Bile acid analysis in biological fluids: a novel approach

Area of Science:

• Biochemistry
• Analytical Chemistry
• Metabolomics

Background:

• Current bile acid analysis methods necessitate pre-separation into distinct groups.
• Accurate quantification and identification of diverse bile acid forms are crucial for diagnosing metabolic disorders.

Purpose of the Study:

• To develop and validate a high-performance liquid chromatography (HPLC) method for simultaneous separation and detection of various bile acids.
• To enable identification of specific bile acids indicative of metabolic errors in bile acid synthesis.

Main Methods:

• Utilized a reverse-phase column with gradient elution HPLC.
• Developed a method for forming fluorescent dimethoxycoumarin esters of bile acids.
• Employed a peak-shift strategy involving cholyglycine hydrolase and solvolysis for enhanced identification.
• Used 7 alpha,12 alpha-dihydroxy-5 beta-cholanoic acid as an internal standard for quantification.

Main Results:

• Successfully separated and detected nonconjugated, glycine-conjugated, and esterified bile acids.
• Identified allylic bile acids associated with metabolic errors under mild ester formation conditions.
• Confirmed bile acid identity through retention time, peak-shift analysis, and internal standardization.

Conclusions:

• The developed HPLC method offers a streamlined approach for comprehensive bile acid analysis.
• This method facilitates the identification of bile acid synthesis defects in biological fluids.
• The technique provides reliable quantification and identification of diverse bile acid species.