Improved synthesis of 3-keto, 4-ene-3-keto, and 4,6-diene-3-keto bile acids

Cholic and deoxycholic acids can be converted into 3-keto derivatives in 75-80% yield, by a four-step synthesis consisting of formylation, selective deformylation of the 3-formoxyl group, oxidation, then deformylation of the remaining formoxyl groups. The intermediate 3-keto formoxyl acids in this sequence were shown to be suitable starting compounds for the synthesis of 4-ene-3-keto acids, in 55-60% yield, via bromination, dehydrobromination, and deformylation. By extending the dehydrobromination reaction, the 7 alpha-formoxyl group of the intermediate 4-ene-3-keto-7 alpha,12 alpha-diformoxyl acid is also lost, hence providing a useful synthetic route to 4,6-diene-3-keto bile acids.     

Electrophysiological evidence for detection and discrimination of pheromonal bile acids by the olfactory epithelium of female sea lampreys (<Emphasis Type="Italic">Petromyzon marinus</Emphasis>)

Electro-olfactograms were used to determine sensitivity and specificity of olfactory organs of female sea lampreys (Petromyzon marinus) to four bile acids: 3-keto petromyzonol sulfate and 3-keto allocholic acid from spermiating males and petromyzonol sulfate and allocholic acid from larvae. Spermiating male bile acids are thought to function as a mating pheromone and larval bile acids as a migratory pheromone. The response threshold was 10^–12 mol l^–1 for 3-keto petromyzonol sulfate and 10^–10 mol l^–1 for the other bile acids. At concentrations above 10^–9 mol l^–1, the sulfated bile acids showed almost identical potency, as did the non-sulfated bile acids. The two sulfated bile acids were more potent than the two non-sulfated ones. In addition, 3-keto petromyzonol sulfate and water conditioned with spermiating males induced similar concentration-response curves and response thresholds. Cross-adaptation experiments demonstrated that the sulfated and non-sulfated bile acids represent different odors to the olfactory epithelium of females. Further exploration revealed that 3-keto petromyzonol sulfate represents a different odor than petromyzonol sulfate, while 3-keto allocholic acid and allocholic acid represent the same odor. Results indicate that male-specific bile acids are potent and specific stimulants to the female olfactory organ, supporting the previous hypothesis that these bile acids function as a pheromone.Abbreviations 3kACA 3-keto allocholic acid - 3kPZS 3-keto petromyzonol sulfate - ACA allocholic acid - ANOVA analysis of variance - ELISA enzyme-linked immunosorbent assay - EOG electro-olfactogram - PIR percent initial response - PZS petromyzonol sulfate - SMW spermiating male washings     

Effects of bile acids on rat hepatic microsomal type I 11β-hydroxysteroid dehydrogenase

The aim of this study was to investigate the effect of various bile acids on hepatic type I 11β-hydroxysteroid dehydrogenase (11β-HSD1) activity in vitro. The rat liver microsome fraction was prepared and 11β-HSD1 activity was assayed using cortisol and corticosterone as substrates for the enzyme reaction. The substrate and various concentrations of bile acids were added to the assay mixture. After incubation, the products were extracted and analyzed using high-performance liquid chromatography. All bile acids tested except deoxycholic acid and 7-keto bile acids inhibited the 11β-HSD1 enzyme reaction to some degree. Ursodeoxycholic acid inhibited the activity less than cholic, chenodeoxycholic, and lithocholic acids. Deoxycholic acid and 7-keto bile acids did not inhibit, but enhanced the enzyme activity. Inhibitions of dehydrogenation by corticosterone were weaker than those by cortisol. Kinetic analysis revealed that the inhibition of 11β-HSD1 was competitive. The inhibition of 11β-HSD1 by bile acids depended on the three-dimensional structural difference in the steroid rings and the presence of the 7α-hydroxy molecule of the bile acids was important for the inhibition of rat hepatic 11β-HSD1 enzyme activity. These results suggest that bile acid administration might modulate 11β-HSD1 enzyme activity.     

