Research Article| Volume 33, ISSUE 5, P617-626, May 1992

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Bile acid solubility and precipitation in vitro and in vivo: the role of conjugation, pH, and Ca2+ ions.

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      The principles governing the in vitro solubility of the common natural conjugated and unconjugated bile acids and salts in relation to pH, micelle formation, and Ca2+ concentration are considered from a theoretical standpoint and then correlated first with experimental observations on model systems and second with the formation of precipitates containing bile acids in health and disease. In vitro, taurine-conjugated bile acids are soluble at strongly acidic pH; glycine-conjugated bile acids are poorly soluble at moderately acidic pH; and many of the common, natural unconjugated bile acids are insoluble at neutral pH. For both glycine-conjugated and unconjugated bile acids, solubility rises exponentially, with increasing pH, until the concentration of the anion reaches the critical micellization concentration (CMC) when micelle formation occurs and solubility becomes practically unlimited. In vivo, in health, conjugated bile acids are present in micellar form in the biliary and intestinal tract. Unconjugated bile acids formed in the large intestine remain at low monomeric concentrations because of the acidic pH of the proximal colon, binding to bacteria, and absorption across the intestinal mucosa. In diseases in which proximal small intestinal content is abnormally acidic, precipitation of glycine-conjugated bile acids (in protonated form) occurs. Increased bacterial formation of unconjugated bile acids occurs with stasis in the biliary tract and small intestine; in the intestine, unconjugated bile acids precipitate in the protonated form. If the precipitates aggregate, an enterolith may be formed. In vitro, the calcium salts of taurine conjugates are highly water soluble, whereas the calcium salts of glycine conjugates and unconjugated bile acids possess limited aqueous solubility that is strongly influenced by bile acid structure. Precipitation occurs extremely slowly from supersaturated solutions of glycine-conjugated bile acids because of metastability, whereas super-saturated solutions of unconjugated bile acids rapidly form precipitates of the calcium salt. In systems containing Ca2+ ions and unconjugated bile acids, pH is important, since it is the key determinant of the anion concentration. For bile acids with relatively soluble calcium salts (or with a low CMC), the concentration of the anion will reach the CMC and micelles will form, thus precluding formation of the insoluble calcium salt. For bile acids, with relatively insoluble calcium salts (or with a high CMC), the effect of increasing pH is to cause the anion to reach the solubility product of the calcium salt before reaching the CMC so that precipitation of the calcium salt occurs instead of micelle formation.(ABSTRACT TRUNCATED AT 400 WORDS)


        • Hofmann A.F.
        • Mysels K.J.
        Bile salts as biological surfactants.
        Colloids Surf. 1988; 30: 145-173
        • Hofmann A.F.
        Enterohepatic circulation of bile acids.
        in: Schultz S.G. Handbook of Physiology. The Gastrointestinal System. American Physiological Society, Bethesda1989: 567-596
        • Lipkin M.
        • Newmark H.
        Effect of added dietary calcium on colonic epithelial-cell proliferation in subjects at high risk for familial colonic cancer.
        N. Engl. J. Med. 1985; 313: 1381-1384
        • van der Meer R.
        • Welberg J.W.M.
        • Kuipers F.
        • Kleibeuker J.H.
        • Mulder N.H.
        • Termont D.S.M.L.
        • Vonk R.J.
        • de Vries H.T.
        • de Vries E.G.E.
        Effects of supplemental dietary calcium on the intestinal association of calcium, phosphate, and bile acids.
        Gastroenterology. 1990; 99: 1653-1659
        • Gu J-J.
        • Hofmann A.F.
        • Ton-Nu H-T.
        • Schteingart C.D.
        • Mysels K.J.
        Solubility of calcium salts of unconjugated and conjugated natural bile acids.
        J. Lipid Res. 1992; 33 (000-000)
        • Jones C
        • Hofmarm A.F.
        • Mysels K.J.
        • Roda A.
        The effect of calcium and sodium ion concentration on the properties of dilute aqueous solutions of glycine-conjugated bile salts.
        J. Colloid Interface Sci. 1986; 114: 452-470
        • Roda A.
        • Fini A.
        Effect of nuclear hydroxy sub-stituents on aqueous solubility and acidic strength of bile acids.
        Hepatology. 1984; 4: 72S-76S
        • Roda A.
        • Festi D.
        • Armanino C.
        • Rizzoli R.
        • Simoni P.
        • Minutello A.
        • Roda E.
        Methodological and clinical aspects of bile acid analysis in biological fluids.
        Prog. Clin. Bwchem. Med. 1989; 8: 129-173
        • Fini A.
        • Roda A.
        Chemical properties of bile acids. IV. Acidity constants of glycine-conjugated bile acids.
        J. Lipid Res. 1987; 28: 755-759
        • Shiftman M.L.
        • Sugerman H.J.
        • Moore E.W.
        Human gallbladder mucosal function. Effect of concentration and acidification of bile on cholesterol and calcium solubility.
        Gastroenterology. 1990; 99: 1452-1459
        • Evans D.F
        • Pye G.
        • Bramley R.
        • Clark A.G.
        • Dyson T.J.
