Determination of 13C Enrichment at Specific Positions of Plasma Glucose by Gas Chromatographic-Mass Spectrometric Analysis of the 1,2:5,6-Diisopropylidene-3-O-Acetyl-α-Furanosyl Derivative

Abstract To determine 13 C enrichment at specific positions of glucose, plasma samples (50-100 μl) were deproteinized using ethyl alcohol, centrifuged, and evaporated to dryness, and the residues were derivatized with acetone/H 2 SO 4 (100/1, v/v). After acetylation, the 1,2:5,6-diisopropylidene-3- O -acetyl-α-furanosyl derivative was analyzed by gas chromatography-mass spectrometry. The method was tested using mixtures of natural and 13 C-labeled glucose molecules. Ions at m / z 113, 114, 143, 144, 287, and 288, corresponding to the C 2 -C 4 , C 1 -C 4 , and C 1 -C 6 parts of natural or 13 C-labeled glucose molecule, respectively, were selectively monitored to determine the 13 C content of these fragments. This method was applied to the study of plasma glucose-labeling pattern following the infusion of [3- 13 C]lactic acid at tracer doses. Assuming equilibration of label through triose-phosphate isomerization in vivo , we determined the 13 C enrichment of carbons 1, 2, and 3 of glucose; the values for E 1 / E 3 and E 2 / E 1 ratios of 8.1 and 0.77, respectively, are consistent with those obtained in studies using radioactive tracers.