Identification of mono- and dihydroxy bile acids in human feces by gas-liquid chromatography and mass spectrometry

The mono- and disubstituted cholanoic acids present in human feces have been investigated. Extracts of feces were fractionated on silicic acid column and individual bile acids were isolated by preparative thin-layer chromatography. The isolated compounds were studied by gas-liquid chromatography of the methyl esters, partial trimethylsilyl ethers, oxidation products, and trifluoroacetates. The probable structures deduced were confirmed by gas chromatography-mass spectrometry and by comparisons with authentic compounds. The following derivatives of 5 Beta-cholanoic acid not previously isolated from human feces were identified: 3,12-diketo, 3-keto-12alpha-hydroxy, 3alpha,12 Beta-dihydroxy, 3 Beta,12 Beta-dihydroxy, 3-keto-7alpha-hydroxy, 3alpha-hydroxy-7-keto, 3 Beta,7alpha-dihydroxy, 3alpha,7alpha-dihydroxy, and 3alpha,7 Beta-dihydroxy. The presence of 3-keto-, 3 Beta-hydroxy-, 3alpha-hydroxy-, 3 Beta-hydroxy-12-keto-, 3alpha-hydroxy-12-keto-, 3 Beta,12alpha-dihydroxy-, and 3alpha,12alpha-dihydroxy-5 Beta-cholanoic acids was confirmed. Evidence was obtained for the presence of two bile acids having at least one hydroxyl group at a carbon atom other than C(3), C(7), or C(12).     

Influence of dietary bile acids on formation of bile acids in rat

Various taurine-conjugated bile acids were fed to rats at the 1%-level in the diet for 3 or 7 days and the effect on several hydroxylations involved in the biosynthesis and metabolism of bile acids was studied. The hydroxylations studied were all catalyzed by the microsomal fraction of liver homogenate fortified with NADPH. The 7α-hydroxylation of cholesterol was inhibited by feeding taurocholic acid, taurocheno-deoxycholic acid and taurodeoxycholic acid for 3 as well as 7 days. No marked inhibition was obtained with taurohyodeoxycholic acid or taurolithocholic acid. The 12α-hydroxylation of 7α-hydroxy-4-cholesten-3-one was inhibited after 3 as well as 7 days by all bile acids except taurohyodeoxycholic acid. With this acid a marked stimulation of 12α-hydroxylation was observed. The effects of the different bile acids on the 7α-hydroxylation of taurodeoxycholic acid were not very marked. The 6β-hydroxylation of lithocholie acid and taurochenodeoxycholic acid was stimulated by taurocholic acid and taurodeoxycholic acid. The reaction was inhibited by taurochenodeoxycholic acid, at least after 7 days. Taurohyodeoxycholic acid inhibited the 6β-hydroxylation slightly and taurolithocholic acid had no effect. The results were discussed in the light of present knowledge concerning mechanisms of regulation of formation and metabolism of bile acids and it was suggested that the mechanisms may be more complex than previously thought.     

Effect of bile acid oxazoline derivatives on microorganisms participating in 7 alpha-hydroxyl epimerization of primary bile acids

We tested bile acid oxazoline derivatives of chenodeoxycholic (CDC-OX), 7-ketolithocholic (7-KLC-OX), ursodeoxycholic (UDC-OX), and deoxycholic (DC-OX) as inhibitors of the 7-epimerization of the primary bile acids cholic acid (CA) and CDC in cultures of four species of bacteria and the human fecal flora. The organisms tested elaborate a 7 alpha- and/or 7 beta-hydroxysteroid dehydrogenase (HSDH); they were Escherichia coli (7 alpha-HSDH), Bacteroides fragilis (7 alpha-HSDH), Clostridium absonum (7 alpha- and 7 beta-HSDH) and Eubacterium aerofaciens (7 beta-HSDH). None of the oxazolines affected 7 alpha-OH oxidation of CA or CDC by E. coli or the growth of the organism. All the oxazolines (except UDC-OX) inhibited the growth of B. fragilis and its 7 alpha-HSDH. In contrast, only DC-OX blocked 7 alpha-OH epimerization of CA by C. absonum. Surprisingly, the other three oxazolines enhanced 7 alpha-OH epimerization of CA, but not that of CDC, which was inhibited (CDC-OX greater than 7-KLC-OX much greater than UDC-OX). Enzymic data suggest that CDC-OX in the presence of CA can induce a greater level of both 7 alpha- and 7 beta-HSDH than CA or CDC-OX alone, CDC-OX being more toxic in the presence of CDC. Formation of urso-bile acid from 7-keto substrates by E. aerofaciens is totally blocked by the oxazolines (except UDC-OX). Similarly, suppression of urso-bile acid formation from primary bile acids by the human fecal flora was evident with DC-OX greater than 7-KLC-OX greater than CDC-OX much greater than UDC-OX, the last being ineffective. The inhibitory activity of the oxazolines on the 7-dehydroxylation of primary bile acids by human fecal flora followed the same order.     

