Interactions of unconjugated bilirubin with vesicles, cyclodextrins and micelles: New modeling and the role of high pKa values

Abstract

BackgroundUnconjugated bilirubin (UCB) is an unstable substance with very low aqueous solubility. Its aqueous pKa values affect many of its interactions, particularly their pH-dependence. A companion paper shows that only our prior solvent partition studies, leading to pKa values of 8.12 and 8.44, met all essential requirements for valid pKa determinations. Other published values, generally lower, some below 5.0, were shown to be invalid. The present work was designed to derive suitable models for interpreting published data on the pH-dependent binding of UCB with four agents, mentioned below, chosen because they are not, themselves, sensitive to changes in the pH range 4-10, and the data, mainly spectrometric, were of reasonable quality.ResultsThese analyses indicated that the high pKa values, dianion dimerization constant and solubilities of UCB at various pH values, derived from our partition studies, along with literature-derived pH- and time-dependent supersaturation effects, were essential for constructing useful models that showed good qualitative, and sometimes quantitative, fits with the data. In contrast, published pKa values below 5.0 were highly incompatible with the data for all systems considered. The primary species of bound UCB in our models were: undissociated diacid for phosphatidylcholine, dianion for dodecyl maltoside micelles and cyclodextrins, and both monoanions and dianion for sodium taurocholate. The resulting binding versus pH profiles differed strikingly from each other.ConclusionsThe insights derived from these analyses should be helpful to explore and interpret UCB binding to more complex, pH-sensitive, physiological moieties, such as proteins or membranes, in order to understand its functions.

Metastable and equilibrium phase diagrams of unconjugated bilirubin IXα as functions of pH in model bile systems: Implications for pigment gallstone formation

“…For example, Cabral and colleagues (7) demonstrated by 13 C-NMR that, at saturation, cholic acid solubilized in NaTC micelles (1:9 mol/mol) increases its pK= a from 4.6 (a value obtained in H 2 O below the critical micellar concentration of cholate) to 5.3. Nonetheless, it is now fairly certain that the derived pK= a of 4.2 and 4.9 for UCB by Lightner and colleagues (27) may be realistic for a hydrophobic environment but not for physiologically relevant highly polar aqueous systems, where the pK= a values of 8.1 and 8.4 proposed by Hahm et al (23) for UCB are more likely correct (32). Both studies are in agreement with this work.…”

“…The fully undissociated form of UCB (H 2 UCB) exhibits only trace aqueous solubility at physiological pH (6,41) by virtue of the involvement of all its polar functions in six internal hydrogen bonds (2,3). The studies of Ostrow, Mukerjee, and colleagues (23,32,33,37,40,44,45) provide detailed information on the quantitative interactions of UCB with different bile salt species and concentrations, as well as UCB ionization, binding capacities, and influence of pH and pK= a values. The studies of Ostrow, Mukerjee, and colleagues (23,32,33,37,40,44,45) provide detailed information on the quantitative interactions of UCB with different bile salt species and concentrations, as well as UCB ionization, binding capacities, and influence of pH and pK= a values.…”

Fluorescence Sensor for the Quantification of Unbound Bilirubin Concentrations

“…Previous studies have reported substantially lower bilirubin solubility levels as well as a substantial dependence of solubility on pH. However, it was also proposed that bilirubin may be supersaturated after preparation but remain soluble long enough to complete the measurements (20 ). We also observed that high-concentration [0.23 mg/dL (4 mol/L)] bilirubin stocks will precipitate over the course of several days but do not show visible signs of precipitation in the first several hours after stock preparation.…”

Chiroptical Properties of Bilirubin‐Serum Albumin Binding Sites

“…In mammals, pigment behavior and functions are dependent on the form in which they appear in the body (bound, unbound, and conjugated). The relation between different BR forms present and the cytotoxic/physiological effects caused is still under debate . Moreover, at physiological or neutral pH, BR is a mixture of mono‐, dianionic, and diacidic ionic states .…”