Arne Norman

Fasting and postprandial serum bile acid concentration with special reference to variations in the conjugate profile

Serum bile acids were group-separated by ion exchange chromatography on diethylaminohydroxypropyl Sephadex LH-20 into unconjugated bile acids and bile acids conjugated with either glycine, taurine, glucuronic acid or sulphuric acid. The conjugate moiety was hydrolysed by treatment with a combination of Helix pomatia and cholylglycine hydrolase and the released bile acids analysed by gas liquid chromatography/mass spectrometry. Analysis of fasting and postprandial serum from six healthy subjects showed that, in addition to the primary bile acids, cholic (C) and chenodeoxycholic acid (CDC), secondary bile acids were present to varying extents. Unconjugated serum bile acids were found in four of the six subjects. Glycine and taurine conjugates of C and CDC and their glucuronides and sulphates were found in all subjects. The postprandial increase of serum bile acids was mainly due to increase of the glycine conjugates of C and CDC. After the meal, the ratio C:CDC in glycine and taurine conjugates shifted to lower values.

Comparison of Bile Acids in Intestinal Contents of Germfree and Conventional Rats.

Summary Two days after administration of cholic acid-24-C14 to germfree and conventional rats the intestinal contents were separated by centrifugation at 25,000 X g. 1-The labeled bile acids throughout the intestinal tract of germfree rats were recovered in the supernatant. 2. In the conventional rats almost all labeled bile acids in the small intestine were recovered in the supernatant whereas 27-44% of the labeled bile acids in cecum were found in the sediment. 3. The labeled bile acids in the sediment after centrifugation of cecal content had a higher per cent of deoxycholic and 3-hydroxy-12-keto-cholanic acid than those present in the supernatant.

Excretion of bile acids in extrahepatic biliary atresia and intrahepatic cholestasis of infancy.

This series included 24 infants, 16 boys and 8 girls, who were admitted to hospital with the diagnosis of obstructive jaundice. Five of the infants were subsequently found to have extra‐hepatic biliary atresia (BA) and the other 19 infants intrahepatic cholestasis of infancy (IHC).

Urinary Bile Acid Excretion in Correlation to Liver Histopathology in Cystic Fibrosis

Urinary bile acid excretion and liver morphology were compared in 25 patients with cystic fibrosis (CF). None showed clinical signs of liver disease. Most of the patients had normal liver function tests. Bile acids were determined in 24-h samples by a modification of the method of Almé. All patients had increased urinary excretion of trihydroxy bile acids, mainly cholic, 3 beta, 7 beta, 12 alpha- and 3 alpha, 7 beta, 12 alpha-trihydroxy-5 beta-cholanoic acids. Lithocholic acid excretion was lower in CF than in normal children. The urinary excretion of 3 beta-hydroxy-5-cholenoic acid was not increased in CF. In three patients with cirrhosis the urinary excretion of chenodeoxycholic acid was increased. The ratio of cholic to 3 beta-hydroxy-5-cholenoic acids was increased in all but three patients, and the ratio of chenodeoxycholic to 3 beta-hydroxy-5-cholenoic acids was increased in those with cirrhosis. These ratios differed more between cirrhotic and non-cirrhotic CF patients in this series than the ratio of cholic to chenodeoxycholic acids.

Excretion of bile acids in healthy children and children with cystic fibrosis.

Urinary bile acid excretion was investigated in twenty-two patients with cystic fibrosis (CF) and in seven healthy children. CF patients with and without antibiotic treatment were compared. Bile acids were determined in 24-h samples after separation into unconjugated, glycine conjugated, taurine conjugated and sulphate conjugated bile acids. In total twenty bile acids were identified of which cholic, chenodeoxycholic, 3 beta-hydroxy-5-cholenoic acid and 24-nor-5 beta-cholan-23-oic acid were routinely present in samples collected from both CF patients and healthy children. None of the other bile acids were preferentially excreted by CF patients. When compared with the normal group, no statistical significance could be attached to the increased total urinary bile acids excreted by the CF patients (due to the large individual variations). The CF patients excreted increased amounts of cholic acid, 3 alpha, 7 beta, 12 alpha-and 3 beta, 7 beta, 12 alpha-trihydroxy-5 beta-cholanoic acids mainly in the unconjugated state. After administration of 24-[14C]cholic acid to thirteen CF patients the isotope excretion in faeces and urine was studied. Most of the patients had a high faecal excretion indicating great losses of bile acids from enterohepatic circulation. Compared to normal adults CF children excreted isotope in increased amounts in the urine.