Xanthomonas maltophilia CBS 897.97 as a source of new 7beta- and 7alpha-hydroxysteroid dehydrogenases and cholylglycine hydrolase: improved biotransformations of bile acids

The paper reports the partial purification and characterization of the 7beta- and 7alpha-hydroxysteroid dehydrogenases (HSDH) and cholylglycine hydrolase (CGH), isolated from Xanthomonas maltophilia CBS 897.97. The activity of 7beta-HSDH and 7alpha-HSDH in the reduction of the 7-keto bile acids is determined. The affinity of 7beta-HSDH for bile acids is confirmed by the reduction, on analytical scale, to the corresponding 7beta-OH derivatives. A crude mixture of 7alpha- and 7beta-HSDH, in soluble or immobilized form, is employed in the synthesis, on preparative scale, of ursocholic and ursodeoxycholic acids starting from the corresponding 7alpha-derivatives. On the other hand, a partially purified 7beta-HSDH in a double enzyme system, where the couple formate/formate dehydrogenase allows the cofactor recycle, affords 6alpha-fluoro-3alpha, 7beta-dihydroxy-5beta-cholan-24-oic acid (6-FUDCA) by reduction of the corresponding 7-keto derivative. This compound is not obtainable by microbiological route. The efficient and mild hydrolysis of glycinates and taurinates of bile acids with CGH is also reported. Very promising results are also obtained with bile acid containing raw materials.     

Human cecal bile acids: concentration and spectrum

To obtain information on the concentration and spectrum of bile acids in human cecal content, samples were obtained from 19 persons who had died an unnatural death from causes such as trauma, homicide, suicide, or drug overdose. Bile acid concentration was measured via an enzymatic assay for 3alpha-hydroxy bile acids; bile acid classes were determined by electrospray ionization mass spectrometry and individual bile acids by gas chromatography mass spectrometry and liquid chromatography mass spectrometry. The 3alpha-hydroxy bile acid concentration (mumol bile acid/ml cecal content) was 0.4 +/- 0.2 mM (mean +/- SD); the total 3-hydroxy bile acid concentration was 0.6 +/- 0.3 mM. The aqueous concentration of bile acids (supernatant after centrifugation) was identical, indicating that most bile acids were in solution. By liquid chromatography mass spectrometry, bile acids were mostly in unconjugated form (90 +/- 9%, mean +/- SD); sulfated, nonamidated bile acids were 7 +/- 5%, and nonsulfated amidated bile acids (glycine or taurine conjugates) were 3 +/- 7%. By gas chromatography mass spectrometry, 10 bile acids were identified: deoxycholic (34 +/- 16%), lithocholic (26 +/- 10%), and ursodeoxycholic (6 +/- 9), as well as their primary bile acid precursors cholic (6 +/- 9%) and chenodeoxycholic acid (7 +/- 8%). In addition, 3beta-hydroxy derivatives of some or all of these bile acids were present and averaged 27 +/- 18% of total bile acids, indicating that 3beta-hydroxy bile acids are normal constituents of cecal content. In the human cecum, deconjugation and dehydroxylation of bile acids are nearly complete, resulting in most bile acids being in unconjugated form at submicellar and subsecretory concentrations.     

Gas-liquid chromatographic determination of human fecal bile acids

A method for the determination of total bile acids in human feces that is suitable for routine application is described and discussed. Bile acids are extracted from freeze-dried feces with acetic acid and toluene, in the presence of the internal standard 23-nordeoxycholic acid. After saponification of the extract, bile acids and the internal standard are methylated and converted by mild chromic acid oxidation into their ketonic derivatives. The resultant mixture of a few stable compounds can be separated and measured quantitatively by gas-liquid chromatography on a methylsiloxane polymer. A reference bile acid mixture including the internal standard is also taken through the entire procedure with each series of samples. It has been demonstrated that, in spite of the omission of the usual purification steps, the method is specific for bile acids.     

Preparation of [3beta-3H] labeled bile acids and bile alcohols.

[3beta-3H]-bile acids and bile alcohols may be useful for metabolic studies in man and animals because the 3-position is invulnerable to bacterial attack. A number of tritium labeled bile acids and bile alcohols were prepared by selective oxidation of the hydroxyl group at carbon-3 followed by reduction with NaBT4. In each case, the bile acids and bile alcohols epimeric at carbon-3 were resolved by analytical and preparative thin-layer chromatography and characterized by gasliquid chromatography. The average yield was 60--65% and specific activities of the final products were in the range of 7.4 x 10(7) dpm/mg.     

