Quantifying intermediary metabolites in whole blood after a simple deproteinization step with sulfosalicylic acid.
We assessed the reproducibility, recovery, and stability of several circulating metabolites--glucose, pyruvate, lactate, alanine, glutamate, glutamine, 3-hydroxybutyrate, acetoacetate, and glycerol--in the presence of sulfosalicylic acid (SSA), which was used to deproteinize blood. The assays, which involved reactions linked to NADH/NAD+, were carried out at 37 degrees C and measured at 355 nm with a Cobas-Bio centrifugal analyzer. The intra- and interbatch CVs were less than 2.1%, except for the interbatch CV for 3-hydroxybutyrate at low concentration (15-30 mumol/L), which was 5.4%. Analytical recovery of metabolites added to blood ranged from 96.4% to 103.0%. Of the metabolites studied, all were stable at -20 degrees C for 90 days in the SSA-blood extract, except for glutamine and acetoacetate, which progressively decreased with time. We conclude that these nine circulating metabolites can be satisfactorily measured after a single deproteinizing step with SSA. This single-step procedure has several advantages over many of the currently used methods.
Khan
K
K
MRC Dunn Nutrition Unit, University of Cambridge, U.K.
Blaak
E
E
Elia
M
M
D000596:Amino Acids / Q000097:blood; D001557:Benzenesulfonates; D001774:Blood Chemical Analysis; D001798:Blood Proteins / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D009247:NADH, NADPH Oxidoreductases; D015203:Reproducibility of Results; D012459:Salicylates / Q000494:pharmacology*
D000596:Amino Acids; D001557:Benzenesulfonates; D001798:Blood Proteins; D012459:Salicylates; D009247:NADH, NADPH Oxidoreductases; C003366:sulfosalicylic acid