The bile acid composition of gastric contents from neonates with high intestinal obstruction.

The study was designed to identify 'atypical' bile acids in gastric contents from three neonates with high intestinal obstruction on the basis that this was likely to represent a rich source of primary bile acids. Cholic acid was the major component, and related 'atypical' bile acids included its C-3 and C-7 oxidation products, its 3 beta-epimer and 2 beta- and 6 alpha-hydroxylation products. Allocholic acid was the only 5 alpha-cholanic acid derivative identified. 7 alpha, 12 alpha-Dihydroxy-3-oxochol-4-en-24-oic acid was found in all three specimens and might be an intermediate in a biosynthetic pathway from cholesterol to cholic acid in which side-chain oxidation precedes at least some of the nuclear changes. Side-chain-hydroxylated derivatives of trihydroxycoprostanic acid were also detected and these may represent intermediates in biosynthetic pathways from cholesterol to cholic acid via 5 beta-cholestan-3 alpha, 7 alpha, 12 alpha-triol. The most abundant bile acid of this type was (25 epsilon)-3 alpha, 7 alpha, 12 alpha, 25-tetrahydroxy-5 beta-cholestan-26-oic acid, which suggested that C-25 hydroxylation may be an important step in the shortening of the C8 side chain of the cholestane triol to the C5 side chain of cholic acid in the neonatal period. Bile acids lacking a substituent at C-12 included chenodeoxycholic acid, its C-3 and C-7 oxidation products, its 3 beta-epimer and its 6 alpha-hydroxylation product (hyocholic acid).     

C26-analogs of naturally occurring bile alcohols-II. Preparation of 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,23-tetrols (23R and 23S) and 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,26-tetrols (25R and 25S) by a hydroboration procedure

A convenient procedure for the synthesis of 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,23-tetrol (23R and 23S) and 24-nor-5 beta-cholestane-3 alpha,7 alpha,12alpha,26-tetrol (25R and 25S) starting from 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,25-tetrol was developed. Dehydration of 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha, 25-tetrol with glacial acetic acid and acetic anhydride yielded a mixture of 24-nor-5 beta-cholest-23-ene-3 alpha,7 alpha,12 alpha-triol and the corresponding delta 25 compound. Hydroboration and oxidation of the mixture of unsaturated nor-triols resulted in the formation of 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,23-tetrols (23R and 23S) and 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,26-tetrols (25R and 25S). In addition, smaller amounts of 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,22 xi-tetrol and 24-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol were also obtained. The C26 bile alcohols epimeric at C-23 and C-25 were resolved by analytical and preparative TLC and characterized by gas-liquid chromatography and mass spectrometry. Provisional assignment of the configurations of the C-23 and C-25 hydroxyl groups were made on the basis of molecular rotation differences. These C26 alcohols will be used to test the stereospecificity of the hepatic enzymes that promote oxidation of the cholesterol side chain.     

Bile acids. LXXIII. Synthesis of analogs of 7 alpha-hydroxy-4-cholesten-3-one as substrates for hepatic steroid 12 alpha-hydroxylase

Analogs of 7 alpha-hydroxy-4-cholesten-3-one were prepared to ascertain structural features necessary for maximal activity of hepatic microsomal 12 alpha-steroid hydroxylase. Methyl 3 alpha,7 alpha-dihydroxy-5 beta-cholane-24-carboxylate derived from chenodeoxycholic acid was oxidized at C-3 with silver carbonate/Celite. The product was hydrolyzed and dehydrogenated with SeO2 to provide 3-oxo-7 alpha-hydroxy-4-cholene-24-carboxylic acid. 5 beta-Cholestane-3 alpha,7 alpha,25-triol and 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25-tetrol were similarly oxidized at C-3 and dehydrogenated to provide 7 alpha,25-dihydroxy-4-cholesten-3-one and 7 alpha,12 alpha,25-trihydroxy-4-cholesten-3-one, respectively. The products were characterized by thin-layer and gas chromatography, ultraviolet, infrared, proton resonance and mass spectrometry.     

