous origin.
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