        • Hardcastle J.D.
        Measurement of gastrointestinal pH profiles in normal ambulant human subjects.
        Gut. 1988; 29: 1035-1041
        • Fallingborg J.
        • Christensen L.A.
        • Ingeman-Neilsen M.
        • Jacobsen B.A.
        • Abildgaard K.
        • Rasmussen H.H.
        pH-Profile and regional transit times of the normal gut measured by radiotelemetry device.
        Aliment. Pharmacol. Ther. 1989; 3: 605-614
        • Poley J.R.
        • Hofmann A.F.
        Role of fat maldigestion in pathogenesis of steatorrhea in ileal resection. Fat digestion after two sequential test meals with and without cholestyramine.
        Gastroenterology. 1976; 71: 38-44
        • Matoba N.
        • Une M.
        • Hoshita T.
        Identification of unconjugated bile acids in human bile.
        J. Lipid Res. 1986; 27: 1154-1162
        • Mallory A.
        • Kern F.
        • Smith J.
        • Savage D.
        Patterns of bile acids and microflora in the human small intestine. I. Bile acids.
        Gastroenterology. 1973; 64: 26-33
        • Northfield T.C
        • McColl I.
        Postprandial concentrations of free and conjugated bile acids down the length of the normal human small intestine.
        Gut. 1973; 14: 513-518
        • Go V.L.W.
        • Poley J.R.
        • Hofmann A.F.
        • Summerskill W.H.J.
        Disturbances of fat digestion induced by acid jejunal pH due to gastric hypersecretion in man.
        Gastroenterology. 1970; 58: 638-664
        • Zentler-Munro P.L.
        • Fine D.R.
        • Batten J.C.
        • Northfield T.C.
        Effect of cimetidine on enzyme inactiva-tion, bile acid precipitation, and lipid solubilisation in pancreatic steatorrhoea due to cystic fibrosis.
        Gut. 1985; 26: 892-901
        • Dutta S.K.
        • Hubbard V.S.
        • Appier M.
        Critical examination of a pH-sensitive enteric-coated pancreatic enzyme preparation in treatment of exocrine pancreatic insufficiency secondary to cystic fibrosis.
        Dig. Dis. Sci. 1988; 33: 1237-1244
        • Setchell K.D.R.
        • Street J.M.
        • Sjövall J.
        Fecal bile acids.
        in: Setchell K.D.R. Kritchevsky D. Nair P.P. The Bile Acids. Methods and Applications. Plenum Press, New York1988: 441-570
        • Newmark H.L.
        • Lupton J.R.
        Determinants and consequences of colonic luminal pH — implications for colon cancer.
        Nutr. Cancer. 1990; 14: 161-173
        • Mcjunkin B.
        • Fromm H.
        • Sarva R.P.
        • Amin P.
        Factors in the mechanism of diarrhea in bile acid malabsorption: fecal pH — a key determinant.
        Gastroenterology. 1981; 80: 1454-1464
        • Hofmann A.F
        • Poley J.R.
        Role of bile acid malabsorption in pathogenesis of diarrhea and steatorrhea in patients with ileal resection. I. Response to cholestyramine or replacement of dietary long chain triglyceride by medium chain triglyceride.
        Gastroenterology. 1972; 62: 918-934
        • Aldini R.
        • Roda A.
        • Festi D.
        • Sama C.
        • Mazzella G.
        • Bazzoli F.
        • Morselli A.M.
        • Roda E.
        • Barbara L.
        Bile acid malabsorption and bile acid diarrhea in intestinal resection.
        Dig. Dis. Sci. 1982; 27: 495-502
        • Tabaqchali S.
        The pathophysiological role of small intestinal bacterial flora.
        Scand. J. Gastroenterol. 1970; 6: 139-163
        • Atwell J.D.
        • Pollock A.J.
        Intestinal calculi.
        Br. J. Surg. 1960; 47: 367-374
        • Bewes P.C
        • Haslewood G.A.D.
        • Roxburgh R.A.
        Bile acid enteroliths and jejunal diverticulosis.
        Br. J. Surg. 1966; 53: 709-711
        • Cabral D.J.
        • Small D.M.
        Physical chemistry of bile.
        in: Schultz S.G. Forte J.G. Rauner B.B. Handbook of Physiology. The Gastrointestinal System. American Physiological Society, Bethesda1989: 621-662
        • Carey M.C.
        Physico-chemical properties of bile acids and their salts.
        in: Danielsson H. Sjovall J. Sterols and Bile Acids. Elsevier Science Publishers B.V., Amsterdam1985: 345-403
        • Mukerjee P.
        • Moroi Y.
        • Murata M.
        • Yang A.Y.S.
        Bile salts as atypical surfactants and solubilizers.
        Hepatology. 1984; 4: 61S-65S
        • Mysels K.J.
        • Mukerjee P.
        Reporting experimental data dealing with critical micellization concentrations (c.m.c.'s) of aqueous surfactant systems.
        Pure Appl. Chem. 1979; 51: 1083-1089
        • Roda A.
        • Hofmann A.F.
        • Mysels K.J.
        The influence of bile salt structure on self-association in aqueous solutions.
