Lactate isotopomer analysis by ¹H NMR spectroscopy: Consideration of long‐range nuclear spin–spin interactions

Abstract

Lactate is a key metabolite and its rates of cellular uptake and release, its production rates from glucose and glycogen, and its interconversion rate with pyruvate are important determinants of cellular energy production. If lactate precursors such as pyruvate and glucose are labeled appropriately with 13 C, 1 H NMR spectroscopy provides a means of quantifying lactate production from each source and allows measurement of all these rates within a single experiment. However, due to the multiplicities of the resonance lines (from nuclear spin-spin couplings) in lactate 13 C isotopomers, the 1 H NMR spectra were found to be more complex than expected, requiring determination of all spin-spin interactions in this anion. All such values were determined for lactate and its precursor pyruvate. The method was then applied to simultaneously measure the rates of exogenous lactate uptake and rates of release of glucose-, glycogen-, and pyruvate-derived lactate in the isolated perfused rat heart.

Prospects for clinical cancer metabolomics using stable isotope tracers

“…Figure 2A shows a 1D NMR spectrum of the medium 24 h after addition of uniformly labeled 13 C glucose to A549 cells. Two forms of lactate are clearly observable, namely those that contain 12 C at the methyl position, and those that contain 13 C. Indeed, the 13 C satellites demonstrate that all three carbons are 13 C (Lloyd et al 2004; Lane et al 2007), as expected for lactate deriving exclusively from uniformly labeled glucose (Lane et al 2008). The unlabeled lactate molecules must have originated from some other (unlabeled) source.…”

Quantification and identification of isotopomer distributions of metabolites in crude cell extracts using 1H TOCSY

“…Extracts are often done to complement in vivo studies Fan et al, 1986Fan et al, , 2005, in which 2D 1 H NMR detection is an attractive complement to 1D direct detect 13 C experiments (Carvalho et al, 1998;Burgess et al, 2001). Furthermore, isotopomers can be simultaneously analyzed in 1 H detected experiments by observing the 13 C satellite peaks (Lewandowski et al, 1991;Vanzijl et al, 1993;Zwingmann et al, 2001;Lloyd et al, 2004).…”

Stable isotope-resolved metabolomics and applications for drug development

“…Such approaches have been successfully applied to many different disease states (Carvalho, Babcock et al 1999; Zwingmann, Richter-Landsberg et al 2001; Lu, Mulder et al 2002; Mason, Petersen et al 2002; Burgess, Weis et al 2003; Mason, Petersen et al 2003; Wang, Lloyd et al 2003; Zwingmann and Leibfritz 2003; Anousis, Carvalho et al 2004; Carvalho, Rodrigues et al 2004; Cline, LePine et al 2004; Des Rosiers, Lloyd et al 2004; Goddard, Mason et al 2004; Jin, Jones et al 2004; Mason and Rothman 2004; Sherry, Jeffrey et al 2004; Des Rosiers and Chatham 2005; Jin, Jones et al 2005; Patel, de Graaf et al 2005; Wang, Lloyd et al 2005; Hausler, Browning et al 2006; Hyder, Patel et al 2006; DeBerardinis, Mancuso et al 2007; Mason, Petersen et al 2007; Fan and Lane 2008; Morrish, Neretti et al 2008; Morrish, Noonan et al 2010). However, the sensitivity and information content is significantly less than using 2D methods with proton detection (Fan 1996; Fan, Higashi et al 1997; Carvalho, Jeffrey et al 1998; Carvalho, Zhao et al 2001; Lloyd, Zeng et al 2004; Lane and Fan 2007). …”