Effect of protein on the determination of total bile acids in serum.

1983 ◽  
Vol 29 (1) ◽  
pp. 171-175 ◽  
Author(s):  
N Q Hanson ◽  
E F Freier
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Hydroxysteroid Dehydrogenase ◽  
Inhibitory Effect ◽  
Normal Serum ◽  
Total Serum ◽  
Enzyme Preparations ◽  
Analytical Recovery ◽  
Fluorescence Light

Abstract We measured total serum bile acids on a fluorescence-light-scattering micro centrifugal analyzer by the direct enzymatic method with 3 alpha-hydroxysteroid dehydrogenase (EC 1.1.1.50) and with resazurin as a fluorogenic electron acceptor. We found that serum protein has an inhibitory effect on the measurement of bile acids, but this effect was eliminated by adding bovine serum albumin to the reaction mixture in a final protein concentration (12.2 g/L) that was high compared with that contributed by a normal serum specimen. The assay is a sensitive method that reaches equilibrium in 5 min. The method is microscale (5 microL of sample, 150 microL of working reagent), is easy to perform, and is accurate (analytical recovery = 104.1%) and precise (CV = 11.1 and 5.7% on specimens with bile acid concentrations of 7.6 and 35.4 mumol/L, respectively). Normal values are 1-12 and less than 9 mumol/L on nonfasting and fasting individuals, respectively. Pure 3 alpha-hydroxysteroid dehydrogenase must be used: we found several enzyme preparations that gave falsely high values for bile acid.

Evaluation of a Commercial Enzymatic Method for the Determination of Total Serum Bile Acids

1980 ◽  
Vol 17 (6) ◽  
pp. 301-306 ◽  
Author(s):  
Per Tobiasson ◽  
Magnus Källberg
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Reference Range ◽  
Reference Value ◽  
Total Serum ◽  
Enzymatic Method ◽  
Normal Range ◽  
Serum Samples ◽  
Serum Bile Acids

An enzymatic method (Sterognost-3α® Flu, Nyegaard & Co, Oslo, Norway) for the determination of total serum bile acids has been evaluated, and reference values for fasting and postprandial total serum bile acids have been established. Precision was rather low in the lower normal range but satisfactory in the upper reference range and for elevated values. The upper reference value for fasting total serum bile acids was 5·0 μmol/l. Maximal postprandial elevations occurred at varying intervals after a test meal and the upper postprandial reference value was 12 μmol/l. Certain serum samples were also analysed by radioimmunological methods for conjugates of cholic and chenodeoxycholic acids. The values obtained with the enzymatic method correlated satisfactorily with those of the radioimmunoassay methods for most of the serum samples analysed. However, a number of samples with slightly elevated bile acid concentrations, as measured by the comparison radioimmunoassay methods, were found to have normal total serum bile acids when measured by the enzymatic method.

Enzyme inactivation in serum before determination of total bile acids.

1983 ◽  
Vol 29 (6) ◽  
pp. 1073-1075 ◽  
Author(s):  
N Q Hanson ◽  
E F Freier
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Final Concentration ◽  
Methanol Solution ◽  
Enzyme Inactivation ◽  
Total Serum ◽  
Sodium Pyruvate ◽  
Lactate Dehydrogenase Activity ◽  
Total Bile Acids

Abstract Endogenous NADH-generating enzymes must be inactivated before total serum bile acids can be measured accurately by the direct enzymic method. To do this, we pretreat the sera with NaOH, in a final concentration of 0.1 mol/L. Consequently, lactate dehydrogenase activity at least as high as 30 000 U/L is destroyed, obviating blank determinations. Values for bile acid in serum, so obtained, agree with values obtained after pretreatment with heat, an alkali-methanol solution, or sodium pyruvate, but our pretreatment has the advantages of ease, speed, economy, and negligible blank values.

scholarly journals Serum Total Bile Acid Levels in Patients Receiving Rifampicin and Isoniazid

1984 ◽  
Vol 21 (3) ◽  
pp. 218-222 ◽  
Author(s):  
Jonathan D Berg ◽  
Harry I Pandov ◽  
Herbert G Sammons
Keyword(s):  
Bile Acid ◽  
Liver Function ◽  
Bile Acids ◽  
Continuous Flow ◽  
Flow System ◽  
Hydroxysteroid Dehydrogenase ◽  
Total Bile Acid ◽  
Skewed Distribution ◽  
Total Serum ◽  
Serum Bile Acid

