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Measurement of the bioequivalence of formulations of chenodiol, a bile acid which is used for gallstone dissolution, is difficult because its high first-pass clearance results in low plasma levels after ingestion of usual dosages. To solve this problem, a new method was developed to determine the bioequivalence of several chenodiol formulations. The method included the following steps: (1) isolation of all bile acids from serum by adsorption to a hydrophobic resin, (2) elution of bile acids from the resin by methanol, (3) separation of the unconjugated bile acid fraction by an ion-exchange procedure, and (4) bioluminescence measurement of the unconjugated 7α-hydroxy bile acids using Sepharose beads containing co-immobilized 7α-hydroxysteroid dehydrogenase, diaphorase, and luciferase. The isolation method gave complete recovery, and the bioluminescence procedure was simple, rapid, and sensitive. The peak level of systemic chenodiol occurred 1 to 2 h following oral ingestion and ranged from 4 to 8 μ.M. This method appears superior to previously reported methods for determining the bioequivalence of chenodiol preparations. In principle, the method is suitable for measurement of the bioequivalence of other bile acids provided the appropriate hydroxysteroid dehydrogenase is available.

Hyperabsorption of dietary oxalate is a major factor in the pathogenesis of enteric hyperoxaluria and a frequent complication of inflammatory bowel disease and ileojejunal bypass surgery. Successful treatment requires reduction in oxalate intake or inhibition of absorption of dietary oxalate by oral ingestion of oxalate binding agents. To identify such agents, oxalate binding by anion-exchange resins, gums, and aluminum hydroxide was measured under conditions that simulated those present in the intestinal lumen. Of the agents tested, those that bound oxalate best were colestipol and aluminum hydroxide. Strongly basic anion-exchange resins readily bound oxalate only in the absence of chloride. These results suggest that colestipol and aluminum hydroxide administration might reduce dietary oxalate absorption in patients with enteric hyperoxaluria.

Duodenal bile obtained from patients with gallstones who were acutely infused with chenodeoxycholic acid, ursodeoxycholic acid, or cholic acid were examined for the propensity toward the formation of a liquid crystalline mesomorphic phase when cholesterol gallstones were incubated in these bile acids. Bile taken from patients infused with ursodeoxycholic acid was found to be enriched in ursodeoxycholic acid; mesophase formation was detected in these samples but not in bile from patients infused with chenodeoxycholic acid or cholic acid.

A physical model approach was utilized to investigate cholesterol monohydrate dissolution kinetics in simulated bile. The static pellet method and the Berthoud theory were employed to assess the contributions of the diffusion-convection mass transfer resistance and those of the interfacial resistance to the overall kinetics. For almost all situations studied, the interfacial resistance was the dominant rate-determining factor. The effects of four bile acids and their concentrations, the bile acid-lecithin ratio, and the added electrolytes and their concentrations on the interfacial resistance were examined. The results were correlated with those obtained with human bile samples, and the indications were that the kinetics of cholesterol dissolution in bile may be explainable on the basis of the principal bile acids, lecithin, and the electrolytes in the bile.

The properties of two polymeric dyes were examined to determine their suitability as nonabsorbable reference markers for intestinal perfusion studies. The first dye was a violet anthroquinone dye, R-478, named polyporphyre; the second was a blue anthroquinone dye, R-411, named polycyanine. The dyes could be determined spectrophotometrically over a concentration range of 0.03-5 μ M (1–200 μ g/mL) and had no appreciable osmotic effect. They were light stable and their absorption spectra did not change in solution over a pH range of 1–11. The dyes had a molecular weight of ~40,000 by gel permeation chromatography; they did not appear to have hydrophobic binding sites because they did not bind added dihydroxy bile acid or fatty acid anions. Determination of the dyes was not influenced by the presence of bilirubin when the latter was present at concentrations reported to occur in small intestinal content during digestion. The dyes did not interfere with an endpoint enzymatic determination of bile acids. The dyes caused little quenching of 3 H- or 14 C-radioactivity in a toluene-based liquid scintillation cocktail. Dyes were easily removed from aqueous solution by passage over a large-pore anion exchange column or by a BaCI 2 :ZnSO 4 protein precipitation procedure. When tested in a rat jejunal perfusion model, changes in dye concentration evoked by hypotonic or hypertonic perfusates were identical to those of polyethylene glycol 4000, a commonly used reference marker. The dyes did not influence the rate of glucose absorption and did not bind to mucus. These studies suggest that these two polymeric dyes have ideal physicochemical and biological properties for use as nonabsorbable reference markers in intestinal perfusion studies in animals. The dyes appear to be an attractive alternative to polyethylene glycol 4000.

Various compounds that might function as cholesterol gallstone dissolution accelerators were studied. The dissolution rates of cholesterol monohydrate pellets in synthetic bile (116 mM sodium cholate-32 mM lecithin) containing the agent at various concentration levels were determined. In the absence of any dissolution rate accelerator, the dissolution kinetics for cholesterol previously were found to be interfacial resistance controlled, and the rates were around 20 times less than the diffusion-controlled rates in the present experiments. Primary, secondary, and tertiary amines and quaternary ammonium compounds were effective accelerators. When the alkyl chain lengths were long enough and/or when the agent concentrations were high enough, the dissolution rates generally approached diffusion-controlled rates. Steroidal amines generally had good activity. Anionic and nonionic surfactants had little or negative activity.

Cholesterol monohydrate dissolution kinetics in human gallbladder bile were studied to determine the magnitudes of the in vitro dissolution rates, the rate resistances in human gallbladder bile, and the extent that the interfacial resistance is the rate-determining factor. Dissolution rate studies also were conducted using human duodenal bile and animal bile for comparison. The dissolution rate resistance, R, ranged from 10(4) sec/cm for chicken bile to 10(4)-10(6) sec/cm for human bile. Interfacial resistance was the rate-determining factor for essentially all results. Where chemical composition data were obtained, the R values for the human bile samples were consistent with predictions made from the simulated bile studies. In two human gallbladder specimens having low bile acid-lecithin molar ratios (i.e., 2.9 and 2.3), very high R values of 1.9 X 10(5) and 4.1 X 10(5) sec/cm were found. These values were in good agreement with the findings in the simulated bile studies and suggest that stone dissolution in patients with low bile acid-lecithin ratios may proceed very slowly, even when the bile is highly undersaturated with respect to cholesterol.

The 37 degree cholesterol solubilities in over 50 solvents, including the homologous n-alkanols through dodecanol and homologous ethyl carboxylates through the undecanoate, and the 37 degree beta-sitosterol solubilities in the n-alkanols through decanol are reported. Additionally, solubility data for cholesterol at 7, 17, and 27 degrees in the alcohol series were obtained. These measurements allowed the calculation of heats of solution for cholesterol in the alkanols, which range from 7.5 kcal for methanol to 4.3 kcal for decanol and which tend to decrease, although irregularly, with increasing alkanol chain length. A solubility maximum in all of these series for both solutes was observed between a chain length of six and seven. A surprisingly irregular, odd-even alternating solubility pattern was noted for cholesterol in the alkanols at all four temperatures. Experimental evidence indicated that this pattern was due to solvent-induced crystalline changes, presumably solvate formation, in each alkanol solvent through C10. Overall, the solubility studies screened solvents for their utility in dissolving cholesterol and, thus, cholesterol gallstones. To these ends, some limited dissolution experiments were performed, which indicated that the solution rate is directly related to the measured solubility in organic solvents. The dissolution behavior is thus different from micellar bile salt solutions, in which a significant interfacial barrier controls kinetics.