Method for the separation of the unconjugates and conjugates of chenodeoxycholic acid and deoxycholic acid by two-dimensional reversed-phase thin-layer chromatography with methyl β-cyclodextrin

Momose, Toshiaki; Mure, Mayumi; Iida, Takashi; Goto, Junichi; Nambara, Toshio

doi:10.1016/s0021-9673(98)00213-1

Cited by 24 publications

(13 citation statements)

References 26 publications

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“…Although the most appropriate mobile phase water content (from 10 to 25 %) differs among compounds, depending on the polarity of the resulting nonionic hydrophobic derivatives, the chromatographic patterns in each group of compounds were very similar; the RF1 and RF2 values increased in the order LCA < CDCA DCA < CA < UDCA with mobile phases 1A-1C and in the order LCA ~ DCA < CDCA < CA < UDCA with mobile phases 2A-2C. These sequences for the doubleconjugated bile acids are similar to those for the unconjugated acids reported previously [6].…”

Section: Resultssupporting

confidence: 85%

“…dDistance between the two spot (see Ref. [6]). eSymbols in parentheses refer to the mobile phases used (see Table III).…”

Section: Resultsmentioning

confidence: 99%

“…2D R P H P T L C was performed as described previously [6]. Briefly, a sample solution was prepared by dissolving the bile acid in the mobile phase and left to stand at room temperature for 30 min.…”

Section: Methodsmentioning

confidence: 99%

“…According to these papers, separation of all of the double-conjugated pairs of CDCA and DCA was unsuccessful. We have previously reported an efficient method for the separation of the five major bile acids in both the unconjugated and conjugated forms [6]. The method involved the use of two-dimensional (2D) reversed-phase (RP) high-performance thin-layer chromatography (HPTLC) [7,8] with aqueous methanol (10-30 %) in the first direction and the same mobile phase containing 5 mM methyl 13-cyclodextrin (Me-J3-CD) in the second direction.…”

mentioning

confidence: 99%

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Separation of double conjugates of bile acids by two-dimensional high-performance thin-layer chromatography with tetra-n-butylammonium phosphate and methyl β-cyclodextrin

et al. 1999

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SummaryThis paper describes a method for the separation, by high-performance thin-layer chromatography (HPTLC), of a series of polar, ionic, and hydrophilic double conjugates of bile acids amidated at the C-24 carboxyl group with glycine or taurine and sulfonated or glucosylated at hydroxyl groups in the 513-steroid nucleus. The method involves two-dimensional (2D) reversed-phase (RP) HPTLC with the combined use of tetra-n-butylammonium phosphate (TBAP) and methyl as mobile phase additives. Complete separation of the hydrophilic bile acid conjugates, particularly of the recalcitrant pairs of chenodeoxycholic acid and deoxycholic acid conjugates in each group, was achieved by 2D inclusion RPHPTLC by developing with methanol-water-0.5 tool L -1 TBAP, 90:10:5-75:25:5 (v/v) in the first dimension and the same mobile phase containing 5 mM Meq3-CD in the second dimension. The method could be usefully applied to biosynthetic and metabolic studies of bile acids in biological materials.necessary. TLC is, therefore, widely used for the preliminary and/or routine analysis of a wide variety of compounds. Several studies have reported the TLC separation of biologically important bile acids in connection with hepatobiliary diseases [1][2][3]. The compounds separated include unconjugated cholic acid (CA), chenodeoxycholic acid (CDCA), ursodeoxycholic acid (UDCA), deoxycholic acid (DCA), and lithocholic acid (LCA), and their conjugates with glycine, taurine or sulfuric acid. As far as we are aware, however, there have been few reports of the TLC analysis of more polar and ionic double conjugates of bile acids, amidated at the C-24 carboxyl group with glycine (or taurine) and sulfonated or glycosylated at the hydroxyl groups in the 513-steroid nucleus, on normal phase (NP) TLC plates with alcohol-acetic acid-water mixtures as the mobile phase [4,5]. According to these papers, separation of all of the double-conjugated pairs of CDCA and DCA was unsuccessful. We have previously reported an efficient method for the separation of the five major bile acids in both the unconjugated and conjugated forms [6]. The method involved the use of two-dimensional (2D) reversed-phase (RP) high-performance thin-layer chromatography (HPTLC) [7,8] with aqueous methanol (10-30 %) in the first direction and the same mobile phase containing 5 mM methyl 13-cyclodextrin (Me-J3-CD) in the second direction. This paper described the application of this 2D inclusion RPHPTLC method to the complete separation of various types of double-conjugated bile acids.

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Section: Resultssupporting

confidence: 85%

“…dDistance between the two spot (see Ref. [6]). eSymbols in parentheses refer to the mobile phases used (see Table III).…”

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Separation of double conjugates of bile acids by two-dimensional high-performance thin-layer chromatography with tetra-n-butylammonium phosphate and methyl β-cyclodextrin

et al. 1999

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“…Reverse-phase TLC has also been used for bile acids ( 200,201 ), but has not achieved popularity because of the much lower capacity of the method, as well as diffi culties in detecting bile acids after separation. Eneroth and Sjövall tabulated R F values using TLC for a large spectrum of bile acids and a number of solvent systems in their review in 1971 ( 195 ).…”

Section: Structure-function Relationshipsmentioning

confidence: 99%

Key discoveries in bile acid chemistry and biology and their clinical applications: history of the last eight decades

Hofmann

Hagey

2014

Journal of Lipid Research

298

284

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conceptions are rare, and provocative judgments stand the test of time. Let us also hope that this effort will be both useful and entertaining. Each of the topics discussed merits at least a full length article, so there will be, of necessity in many instances, consideration of only what we perceive to be the highlights. THE EBB AND FLOW OF MEDICAL INTEREST IN BILE ACIDSAfter the elucidation of the true chemical structure of bile acids in 1932 (see below), there was little interest in bile acids in the Western world. One exception to this statement was the laboratory of Siegfried Thannhauser, who wrote the fi rst textbook of metabolic biochemistry in Germany. He studied cholesterol and bile acid balance in the biliary fi stula dog ( 1 ). During this time, bile acids were sold as liver tonics and laxatives, but there were no placebo-controlled studies showing effi cacy. Indeed, bile acids were considered by the medical profession to have no useful therapeutic properties. The tri-oxo derivative of cholic acid (called "dehydrocholic acid") was known to induce bile fl ow in animals ( 2 ), and was occasionally used to stimulate bile fl ow in patients; but again there were no controlled studies showing effi cacy in hepatobiliary disease.

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High-Performance Thin-Layer Chromatography (HPTLC)

Srivastava¹

2011

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Method for the separation of the unconjugates and conjugates of chenodeoxycholic acid and deoxycholic acid by two-dimensional reversed-phase thin-layer chromatography with methyl β-cyclodextrin

Cited by 24 publications

References 26 publications

Separation of double conjugates of bile acids by two-dimensional high-performance thin-layer chromatography with tetra-n-butylammonium phosphate and methyl β-cyclodextrin

Separation of double conjugates of bile acids by two-dimensional high-performance thin-layer chromatography with tetra-n-butylammonium phosphate and methyl β-cyclodextrin

Key discoveries in bile acid chemistry and biology and their clinical applications: history of the last eight decades

High-Performance Thin-Layer Chromatography (HPTLC)

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