Comparative studies of bile salts. Bile salts of the lamprey Petromyzon marinus L.
ABSTRACT: 1. Bile salts of Petromyzon marinus L. ammocoetes appeared to consist solely or chiefly of a crystalline substance, whose chromatographic and i.r.-spectral characteristics suggested that it was a monosulphate ester of a bile alcohol having the 3alpha,7alpha,12alpha-trihydroxy pattern of substitution in a 5alpha-steroid nucleus. 2. This substance on cleavage with dioxan-trichloroacetic acid gave petromyzonol, n.m.r. and mass-spectral examination of which suggested the structure 5alpha-cholane-3alpha,7alpha,12alpha,24-tetrol. 3. 3alpha,7alpha,12alpha-Trihydroxy-5alpha-cholanoic acid (allocholic acid) from the lizards Anolis lineatopus lineatopus Gray and Cyclura carinata Harlan (family Iguanidae) was esterified with propan-1-ol and reduced by lithium aluminium hydride to 5alpha-cholane-3alpha,7alpha,12alpha,24-tetrol, identical with petromyzonol. 4. Chromic acid oxidation of petromyzonol sulphate from lamprey bile, followed by acid hydrolysis, gave 24-hydroxy-5alpha-cholane-3,7,12-trione; hence the sulphate ester group is at C-24. 5. Petromyzonol sulphate is both primitive and unique: a study of its biogenesis might improve our understanding of evolution at the molecular level.
Project description:1. G.l.c. examination of bile alcohols prepared from the sucker Catostomus commersoni Lacépède (family Catostomidae) showed that although 5alpha-cyprinol (5alpha-cholestane-3alpha,7alpha,12alpha,26,27-pentol) was a minor constituent, the principal bile alcohol was an undescribed substance, probably present in the bile as the C-26 sulphate ester, whose i.r., n.m.r. and mass spectra agreed with the structure 5alpha-cholestane-3alpha,7alpha,12alpha,24,26-pentol. 2. M(D) studies suggest that this 5alpha-chimaerol is the 24(+), 25S enantiomer and that 5beta-chimaerol (chimaerol) from Chimaera monstrosa bile also has the 24(+), 25S configuration. These findings imply that bile alcohol biosynthesis in suckers and chimaeras includes stereospecific oxidation of cholesterol at C-26. 3. C. commersoni bile acids (present in minor amounts) probably consist largely of 3alpha,7alpha,12alpha-trihydroxy-5alpha-cholan-24-oic acid (allocholic acid). 4. 5alpha-Chimaerol sulphate and 5alpha-cyprinol sulphate are probably biochemically equivalent as bile salts, and can be considered as arising by parallel evolution.
Project description:1. The bile of germ-free domestic fowl contains taurine conjugates of 3alpha,7alpha-dihydroxy-5beta-cholan-24-oic acid (chenodeoxycholic acid), 3alpha,7alpha,12alpha-trihydroxy-5beta-cholan-24-oic acid (cholic acid) and its 5alpha-epimer (allocholic acid): that of germ-free pigs contains glycine and taurine conjugates of chenodeoxycholic acid, 3alpha,6alpha-dihydroxy-5beta-cholan-24-oic acid (hyodeoxycholic acid), 3alpha,6alpha,7alpha-trihydroxy-5beta-cholan-24-oic acid (hyocholic acid) and (probably) cholic acid. Keto acids were not found. 2. Allocholic acid and hyodeoxycholic acid are thus proved to be primary bile acids in intact animals. 3. The evolutionary and biochemical implications of these findings are briefly considered.
Project description:1. Both 25-d- and 25-l-3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoic acid were isolated from the gall-bladder bile of Crocodylus niloticus. 2. The catabolism of cholesterol to 25-d- and 25-l-3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoic acid respectively was studied by using a rat liver preparation in vitro. The results show that rat liver can metabolize cholesterol to both forms of 3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoic acid. However, a preference was noted for the formation from [4-(14)C]cholesterol of 3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoic acid (25-d), which was isolated from the incubations with a specific radioactivity about four times that of 3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoic acid (25-l). 3. The results indicate that 3alpha,7alpha,12alpha-trihydroxy-5beta-cholestanoic acid is a normal intermediate in the biosynthesis of bile acids from cholesterol in the rat.
Project description: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.
Project description:1. Arapaima gigas bile salts were hydrolysed by alkali or cleaved with dioxan-trichloroacetic acid to give cholic acid, arapaimic acid, arapaimol-A and arapaimol-B. 2. I.r., n.m.r. and mass spectroscopy and [alpha](D) measurements indicated that arapaimic acid and arapaimol-A and -B are respectively 2alpha,3alpha,7alpha,12alpha-tetrahydroxy-5beta,25in-cholestan-26-oic acid, 5beta,25R-cholestane-2beta,3alpha,7alpha,12alpha,26-pentol and 5beta-cholestane-2beta,3alpha,7alpha,12alpha,26,27-hexol. 3. Partial synthesis of 2beta,3alpha,7alpha,12alpha-tetrahydroxy-5alpha- and -5beta-cholan-24-oic acid and their spectral examination fully confirmed these conclusions. 4. A. gigas bile salts show primitive features in that they comprise alcohol sulphates and a C(27) acid; they are also specialized in showing 2beta-hydroxylation.