        J. Biol. Chem. 1983; 258: 6362-6370
        • Hofmann A.F.
        The function of bile salts in fat absorption: the solvent properties of dilute micellar solutions of conjugated bile salts.
        Biochem. J. 1963; 89: 57-68
        • Shinoda K.
        • Nakagawa T.
        • Tamamushi B.
        • Isemure T.
        Colloidal Surfactants, Some Physical Chemical Properties. Academic Press, New York1963
        • Pearson J.T
        • Lawrence A.S.C.
        Behaviour of calcium ions in micellar sodium dodecyl sulphate solution containing solubilized polar organic compounds.
        Trans. Faraday Soc. 1967; 63: 488-494
        • Krafft F.
        • Wiglow H.
        Über das Verhalten der Fettsauren Alkalien und der Seifen in Gegenwart von Wasser. III. Die Seifen also Krystalloide. IV. Die Seifen als Cholloide.
        Berichte Dtsch. Chem. Gesellsch. 1985; 28: 2566-2573
        • Hay D.W.
        • Carey M.C.
        Chemical species of lipids in bile.
        Hepatology. 1990; 12: 6S-16S
        • Hofmann A.F.
        • Borgstrom B.
        The intraluminal phase of fat digestion in man: the lipid content of the micellar and oil phases of intestinal content obtained during fat digestion and absorption.
        J. Clin. Invest. 1964; 43: 247-257
        • Mansbach C.M.
        • Cohen R.S.
        • LefF P.B.
        Isolation and properties of the mixed lipid micelles present in intestinal content during fat digestion in man.
        J. Clin. Invest. 1975; 56: 781-791
        • Hofmann A.F.
        Fat digestion: the interaction of lipid digestion products with micellar bile acid solutions.
        in: Rommel K. Goebell H. Lipid Absorption: Biochemical and Clinical Aspects. MTP Press, Lancaster1976: 3-18
        • Duane W.C.
        The intermicellar bile salt concentration in equilibrium with the mixed micelles of human bile.
        Biochim. Biophys. Ada. 1975; 398: 275-286
        • Lindheimer M.
        • Montet J.C.
        • Bontemps R.
        • Rouviewe J.
        • Brun B.
        Self-diffusion study of bile salt-monoolein micelles. Determination of the intermicellar bile salt concentration.
        J. Chim. Phys. Phys. -Chim. Biol. 1983; 80: 315-323
        • Duane W.C.
        Taurocholate- and taurochenodeoxy-cholate-lecithin micelles.
        Biochem. Biophys. Res. Commun. 1977; 74: 223-229
        • Higuchi W.I.
        • Arakawa M.
        • Lee P.H.
        • Noro S.
        Simple micelle-mixed micelle coexistence equilibria for the taurocholate-, taurochenodeoxycholate-, and taurourso-deoxycholate-lecithin systems.
        J. Colloid Interface Sci. 1987; 119: 30-37
        • Hofmann A.F.
        Pathogenesis of cholesterol gallstones. In Proceedings of the Symposium on Current Concepts in the Management of Cholelithiasis.
        J. Clin. Gastroenterol. 1988; 10: S1-S11
        • Akiyoshi T
        • Nakayama F.
        Bile acid composition in brown pigment stones.
        Dig. Dis. Sci. 1990; 35: 27-32
        • Hofmann A.F.
        • Mosbach E.H.
        Identification of allodeoxycholic acid as the major component of gallstones induced in the rabbit by 5α-cholestan-3β-ol.
        J. Biol. Chem. 1964; 239: 2813-2821
        • Cohen B.I.
        • Ayyad N.
        • Mosbach E.H.
        • McSherry C.K.
        • Matoba N.
        • Hofmann A.F.
        • Ton-Nu H-T.
        • Peng Y.
        • Schteingart C.D.
        • Stenger R.J.
        Replacement of cholesterol gallstones by murideoxycholyl taurine gallstones in prairie dogs fed murideoxycholic acid.
        Hepatology. 1991; 14: 158-168
        • Zaki F.G.
        • Carey Jr., J.B.
        • Hoffbauer F.W.
        • Nokolo C.
        Biliary reaction and choledocholithiasis induced in the rat by lithocholic acid.
        J. Lab. Clin. Med. 1967; 69: 737-748
        • Palmer R.H.
        • Hruban Z.
        Production of bile duct hyperplasia and gallstones by lithocholic acid.
        J. Clin. Invest. 1966; 45: 1255-1267
        • Mai M.L.
        • Emmett M.
        • Sheikh M.S.
        • Santa Ana C.A.
        • Schiller L.R.
        • Fordtran J.S.
        Calcium acetate, an effective phosphorus binder in patients with renal failure.
        Kidney Int. 1989; 36: 690-695
        • Saunders D.
        • Sillery J.
        • Chapman R.
        Effect of calcium carbonate and aluminum hydroxide on human intestinal function.
        Dig. Dis. Sci. 1988; 33: 409-413
        • Van der Meer R.
        • Vries H.T.
        Differential binding of glycine- and taurine-conjugated bile acids to insoluble calcium phosphate.
        Biochem. J. 1985; 229: 265-268