Optimized metabolite extraction from blood serum for 1H nuclear magnetic resonance spectroscopy.
Blood serum is commonly used for clinical diagnostics because its protein composition bears a wealth of information about the health of an organism. More recently the analysis of the small molecule composition, the metabolome, has received increased attention because the metabolite composition is influenced by many diseases, by the administration of drugs and toxins, and by the diet and life style of an individual. When nuclear magnetic resonance spectroscopy is used as an analytical tool it is often preferable to remove catalytically active proteins, in particular for longer measurements, because metabolite concentrations are otherwise in constant flux. Here we have compared different protocols for the separation of proteins and metabolites, including precipitation methods and ultrafiltration. Whereas most extraction methods involving protein precipitation deplete some metabolites, ultrafiltration is superior in retaining metabolite concentrations and offers excellent reproducibility. We also describe a new method to recover the hydrophobic fraction for ultrafiltration with good reproducibility.
Tiziani
Stefano
S
Henry Wellcome Building for Biomolecular NMR Spectroscopy, CR UK Institute for Cancer Studies, University of Birmingham, Birmingham B15 2TT, UK.
Emwas
Abdul-Hamid
AH
Lodi
Alessia
A
Ludwig
Christian
C
Bunce
Christopher M
CM
Viant
Mark R
MR
Günther
Ulrich L
UL
D000183:Actihaemyl; D000818:Animals; D001774:Blood Chemical Analysis / Q000379:methods*; D001798:Blood Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D002417:Cattle; D011232:Chemical Precipitation; D006859:Hydrogen / Q000737:chemistry; D057927:Hydrophobic and Hydrophilic Interactions; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D015203:Reproducibility of Results; D014462:Ultrafiltration
D001798:Blood Proteins; C088695:deproteinized calf blood serum; D000183:Actihaemyl; D006859:Hydrogen