A doubly labelled tracer molecule of glucose, [6,6-2HJglucose, has been used to measure the rate of glucose appearance in blood due to endogenous glucose production (by the liver and kidneysj.l,2 Considered \u27non-recirculating\u27, this tracer generally provides a realistic estimate of glucose kinetics.2,3 Used as a dual tracer in conjunction with [l-13C]glucose, the extent of glucose recycling can be quantitated.2-4 but the extent of possible recirculation of a single deuterium atom on newly formed glucose must then be considered

Brooks, George A.

Casazza, Gretchen A.

Colberg, Sheri R.

Horning, Michael A.

English

Old Dominion University

Old Dominion UniversityODU Digital CommonsEducational Foundations & Leadership FacultyPublications Educational Foundations & Leadership200Recycling of Deuterium From DideuteratedGlucose During Moderate ExerciseMichael A. HorningSheri R. ColbergOld Dominion UniversityGretchen A. CasazzaGeorge A. BrooksFollow this and additional works at: https://digitalcommons.odu.edu/efl_fac_pubsPart of the Exercise Physiology Commons, and the Exercise Science CommonsThis Article is brought to you for free and open access by the Educational Foundations & Leadership at ODU Digital Commons. It has been acceptedfor inclusion in Educational Foundations & Leadership Faculty Publications by an authorized administrator of ODU Digital Commons. For moreinformation, please contact digitalcommons@odu.edu.Repository CitationHorning, Michael A.; Colberg, Sheri R.; Casazza, Gretchen A.; and Brooks, George A., "Recycling of Deuterium From DideuteratedGlucose During Moderate Exercise" (200). Educational Foundations & Leadership Faculty Publications. 52.https://digitalcommons.odu.edu/efl_fac_pubs/52Original Publication CitationHorning, M. A., Colberg, S. R., Casazza, G. A., & Brooks, G. A. (2000). Recycling of deuterium from dideuterated glucose duringmoderate exercise. Annals of Clinical Biochemistry, 37, 540-542. doi:10.1258/0004563001899546Short Report Ann Clin Biochem 2000; 37: 540-542Recycling of deuterium from dideuterated glucose duringmoderate exerciseMichael A Horning', Sheri R Colberg-, Gretchen A Casazza'and George A Brooks'From the' Exercise Physiology Laboratory, Department ofIntegrative Biology, University ofCalifornia,Berkeley, CA 94720, and the 2Department of Exercise Science, Physical Education, and Recreation,Old Dominion University, Norfolk, VA 23529, USAA doubly labelled tracer molecule of glucose,[6,6-2HJglucose, has been used to measure therate of glucose appearance in blood due toendogenous glucose production (by the liver andkidneysj.l? Considered 'non-recirculating', thistracer generally provides a realistic estimate ofglucose kinetics.s-' Used as a dual tracer inconjunction with [l.13C]glucose, the extent ofglucose recycling can be quantitated.r" but theextent of possible recirculation of a singledeuterium atom on newly formed glucose mustthen be considered.The purpose of this study was to examinewhether recirculation of a singly-deuteratedmolecule ([6-2H)glucose) does occur to a sig-nificant extent during a constant infusion ofdideuterated tracer ([6,6.2HJglucose) and toquantify the extent to which it may causeoverestimation of the glucose (M + I) signaland, therefore, glucose recycling determinedwith dual glucose tracers. Twelve men (age28·0± 1·4 years) were studied in the post-absorptive state. Using a forearm vein catheter,a primed, continuous infusion of [6,6-2HJglucosewas administered at a rate of 2 rug/min during90min of rest and 6 mg/min during 60min ofcycle exercise at 50% of the pre-determinedoxygen consumption (\10 2) peak in six subjects,while the other six completed the same protocolwithout infusion of any tracer to determine thecontribution to the (M + I) signal from naturalisotopic abundance (both [l3C] and [2HJsources).Arterialized blood samples were obtainedfrom a heated hand vein in the contra-lateralarm from tracer infusion. These were analysedfor glucose isotopic enrichment (measuringatoms per cent excess; APE) by gas chromato-graph mass spectroscopy (GC(MS; HewlettCorrespondence: Dr Sheri R Colberg.Reprint requests: Dr G A Brooks.540Packard GC Model 5890, Series II, and an MSModel 5989A; Hewlett Packard, Palo Alto CA,USA) in the chemical ionization (CI) mode. Apenta-acetate derivative of glucose allowed ionsto be selectively monitored at mlz 331·2, 332·2,and 333,2.1Steady-state conditions for enrichment of the(M + 2) tracer were achieved during the final30min of rest and exercise (l·795±0·181 and2·429± 0,198, respectively). The increase inenrichment of the (M + I) signal was0·107± 0·053% above baseline at rest and0·203± 0·043% during moderate exercise duringinfusion of the glucose