Glycine Conjugates Of Bile Acids Research Articles

Absence of glycochenodeoxycholic acid (GCDCA) in human bile is an indication of cholestasis: A 1H MRS study

The utility of (1)H MR spectroscopy in detecting chronic cholestasis has been investigated. The amide proton region of the (1)H MR spectrum of human bile plays a major role in differentiating cholestatic (Ch) patterns from the normal ones. Bile obtained from normal bile ducts contains both taurine and glycine conjugates of bile acids--cholic acid (CA), chenodeoxycholic acid (CDCA), and deoxycholic acid (DCA). Absence of a glycine-conjugated bile acid glycochenodeoxycholic acid (GCDCA) has been observed in bile samples obtained from primary sclerosing cholangitis (PSC) patients. A total of 32 patients with various hepatobiliary diseases were included in the study. Twenty-one patients had PSC and 11 had normal cholangiograms. One PSC patient was excluded from the study because of a bad spectrum. Seventeen out of the 20 PSC patients showed an absence of GCDCA in their (1)H MR spectrum of bile. Six of the 11 reference patients with normal cholangiogram also showed spectra similar to those of PSC, indicating the possibility of cholestasis. DQF-COSY and TOCSY experiments performed on bile samples from PSC patients also revealed absence of phosphatidylcholine (PC) in some of the bile samples, suggesting possible damage to the cholangiocytes by the toxic bile. These observations suggest that analysis of human bile by (1)H MRS could be of value in the diagnosis of chronic Ch liver disorders.

Amidation of Salicyluric Acid and Gentisuric Acid: A Possible Role for Peptidylglycine α-Amidating Monooxygenase in the Metabolism of Aspirin

Bifunctional peptidylglycine α-amidating monooxygenase (PAM) catalyzes the copper-, ascorbate-, and O2-dependent cleavage of C-terminal glycine-extended peptides, N-acylglycines, and the bile acid glycine conjugates to the corresponding amides and glyoxylate. Two known metabolites of aspirin, salicyluric acid and gentisuric acid, are also substrates for PAM, leading to the formation of salicylamide and gentisamide. The time course for O2 consumption and glyoxylate production indicates that salicylurate amidation is a two-step reaction. Salicylurate is first converted to N-salicyl-α-hydroxyglycine, which is ultimately dealkylated to salicylamide and glyoxylate. The enzymatically generated salicylamide and N-salicyl-α-hydroxyglycine were characterized by mass spectrometry and two-dimensional 1H–13C heteronuclear multiple quantum coherence NMR.

The Enzymatic Formation of Novel Bile Acid Primary Amides

Bifunctional peptidylglycine α-amidating monooxygenase (PAM) catalyzes the copper-, ascorbate-, and O2-dependent cleavage of C-terminal glycine-extended peptides and N-acylglycines to the corresponding amides and glyoxylate. The α-amidated peptides and the long-chain acylamides are hormones in humans and other mammals. Bile acid glycine conjugates are also substrates for PAM leading to the formation of bile acid amides. The (VMAX/Km)app values for the bile acid glycine conjugates are comparable to other known PAM substrates. The highest (VMAX/Km)app value, 3.1 ± 0.12 × 105 M−1 s−1 for 3-sulfolithocholylglycine, is 6.7-fold higher than that for d-Tyr–Val–Gly, a representative peptide substrate. The time course for O2 consumption and glyoxylate production indicates that bile acid glycine conjugate amidation is a two-step reaction. The bile acid glycine conjugate is first converted to an N-bile acyl-α-hydroxyglycine intermediate which is ultimately dealkylated to the bile acid amide and glyoxylate. The enzymatically produced bile acid amides and the carbinolamide intermediates were characterized by mass spectrometry and two-dimensional 1H–13C heteronuclear multiple quantum coherence NMR.

Capillary gas chromatographic analysis of oxo, oxo-hydroxy and unsaturated bile acid glycine conjugates

A method for the direct gas chromatographic analysis without prior hydrolysis has been developed for a series of glycine-conjugated bile acids having oxo or multiple functional groups, including oxo and hydroxyl groups and double bonds. The glycine conjugates without and with the hydroxyl groups were derivatized to their ethyl or methyl esters and ethyl ester-trimethylsilyl or methyl ester-dimethylethylsilyl ethers, respectively, and chromatographed on an aluminum-clad, flexible, fused-silica capillary column coated with a thin film (0.1 μm) of chemically bonded and cross-linked methyl polysiloxane. The change of methylene unit value exerted by derivatization and glycination for each compound is discussed.

