title: Aqueous dissociation constants of bile pigments and sparingly soluble carboxylic acids by 13C NMR in aqueous dimethyl sulfoxide: effects of hydrogen bonding.
abstract: pKas for the acid dissociation of the carboxyl groups of bilirubin in water have been reported recently to be 8.1-8.4, or higher. These high values were attributed to intramolecular hydrogen bonding. They have led to suggestions that monoanions of bilirubin predominate at physiologic pH and are the species transported most readily into hepatocytes by carriers. Such high aqueous pKas are inconsistent with recent 13C nuclear magnetic resonance (NMR) measurements on mesobilirubin XIII alpha, done on aqueous solutions containing dimethyl sulfoxide. To investigate whether the presence of dimethyl sulfoxide leads to unreliable values when using 13C NMR spectroscopy to determine pKas of carboxylic acids that can undergo intramolecular hydrogen bonding, we measured the pKas of 13C-labeled fumaric, maleic, and phthalic acids in solutions containing up to 27 vol% dimethyl sulfoxide. In addition, we used 13C NMR to estimate the pKas of 2,2'-methylenebis[5-carbomethoxy-4-methylpyrrole-3-[1-13C] propanoic acid], a model for the two central rings of bilirubin. Our results show that 13C NMR of aqueous dimethyl sulfoxide solutions can be used with confidence to measure pKas of intramolecularly hydrogen-bonded carboxylic acids. They support our previous estimates for the pKas of bilirubin and confirm that intramolecular hydrogen bonding has little effect on the acidity of bilirubins in water. Together with previous studies and chemical arguments they strongly suggest that reported aqueous pKas of > 8, or even > 6, for the carboxyl groups of bilirubin are incorrect and that arguments used to rationalize them are questionable.
authors.affiliation: Department de Quimica Organica, Universitat de Barcelona, Catalunya, Spain.
authors.forename: F R
authors.initials: FR
authors.lastname: Trull
authors.forename: S
authors.initials: S
authors.lastname: Boiadjiev
authors.forename: D A
authors.initials: DA
authors.lastname: Lightner
authors.forename: A F
authors.initials: AF
authors.lastname: McDonagh
mesh_terms: D001654:Bile Pigments / Q000737:chemistry*; D001663:Bilirubin / Q000737:chemistry; D001664:Biliverdine / Q000737:chemistry; D002247:Carbon Isotopes; D002264:Carboxylic Acids / Q000737:chemistry*; D004121:Dimethyl Sulfoxide / Q000737:chemistry*; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D009682:Magnetic Resonance Spectroscopy; D009994:Osmolar Concentration
chemical_list: D001654:Bile Pigments; D002247:Carbon Isotopes; D002264:Carboxylic Acids; D001664:Biliverdine; D001663:Bilirubin; D004121:Dimethyl Sulfoxide