Identification of unconjugated bile acids in human bile

Unconjugated bile acids in the bile of healthy and diseased humans were determined qualitatively and quantitatively by means of gas-liquid chromatography and gas-liquid chromatography-mass spectrometry, after their isolation by ion-exchange chromatography. In a healthy person and three patients with cholelithiasis, unconjugated bile acids comprised 0.1-0.4% of total biliary bile acids. The bile acid composition of the unconjugated fraction was quite different from that of the glycine- or taurine-conjugate fraction, in that it contained a relatively large proportion of unusual bile acids including C23 and C27 bile acids. In two patients with cerebrotendinous xanthomatosis, C22 and C23 bile acids were the major constituents of the biliary unconjugated bile acids, and comprised about 0.8% of total bile acids; no detectable amounts of C27 bile acids were found in their bile. The analysis of biliary unconjugated bile acids may be useful for the diagnosis of metabolic diseases concerning bile acids, particularly the accumulation or disappearance of unusual bile acids.     

AKR1B7 is induced by the farnesoid X receptor and metabolizes bile acids

Although bile acids are crucial for the absorption of lipophilic nutrients in the intestine, they are cytotoxic at high concentrations and can cause liver damage and promote colorectal carcinogenesis. The farnesoid X receptor (FXR), which is activated by bile acids and abundantly expressed in enterohepatic tissues, plays a crucial role in maintaining bile acids at safe concentrations. Here, we show that FXR induces expression of Akr1b7 (aldo-keto reductase 1b7) in murine small intestine, colon, and liver by binding directly to a response element in the Akr1b7 promoter. We further show that AKR1B7 metabolizes 3-keto bile acids to 3β-hydroxy bile acids that are less toxic to cultured cells than their 3α-hydroxy precursors. These findings reveal a feed-forward, protective pathway operative in murine enterohepatic tissues wherein FXR induces AKR1B7 to detoxify bile acids.     

12 beta-dehydrogenation of bile acids by Clostridium paraputrificum, C. tertium, and C. difficile and epimerization at carbon-12 of deoxycholic acid by cocultivation with 12 alpha-dehydrogenating Eubacterium lentum

12 beta-Hydroxysteroid dehydrogenating activities were detected in 13 strains of Clostridium paraputrificum, 1 strain of C. tertium, and 1 strain of C. difficile, together with a 3 alpha- and 3 beta-hydroxysteroid dehydrogenase system in many strains. Redox reactions a C-12 of disubstituted and trisubstituted bile acids were performed unspecifically by representative strains of C. paraputrificum. 3 alpha,12 beta-, 3 beta,12 beta-Dihydroxy-, 3 alpha, 7 alpha, 12 beta-trihydroxy-, and 3-keto,12 beta-hydroxy-5 beta-cholanoic acids, so far not known as bacterial bile acid metabolites, were identified. Epimerization of the 12 alpha-hydroxyl group of deoxycholate via the 12-keto intermediate was achieved by cocultivation of C. paraputrificum and Eubacterium lentum, elaborating a 12 alpha-hydroxysteroid dehydrogenase only. In addition, epimerization at C-12 was demonstrated with mixed human fecal cultures.     

12 beta-dehydrogenation of bile acids by Clostridium paraputrificum, C. tertium, and C. difficile and epimerization at carbon-12 of deoxycholic acid by cocultivation with 12 alpha-dehydrogenating Eubacterium lentum.

12 beta-Hydroxysteroid dehydrogenating activities were detected in 13 strains of Clostridium paraputrificum, 1 strain of C. tertium, and 1 strain of C. difficile, together with a 3 alpha- and 3 beta-hydroxysteroid dehydrogenase system in many strains. Redox reactions a C-12 of disubstituted and trisubstituted bile acids were performed unspecifically by representative strains of C. paraputrificum. 3 alpha,12 beta-, 3 beta,12 beta-Dihydroxy-, 3 alpha, 7 alpha, 12 beta-trihydroxy-, and 3-keto,12 beta-hydroxy-5 beta-cholanoic acids, so far not known as bacterial bile acid metabolites, were identified. Epimerization of the 12 alpha-hydroxyl group of deoxycholate via the 12-keto intermediate was achieved by cocultivation of C. paraputrificum and Eubacterium lentum, elaborating a 12 alpha-hydroxysteroid dehydrogenase only. In addition, epimerization at C-12 was demonstrated with mixed human fecal cultures.     

