F. Mosora

Glucose Naturally Labeled with Carbon-13: Use for Metabolic Studies in Man

The ratio of carbon-13 to carbon-12 is much higher in most commerical preparations of glucose used for oral glucose tolerance tests than it is in carbon dioxide in expired air. This recent discovery provided a novel and potentially significant means of studying glucose metabolism. The changes in the ratio of carbon-13 to carbon-12 in carbon dioxide expired after oral glucose administration were determined by mass spectrometry. In six healthy male volunteers, the administration of glucose resulted in a marked, reproducible rise in the isotopic ratio in expired carbon dioxide; the ratio reached its maximum at 4 hours and then declined progressively.

Quantitative Evaluation of the Oxidation of an Exogenous Glucose Load Using Naturally Labeled 13C-Glucose

Maize glucose was used as a natural tracer for studies of metabolism. It is richer in 13C than common vegetables and foods derived from these, and the C02 formed from it is consequently richer in 13C than the CO2 expired by man fed on a diet of common vegetables. The quantitative results, obtained by measurement of delta 13C of the expired CO2 and of VCO2 during the oxidation of an exogenous glucose load (about 100 g) in eight normal subjects over 7 hr, have shown a consumption of 28.64 +/- 1.44 g of glucose (mean +/- SEM), which represents about 30% of the load given. A comparison is made with the results obtained from other methods and the originality and usefulness of this new quantitative procedure is outlined.

Effect of glucose ingestion on energy substrate utilization during prolonged muscular exercise

SummaryThe distribution of substrates utilized during prolonged exercise was investigated in normal human volunteers with and without ingestion of 100 g exogenous glucose. The energy provided by protein oxidation was derived from urinary nitrogen excretion and the total energy provided by carbohydrates and lipids was calculated from respiratory quotient (RQ) determinations. The contribution of exogenous glucose to the energy supply was determined by an original procedure using “naturally labeled 13C-glucose” as metabolic tracer. Protein oxidation provided between 1 and 2% of the total energy requirement; this amount was not affected by glucose ingestion. In the absence of exogenous glucose ingestion, carbohydrate were progressively replaced by lipids as source of energy. Exogenous glucose contributed markedly to total carbohydrate oxidation and decreased the percentage of energy derived from lipids. In addition, ingestion of exogenous glucose resulted in a significant economy of endogenous carbohydrates and permitted to prolong the duration of exercise.

Glucose Utilization During Exercise in Normal and Diabetic Subjects: The Role of Insulin

Due to selective isotopic effects occurring during photosynthesis, certain natural sugars are enriched in 13C Using such “naturally labeled 13C-glucose,” we studied glucose oxidation during exercise in seven normal volunteers and in six insulin-dependent diabetics after an overnight fast. In the diabetics, blood glucose was monitored the night before the test and adjusted to about 100 mg/dl by intravenous insulin infusion. The insulin infusion was withheld 15 min before exercise in four diabetics and maintained at 0.9 U/h for 2 h; then it was maintained at 0.6 U/h for 2 h in five diabetics. Three patients underwent both tests. All subjects exercised on a treadmill for 4 h at about 45% of their max. After 15 min adaptation, all received 100 g 13C-labeled glucose orally. Total glucose oxidation was derived from non-protein RQ and exogenous glucose oxidation evaluated as previously described. The diabetics had no residual B-cell function as indicated by negligible plasma C-peptide values and a lack of Cpeptide response to the oral glucose challenge. Total glucose oxidation averaged 230 ± 14 g/4 h in the normal subjects. It was similar (238 ± 19 g/4 h) in the diabetics receiving an intravenous insulin infusion, but decreased to 176 ± 14 g/4 h when no insulin was infused. Exogenous glucose oxidation was 92 ± 3 g/4 h and 84 ± 8 g/4 h (not statistically different) in the controls and in the insulin-infused diabetics, respectively. It was 43 ± 11 g/4 h in the diabetics exercising without being infused with insulin. We conclude that (1) in well-insulinized diabetic patients, prolonged muscular exercise can be performed under metabolic conditions which are basically similar to those of normal subjects; (2) during prolonged exercise, well-insulinized diabetic patients are able to oxidize up to 85–90% of a 100-g exogenous glucose load given orally and oral glucose can thus be ingested during prolonged exercise in well-controlled juvenile insulintreated diabetics; (3) even in the absence of insulin administration during exercise, juvenile diabetics who start exercising when blood glucose is near normal are able to perform a 4-h exercise at 45–50% of their max. Under these conditions, however, they are unable to utilize more than 40–45% of a 100-g glucose load given orally. They rely more upon lipid stores than the normal subjects or the well-insulinized diabetics.

Naturally labeled 13C-glucose. Metabolic studies in human diabetes and obesity.

The 13C/12C ratio in most commercial preparations of glucose used in clinical investigation is much higher than that of expired air. Variations in expired air 13C, after glucose load, are indicative of the catabolism of this exogenous glucose. The changes in the 13C/12C ratio of CO2 in expired air after oral administration of glucose were determined by mass spectrometry. Results in eleven healthy volunteers and seven obese subjects with normal oral glucose tolerance tests (OGTT) are reported. In all cases, the administration of glucose resulted in a marked rise in the 13C/12C ratio of expired CO2, reaching its maximum at the fourth hour and then declining progressively. In seven obese patients with chemical diabetes and in five insulin-dependent diabetics, the 13C/12C ratio of expired CO2 during OGTT was significantly reduced, in comparison with the results obtained from the control groups. This study demonstrates the potential of this procedure using “naturally labeled” 13C glucose for the metabolic studies in man as anticipated from previous studies of Duchesne and his coworkers.

