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APPLIED SCIENCES

Carbohydrate Supplementation and the Influence of Breakfast on Fuel Use in Hypoxia

GRIFFITHS, ALEX1; DEIGHTON, KEVIN1; BOOS, CHRISTOPHER J.1,2; ROWE, JOSHUA1; MORRISON, DOUGLAS J.3; PRESTON, TOM3; KING, RODERICK1; O’HARA, JOHN P.1

Author Information
Medicine & Science in Sports & Exercise: April 2021 - Volume 53 - Issue 4 - p 785-795
doi: 10.1249/MSS.0000000000002536

Abstract

Hypoxia experienced at altitude induces a curvilinear decrement in endurance performance (1). As such, methods of overcoming this impairment in performance warrant investigation. The effect of carbohydrate supplementation on substrate oxidation in sea level conditions is well established (2–4); however, differing effects have been observed in hypoxia (5–7). These responses may be explained by the contrasting use of preexercise nutritional status within experimental design (8,9). The hormonal and substrate storage responses to breakfast consumption/omission, which induce the aforementioned effect (10), likely also have implications for substrate oxidation during exercise with carbohydrate supplementation. As such, the effect of carbohydrate supplementation and the influence of preexercise nutritional status on substrate oxidation in hypoxia warrant further investigation.

After preexercise breakfast consumption, Péronnet et al. (6) observed a greater relative contribution of carbohydrate oxidation to energy expenditure during exercise matched for relative intensities (77% altitude-specific V˙O2max) in acute hypobaric hypoxia (445 mm Hg or 4300 m) compared with normoxia after glucose ingestion (1.75 g·min−1). This was attributed to a greater reliance on endogenous carbohydrate oxidation in hypoxia compared with normoxia. By contrast, O’Hara et al. (5) employed participants after preexercise breakfast omission and observed a significantly lower contribution of whole-body carbohydrate oxidation to energy expenditure during exercise matched for relative intensities (~74% altitude-specific V˙O2max) in hypoxia (terrestrial altitude ~3375 m) compared with normoxia after carbohydrate supplementation (1.2 g·min−1 glucose, 0.6 g·min−1 fructose). A significant reduction in endogenous carbohydrate oxidation was observed in hypoxia compared with normoxia, derived from a reduced reliance on muscle glycogen. This varied response between studies appears to be derived from an altered reliance on endogenous carbohydrate stores in hypoxia, which may be determined by preexercise nutritional status. Exogenous carbohydrate oxidation was not different between conditions in these studies, but reduced rates have been observed in hypoxia in females recently (11). Data that challenge the concept that breakfast may explain discrepancies in the literature have been published recently (12). Margolis et al. (12) employed participants after a 12-h fast and observed an increased reliance on carbohydrate oxidation (derived from endogenous sources) during exercise in acute hypoxia compared with normoxia. These findings contrast those observed by O’Hara et al. (5), who also employed fasted participants. However, Margolis et al. (12) used exercise matched for absolute intensities in hypoxia compared with normoxia, during which it may be expected that an increased relative exercise intensity, rather than hypoxia per se, may induce an increased reliance on carbohydrate oxidation (9,13).

To the authors’ knowledge, only one study has investigated the effect of carbohydrate supplementation on substrate oxidation in hypoxia in a placebo-controlled fashion, using 13C tracer methods (7). Young et al. (7) observed an increase in total carbohydrate oxidation during exercise (55% V˙O2max) with carbohydrate supplementation (1.00 g·min−1 glucose, 0.82 g·min−1 fructose) compared with placebo in acute hypoxia (terrestrial altitude ~4300 m). As expected, this increased reliance on carbohydrate oxidation in the carbohydrate group was derived from exogenous sources. However, as Young et al. (7) employed participants in the fasted state, the effects of carbohydrate supplementation are likely more pronounced than in fully fed participants. Interestingly, and in contrast to previous literature (5,6) they also observed a reduced reliance on exogenous carbohydrate oxidation during exercise matched for absolute intensities in hypoxia compared with normoxia, questioning the efficacy of carbohydrate supplementation in such conditions. This finding has also been replicated in fasted participants recently (12). This is especially surprising given the use of exercise matched for absolute intensities in hypoxia versus normoxia and the increased propensity for carbohydrate oxidation during exercise at greater exercise intensities (14).

