Hepatic UDP‐glucose ¹³C isotopomers from [U‐¹³C]glucose: A simple analysis by ¹³C NMR of urinary menthol glucuronide

Abstract

Menthol glucuronide was isolated from the urine of a healthy 70-kg female subject following ingestion of 400 mg of peppermint oil and 6 g of 99% [U-13 C]glucose. Glucuronide 13 C-excess enrichment levels were 4 -6% and thus provided high signalto-noise ratios (SNRs) for confident assignment of 13 C-13 C spincoupled multiplet components within each 13 C resonance by 13

Sources of hepatic glycogen synthesis during an oral glucose tolerance test: Effect of transaldolase exchange on flux estimates

“…Thus, the probability of forming [U- 13 C]glucuronide isotopomers by indirect pathway is negligible (6). Hence, quantification of the fractional 13 C-enrichment of [U- 13 C]glucuronide divided by the 13 C-enrichment of the plasma [U- 13 ϫ 100(%) [3] Indirect pathway ϭ 100-direct pathway(%) [ Transaldolase (TA) catalyzes the exchange of the 4-, 5-, and 6-carbon fragment of fructose-6-phosphate (F6P) and glyceraldehyde-3-phosphate (G3P), this exchange being independent of oxidative pentose phosphate pathway (PPP) flux (16) (Fig.…”

“…In humans, when a xenobiotic agent (i.e., acetaminophen or peppermint oil) is provided, the enrichment distributions of hepatic UDP-glucose are also reflected in the respective urinary glucuronide conjugate (i.e., acetaminophen or menthol glucuronide (3)(4)(5)(6)) because the glucose moiety of UDP-glucose is converted to glucuronide by a single dedicated pathway. To date, contributions of direct and indirect pathways to hepatic glycogen synthesis have been estimated with [1- 13 C]glucose, [1][2][3][4][5][6][7][8][9][10][11][12][13][14] C]glucose or deuterated water ( 2 H 2 O) in conjunction with acetaminophen.…”

“…Direct and indirect pathway contributions were estimated by 13 C and 2 H nuclear magnetic resonance (NMR) analysis of plasma glucose and urinary menthol glucuronide excess enrichments after ingestion of peppermint oil and an oral glucose load enriched with stable isotope tracers. Compared with the rather laborious purification and derivatization methods required for analysis of acetaminophen glucuronide (12,13), menthol glucuronide can be directly analyzed by NMR spectroscopy after a simple isolation protocol (5,6).…”

“…The main disadvantages of [1-13 C]glucose are that 1) a relatively large amount (ϳ10 g) is required to generate sufficient excess enrichment of the UDP-glucose and glucuronide pools so that glucuronide 13 C-enrichment can be reliably quantified above the background 1.11% 13 C-enrichment level, and 2) the effects of glucose recycling on glucose and glucuronide 13 C-enrichments have to be accounted for because [1][2][3][4][5][6][7][8][9][10][11][12][13] Therefore, the use of two tracers that effectively eliminate background effects and are nonrecyclable was focused on here. With the first tracer, [U- 13 C]glucose, the background 13 C-contribution becomes negligible because of the tiny probability-(0.011) 6 -of a naturally occurring glucose molecule containing six 13 C-atoms. The [U- 13 C]glucose precursor and [U- 13 C]glucuronide product generate resolvable signals in the 13 C spectrum, allowing direct quantification by 13 C NMR, and whole-body glucose recycling does not regenerate significant levels of [U- 13 C]glucose (6).…”

“…With the first tracer, [U- 13 C]glucose, the background 13 C-contribution becomes negligible because of the tiny probability-(0.011) 6 -of a naturally occurring glucose molecule containing six 13 C-atoms. The [U- 13 C]glucose precursor and [U- 13 C]glucuronide product generate resolvable signals in the 13 C spectrum, allowing direct quantification by 13 C NMR, and whole-body glucose recycling does not regenerate significant levels of [U- 13 C]glucose (6). Moreover, this tracer is already widely used in plasma kinetic studies of glucose disposal and is less expensive than [1- 13 13 C]glucose metabolism or vice versa, enrichment from both tracers can be analyzed from a common blood or urine sample and therefore studied in the same experiment.…”