Mass Spectrometry Core Laboratory

1
Mass Spectrometry Core Laboratory Director
Brian R Walker Technician Wendy Barron Site 2nd
floor of Out-Patient Building, Western General
Hospital
).

• Stable isotope tracers
• Conventional clinical chemistry tests measure
  concentrations of hormones, small molecules,
  lipids etc in body fluids.
• Concentrations tell us how much is present, but
  not the rate of flux, i.e. how much is being
  released into, and cleared from, the fluid
  concerned.
• Clinical researchers often need to know the
  production (Ra) and clearance (Rd) rates of
  hormones and chemicals in body compartments. This
  requires a tracer.
• A tracer is a labelled version of the molecule of
  interest. Labelling can be radioactive or
  non-radioactive (ie stable, eg 2H, 13C, 15N,
  18O). GCMS is needed to measure non-radioactive
  stable isotopomers.

• What is a Mass Spectometer?
• A mass spectrometer separates molecules according
  to their charge and mass.
• Every molecule has a characteristic pattern of
  charges at different masses. A mass spectrometer
  is therefore highly specific.
• To make the instrument sensitive, samples can be
  prepared so that molecules accept more charges
  (after derivatisation) and are separated from
  each other (by sample extraction and/or
  chromatography).

• What is a Mass Spectometer used for?
• Measurement of concentrations of a wide variety
  of known molecules, from gases to proteins.
• Characterisation of unknown molecules.
• Discriminating isotopomers of molecules. Most
  elements have naturally abundant atoms of several
  different masses. Some are radioactive, but many
  are not. Non-radioactive isotopes are called
  stable isotopes (eg 2H2 deuterium heavy
  hydrogen 13C Carbon 13). Organic molecules
  will sometimes contain atoms of higher mass. The
  mass spectrometer can detect these and measure
  the proportion of each mass. This is crucial to
  use of tracers in clinical research.

At steady state Ra rate of tracer infusion -
(rate of tracer infusion) enrichment Rd Ra
(rate of tracer infusion) Enrichment is measured
by the proportion of molecules carrying the label
by mass spectrometry
Distinguishing isotopomers of cortisol (F).
Endogenous F is detected by monitoring for an ion
with mass/charge ratio of 605. This is
distinguishable from infused labelled d4F and its
metabolite D3F only by differences in mass, thus
allowing calculation of enrichment
Example 1 Concentration 10 dots/box Enrichment
20 Ra (1/0.2)-1 4 dots/min Rd 5 dots/min
Tracer infusion _at_1 dots/min
Rd of hormone and tracer
Ra of hormone _at_? dot/min
POOL
Example 2 Concentration 10 dots/box Enrichment
40 Ra (1/0.4)-1 1.5 dots/min Rd 2.5
dots/min
Tracer infusion _at_1 dot/min
Rd of hormone and tracer
Ra of hormone _at_? dot/min
POOL

• Uses of GCMS in the Clinical Research Facility
• Current
• Glucose production and clearance (2H2-glucose)
• Gluconeogenesis (13C6-glucose)
• Cortisol metabolism (2H4-cortisol, d4F)
• Future
• Protein synthesis
• NO synthesis
• Fatty acid turnover
• etc