Carbohydrate supplementation has been demonstrated to improve time trial performance of sea level residents during energy deficit in hypoxia (15). However, later research by the same group observed no difference in time trial performance after carbohydrate supplementation in fed participants (albeit in moderate altitude residents) (16). In addition, recent literature has demonstrated no effect of carbohydrate supplementation on time trial performance in acute or chronic hypoxia when fasted and in energy deficit, respectively (17). The effects of carbohydrate supplementation in both the fasted and fed state are yet to be determined using a within-study design and an ecologically valid mode of exercise in relation to the severity of hypoxia. Therefore, the purpose of this study was to investigate the effect of carbohydrate supplementation on substrate use (carbohydrate [exogenous and endogenous—muscle and liver glycogen] and fat oxidation) and endurance performance in hypoxia after both breakfast consumption and omission. As a methodologically novel approach to this study, both placebo and carbohydrate beverages were enriched with 13C glucose to allow the comparison of endogenous (muscle and liver) carbohydrate contributions between trials.

METHODS

Participants

Eleven, physically active, healthy male volunteers (23 ± 3 yr, 178.0 ± 7.0 cm, 76.6 ± 7.0 kg) provided written, informed consent to participate in this study. The study received institutional ethical approval (Leeds Beckett research ethics committee, application reference 46180) and was conducted in accordance with the Declaration of Helsinki. All participants were nonsmokers, normotensive, free from food allergies, and not taking any medication. None of the participants had traveled to an altitude of >1500 m within the previous 3 months and were all currently residing at an altitude of <500 m.

A priori power analysis revealed that eight participants provided 80% power to detect differences in absolute whole-body carbohydrate oxidation between carbohydrate and placebo groups during exercise in hypoxia, assuming an effect size of 0.97 (7) and an alpha of 0.05. In addition, a priori power analysis also revealed that eight participants provided 80% power to detect differences in absolute whole-body carbohydrate oxidation between breakfast consumption and omission trials during exercise in hypoxia, assuming an effect size of 1.30 (8) and an alpha level of 0.05. Although the effect sizes extracted from previous literature were studies using two trials, effect sizes for relevant variables are likely similar, and a correction for four trials was made to match the experimental design of the present study. In excess of the required sample size, 12 participants were initially recruited to this study; however, one participant dropped out due to injury. Therefore, 11 participants completed the study.

Experimental Design

Participants were required to make a total of seven visits to the laboratory. The first visit involved preexercise screening, anthropometry, verbal familiarization with testing procedures, sickle cell trait test, and a baseline 12-lead ECG test (18). Further exclusion criteria included diabetes and thyroid disorders. The second and third visit required participants to be acutely exposed to normobaric hypoxia (fraction of inspired oxygen [FiO2]: ~11.7% when considering water vapor partial pressure and daily fluctuations in barometric pressure [19]) equivalent to 4300 m (partial pressure of inspired oxygen [PiO2]: 83 mm Hg) in an environmental chamber (TISS, Alton, UK, and Sporting Edge, Sheffield on London, UK). On visit 2, participants completed a submaximal and maximal exercise test to calculate walking speed required to elicit 50% V˙O2max in normobaric hypoxia. On visit 3, participants completed two 30-min submaximal walking tests at 50% V˙O2max (before and after breakfast) and a 3-km familiarization time trial (see experimental trials). The 30-min submaximal walking tests were used to measure background 13C enrichment of expired CO2 observed in response to exercise (no glucose or placebo beverage ingested) for use in the calculation of exogenous carbohydrate oxidation (see Calculations section). These two preliminary trials were separated by >48 h.