Synthesis of four diastereoisomers at carbons 24 and 25 of 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid, intermediates of bile acid biosynthesis

The synthesis of four stereoisomers at C-24 and C-25 of 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid is described. Pyridium chlorochromate oxidation of 3 alpha,7 alpha,12 alpha-triacetoxy-5 beta-cholan-24-ol (II) prepared from cholic acid (I) afforded 3 alpha,7 alpha,12 alpha-triacetoxy-5 beta-cholan-24-al (III) which was converted to a mixture of the four stereoisomers (IV-VII) by a Reformatsky reaction with ethyl DL-alpha-bromopropionate followed by alkaline hydrolysis. Separation of these isomers (IV-VII) was achieved by silica gel column chromatography, and subsequent reversed-phase partition column chromatography. The configurations at C-24 were elucidated by conversion of each isomer into (24R)- or (24S)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24-tetrol (XII or XI) by Kolbe electric coupling, the C-24 configurations of which were determined by modified Horeau's method and 13C-nuclear magnetic resonance spectroscopy. The stereochemistries at C-25 were deduced by comparison of IV-VII with the products of the hydroboration followed by oxidation with alkaline hydrogen peroxide of (24E)-3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-en-26-oic acid (XIII).     

Synthesis of C-22, C-23-3H-labeled 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestane

This report describes an efficient synthesis of C-22, C-23-(3)H-labeled 3alpha,7alpha,12alpha-trihydroxy-5beta-cholestane. - Somanathan, R., and S. Krisans. Synthesis of C-22, C-23-(3)H-labeled 3alpha,7alpha,12alpha-trihydroxy-5beta-cholestane.     

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.     

Metabolism of bile alcohols in the perfused rabbit liver.

The mechanism and sequence of side chain hydroxylation of cholesterol in bile acid synthesis was studied in the isolated perfused rabbit liver. A comparison was made between the importance of 26- and 25-hydroxylation in cholic acid biosynthesis in the rabbit. The formation of [G-3H]cholic acid was observed when the liver was perfused with 5beta-[G-3H]cholestane-3alpha, 7alpha-diol, 5beta-[G-3H]cholestane-3alpha, 7alpha-12alpha-triol, and 5beta-[G-3H]cholestane-3alpha, 7alpha, 26-triol. No [G-3H]chenodeoxycholic acid was detected in the bile. These findings indicate that potential precursors of chenodeoxycholic acid were hydroxylated at position 12alpha either subsequent to or before hydroxylation of the cholesterol side chain. In addition, no other intermediates (tetrahydroxy or pentahydroxy bile alcohols) were found in the bile when these compounds were perfused in the liver. Bile acid precursors were detected in bile when the rabbit liver was perfused with 5beta-[24-14C]cholestane-3alpha, 7alpha, 25-triol. The 5beta-[24-14C]cholestane-3alpha, 7alpha, 25-triol was hydroxylated in the liver at the 12alpha position to yield the corresponding 5beta-cholestane-3alpha, 7alpha, 12alpha, 25-tetrol. The tetrol was further metabolized to a series of pentols (5beta-cholestane-3alpha, 7alpha, 12alpha, 22, 25-pentol; 5beta-cholestane-3alpha, 7alpha, 12alpha, 23, 25-pentol; 5beta-cholestane-3alpha, 7alpha, 12alpha, 24, 25-pentol; and 5beta-cholestane-3alpha, 7alpha, 12alpha, 25, 26-pentol). The major bile acid obtained from the perfusion of the 5beta-cholestane-3alpha, 7alpha, 25-triol was cholic acid. The experiments indicated that in the rabbit liver 12alpha-hydroxylation can occur after hydroxylation of the cholesterol side chain at either C-25 (5 beta-cholestane-3alpha, 7alpha, 25-triol) or C-26 (5beta-cholestane-3alpha, 7alpha-26-triol). Apparently, the rabbit can form cholic acid via the classical 26-hydroxylation pathway as well as via 25-hydroxylated intermediates.     