tracer (see Fig. I). Incontrol subjects, the (M + I) signal at each timepoint was unchanged from baseline values.Data from the current study were used torecalculate glucose recycling rates from ourprevious work." The corrected values were6·88 ± 3·28% during rest instead of 8·60± 3·85,and 9·93± 5·21% during exercise instead of1l·90±5·22, representing a 20·0 and 17·0%previous overestimation of recycling during restand exercise conditions, respectively.In this study, infusion of a dideuteratedglucose isotopic tracer resulted in a measurableamount of tracer recycling into newly formeddeuterated glucose, thus increasing the (M + I)signal. This recycling caused underestimation oftotal glucose production calculated previouslywith a [l-13C]glucose tracer as well as over-estimation of total glucose recycling calculatedfrom the simultaneous use of [1_I3C]glucose and[6,6-2H]glucose under this protocol.In many studies, both stable isotope carbonand hydrogen glucose tracers are infusedsimultaneously in order to assess glycolyticcarbon recycling through gluconeogenesis inthe liver.? In the gluconeogenic process, labelledlactate is reconverted to glucose, thereby raisingthe measured enrichment of the carbon tracer bythe amount of recycled [13C] label. Deuterium onDeuterium recycling 5410·40·3W 0·2a.sIIIIII2l 0·1)(Q)E~ 0Q)0-IIIE0:(-0·1-0·2....... Infusion---No infusion-0·3-100 -50 0 50 100Time (min)FIGURE I. Increase in (M + I) enrichment (APE) with [6.6- 2HJglucose infusion during rest (- 90 to Omin ) andexercise (0 to 60 min). Data are given as mean ± standard error oj"the meanfor six subjects at each time point.the sixth carbon is not lost during glycolysis persc, but rather at two possible sites duringgluconeogenesis. One deuterium is lost in thepyruvate carboxylase reaction converting pyr-uvate to oxaloacetate.' A deuterium may also belost from the fifth carbon in the equilibrationwith the mitochondrial hydrogen pool duringequilibration between oxaloacetatc, malate andfumarate in the Krebs cycle. If only onedeuterium is lost from the dideuterated tracerinstead of both, then newly formed glucose willactually be [6-2H]glucose instead of unlabelledglucose, thus increasing the (M + I) signal as ourdata indicate. In accordance with our results,Kathan" presented data from one subjectshowing an increase in the (M + I) enrichmentin blood during a constant infusion of [6,6-2H]glucose at rest, indicating recycling of asingle deuterium atom.In summary, our data demonstrate thatrecirculation of a singly-deuterated glucoseduring a constant infusion of [6,6-2H]glucosewill cause an overestimation of the glucose(M + I) signal. While all calculated recyclingvalues from our previous study" were in rangecomparable to published research using similartracers at rest;' we have now demonstrated thatglucose recycling in our study was actuallyoverestimated with the simultaneous use of thesetwo tracers. These results arc applicable to allstudies incorporating dual isotopic tracers whereone of the tracers is [6,6-2H]glucose and theother tracer increases the (M + I) signal. Ournew data allow for the actual quantification andcorrection of recycling under these specificconditions and rates of tracer infusion. How-ever, another solution to this problem may be touse an alternate methodology for analysis thatmay be able to effectively account for therecycling of deuterium.AcknowledgementsThis work was supported by a grant from theNational Institutes of Health.REFERENCE."lBrooks GA. ButterfieldGE, Wolfe RR. Groves BM.Mazzeo RS. Sulton JR. Wolfcl EE, Reeves JT.Increased dependency on hlood glucose afteracclimatization to 4,300111. Appl Phvsiol 1991; 70:919 27All" C!ill Biochem 2000: 37542 Horning et al.2 Kalhan SC, Bier OM, Savin SM, Adam PAJ.Estimation of glucose turnover and 13C recyclingin the human newborn by simultaneous [1-13C]glu-cose and [6,6-2H21glucose tracers. J Clin EndocrinolMetab 1980;50: 456--603 Reinauer H, Gries FA, Hubinger A, Knode 0,Severing K, Susanto F. Determination of glucoseturnover and glucose oxidation rates in man withstable isotope tracers. J Clin Chern Clin Biochern1990;28: 505-11Ann Clin Biochem 2000: 374 Colberg SR, Casazza GA, Horning MA, BrooksGA. Increased dependence on blood glucose insmokers during rest and sustained exercise. J ApplPhysiol1994; 76: 26-325 Kalhan Sc. Stable isotope tracers for studies ofglucose metabolism. J Lab Clin Med 1990;116: 615-22Accepted for publication 1 April 2000

Recycling of Deuterium From Dideuterated Glucose During Moderate Exercise

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Recycling of Deuterium From Dideuterated Glucose During Moderate Exercise

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