MS/MS Mass Spectrometry Filtering Tree for Bile Acid Regio- and Stereoisomer Annotation

Bile acids are essential
steroids with a wide range of biological
roles, including the regulation of immunity, nutrient absorption,
insulin signaling, appetite, and body temperature. However, due to
similarities in their MS/MS spectra, spectral matching with reference
MS/MS libraries generally fails to differentiate between isomers.
This study introduces a proof-of-concept workflow that uses a mass
spectrometry query language filtering tree to distinguish isomeric
bile acids in untargeted LC-MS/MS data by leveraging intensity ratios
of ions that are close to one another in the MS/MS spectrum. It can
be retrospectively applied to existing LC-MS/MS data in data repositories.
The filtering tree concept provides the opportunity to annotate and
distinguish previously unknown bile acid isomers across LC-MS/MS data
sets. To facilitate the ease of applying these filters to LC-MS/MS
data sets, we developed a web-based application that simplifies the
stepwise filtering tree workflow, removing the need for coding expertise.
Here, we apply the multistep filtering application to a representative
public data set, which revealed distinct patterns of bile acids associated
with different diet types across diverse mammalian species. We further
identified the previously uncharacterized bile acid deoxycholyl-N-acetyl-putrescine, which was elevated in carnivores.