Ketonic bile acids in urine of infants during the neonatal period

Ketonic bile acids have been found to be quantitatively important in urine of healthy infants during the neonatal period. In order to determine their structures, the bile acids in urine from 11 healthy infants were analyzed by gas-liquid chromatography-mass spectrometry (GLC-MS) and three samples with particularly high levels of ketonic bile acids were selected for detailed studies by ion exchange chromatography, fast atom bombardment mass spectrometry, microchemical reactions, and GLC-MS. The major ketonic bile acid was identified as 7 alpha, 12 alpha-dihydroxy-3-oxo-5 beta-chol-1-enoic acid, not previously described as a naturally occurring bile acid. The positional isomer 7 alpha, 12 alpha-dihydroxy-3-oxo-4-cholenoic acid, recently described as a major urinary bile acid in infants with severe liver diseases, was also excreted by most infants. Three acids related to cholic acid were identified: 7 alpha, 12 alpha-dihydroxy-3-oxo-, 3 alpha, 12 alpha-dihydroxy-7-oxo-, and 3 alpha, 7 alpha-dihydroxy-12-oxo-5 beta-cholanoic acids. Five bile acids having one oxo and three hydroxy groups were also present. Based on mass spectra and biological considerations two of these were tentatively given the structures 1 beta, 7 alpha, 12 alpha-trihydroxy-3-oxo- and 1 beta, 3 alpha, 12 alpha-trihydroxy-7-oxo-5 beta-cholanoic acids. Some of the others had a hydroxy group at C-4 or C-2. The levels of ketonic bile acids were higher on the third than on the first day of life, and lower after 1 month. The formation and excretion especially of 3-oxo bile acids is proposed to result from changes of the redox state in the liver in connection with birth.     

Selective reduction of oxo bile acids: synthesis of 3 beta-, 7 beta-, and 12 beta-hydroxy bile acids

Preparation of some biologically important keto bile acids is described. Advantage is taken of the preferential ketalization of 3-oxo group in bile acids over 7- and 12-oxo groups for the selective reduction of these keto groups. The method was found to be specially useful for preparation of 7 beta-, 12 alpha, and 12 beta-[3H]-3-oxo bile acids. Improved methods are also described for the preparation of epimers of naturally occurring bile acids at C-3, C-7, and C-12. 3 beta-Hydroxy bile acids (iso-bile acids) were prepared with the use of diethylazodicarboxylate/triphenylphosphine/formic acid. Iso-bile acids were obtained in excellent yields (80-95%) except during synthesis of isoursodeoxycholic acid (yield, 50%). Isoursodeoxycholic acid was, however, prepared in very good yield via epimerization of 3 alpha-hydroxyl group in 7-oxolithocholic acid followed by stereoselective reduction of 7-oxo group. A highly efficient method for the reduction of 7-oxo and 12-oxo groups was developed. Thus, 7-oxolithocholic acid and 7-oxoisolithocholic acid on reduction with potassium/tertiary amyl alcohol yielded ursodeoxycholic acid and isoursodeoxycholic acid in yields of 96% and 94%, respectively, while reduction of 7-oxodeoxycholic acid resulted in ursocholic acid in 93% yield. In a similar manner, reduction of 12-oxolithocholic acid and 12-oxochenodeoxycholic acid yielded 3 alpha, 12 beta-dihydroxy-5 beta-cholanoic acid (lagodeoxycholic acid; 92% yield) and 3 alpha, 7 alpha, 12 beta-trihydroxy-5 beta-cholanoic acid (lagocholic acid, 86% yield).     

Bile acid sulfates in serum bile acids determination.

Some bile acid sulfates were synthesized and characterized. The configuration of sulfate groups at C-3, C-7 and C-12 positions was confirmed by Nuclear Magnetic Resonance analysis. These sulfates were utilized in a study of their chemical behaviour in different analytical procedures currently used for serum bile acids determination. Procedures for bile acids extraction from serum with ethanol or Amberlite XAD-2 result in an important loss of the most polar sulfated bile acids. Complete separation of unsulfated from sulfated bile acids on Sephadex LH-20 is not achieved when deconjugation of the most polar bile acid sulfate is slow but does not produce artifacts. Enzymatic determination of bile acids gives positive response with some bile acid sulfates. The current procedures of serum bile acids determination are discussed in consideration of these results.     