Serum bile acid levels in 61 patients receiving daily doses of rifampicin and isoniazid for the treatment of tuberculosis have been investigated. Bile acids were measured using 3α-hydroxysteroid dehydrogenase in a continuous-flow system. Abnormally elevated levels were found in 44 patients (72%) during the period of study up to 80 days after onset of treatment. The results showed a mean of 24·9 μmol/l and a positively skewed distribution. Whilst marginally raised levels of bilirubin were seen in some samples (mean 8·2 μmol/l), these did not reflect the marked changes observed in bile acids. Patients receiving rifampicin and isoniazid may therefore have markedly elevated levels of total serum bile acids, while other tests used to assess liver function can remain normal.

Role of amidation in bile acid effect on DNA synthesis by regenerating mouse liver

1995 ◽  
Vol 268 (6) ◽  
pp. G1051-G1059
Author(s):  
E. R. Barbero ◽  
M. C. Herrera ◽  
M. J. Monte ◽  
M. A. Serrano ◽  
J. J. Marin
Keyword(s):  
Bile Acid ◽  
Dna Synthesis ◽  
Bile Acids ◽  
High Performance ◽  
Cell Injury ◽  
Intraperitoneal Administration ◽  
Inhibitory Effect ◽  
Toxicity Tests ◽  
Maximal Effect

Effect of bile acids on DNA synthesis by the regenerating liver was investigated in mice in vivo after partial hepatectomy (PH). Radioactivity incorporation into DNA after [14C]thymidine intraperitoneal administration peaked at 48 h after PH. At this time a significant taurocholate-induced dose-dependent reduction in DNA synthesis without changes in total liver radioactivity content was found (half-maximal effect at approximately 0.1 mumol/g body wt). Effect of taurocholate (0.5 mumol/g body wt) was mimicked by chocolate, ursodeoxycholate, deoxycholate, dehydrocholate, tauroursodeoxycholate, taurochenodeoxycholate, and taurodeoxycholate. In contrast, chenodeoxycholate, glycocholate, glycochenodeoxycholate, glycoursodeoxycholate, glycodeoxycholate, 5 beta-cholestane, bromosulfophthalein, and free taurine lacked this effect. No relationship between hydrophobic-hydrophilic balance and inhibitory effect was observed. Analysis by high-performance liquid chromatography indicated that inhibition of thymidine incorporation into DNA was not accompanied by an accumulation of phosphorylated DNA precursors in the liver but rather by a parallel increase in nucleotide catabolism. Bile acid-induced modifications in DNA synthesis were observed in vivo even in the absence of changes in toxicity tests, which suggests that the inhibitory effect shared by most unconjugated and tauroconjugated bile acids but not by glycoconjugated bile acids should be accounted for by mechanisms other than nonselective liver cell injury.

scholarly journals Taurolithocholate-induced Ca2+ release is inhibited by phorbol esters in isolated hepatocytes

1992 ◽  
Vol 287 (3) ◽  
pp. 891-896 ◽  
Author(s):  
L Combettes ◽  
B Berthon ◽  
M Claret
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Phorbol Ester ◽  
Phorbol Esters ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Inhibitory Effect ◽  
Cell Uptake ◽  
Intracellular Binding ◽  
Pkc Activation

The monohydroxy bile acid taurolithocholate (TLC) causes a rapid and transient increase in free cytosolic Ca2+ concentration ([Ca2+]i) in suspensions of rat hepatocytes similar to that elicited by the InsP3-dependent hormone vasopressin. The effect of the bile acid is due to a mobilization of Ca2+, independent of InsP3, from the endoplasmic reticulum (ER). Short-term preincubation of cells with the phorbol ester 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (PMA), which activates protein kinase C (PKC), blocked the increase in [Ca2+]i induced by TLC, but did not alter that mediated by vasopressin. We obtained the following results, indicating that the effect of PMA is mediated by the activation of PKC. (1) Phorbol esters were effective over a concentration range where they activate PKC (IC50 = 0.5 nM); (2) phorbol esters that do not activate PKC did not inhibit the effects of TLC; (3) the permeant analogue oleoylacetylglycerol mimicked the inhibitory effect of PMA; (4) lastly, the inhibition of the TLC-induced Ca2+ mobilization by phorbol esters was partially prevented by preincubating the cells with the PKC inhibitors H7 and AMG-C16. Preincubating hepatocytes with PMA had no effect on the cell uptake of labelled TLC, indicating that the phorbol ester does not interfere with the transport system responsible for the accumulation of bile acids. In saponin-treated liver cells, PMA added before or after permeabilization failed to abolish TLC-induced Ca2+ release from the ER. The possibility is discussed that PMA, via PKC activation, may alter the intracellular binding or the transfer of bile acids in the liver.