Capillary gas-liquid chromatography of glycine-conjugated bile acids without prior hydrolysis.

A method is described for the gas-liquid chromatographic (GLC) analysis of intact glycine conjugates of the major bile acids present in human plasma. It is, therefore, now possible to analyze glycine-conjugated and unconjugated bile acids together on a single GLC column without the necessity for a hydrolytic step. A large number of derivatives of bile acid glycine conjugates were examined, but only acetate- and silyl ether-derivatives of carboxylic acid methyl esters were found initially to be suitable. It was not possible to make acetates consistently, and trimethylsilyl ethers did not allow resolution of the glycine conjugates of cholic and chenodeoxycholic acids. Dimethylethylsilyl ether methyl ester derivatives were subsequently found to give the best results. Chromatographic conditions for successful analysis of these derivatives were examined and it was found to be necessary to use wall-coated capillary columns of thin film thickness (0.12 micron) and very high carrier gas flow rates (ca. 20 ml/min hydrogen). Using acetonitrile and Bond Elut extraction, fractionation on Sep-Pak SIL cartridges, and derivatization as dimethylethylsilyl ether methyl esters, the capillary gas-liquid chromatography of intact glycine-conjugated bile acids from human plasma was demonstrated for the first time.

A new hydrolase specific for taurine-conjugates of bile acids.

Through the investigation of the bile acid-deconjugation activities of human intestinal anaerobes, a new enzyme was discovered in Peptostreptococcus intermedius which hydrolyzed specifically the taurine-conjugates, but not the glycine-conjugates of bile acids. However, the enzymes in Streptococcus faecalis and Lactobacillus brevis hydrolyzed chiefly the glycine-conjugates.

Bile acid sulfates: II. Synthesis of 3‐monosulfates of bile acids and their conjugates

AbstractBile acid 3‐monosulfates were synthesized by the sulfation of 3‐hydroxy formyloxy bile acids with sulfur trioxide‐triethylamine in dimethylformamide at 25 C for 0.5 hr. The protecting formyl groups were then hydrolyzed under mild alkaline conditions, and the deformylated products were isolated as p‐toluidinium salts. These p‐toluidinium salts were converted easily to the corresponding disodium salts by methanolic sodium hydroxide. Disodium salts were then isolated by precipitation from methanolether. The corresponding glycine conjugates were similarly synthesized by the sulfation of ethyl esters of 3‐hydroxy formyloxy bile acid glycine conjugates. However, the taurine‐conjugated bile acid sulfates were obtained by conjugating bile acid 3‐monosulfates, either as triethylammonium salt or as disodium salt, with taurine in dimethylformamide in the presence of N‐ethoxycarbonyl‐2‐ethoxy‐1,2‐dihydroquinoline. These procedures produced the desired bile acid sulfates in high yield (typically above 90%) with minimum manipulation. No time‐consuming and capacity‐limited chromatographic purification was needed to isolate the pure sulfates. The thin layer chromatographic mobilities, the infrared spectra, and some of the preliminary studies of the properties of these synthesized 3‐monosulfates are also discussed.

Serum concentration of bile acids in relation to the normal menstrual cycle, the administration of oral contraceptives, and pregnancy

The effects of endogenous and exogenous steroids on serum levels of bile acid glycine conjugates were investigated by measuring these serum levels in 10 healthy women during normal menstrual cycle, in 10 healthy women during a cycle of combination oral contraceptives (OCs) (50-100 mcg of ethinyl estradiol or mestranol), and in 27 healthy women during normal pregnancy. Peripheral venous blood was obtained from nonpregnant patients on Days 2, 6, 12, 18, and 24 after onset of uterine bleeding. No significant differences in glycocholic and glycochenodeoxycholic acid serum levels occurred at any point in the cycle. Similarly, no significant differences were noted between patients receiving OCs and those with normal menstrual cycles. Fasting peripheral blood samples from the 27 pregnant volunteers (collected throughout the 3 trimesters) also revealed no significant serum level differences during any trimester. Physiologic changes in endogenous estrogens and progesterones have no appreciable effect on the bile acid serum levels measured; nor, apparently, do moderate doses of synthetic estrogens in OCs alter bile acid serum levels in healthy women.