Identification of new C27 and C24 bile acids in the bile of Alligator mississippiensis

Biliary bile acids of Alligator mississippiensis were analyzed by gas-liquid chromatography-mass spectrometry after fractionation by silica gel column chromatography. It was shown that the alligator bile contained 12 C27 bile acids and 8 C24 bile acids. In addition to the C27 bile acids, such as 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoic acid, 3 alpha,7 alpha,12 alpha-trihydroxy-5 alpha-cholestanoic acid, 3 alpha,7 alpha-dihydroxy-5 beta-cholestanoic acid, 3 alpha,12 alpha-dihydroxy-5 beta-cholestanoic acid, 7 alpha,12 alpha-dihydroxy-3-oxo-5 beta-cholestanoic acid, and 3 alpha,12 alpha-dihydroxy-7-oxo-5 beta-cholestanoic acid, identified previously in the bile of A. mississippiensis, 3 alpha,7 beta-dihydroxy-5 beta-cholestanoic acid, 3 alpha,7 beta,12 alpha-trihydroxy-5 beta-cholestanoic acid, 7 beta,12 alpha-dihydroxy-3-oxo-5 beta-cholestanoic acid, 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestanoic acid, 3 alpha,7 alpha,12 alpha,26-tetrahydroxy-5 beta-cholestanoic acid, and 1 beta,3 alpha,7 alpha,12 alpha-tetrahydroxy-5 beta-cholestanoic acid were newly identified. And in addition to the C24 bile acids, such as chenodeoxycholic acid, ursodeoxycholic acid, cholic acid, and allocholic acid, identified previously, deoxycholic acid, 3 alpha,7 alpha-dihydroxy-5 beta-chol-22-enoic acid, 3 alpha,7 alpha,12 alpha-trihydroxy-5 alpha-chol-22-enoic acid, and 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-chol-22-enoic acid were newly identified.     

Identification of bile acids in the serum and urine in cholestasis. Evidence for 6alpha-hydroxylation of bile acids in man.

In this qualitative study of the pattern of bile acid excretion in cholestasis, methods are described for the isolation of bile acids from large volumes of urine and plasma. The bile acids were subjected to a group separation and identified by combined gas chromatography-mass spectrometry. The techniques were developed to allow identification of the minor components of the bile acid mixture. Four bile acids that have not previously been described in human urine and plasma were detected, namely 3beta, 7alpha-dihydroxy-5beta-cholan-24-oic acid, 3alpha, 6alpha-dihydroxy-5beta-cholan-24-oic acid (hyodeoxycholic acid), 3alpha, 6alpha, 7alpha-trihydroxy-5beta-cholan-24-oic acid (hyocholic acid) and 3alpha, 7beta, 12alpha-trihydroxy-5beta-cholan-24-oic acid. In addition three C27 steroids were found; 26-hydroxycholesterol and a trihydroxy cholestane, probably 5 beta-cholestane-3alpha, 7alpha, 26-triol were found in the sulphate fraction of plasma and urine. In the plasma sample, a sulphate conjugate of 24-hydroxycholesterol was found. The presence of these compounds probably reflects the existence of further pathways for bile acid metabolism. It is not yet known whether this is a consequence of the cholestasis or whether they are also present in normal man, at much lower concentrations.     

Synthesis of N-acetylglucosaminides of unconjugated and conjugated bile acids.

3-N-Acetylglucosaminides of unconjugated, glycine- and taurine-conjugated bile acids have been synthesized. Bile acids appropriately protected were condensed with acetochloroglucosamine through the 3 alpha-hydroxyl group by means of the Koenigs-Knorr reaction using cadmium carbonate as a catalyst. Subsequent borohydride reduction and/or alkaline hydrolysis provided desired 3-N-acetylglucosaminides of unconjugated bile acids. Glycine-conjugates were obtained from N-acetylglucosaminides of unconjugated bile acids and ethyl glycinate by the carbodiimide method. The preparation of N-acetylglucosaminides of taurine-conjugates was attained by the Koenigs-Knorr reaction of bile acid p-nitrophenyl esters followed by condensation with taurine. 7-N-Acetylglucosaminides of ursodeoxycholates were prepared in a similar fashion. The convenient synthesis of 3-N-acetylglucosaminides of unconjugated bile acids is also described.