Enzymatic oxidation of unconjugated bilirubin to assess its interactions with taurocholate.
The rate of peroxidation of unconjugated bilirubin (UCB), catalyzed by horseradish peroxidase (HRP), has been employed by Jacobsen (1969. FEBS Lett. 5: 112-114) to assess the fraction of unbound UCB in the presence of serum albumin. We used this method to examine the interactions of UCB with taurocholate (TC) at pH 8.2, assuming solubilization of UCB by TC is due to pigment binding and/or to partitioning into the micelle, thus rendering UCB unavailable for peroxidation. Inhibition of UCB peroxidation conformed with predictions based on these assumptions and demonstrated significant interaction of UCB with both monomeric and micellar TC. Although significant inhibition of UCB peroxidation was seen with TC monomer, inhibition was even greater with TC micelles. In contrast, pyrogallol, another substrate of HRP, acted very differently in the presence of TC. Inhibition of pyrogallol peroxidation by TC was much less than with UCB and occurred primarily with monomeric TC, with little further inhibition in the micellar range. The results of this study suggest that at taurocholate concentrations above 50 mM, similar to the physiologic bile salt concentrations in human bile, at least 99% of UCB is bound to bile salt, dramatically decreasing the concentration of unbound UCB. Since bile salts also bind Ca2+, they play a dual role in protection against the precipitation of calcium bilirubinate from bile. Therefore, bile salts are a major factor in the prevention of the formation and growth of pigment gallstones.
D001663:Bilirubin / Q000378:metabolism*; D006735:Horseradish Peroxidase / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008823:Micelles; D008956:Models, Chemical; D010084:Oxidation-Reduction; D011748:Pyrogallol / Q000378:metabolism; D013656:Taurocholic Acid / Q000378:metabolism* / Q000494:pharmacology
D008823:Micelles; D011748:Pyrogallol; D013656:Taurocholic Acid; D006735:Horseradish Peroxidase; D001663:Bilirubin