Bile acids conjugation in human bile is not random: new insights from (1)H-NMR spectroscopy at 800 MHz

Abstract

Bile acids constitute a group of structurally closely related molecules and represent the most abundant constituents of human bile. Investigations of bile acids have garnered increased interest owing to their recently discovered additional biological functions including their role as signaling molecules that govern glucose, fat and energy metabolism. Recent NMR methodological developments have enabled single-step analysis of several highly abundant and common glycine- and taurine- conjugated bile acids, such as glycocholic acid, glycodeoxycholic acid, glycochenodeoxycholic acid, taurocholic acid, taurodeoxycholic acid, and taurochenodeoxycholic acid. Investigation of these conjugated bile acids in human bile employing high field (800 MHz) (1)H-NMR spectroscopy reveals that the ratios between two glycine-conjugated bile acids and their taurine counterparts correlate positively (R2 = 0.83-0.97; p = 0.001 x 10(-2)-0.006 x 10(-7)) as do the ratios between a glycine-conjugated bile acid and its taurine counterpart (R2 = 0.92-0.95; p = 0.004 x 10(-3)-0.002 x 10(-10)). Using such correlations, concentration of individual bile acids in each sample could be predicted in good agreement with the experimentally determined values. These insights into the pattern of bile acid conjugation in human bile between glycine and taurine promise useful clues to the mechanism of bile acids' biosynthesis, conjugation and enterohepatic circulation, and may improve our understanding of the role of individual conjugated bile acids in health and disease.

Fig. 1

Typical ¹H-NMR spectrum at 800 MHz of human gallbladder bile (100 μL bile diluted to 600 μL)

Fig. 2

Left Portions of the 800 MHz ¹H-NMR spectra of gallbladder bile from five different patients (four from non-liver disease controls and one from hepatitis C patient). The spectra were obtained by decoupling methylene protons of glycine-/taurine-conjugated bile acids. Right The corresponding deconvoluted spectra obtained for estimating the integrals. GCA glycocholic acid; GCDCA glycochenodeoxycholic acid; GDCA glycodeoxycholic acid; TCA taurocholic acid; TCDCA: taurochenocholic acid and TDCA taurodeoxycholic acid

Fig. 3

Concentration of individual bile acids (in mmol/L) for different bile samples. Abbreviations are as described before in Fig. 2 caption

Fig. 4

Ratios of a glycine- to taurine-conjugated bile acids and b–d glycine to the corresponding taurine-conjugated bile acid. Abbreviations are as described before in Fig. 2 caption

Fig. 5

Plots showing the linear correlation of the ratios a–c between a pair of glycine-conjugated bile acids and between the corresponding taurine-conjugated bile acids; and d–f between a glycine-conjugated bile acid and its taurine counterpart and between another glycine-conjugated bile acid and its taurine counterpart. Abbreviations are as described before in Fig. 2 caption