METABOLIC LOSS OF DEUTERIUM FROM ISOTOPICALLY LABELED GLUCOSE

The isotopically substituted molecule (6-C-13, 1, 6, 6-H-2(3))glucose was evaluated to determine whether metabolic H-2 loss would prevent its use in quantitating pentose phosphate pathway (PPP) activity. PPP activity causes the C1 of glucose to be lost as CO2, while C6 can appear in lactate. H-2 NMR analysis of the lactate produced from this glucose can distinguish (3-H-2)-lactate (from C1 of glucose) from (3-C-13, 3, 3-H-2(2))lactate (from C6 of glucose). H-2 NMR spectroscopic analysis of medium containing (6-C-13, 1, 6, 6-H-2(3))glucose after incubation with cultured rat 9L glioma cells suggested a 30.8 +/- 2.1% PPP activity as compared with 6.0 +/- 0.8% from separate, parallel incubations with (1-C-13)glucose and (6-C-13)glucose. Subsequent experiments with other isotopically labeled glucose molecules suggest that this discrepancy is due to selective loss of H-2 from the C1 position of glucose, catalyzed by phosphomannose isomerase. Failure to consider H-2 exchange from the C1 and C6 positions of glucose can lead to incorrect conclusions in metabolic studies utilizing this and other deuterated or tritiated glucose molecules.