RAPID ASSOCIATION OF UNCONJUGATED BILIRUBIN WITH AMORPHOUS CALCIUM-PHOSPHATE

The association of unconjugated bilirubin (UCB) with amorphous calcium phosphate was studied in vitro. To this end UCB, solubilized in diffe rent micellar bile salt solutions, was incubated with freshly prepared calcium phosphate precipitate. It was demonstrated that amorphous cal cium phosphate (ACP) rapidly binds and precipitates UCB in a dose-depe ndent way. The results indicate that binding of UCB to ACP is specific : binding to barium phosphate was negligible and addition of low amoun ts of Mg2+ before formation of the calcium phosphate precipitate (Ca:M g = 5:1) inhibited binding by 80%. Free Ca2+ stimulated binding, where as free phosphate ions inhibited binding of UCB in taurocholate soluti ons and to a lesser extent in glycocholate solutions. The apparent aff inity of UCB for amorphous calcium phosphate was different in the vari ous bile salt solutions. Binding of UCB decreased at pH > 8.5 in tauro cholate solutions, but not in glycocholate solutions where binding of UCB was constant from pH 7.5-10.5. We propose a model in which UCB dir ectly binds to amorphous calcium phosphate in the presence of bile sal ts that weakly interact with ACP, like taurocholate. In the presence o f bile salts that strongly interact with ACP, such as glycochenodeoxyc holate, binding of UCB may also occur via the bile salt. In conditions of unconjugated hyperbilirubinemia, such as the Crigler-Najjar syndro me, neonatal jaundice, and in the Gunn rat, considerable amounts of UC B diffuse across the intestinal mucosa. Binding of UCB to calcium phos phate in the intestine may stimulate its excretion and thereby constit ute a relevant mechanism of excretion.