Simplified analysis of acetaminophen glucuronide for quantifying gluconeogenesis and glycogenolysis using deuterated water.

Measurement of acetaminophen glucuronide (AG) (2)H enrichment from deuterated water ((2)H2O) by (2)H nuclear magnetic resonance (NMR) analysis of its monoacetone glucose (MAG) derivative provides estimation of gluconeogenic and glycogenolytic contributions to endogenous glucose production (EGP). However, AG derivatization to MAG is laborious and unsuitable for high-throughput studies. An alternative derivative, 5-O-acetyl monoacetone glucuronolactone (MAGLA), was tested. Eleven healthy subjects ingested (2)H2O to 0.5% body water enrichment and 500 mg of acetaminophen. Plasma glucose and urinary glucuronide positional (2)H enrichments were measured by (2)H NMR spectroscopy of MAG and MAGLA, respectively. A Bland-Altman analysis indicated agreement at the 95% confidence level between glucose and glucuronide estimates.

Sources of hepatic glucose production by 2H2O ingestion and Bayesian analysis of 2H glucuronide enrichment.

The contribution of gluconeogenesis to hepatic glucose production (GP) was quantified after (2)H(2)O ingestion by Bayesian analysis of the position 2 and 5 (2)H-NMR signals (H2 and H5) of monoacetone glucose (MAG) derived from urinary acetaminophen glucuronide. Six controls and 10 kidney transplant (KTx) patients with cyclosporine A (CsA) immunosuppressant therapy were studied. Seven KTx patients were lean and euglycemic (BMI = 24.3 +/- 1.0 kg/m(2); fasting glucose = 4.7 +/- 0.1 mM) while three were obese and hyperglycemic (BMI = 30.5 +/- 0.7 kg/m(2); fasting glucose = 7.1 +/- 0.5 mM). For the 16 spectra analyzed, the mean coefficient of variation for the gluconeogenesis contribution was 10% +/- 5%. This uncertainty was associated with a mean signal-to-noise ratio (SNR) of 79:1 and 45:1 for the MAG H2 and H5 signals, respectively. For control subjects, gluconeogenesis contributed 54% +/- 7% of GP as determined by the mean and standard deviation (SD) of individual Bayesian analyses. For the lean/normoglycemic KTx subjects, the gluconeogenic contribution to GP was 62% +/- 7% (P = 0.06 vs. controls), while hyperglycemic/obese KTx patients had a gluconeogenic contribution of 68% +/- 3% (P < 0.005 vs. controls). These data suggest that in KTx patients, an increased gluconeogenic contribution to GP is strongly associated with obesity and hyperglycemia.

Sources of endogenous glucose production in the Goto-Kakizaki diabetic rat.

Plasma glucose, insulin and glucose tolerance were quantified in diabetic Goto-Kakizaki (GK) rats (342+/-45 g, n = 5) and compared with weight-matched non-diabetic Wistars (307+/-30 g, n = 8). Compared to Wistars, GK rats had higher fasting plasma insulin (219+/-50 versus 44+/-14 pmol/l, P<0.002) and glucose (9.2+/-2.3 versus 5.5+/-0.5 mmol/l, P<0.025). GK rats showed impaired glucose tolerance (IPGTT 2 h plasma glucose=14+/-1.5 versus 6.4+/-0.1 mmol/l, P<0.001). Endogenous glucose production (EGP) from glycogenolysis, phosphoenolpyruvate (PEP) and glycerol after 6 hours of fasting was quantified by a primed infusion of [U-(13)C]glucose and (2)H(2)O tracers and (2)H/(13)C NMR analysis of plasma glucose. EGP was higher in GK compared to Wistar rats (191+/-16 versus 104+/-27 mumol/kg per min, P<0.005). This was sustained by increased gluconeogenesis from PEP (85+/-12 versus 35+/-4 mumol/kg per min, P<0.02). Gluconeogenesis from glycerol was not different (20+/-3 in Wistar versus 30+/-6 mumol/kg per min for GK), and glycogenolysis fluxes were also not significantly different (76+/-23 mumol/kg per min for GK versus 52+/-19 mumol/kg per min for Wistar). The Cori cycle accounted for most of PEP gluconeogenesis in both Wistar and GK rats (85+/-15% and 77+/-10%, respectively). Therefore, increased gluconeogenesis in GK rats is largely sustained by increased Cori cycling while the maintenance of glycogenolysis indicates a failure in hepatic autoregulation of EGP.