Enzymatic determination of serum 12α-hydroxy bile acid concentration with 12α-hydroxysteroid dehydrogenase

1988 ◽  
Vol 23 (6) ◽  
pp. 646-651 ◽  
Author(s):  
Naoki Tamasawa ◽  
Masashi Yoneda ◽  
Isao Makino ◽  
Kazuo Takebe ◽  
Shigeru Ueda ◽  
...  
Keyword(s):  
Bile Acid ◽  
Acid Concentration ◽  
Hydroxysteroid Dehydrogenase ◽  
Bile Acid Concentration ◽  
Enzymatic Determination

Radioimmunoassay of conjugated cholic acid, chenodeoxycholic acid, and deoxycholic acid from human serum, with use of 125I-labeled ligands.

1979 ◽  
Vol 25 (2) ◽  
pp. 264-268 ◽  
Author(s):  
O Mäentausta ◽  
O Jänne
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Cholic Acid ◽  
Chenodeoxycholic Acid ◽  
Deoxycholic Acid ◽  
Serum Samples ◽  
Analytical Recovery ◽  
Hepatobiliary Diseases ◽  
Polyethylene Glycol Precipitation ◽  
Free Serum

Abstract We describe a method for radioimmunoassay of conjugated cholic acid, chenodeoxycholic acid, and deoxycholic acid in serum. In the method, 125I-labeled bile acid conjugates are used as the tracers along with antibodies raised against individual bile acid-bovine serum albumin conjugates. Antibody-bound and free bile acids were separated by polyethylene glycol precipitation (final concentration, 125 g/L). Before radioimmunoassay, 0.1-mL serum samples were precipitated with nine volumes of ethanol, and portions from the supernate were used in the assays. The lowest measurable amounts of the bile acids, expressed as pmol/tube, were: cholic acid conjugates, 2; chenodeoxycholic acid conjugates, 0.5; and deoxycholic acid conjugates. 2. Analytical recovery of bile acids added to bile acid-free serum ranged from 85 to 110%; intra-assay and inter-assay CVs ranged from 3.2 to 5.3% and from 5.3 to 12.2%, respectively. Concentrations (mean +/- SD) of the bile acid conjugates in serum from apparently healthy women and men (in mumol/L) were: cholic acid conjugates, 0.43 +/- 0.17 (n = 126); chenodeoxycholic acid conjugates, 0.47 +/- 0.23 (n = 111); and deoxycholic acid conjugates, 0.33 +/- 0.11 (n = 96). The values for primary bile acids were greatly increased in patients with various hepatobiliary diseases.

A Direct Enzymic Assay for 7α-Hydroxy Bile Acids and Their Conjugates

1973 ◽  
Vol 44 (1) ◽  
pp. 95-98 ◽  
Author(s):  
G. A. D. Haslewood ◽  
G. M. Murphy ◽  
Judith M. Richardson
Keyword(s):  
Thin Layer ◽  
Bile Acids ◽  
Thin Layer Chromatography ◽  
Biological Fluids ◽  
Hydroxysteroid Dehydrogenase ◽  
Layer Chromatography ◽  
Enzymic Assay ◽  
Bile Acids And Salts ◽  
Secondary Bile Acids

1. A 7α-hydroxysteroid dehydrogenase preparation isolated from a strain of Escherichia coli provides a specific and rapid means for the determination of 7α-hydroxy bile acids and their conjugates. 2. When used in conjunction with 3α-hydroxysteroid dehydrogenase the method enables the concentrations of primary and secondary bile acids and salts in biological fluids to be determined directly without the use of thin-layer chromatography. 3. When used with thin-layer chromatography the method allows the specific quantitative determination of chenodeoxycholic acid or its conjugates in the presence of deoxycholic acid and its derivatives.

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