Novel Strategy for Untargeted Chiral Metabolomics using Liquid Chromatography-High Resolution Tandem Mass Spectrometry

Stereospecific
recognition of metabolites plays a significant role
in the detection of potential disease biomarkers thereby providing
new insights in diagnosis and prognosis. D-Hdroxy/amino acids
are recognized as potential biomarkers in several metabolic disorders.
Despite continuous advances in metabolomics technologies, the simultaneous
measurement of different classes of enantiomeric metabolites in a
single analytical run remains challenging. Here, we develop a novel
strategy for untargeted chiral metabolomics of hydroxy/amine groups
(−OH/–NH2) containing metabolites, including
all hydroxy acids (HAs) and amino acids (AAs), by chiral derivatization
coupled with liquid chromatography-high resolution tandem mass spectrometry
(LC-HR-MS/MS). Diacetyl-tartaric anhydride (DATAN) was used for the
simultaneous derivatization of–OH/–NH2 containing
metabolites as well as the resulting diastereomers, and all the derivatized
metabolites were resolved in a single analytical run. Data independent
MS/MS acquisition (DIA) was applied to positively identify DATAN-labeled
metabolites based on reagent specific diagnostic fragment ions. We
discriminated chiral from achiral metabolites based on the reversal
of elution order of D and L isomers derivatized
with the enantiomeric pair (±) of DATAN in an untargeted manner.
Using the developed strategy, a library of 301 standards that consisted
of 214 chiral and 87 achiral metabolites were separated and detected
in a single analytical run. This approach was then applied to investigate
the enantioselective metabolic profile of the bone marrow (BM) and
peripheral blood (PB) plasma samples from patients with acute myeloid
leukemia (AML) at diagnosis and following completion of the induction
phase of chemotherapeutic treatment. The sensitivity and selectivity
of the developed method enabled the detection of trace levels of the D-enantiomer of HAs and AAs in primary plasma patient samples.
Several of these metabolites were significantly altered in response
to chemotherapy. The developed LC-HR-MS method entails a valuable
step forward in chiral metabolomics.