Bile acids of snakes of the subfamily Viperinae and the biosynthesis of C-23-hydroxylated bile acids in liver homogenate fractions from the adder, Vipera berus (Linn.).

1. Analysis of bile salts of four snakes of the subfamily Viperinae showed that their bile acids consisted mainly of C-23-hydroxylated bile acids. 2. Incubations of 14C-labelled sodium cholate (3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholan-24-oate) and deoxycholate (3 alpha, 12 alpha-dihydroxy-5 beta-cholan-24-oate) with whole and fractionated adder liver homogenates were carried out in the presence of molecular oxygen and NADPH or an NADPH-generating system. The formation of C-23-hydroxylated bile acids, namely bitocholic acid (3 alpha, 12 alpha, 23xi-trihydroxy-5 beta-cholan-24-oic acid) and 3 alpha, 7 alpha, 12 alpha, 23 xi-tetrahydroxy-cholanic acid (3 alpha, 7 alpha, 12 alpha, 23 xi-tetrahydroxy-5 beta-cholan-24-oic acid), was observed mainly in the microsomal fraction and partly in the mitochondrial fraction. 3. Biosynthetic pathways of C-23-hydroxylated bile acids are discussed.     

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.     

Absolute configuration at C-24 of 5 beta-ranol, a principal bile alcohol of the bullfrog

The stereochemistry of the hydroxyl group at C-24 in 5 beta-ranol (27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,26-pentol) a principal bile alcohol of the bullfrog which is structurally related to the major human urinary bile alcohol, 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentol, is described. Two isomers (IIIa and IIIb) at C-24 of 27-nor-5 beta-cholest-25-ene-3 alpha,7 alpha,12 alpha, 24-tetrol were synthesized from cholic acid (I) by the conversion to 3 alpha, 7 alpha, 12 alpha-triacetoxy-5 beta-cholan-24-al (II) followed by a Grignard reaction with vinylmagnesium bromide. The absolute configurations at C-24 of the unsaturated tetrols (IIIa and IIIb) were elucidated as S and R, respectively, by means of the difference of the reactivity to Sharpless oxidation, a stereoselective epoxidation. Catalytic hydrogenation of each delta 25-tetrol (IIIa or IIIb) gave (24R)- or (24S)-27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha, 24-tetrol (IVa or IVb). The configurations at C-24 of two isomeric 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-27-nor-5 beta-cholestan-26-oic acids (Va and Vb) were determined as S and R, respectively, by means of their conversion into the saturated tetrols (IVa and IVb) of known absolute configurations by a Kolbe electrolytic coupling with acetic acid. The lithium aluminum hydride reduction product of the 24R-acid (Vb) was identical with the naturally occurring 5 beta-ranol, hence 5 beta-ranol has the 24R configuration.     

Structural and biosynthetic studies of a principal bile alcohol, 27-nor-5beta-cholestane-3alpha,7alpha,12alpha,24,25-pentol, in human urine

The stereochemistry at C-24 and C-25 of 27-nor-5beta-cholestane-3alpha,7alpha,12alpha,24 ,25-pentol, a principal bile alcohol in human urine, and its biosynthesis are studied. Four stereoisomers of the C(26)-24,25-pentols were synthesized by reduction with LiAlH(4) of the corresponding epoxides prepared from (24S)- or (24R)-27-nor-5beta-cholest-25-ene-3alpha, 7alpha,12alpha,24-tetrol. The stereochemistries at C-25 were deduced by comparison of the C(26)-24,25-pentols with the oxidation products of (24Z)-27-nor-5beta-cholest-24-ene-3alpha,7alpha, 12alpha-triol with osmium tetraoxide. On the basis of this assignment, the principal bile alcohol excreted into human and rat urine was determined to be (24S,25R)-27-nor-5beta-cholestane-3alpha,7alpha, 12alpha,24,25-pentol, accompanied by a lesser amount of (24R, 25R)-isomer. To elucidate the biosynthesis of the C(26)-24,25-pentol, a putative intermediate, 3alpha,7alpha, 12alpha-trihydroxy-27-nor-5beta-cholestan-24-one, derived from 3alpha,7alpha, 12alpha-trihydroxy-24-oxo-5beta-cholestanoic acid by decarboxylation during the side-chain oxidation of 3alpha,7alpha, 12alpha-trihydroxy-5beta-cholestanoic acid, was incubated with rat liver homogenates. The 24-oxo-bile alcohol could be efficiently reduced to yield mainly (24R)-27-nor-5beta-cholestane-3alpha,7alpha, 12alpha,24-tetrol. If a 25R-hydroxylation of the latter steroid occurs, it should lead to formation of (24S,25R)-C(26)-24,25-pentol. Now it has appeared that a major bile alcohol excreted into human urine is (24S,25R)-27-nor-5beta-cholestane-3alpha,7alpha, 12alpha, 24, 25-pentol, which might be derived from 3alpha,7alpha, 12alpha-trihydroxy-27-nor-5beta-cholestan-24-one via (24R)-27-nor-5beta-cholestane-3alpha, 7alpha,12alpha,24-tetrol.     

