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LC-MS/MS Analysis of Bile Acids using Ascentis® Express C18

Cory Muraco

MilliporeSigma, Bellefonte, PA

Introduction

An Ascentis® Express C18 column was used for the LC-MS/MS analysis of bile acids and their conjugates. The separation of multiple isobaric compounds was demonstrated in addition to excellent linearity, enabling clinically relevant concentrations to be analyzed.

Experimental Conditions for Analysis of Bile Acids

Table 1.Chromatographic Conditions for Analysis of Bile Acids on Ascentis® Express C18


Performance results for the Separation of Bile Acids on Ascentis® Express C18

Chromatogram showing the LC-MS separation of 13 bile acids and conjugates using an Ascentis® Express C18 column

Figure 1.LC-MS Results for the Separation of Bile Acids on Ascentis® Express C18. Numbering corresponds to elution order as indicated on the right.

Peak Identities

  1. Sodium tauroursodeoxycholate (TUDC)
  2. Glycoursodeoxycholic acid (GDC)
  3. Taurocholic acid sodium salt hydrate (TCA)
  4. Glycocholic acid hydrate (GCA)
  5. Sodium taurochenodeoxycholate (TCDC)
  6. Ursodeoxycholic acid (UDC)
  7. Sodium taurodeoxycholate hydrate (TDC)
  8. Sodium glycochenodeoxycholate (GCDC)
  9. Cholic acid (CA)
  10. Sodium taurolithocholate (TLC)
  11. Chenodeoxycholic acid (CDC)
  12. Deoxycholic acid (DC)
  13. Lithocholic acid (LC)[MN1]
Table 2.MS Transitions for Each Compound Identified in Figure 1

Linearity of Assay for Each Compound

Linearity of Assay for the bile acids and conjugates used in their LC-MS analysis

Sodium tauroursodeoxycholate (TUDC)

Figure 2.Sodium tauroursodeoxycholate (TUDC)

Taurocholic acid sodium salt hydrate (TCA)

Figure 3.Taurocholic acid sodium salt hydrate (TCA)

Glycoursodeoxycholic acid (GDC)

Figure 4.Glycoursodeoxycholic acid (GDC)

Glycocholic acid hydrate (GCA)

Figure 5.Glycocholic acid hydrate (GCA)

Sodium taurochenodeoxycholate (TCDC)

Figure 6.Sodium taurochenodeoxycholate (TCDC)

Sodium taurodeoxycholate hydrate (TDC)

Figure 7.Sodium taurodeoxycholate hydrate (TDC)

Ursodeoxycholic acid (UDC)

Figure 8.Ursodeoxycholic acid (UDC)

Sodium glycochenodeoxycholate (GCDC)

Figure 9.Sodium glycochenodeoxycholate (GCDC)

Cholic acid (CA)

Figure 10.Cholic acid (CA)

Chenodeoxycholic acid (CDC)

Figure 11.Chenodeoxycholic acid (CDC)

Sodium taurolithocholate (TLC)

Figure 12.Sodium taurolithocholate (TLC)

Deoxycholic acid (DC)

Figure 13.Deoxycholic acid (DC)

Lithocholic Acid (LC)

Figure 14.Lithocholic Acid (LC)


Conclusion

Excellent resolution and linearity were obtained for the isobaric bile acids on the Ascentis® Express C18 column.

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