Low solubility of unconjugated bilirubin in dimethylsulfoxide – water systems: implications for pKa determinations
BackgroundAqueous pKa values of unconjugated bilirubin are important determinants of its solubility and transport. Published pKa data on an analog, mesobilirubin-XIIIα, studied by 13C-NMR in buffered solutions containing 27 and 64 vol% (C2H3)2SO because of low aqueous solubility of mesobilirubin, were extrapolated to obtain pKa values in water of 4.2 and 4.9. Previous chloroform-water partition data on bilirubin diacid led to higher estimates of its pKa, 8.12 and 8.44, and its aqueous solubility. A thermodynamic analysis, using this solubility and a published solubility in DMSO, suggested that the systems used to measure 13C-NMR shifts were highly supersaturated. This expectation was assessed by measuring the residual concentrations of bilirubin in the supernatants of comparable DMSO-buffer systems, after mild centrifugation to remove microprecipitates.ResultsExtensive sedimentation was observed from numerous systems, many of which appeared optically clear. The very low supernatant concentrations at the lowest pH values (4.1-5.9) were compatible with the above thermodynamic analysis. Extensive sedimentation and low supernatant concentrations occurred also at pH as high as 7.2.ConclusionsThe present study strongly supports the validity of the aqueous solubility of bilirubin diacid derived from partition data, and, therefore, the corresponding high pKa values. Many of the mesobilirubin systems in the 13C-NMR studies were probably supersaturated, contained microsuspensions, and were not true solutions. This, and previously documented errors in pH determinations that caused serious errors in pKa values of the many soluble reference acids and mesobilirubin, raise doubts regarding the low pKa estimates for mesobilirubin from the 13C-NMR studies.
Bile pigment pharmacokinetics and absorption in the rat: therapeutic potential for enteral administration
BACKGROUND AND PURPOSE Bilirubin and biliverdin possess antioxidant and anti‐inflammatory properties and their exogenous administration protects against the effects of inflammation and trauma in experimental models. Despite the therapeutic potential of bile pigments, little is known about their in vivo parenteral or enteral absorption after exogenous administration. This study investigated the absorption and pharmacokinetics of bile pigments after i.v., i.p. and intraduodenal (i.d.) administration in addition to their metabolism and routes of excretion.
Resonance Raman study of the chromophore-specific vibrational and electronic of bilirubin
UV–VIS absorption and resonance Raman (RR) studies have been used to elucidate the chromophore-specific electronic and vibrational spectroscopic bands of bilirubin in chloroform and in aqueous solution at pH 10 and 7.4. RR excitation profiles reveal two sets of chromophore-specific bands in each case, arising from the AB and CD pyrromethenone chromophores. A band-shape analysis of the UV–VIS absorption spectra in combination with these excitation profiles provides the absorption maxima of the individual chromophores. In the neutral species which is present in chloroform, the AB chromophore absorbs at a longer wavelength than the CD chromophore. However, in alkaline aqueous solution, where the dianion is formed, the AB chromophore absorbs at the shorter wavelength as a consequence of a large blue shift of this band. In pH 7.4 aqueous solution, the AB chromophore absorbs at the longer wavelength owing to the formation of a monoanion in which the strong internal hydrogen bonding of the lactam A group is retained, while the proton which is hydrogen bonded to the lactam D group is removed. The RR spectra support this interpretation.