Identification of 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24 xi, 25 xi,26-hexol and partial characterization of the bile alcohol profile in urine

The nature of the bile alcohols present in urine of an infant with neonatal cholestasis has been investigated. Urine was extracted with Sep-Pak C18 cartridges and a glucuronide fraction was isolated by ion exchange chromatography on Lipidex-DEAP. Following enzymatic hydrolysis and purification on Lipidex-DEAP, the bile alcohols were isolated by high performance liquid chromatography. Fourteen compounds were studied by a combination of microchemical reactions and capillary column gas-liquid chromatography-mass spectrometry. Both C26 and C27 bile alcohols were present. Among the former, three additional isomers of the previously identified 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24 xi,25 xi-pentol were detected. A new C26 bile alcohol, 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24 xi,25 xi,26 -hexol, was identified, and a 27-norcholestane-pentolone with hydroxyl groups at C-24 and C-25 and a keto group in the ring system was partially characterized. The C27 bile alcohols consisted of cholestanepentols, -tetrolones, and -pentolones. 5 beta-Cholestane-3 alpha,7 alpha,12 alpha,25,26-pentol (5 beta-bufol), one of its isomers and an isomer of cholestane-3,7,12,24,26-pentol were present. Two cholestanetetrolones and two cholestanepentolones having the keto group in the ring system were partially characterized. The hydroxyl groups in the side chain of the tetrolones were at C-24,26 and C-25,26, respectively, whereas the pentolones had hydroxyl groups at C-24,25 and C-25,26, respectively. The excretion of glucuronidated bile alcohols in urine is suggested to reflect an alternative metabolism of intermediates in the normal biosynthesis of bile acids.     

Synthesis of 3 alpha,6 beta,7 alpha,12 beta- and 3 alpha,6 beta,7 beta,12 beta-tetrahydroxy-5 beta-cholanoic acids.

Chemical synthesis of 3 alpha,6 beta,7 alpha,12 beta- and 3 alpha,6 beta,7 beta,12 beta-tetrahydroxy-5 beta-cholan-24-oic acids is described. 3 alpha,12 beta-Dihydroxy-5 beta-chol-6-en-24-oic acid used as the starting material in the synthesis was prepared via oxidation of 3 alpha,12 alpha-dihydroxy-5 beta-chol-6-en-24-oic acid 3-hemisuccinate at C-12 followed by reduction with potassium/tertiary amyl alcohol. alpha-Epoxidation of the ester diacetate of 3 alpha,12 beta-dihydroxy-5 beta-chol-6-en-24-oic acid with m-chloroperbenzoic acid followed by cleavage of the epoxide with acetic acid and alkaline hydrolysis yielded 3 alpha,6 beta,7 alpha,12 beta-tetrahydroxy-5 beta-cholan-24-oic acid (overall yield 25%). N-Methylmorpholine-N-oxide-catalyzed osmium tetroxide oxidation of the ester diacetate of 3 alpha,12 beta-dihydroxy-5 beta-chol-6-en-24-oic acid followed by alkaline hydrolysis yielded 3 alpha,6 beta,7 beta,12 beta-tetrahydroxy-5 beta-cholan-24-oic acid (overall yield 33%). The structures of the synthesized bile acids were confirmed from their proto nuclear magnetic resonance and mass spectral fragmentation patterns.     