Bile acid transport by serum lipoproteins in patients with extrahepatic cholestasis

Individual bile acid conjugates were determined by gas-liquid chromatography in very low density (VLDL), low density (LDL) and high density (HDL) lipoprotein fractions obtained by sequential ultracentrifugation of serum from cholestatic patients. In the lipoproteins were found 16-48% of the serum bile acids. In LDL, the amount found was twice that in VLDL and HDL together. More cholic than chenodeoxycholic acid was detected in the lipoproteins. The occurrence of bile acids in lipoproteins might be of importance for the transport of bile acids from blood to peripheral tissues during cholestasis.

Bile acid conjugates present in tissues during extrahepatic cholestasis

Bile acid concentrations in the muscle, subcutaneous fat and skin of patients with extrahepatic cholestasis were compared to those in serum. High concentrations were found in most tissue samples. In tissues from patients administered [24-14C]chenodeoxycholic acid, the specific activity was lower than that in serum, indicating a storage of bile acids in these tissues. In some patients, higher concentrations of bile acids were found in tissue samples than in serum. This was mainly due to increased concentrations of cholic acid. Monohydroxy bile acids were not found in increased concentrations. Bile acids covalently bound to tissues were not detected.

URINARY BILE ACID CONJUGATES IN EXTRAHEPATIC BILIARY ATRESIA

The urinary excretion of bile acid conjugates was studied after the administration of 24‐14C‐labelled chenodeoxycholic acid and cholic acid and their corresponding glycine conjugates labelled with glycine‐l‐14T in 5 infants with extrahepatic biliary atresia. Chenodeoxycholic acid‐24‐14C and chenodeoxycholyl[l‐14C]glycine were mainly excreted in the form of labelled metabolites with the TLC behaviour of monosulphates of tauro‐ and glycochenodeoxycholate and glycochenodeoxycholate, respectively. Most of the cholic acid‐24‐14C and the cholyl[l‐14C]glycine were found to be excreted in the form of glycocholate. All labelled bile acids were excreted in conjugated form.

METABOLISM OF LITHOCHOLIC ACID-24–14C IN EXTRAHEPATIC BILIARY ATRESIA

Lithocholic acid‐24‐14C was given to 2 infants with extrahepatic biliary atresia. After oral administration about half of the amount of isotope was found to be adsorbed. After par‐ 9. Makino, I., Sjovall, J., Norman, A. & Strandvik, enteral administration about half of the amount of isotope was excreted in the urine for 3 days. Lithocholic acid was shown to be transformed to a small extent to more polar compounds. These metabolites were not cholic, chenodeoxycholic, α‐ or,β‐muricholic, hyocholic, hyodeoxycholic or ursodeoxycholic acids. Most of the lithocholic acid was excreted as the sulphate esters of tauro‐ and glycolithocholic acid. No sulphate ester of lithocholic or free lithocholic acid was excreted.

Conjugation, metabolism and excretion of 24-14C chenodeoxycholic acid in patients with extrahepatic cholestasis before and after biliary drainage-analysis of conjugated bile acids by HPLC

[24-14C] chenodeoxycholic acid (CDC) was given to patients with total extrahepatic cholestasis two or three days before an external drainage was made, and excretion of the isotope in urine and bile followed. Bile acids were group-separated by anion exchange chromatography on DEAP-Sephadex LH-20 and the individual conjugates isolated by HPLC. 51.0-75.4% of the administered isotope was excreted; 16.2-29.9% as sulphates, 0.1-2.4% as glucuronides and 20.7-58.7% as glycine and taurine conjugates. 5.2-21.0% of excreted isotope consisted of transformation products of CDC, mainly cholic acid, hyocholic acid and ursodeoxycholic acid. Labelled urinary sulphates were the 3-sulphates of glycochenodeoxycholic and taurochenodeoxycholic acid. During cholestasis the renal clearance was about ten times higher for the sulphates compared with the non-sulphated conjugate. The clearance of glycine conjugates and their sulphates was of the same magnitude as that of the corresponding taurine conjugates. During the biliary drainage period, most of the labelled sulphates were excreted in urine, while most of the glycine and taurine conjugates were excreted in bile.