Utilization of Oral Sucrose Load During Exercise in Humans: Effect of the α-Glucosidase Inhibitor Acarbose

We investigated the hormonal and metabolic response to a 100-g sucrose load given 15 min after adaptation to moderate-intensity (50% Vmaxo2) long-duration (4-h) exercise in healthy volunteers. The effect of a 100-mg dose of the α-glucosidase inhibitor Acarbose ingested with the sucrose load was also investigated. “Naturally labeled [13C] sucrose” was used to follow the conversion to expired-air CO2 of the sugar ingested by isotope-ratio mass spectrometry. Circulating hormone and metabolite data were obtained in nine subjects, and indirect calorimetry and stable isotope methodology were applied to six of them. Under placebo, 93 ± 4 g sucrose were entirely oxidized during the 4 h of exercise, total carbohydrate utilization was 235 ± 14 g, endogenous carbohydrate utilization was 142 ± 13 g, and total lipid oxidation was 121 ± 7 g. A single oral dose of 100 mg Acarbose ingested with the sucrose load did not significantly modify total carbohydrate (239 ± 2 g/4 h) or lipid (122 ± 6 g/4 h) oxidation. In contrast, sucrose oxidation was reduced to 53 ± 6 g/4 h and endogenous carbohydrate utilization increased to 186 ± 7 g/4 h. Reduction of the rises in blood glucose and fructose and of the increases in plasma insulin and C peptide under Acarbose confirmed these effects, whereas lower circulating levels of alanine suggested a higher rate of gluconeogenesis. These data show that a 100-g glucose load ingested soon after initiation of exercise is a perfect available metabolic substrate. Furthermore, the simultaneous ingestion of 100 mg Acarbose significantly reduces the availability of sucrose during exercise, a finding that has to be considered if this or other compounds with similar properties are envisaged for the treatment of diabetic patients.

Availability of glucose ingested during muscle exercise performed under acipimox-induced lipolysis blockade

This study investigated the percentage of carbohydrate utilization than can be accounted for by glucose ingested during exercise performed after the ingestion of the potent lipolysis inhibitor Acipimox. Six healthy male volunteers exercised for 3 h on a treadmill at about 45% of their maximal oxygen uptake, 75 min after having ingested 250 mg of Acipimox. After 15-min adaptation to exercise, they ingested either glucose dissolved in water, 50 g at time 0 min and 25 g at time 60 and 120 min (glucose, G) or sweetened water (control, C). Naturally labelled [13C]glucose was used to follow the conversion of the ingested glucose to expired-air CO2. Acipimox inhibited lipolysis in a similar manner in both experimental conditions. This was reflected by an almost complete suppression of the exercise-induced increase in plasma free fatty acid and glycerol and by an almost constant rate of lipid oxidation. Total carbohydrate oxidation evaluated by indirect calorimetry, was similar in both experimental conditions [C, 182, (SEM 21); G, 194 (SEM 16) g · 3 h−1], as was lipid oxidation [C, 57 (SEM 6); G, 61 (SEM 3) g · 3 h−1]. Exogenous glucose oxidation during exercise G, calculated by the changes in13C:12C ratio of expired air CO2, averaged 66 (SEM 5) g · 3 h−1 (19% of the total energy requirement). Consequently, endogenous carbohydrate utilization was significantly smaller after glucose than after placebo ingestion: 128 (SEM 18) versus 182 (SEM 21) g · 3 h−1, respectively (P < 0.05). Symptoms of intense fatigue and leg cramps observed with intake of sweet placebo were absent with glucose ingestion.In conclusion, we found glucose ingestion during 3-h exercise with lipolysis blockade could provide metabolic substrate permitting a significant sparing of endogenous carbohydrate and consequently an improvement in performance.

Oxidation of an exogenous glucose load using naturally labelled 13C-glucose

SummaryThe effect of a 14 day-administration of butylbiguanide was investigated in a group of 10 obese patients with mild-to-moderate glucose intolerance. Glucose tolerance was significantly improved, while fasting blood glucose and plasma levels of free fatty acids, insulin and glucagon remained unchanged. The estimation of the amount of the oral glucose load oxidized into CO2 was performed by means of a recently described procedure using “naturally labelled 13C-glucose” as tracer. The curves depicting the oxidation of the exogenous glucose load were similar in shape and magnitude before and after administration of the biguanide; in the latter case, however, slightly higher rates of oxidation of exogenous glucose were recorded during the 2nd, 3rd and 4th hours of the test. These data do not provide evidence that the biguanide-induced improvement in glucose tolerance in patients with mild-to-moderate glucose intolerance is associated with any inhibiting or delaying effect of this type of drug on intestinal absorption (and subsequent oxidation) of the exogenous glucose load. On the contrary, a slight, but statistically significant, increase in the oxidation of exogenous glucose has been observed after butylbiguanide.

Experimental Studies of Variations of the State of Water in Living Cells

The high resolution NMR spectra and the nuclear magnetic relaxation times of water protons in frog sciatic nerves untreated and treated with 5 mM DNP, as well as the high resolution NMR spectra and the nuclear magnetic relaxation times of water protons in nerves from which this inhibitor was washed away, showed marked differences as a consequence of the change occuring in water state. Considering these results in the light of our present knowledge concerning the physical state of water in living cells, a tentative interpretation is proposed.