Potential bile acid metabolites. XVIII. Synthesis of stereoisomeric 3,6,12 alpha-trihydroxy-5 beta-cholanoic acids.

Two new 6-hydroxylated bile acids, 3 beta, 6 alpha, 12 alpha- and 3 beta, 6 beta, 12 alpha-trihydroxy-5 beta-cholanoic acids, were synthesized from deoxycholic acid. In addition, their C-3 epimers, 3 alpha, 6 alpha, 12 alpha- and 3 alpha, 6 beta, 12 alpha-trihydroxy acids, were prepared by a new route. The principal reactions used were 1) 6 beta-hydroxylation of 3-methoxy-3,5-dienes with m-chloroperbenzoic acid in aqueous dioxane; 2) catalytic hydrogenation of the resulting 6 beta-hydroxy-3-oxo-4-enes to the 6 beta-hydroxy-3-oxo-5 beta compounds with palladium on calcium carbonate catalyst in ethanol; and 3) stereoselective reduction of appropriate 3-oxo derivatives with potassium tri-sec-butylborohydride and tert-butylamine-borane complex. The thin-layer chromatographic, gas-liquid chromatographic, and high performance liquid chromatographic mobilities, and 1H- and 13C-nuclear magnetic resonance spectroscopic data of the four stereoisomers are presented. With this work all the 6-hydroxylated derivatives of lithocholic, deoxycholic, chenodeoxycholic, ursodeoxycholic, and cholic acids in the 5 beta series are now known and have been synthesized.     

Specificity of bile salt sulfatase activity from Clostridium sp. strains S1.

Clostridium sp. strain S1, an unnamed bile acid-desulfating strain from rat intestinal microflora (S.M. Huijghebaert, J. A. Mertens, and H. J. Eyssen, Appl. Environ. Microbiol. 43:185-192, 1982), was examined for its ability to desulfate different bile acid sulfates and steroid sulfates in growing cultures. Clostridium sp. strain S1 desulfated the 3 alpha-monosulfates of chenodeoxycholic, deoxycholic, and cholic acid, but not their 7 alpha- or 12 alpha-monosulfates. Among the 3-sulfates of the 5 alpha- and 5 beta-bile acids, only bile acid-3-sulfates with an equatorial sulfate group were desulfated. Hence, Clostridium sp. strain S1 desulfated the 3-sulfates of bile acids with a 3 alpha, 5 beta-, a 3 beta, 5 alpha- or a 3 beta, delta 5-structure. In contrast, the bile acid-3-sulfates with a 3 beta, 5 beta- or a 3 alpha, 5 alpha-structure were not desulfated. In addition, Clostridium sp. strain S1 did not hydrolyze the equatorial 3-sulfate esters of C19 and C21 steroids and cholesterol or the phenolic 3-sulfate esters of estrone and estradiol. 23-Nordeoxycholic acid with a C-23 carboxyl group was also not desulfated, in contrast to the 5 beta-bile acid 3 alpha-sulfates with a C-24 or C-26 carboxyl group. Therefore, the specificity of the sulfatase of Clostridium sp. strain S1 is related to the location of the sulfate group on the bile acid molecule, the equatorial orientation of the sulfate group, and the structure of the C-17 side chain, its carboxyl group, and chain length.     

Absolute configuration at carbon 23 of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol excreted by patients with cerebrotendinous xanthomatosis

Absolute configuration at C-23 of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,23,25-pentol, one of the bile alcohols isolated from the patients with cerebrotendinous xanthomatosis, was unequivocally determined as 23S by conversion of a key intermediate, (23S)-5 beta-cholest-25-ene-3 alpha,7 alpha,12 alpha,23-tetrol to either the bile alcohol of known absolute configuration, (23R)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,23-tetrol, or the naturally occurring 23,25-pentol.     

