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Measurement of gluconeogenesis by deuterated water: the effect of equilibration time and fasting period

¹Department of Endocrinology and Metabolism, and ²Department of Clinical Chemistry, Laboratory of Endocrinology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands

Submitted 21 June 2005 ; accepted in final form 8 January 2006

Fasting gluconeogenesis (GNG) is often quantified using the ²H₂O technique, which is based on plasma ²H₂O enrichment and ensuing enrichment of plasma glucose at the C5 and C2 positions. Fractional (fr)GNG can be calculated using the ratio of C5 to C2 enrichment or the ratio of C5 to plasma ²H₂O enrichment. For the latter, equilibration of ²H₂O and C2 is required. The optimal equilibration period of ²H₂O and C2 remains to be elucidated. In six healthy male subjects fasted for 18 h, we studied the effects of 3-, 5-, and 15-h ²H₂O incubation periods on 1) the equilibration of plasma ²H₂O and C2 glucose enrichment, 2) the measurement of frGNG, and 3) C5 labeling of hepatic glycogen after 1 mg of glucagon administration. After 3-h ²H₂O incubation, plasma ²H₂O and C2 were not equilibrated, frGNG C5/²H₂O and C5/C2 were also different as was gluconeogenesis calculated with C5/²H₂O and C5/C2. After 5- and 15-h ²H₂O incubation, plasma ²H₂O and C2 were equilibrated, and frGNG C5/²H₂O and C5/C2 were similar, as was GNG calculated with C5/²H₂O and C5/C2. After glucagon administration, no difference of C5 enrichment was found between 3, 5, and 15 h of ²H₂O incubation. In conclusion, for reliable measurement of GNG in healthy subjects with C5/²H₂O incubation periods longer than 3 h are required. After 5- and 15-h ²H₂O incubation, GNG can be reliably measured with C5/²H₂O. Gluconeogenetic labeling of glycogen did not affect the results after 3, 5, or 15 h of ²H₂O incubation.

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