Project description:On the basis of i.r. and chromatographic evidence 5alpha-cholestane-3alpha,7alpha,12alpha,24xi,26-pentol(I) and 24xi,26-oxido-5alpha-cholestane-3alpha,7alpha,12alpha-triol (IV) structures have been proposed for the principal cleavage and alkaline-hydrolysis products of the bile salts from Catostomus commersoni (Anderson & Haslewood, 1969). N.m.r.- and mass-spectral analyses of these substances and the tetra- (II) and penta-acetate (III) derivatives of (I) provided supporting evidence for the structural assignments.
Project description:1. Bile salts of the sturgeons Acipenser guldenstaedti Brandt, Acipenser stellatus Pall and Huso huso L. and of the paddlefish Polyodon spathula Walbaum are shown to be closely similar, consisting mainly of taurocholate with minor amounts of tauroallocholate and the monosulphates of bile alcohols. The bile alcohols, comprising less than 10% of the bile salts, are mixtures with high proportions of substances resembling C(27) tetrols and of C(27) pentols, including 5beta-cyprinol and (probably) 5alpha-cyprinol. 2. 5beta-Cyprinol (3alpha,7alpha,12alpha,26,27-pentahydroxy-5beta-cholestane) was made from cholic acid via 3alpha,7alpha,12alpha-triacetoxy-5beta-cholan-24-ol in an overall yield of about 0.8%. 3. The chemical nature of chondrostean bile salts agrees with the systematic position of the fishes and suggests further correspondence between evolution at the morphological and molecular levels.
Project description:1. The metabolism of 4-[4-14C]androstene-3,17-dione, 4-[4-14C]pregnene-3,20-dione, 5alpha-[4-14C]androstane-3alpha,17beta-diol, [4-14C]cholesterol, 7alpha-hydroxy-4-[6beta-3H]cholesten-3-one, 5beta-[7beta-3H]cholestane-3alpha,7alpha-diol and [3H]lithocholic acid was studied in the microsomal fraction of livers from control and orotic acid-treated male rats. 2. As a result of the treatment the orotic acid-fed rats had fatty livers and subnormal concentrations of cholesterol and triglycerides in serum. 3. The 6beta- and 7alpha-hydroxylation of 4-androstene3,17-dione, and the 2alpha-, 2beta- and 18-hydroxylation of 5alpha-androstane-3alpha,17beta-diol, and the 5alpha-reduction of 4-androstene-3,17-dione and 4-pregnene-3,20-dione were decreased by 40--50% in orotic acid-fed rats. Other oxidative and reductive reactions of the steroid hormones were not significantly affected. 4. The 12alpha-hydroxylation of 7alpha-hydroxy-4-cholesten-3-one was decreased by about 50%, whereas the 7alpha-hydroxylation of cholesterol and the 26-hydroxylation of 5beta-cholestane-3alpha,7alpha-diol were not significantly decreased. The 6beta-hydroxylation of lithocholic acid was stimulated by 40%. 5. The results are discussed in relation to present knowledge of the heapatic drug-metabolizing enzymes and to the recent findings of an abnormal bile acid metabolism in liver disease.
Project description:According to current views, the second peroxisomal beta-oxidation pathway is responsible for the degradation of the side chain of bile acid intermediates. Peroxisomal multifunctional enzyme type 2 [peroxisomal multifunctional 2-enoyl-CoA hydratase/(R)-3-hydroxyacyl-CoA dehydrogenase; MFE-2] catalyses the second (hydration) and third (dehydrogenation) reactions of the pathway. Deficiency of MFE-2 leads to accumulation of very-long-chain fatty acids, 2-methyl-branched fatty acids and C(27) bile acid intermediates in plasma, but bile acid synthesis is not blocked completely. In this study we describe an alternative pathway, which allows MFE-2 deficiency to be overcome. The alternative pathway consists of alpha-methylacyl-CoA racemase and peroxisomal multifunctional enzyme type 1 [peroxisomal multifunctional 2-enoyl-CoA hydratase/(S)-3-hydroxyacyl-CoA dehydrogenase; MFE-1]. (24E)-3alpha,7alpha,12alpha-Trihydroxy-5beta-cholest-24-enoyl-CoA, the presumed physiological isomer, is hydrated by MFE-1 with the formation of (24S,25S)-3alpha,7alpha,12alpha,24-tetrahydroxy-5beta-cholestanoyl-CoA [(24S,25S)-24-OH-THCA-CoA], which after conversion by a alpha-methylacyl-CoA racemase into the (24S,25R) isomer can again be dehydrogenated by MFE-1 to 24-keto-3alpha,7alpha,12alpha-trihydroxycholestanoyl-CoA, a physiological intermediate in cholic acid synthesis. The discovery of the alternative pathway of cholesterol side-chain oxidation will improve diagnosis of peroxisomal deficiencies by identification of serum 24-OH-THCA-CoA diastereomer profiles.
Project description:1. Material containing the less polar sulphate previously noticed in hagfish bile salts gave, after dioxan-trichloroacetic acid cleavage, 16-deoxymyxinol [3beta,7alpha,-26(27)-trihydroxy-5alpha-cholestane]. 2. Anodic coupling of 3beta-hydroxy-5beta-cholanoic acid and the mixed half esters of dl-methylsuccinic acid, followed by lithium aluminium hydride reduction, yielded 3beta,26(27)-dihydroxy-5beta-cholestane. 3. 16-Deoxymyxinol, the third known bile alcohol having the 3beta-hydroxy-5alpha-hydrogen configuration, poses again the question of how the 3beta-hydroxyl group of cholesterol can be ;retained' in biosynthesis of primitive bile salts.