Configuration of the 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-enoic acid, an intermediate in the peroxisomal conversion of 3 alpha,7 alpha,12-trihydroxy-5 beta-cholestanoic acid to cholic acid in rat liver.

3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-enoic acid, formed in the peroxisomal oxidation of 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoyl-CoA by the THCA-CoA oxidase in rat liver, was isolated and purified on reverse phase HPLC. The configuration of the C-24/25 double bond was determined to be trans (E) by using 1H-NMR spectrometry.     

Stereospecific formation of (24E)-3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-en-26-oic acid and (24R,25S)-3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid from either (25R)- or (25S)-3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestan-26-oic acid by rat liver homogenate

Studies of the stereochemistry of the intermediates, 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholest-24-en-26-oic acid and 3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid, in the biosynthetic sequence between 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestan-26-oic acid and cholic acid have been undertaken. (25R)- or (25S)-3 alpha,7 alpha, 12 alpha-Trihydroxy-5 beta-cholestan-26-oic acid was incubated with rat liver homogenates. The reaction products were converted to p-bromophenacyl ester derivatives and the esters were analyzed by high-performance liquid chromatography. By comparison with authentic samples of two (24E)- and (24Z)-isomers of the alpha, beta-unsaturated acid and of four isomers at C-24 and C-25 of the beta-hydroxy acid, (24E)-3 alpha,7 alpha, 12 alpha-trihydroxy-5 beta-cholestan-26-oic acid and (24R,25S)-3 alpha,7 alpha,12 alpha,24-tetrahydroxy-5 beta-cholestan-26-oic acid were found to be formed from either (25R)- or (25S)-3 alpha,7 alpha, 12 alpha-trihydroxy-5 beta-cholestan-26-oic acid. No formation of the (24Z)-isomer of the trihydroxycholestenoic acid or the other three isomers of the tetrahydroxycholestanoic acid was detected. The findings are discussed in relation to the assumed pathway for side chain cleavage in cholic acid biosynthesis.     

Multifunctionalized alpha,beta-cyclopentenones from C-2 and C-4-ulopyranosyl compounds: a stereospecific rearrangement initiated by base

Base treatment of O-benzyl protected C-2- or C-4-ulopyranosyl compounds (4 alpha, 4 beta, and 11) by either 10% Et(3)N or 1% K(2)CO(3) in MeOH initiated a beta elimination to afford alpha,beta-unsaturated C-ulopyranosyl compounds (5 alpha, 5 beta, and 12), which further rearranged in a stereocontrolled manner to multifuctionalized alpha,beta-cyclopentenones (6 and 14) in 70-80% yield. Both C-alpha- and C-beta-2-ulosides (5 alpha and 5 beta) produced the same cyclopentenone 6, indicating that a 1,2-enolate is formed prior to the cleavage of the C-5--O bond. Because 6 is racemic, it was probably formed by the intramolecular cycloaldolization of two equally populated enantiomeric intermediates. When treated with 90% Et(3)N in MeOH, 5 alpha yielded almost exclusively 15 (isomer of 6), which was formed by a migration of the double bond in 5 alpha during the previously described rearrangement. Thus either 6 or 15 was the major product, depending on the base used.     

Synthesis and structure of 26 (or 27)-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24S,25 xi-pentol isolated from the urine and feces of a patient with sitosterolemia and xanthomatosis

The urine and feces of a patient with the rare inherited lipid storage disease, sitosterolemia and xanthomatosis, were analyzed. Substantial quantities of C26-bile alcohol, 26 (or 27)-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24S,25 xi-pentol along with 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24-tetrol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25-tetrol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24R,25-pentol, and 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25,26-pentol were found. The structure of the C26-bile alcohol was confirmed by direct comparison (gas-liquid chromatography-mass spectrometry and thin-layer chromatography) with a standard sample synthesized from cholic acid. The configurational assignment at C-24 was determined by lanthanide-induced circular dichroism Cotton effect measurements. The increased excretion of these C26- and C27-bile alcohols suggests an abnormality of bile acid biosynthesis in this disease.