title: Determination of low levels of 2H-labeling using high-resolution mass spectrometry: application in studies of lipid flux and beyond.
abstract: BACKGROUND
  The ability to measure low levels of (2)H-labeling is important in studies of metabolic flux, e.g. one can estimate lipid synthesis by administering (2)H2O and then measuring the incorporation of (2)H into fatty acids. Unfortunately, the analyses are complicated by the presence of more abundant naturally occurring stable isotopes, e.g. (13)C. Conventional approaches rely on coupling gas chromatographic separation of lipids with either quadrupole-mass spectrometry (q-MS) and/or pyrolysis-isotope ratio mass spectrometry (IRMS). The former is limited by high background labeling (primarily from (13)C) whereas the latter is not suitable for routine high-throughput analyses.
  
  
  METHODS
  We have contrasted the use of continuous flow-pyrolysis-IRMS against high-resolution mass spectrometry (i.e. Qq-FT-ICR MS) for measuring the (2)H-enrichment of fatty acids and peptides.
  
  
  RESULTS
  In contrast to IRMS, which requires ~30 min per analysis, it is possible to measure the (2)H-enrichment of palmitate via direct infusion high-resolution mass spectrometry (HRMS) in ~3 min per sample. In addition, Qq-FT-ICR MS enabled measurements of the (2)H-enrichment of peptides (which is not possible using IRMS).
  
  
  CONCLUSIONS
  High-resolution mass spectrometry can be used to measure low levels of (2)H-labeling so we expect that this approach will enhance studies of metabolic flux that rely on (2)H-labeled tracers, e.g. (2)H2O. However, since the high-resolution analyses require greater amounts of a given analyte one potential limitation centers on the overall sensitivity. Presumably, future advances can overcome this barrier.
authors.affiliation: Merck Research Laboratories, Rahway, NJ, USA.
authors.forename: Kithsiri B
authors.initials: KB
authors.lastname: Herath
authors.forename: Wendy
authors.initials: W
authors.lastname: Zhong
authors.forename: Jiong
authors.initials: J
authors.lastname: Yang
authors.forename: Ablatt
authors.initials: A
authors.lastname: Mahsut
authors.forename: Rory J
authors.initials: RJ
authors.lastname: Rohm
authors.forename: Vinit
authors.initials: V
authors.lastname: Shah
authors.forename: Jose
authors.initials: J
authors.lastname: Castro-Perez
authors.forename: Haihong
authors.initials: H
authors.lastname: Zhou
authors.forename: Athula B
authors.initials: AB
authors.lastname: Attygalle
authors.forename: Ling
authors.initials: L
authors.lastname: Kang
authors.forename: Sheo
authors.initials: S
authors.lastname: Singh
authors.forename: Douglas G
authors.initials: DG
authors.lastname: Johns
authors.forename: Michele A
authors.initials: MA
authors.lastname: Cleary
authors.forename: Brian K
authors.initials: BK
authors.lastname: Hubbard
authors.forename: Stephen F
authors.initials: SF
authors.lastname: Previs
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Roddy
mesh_terms: D000818:Animals; D002522:Chlorocebus aethiops; D003903:Deuterium / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D017666:Deuterium Oxide / Q000008:administration & dosage; D005227:Fatty Acids / Q000737:chemistry* / Q000378:metabolism; D005260:Female; D007553:Isotope Labeling / Q000379:methods*; D016014:Linear Models; D008253:Macaca mulatta; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism
chemical_list: D005227:Fatty Acids; D010455:Peptides; D003903:Deuterium; D017666:Deuterium Oxide