Determination of carbon-13 labeled lactate in blood by gas chromatography/mass spectrometry

Tserng, Kou‐Yi; Gilfillan, Carol; Kalhan, Satish C.

doi:10.1021/ac00267a049

Cited by 56 publications

(36 citation statements)

References 28 publications

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“…The deuterium enrichment of the lactate was determined by gas chromatograph-mass spectrometric analysis (23). The reproducibility of the mass spectrometric analysis has been reported previously (23). Preparation of lactate from glucose by the coupled enzymatic reaction gave a coefficient of variation of 13-15% at the low, < 0.5%, enrichments.…”

Section: Methodsmentioning

confidence: 85%

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Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

et al. 1995

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A method is introduced for estimating the contribution of gluconeogenesis to glucose production. 2H20 is administered orally to achieve 0.5% deuterium enrichment in body water. Enrichments are determined in the hydrogens bound to carbons 2 and 6 of blood glucose and in urinary water. Enrichment at carbon 6 of glucose is assayed in hexamethylenetetramine, formed from formaldehyde produced by periodate oxidation of the glucose. Enrichment at carbon 2 is assayed in lactate formed by enzymatic transfer of the hydrogen from glucose via sorbitol to pyruvate. The fraction gluconeogenesis contributes to glucose production equals the ratio of the enrichment at carbon 6 to that at carbon 2 or in urinary water. Applying the method, the contribution of gluconeogenesis in healthy subjects was 23-42% after fasting 14 h, increasing to 59-84% after fasting 42 h. Enrichment at carbon 2 to that in urinary water was 1.12±0.13. Therefore, the assumption that hydrogen equilibrated during hexose-6-P isomerization was fulfilled. The 3H/14C ratio in glucose formed from [3-3H,3-'4C]lactate given to healthy subjects was 0.1 to 0.2 of that in the lactate. Therefore equilibration during gluconeogenesis of the hydrogen bound to carbon 6 with that in body water was 80-90% complete, so that gluconeogenesis is underestimated by 10-20%. Glycerol's contribution to gluconeogenesis is not included in these estimates. The method is applicable to studies in humans of gluconeogenesis at safe doses of 21H20. (J. Clin. Invest. 1995. 95:172-178.)

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Section: Methodsmentioning

confidence: 85%

“…). The deuterium enrichment of the lactate was determined by gas chromatograph-mass spectrometric analysis (23). The reproducibility of the mass spectrometric analysis has been reported previously (23).…”

Section: Methodsmentioning

confidence: 93%

Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

et al. 1995

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“…Lactate was extracted from 0.2 mL of plasma by ethyl acetate after acidification with HCl. Lactate was then derivatized to its propylamine-heptafluorobutyrate (HFB) form, and the m/z 328 to m/z 331 (carbons 1 to 3 of lactate, chemical ionization) was monitored for the detection of m1 (recycled lactate through the PPC) and m2 (lactate produced by glycolysis) for the estimation of pentose cycle activity (Tserng et al, 1984). Mass spectral data were obtained on the HP5973 mass selective detector connected to an HP6890 gas chromatograph.…”

Section: Sample Analysismentioning

confidence: 99%

Increased Pentose Phosphate Pathway Flux after Clinical Traumatic Brain Injury: A [1,2-¹³C₂]glucose Labeling Study in Humans

Dusick

Glenn

Lee

et al. 2007

J Cereb Blood Flow Metab

115

134

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Patients with traumatic brain injury (TBI) routinely exhibit cerebral glucose uptake in excess of that expected by the low levels of oxygen consumption and lactate production. This brings into question the metabolic fate of glucose. Prior studies have shown increased flux through the pentose phosphate cycle (PPC) during cellular stress. This study assessed the PPC after TBI in humans.[1,2-13 C 2 ]glucose was infused for 60 mins in six consented, severe-TBI patients (GCS < 9) and six control subjects. Arterial and jugular bulb blood sampled during infusion was analyzed for 13 C-labeled isotopomers of lactate by gas chromatography/mass spectroscopy. The product of lactate concentration and fractional abundance of isotopomers was used to determine blood concentration of each isotopomer. The difference of jugular and arterial concentrations determined cerebral contribution. The formula PPC = (m1/m2)/(3 + (m1/m2)) was used to calculate PPC flux relative to glycolysis. There was enrichment of [1,2-13 C 2 ]glucose in arterial-venous blood (enrichment averaged 16.6% in TBI subjects and 28.2% in controls) and incorporation of 13 C-label into lactate, showing metabolism of labeled substrate. The PPC was increased in TBI patients relative to controls (19.6 versus 6.9%, respectively; P = 0.002) and was excellent for distinguishing the groups (AUC = 0.944, P < 0.0001). No correlations were found between PPC and other clinical parameters, although PPC was highest in patients studied within 48 h of injury (averaging 33% versus 13% in others; P = 0.0006). This elevation in the PPC in the acute period after severe TBI likely represents a shunting of substrate into alternative biochemical pathways that may be critical for preventing secondary injury and initiating recovery.

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“…This will be done through the homogenization of samples until obtaining a perchloric acid-pH adjusted supernatant likewise to that described for the determination of G 6-P levels (Fillat et al, 1992). Once obtained this extract, samples will be treated as described in Tserng et al (1984); Lee et al (1996) and Kurland et al (2000). For this purpose samples will be taken to dryness and they will be subsequently resuspended in 400 µL of 0.5 % (w:v) hydroxylamine hydrochloride in pyridine.…”

Section: Determination Of the Rhythm Of Formation Of Intermediate Metmentioning

confidence: 99%

Spermiomics: A New Term Describing the Global Survey of the Overall Sperm Function by the Combined Utilization of Immunocytochemistry, Metabolomics, Proteomics and Other Classical Analytical Techniques

Rodríguez‐Gil¹

2012

Applications of Immunocytochemistry

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Determination of carbon-13 labeled lactate in blood by gas chromatography/mass spectrometry

Cited by 56 publications

References 28 publications

Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

Increased Pentose Phosphate Pathway Flux after Clinical Traumatic Brain Injury: A [1,2-¹³C₂]glucose Labeling Study in Humans

Spermiomics: A New Term Describing the Global Survey of the Overall Sperm Function by the Combined Utilization of Immunocytochemistry, Metabolomics, Proteomics and Other Classical Analytical Techniques

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Determination of carbon-13 labeled lactate in blood by gas chromatography/mass spectrometry

Cited by 56 publications

References 28 publications

Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

Increased Pentose Phosphate Pathway Flux after Clinical Traumatic Brain Injury: A [1,2-13C2]glucose Labeling Study in Humans

Spermiomics: A New Term Describing the Global Survey of the Overall Sperm Function by the Combined Utilization of Immunocytochemistry, Metabolomics, Proteomics and Other Classical Analytical Techniques

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Product

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Increased Pentose Phosphate Pathway Flux after Clinical Traumatic Brain Injury: A [1,2-¹³C₂]glucose Labeling Study in Humans