Characterization of p-hydroxy-hippuric acid as an inhibitor of Ca2+-ATPase in end-stage renal failure.
Characterization of p-hydroxy-hippuric acid as an inhibitor of Ca2+-ATPase in end-stage renal failure. In patients with end-stage renal failure (ESRF), disturbances of Ca2+ metabolism are common. Besides hormonal changes, inhibition of cellular Ca2+-ATPase was postulated to contribute to uremic toxicity. We purified a potent inhibitor of the Ca2+-ATPase from the ultrafiltrate of patients with ESRF by multiple steps of high-performance liquid chromatography to homogeneity, and identified the isolated inhibitor by mass spectrometric methods as p-hydroxy-hippuric acid. The enzyme used for the Ca2+-ATPase assay system was isolated from red blood cells by cross-flow filtration. The activity of the Ca2+-ATPase was measured spectrophotometrically as the difference in hydrolysis of adenosine 5'-triphosphate (ATP) in the presence and absence of Ca2+ with different concentrations of ATP and p-hydroxyhippuric acid. The Ca2+-ATPase was found to be inhibited by p-hydroxy-hippuric acid at a concentration above 11.7 micromol/L. p-Hydroxyhippuric acid inhibited the erythrocyte Ca2+-ATPase by reducing Vmax and increasing the Km value. The EC50 (log mol/L; mean +/- SEM) for p-hydroxy-hippuric acid was calculated as 4.82 +/- 0.14. In conclusion, p-hydroxy-hippuric acid may play a role in disturbed Ca2+ metabolism in end-stage renal failure.
Jankowski
J
J
Medizinische Klinik I, Universitäst-Klinik Marienhospital, Ruhr-Universität Bochum, Herne, Germany.
Tepel
M
M
Stephan
N
N
van der Giet
M
M
Breden
V
V
Zidek
W
W
Schlüter
H
H
D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors* / Q000097:blood; D016022:Case-Control Studies; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000201:enzymology; D005260:Female; D006440:Hemofiltration; D006626:Hippurates / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D007676:Kidney Failure, Chronic / Q000378:metabolism* / Q000628:therapy; D008297:Male
D004791:Enzyme Inhibitors; D006626:Hippurates; C056283:4-hydroxyhippuric acid; D000252:Calcium-Transporting ATPases