On visits 4–7 (normobaric hypoxia equivalent to 4300 m), participants completed a 4-h 30-min experimental trial that included rest, followed by a 90-min submaximal walking test (50% V˙O2max), a 3-km time trial, and a postexercise rest period (Fig. 1). Two trials involved preexercise breakfast consumption followed by ingestion of a carbohydrate (B-CHO) or placebo beverage (B-PLA), and the other two trials involved preexercise breakfast omission followed by ingestion of a carbohydrate (F-CHO) or placebo beverage (F-PLA) during the 1-h 30-min submaximal walking test. The carbohydrate beverage trials involved ingestion of 1.2 g·min−1 (108 g) of glucose (D-glucose; Thornton and Ross Ltd., Huddersfield, UK). Stock glucose was enriched using 0.18 g of U-13C6 D-glucose (Cambridge Isotope Laboratories Inc., Tewksbury, MA), achieving an enrichment of δ13C = 115.6‰. As a methodological approach novel to hypoxia, the placebo beverage trials involved ingestion of a water solution, also enriched with 0.18 g U-13C6 D-glucose (99 atom %). This allowed for the comparison of endogenous (muscle and liver) carbohydrate contributions between trials. All δ13C measurements are quoted with reference to the internationally accepted standard for carbon isotope measurements, VPDB. The 13C abundance of stock glucose and 13C enrichment of spiked glucose was determined using liquid chromatography coupled to isotope ratio mass spectrometry (LC-IRMS; Isoprime, Cheadle, UK) (20). The enriched water solution has been shown to have negligible effects on substrate oxidation (21). Each beverage contained 25.7 mmol·L−1 sodium chloride (2.25 g). These visits were separated by ≥7 d, and preexercise nutritional status (breakfast consumption or omission) was randomized in a single blind fashion. The order of beverage ingestion was randomized in a double-blind fashion by a researcher independent to the study.

F1
FIGURE 1:
Schematic of full experimental trial.

Diet and Physical Activity before Testing

Participants recorded their food intake for the 24 h before the second preliminary trial (visit 3) and were instructed to replicate this for each experimental trial. During this time, participants were asked not to perform strenuous activity or consume caffeine or alcohol. Participants consumed a standardized meal before the second preliminary trial and all experimental trials, which contained fusilli pasta, pasta sauce, cheese, semiskimmed milk, and an apple (1043 kcal, 55% carbohydrate, 30% fat, 15% protein). This meal was consumed to minimize the possibility of a second meal effect confounding glycemic control or any other measured variables (22). A week before and throughout the duration of the study, participants were also asked to refrain from consuming carbohydrates derived from plants that use the C4 photosynthetic cycle, in which there is a higher natural abundance of 13C (23). This ensured that background 13CO2 abundance was less likely to be perturbed from oxidation of endogenous and dietary substance stores from naturally “enriched” C4 origin.

Preliminary Testing

On visit 2, participants completed a submaximal and maximal exercise test in normobaric hypoxia, as described previously (8). Briefly, the submaximal test involved walking at 10% gradient, with a 10-kg backpack at a range of walking speeds. The maximal test involved running at a constant speed, dependent on fitness, aiming for an RPE of 12. The test began at 1% gradient and increased every minute until volitional exhaustion. These data were used to establish walking speeds that would elicit 50% V˙O2max while carrying a 10-kg rucksack at a 10% gradient.

On visit 3, participants arrived at the environmental chamber fasted (~12 h overnight) and subsequently completed a 30-min submaximal walking test. Participants were then permitted 15 min to consume a standardized breakfast (535 kcal, 58% carbohydrate, 24% fat, and 18% protein) as detailed previously (8). After breakfast consumption, participants rested for 1 h and then repeated the 30-min submaximal walking test. Expired gas was collected (see Experimental Trials section) at the end of each submaximal walking test to measure background 13C enrichment of expired gas without ingestion of the carbohydrate or placebo drink for calculation of exogenous carbohydrate oxidation (see Calculations section). After a 5-min rest period, participants then completed a 3-km time trial to assess performance (see Experimental Trials section).

Experimental Trials

Participants entered the environmental chamber at 7:30 am, after a 12-h fast. Participants then rested for 30 min. At 30 min in the B-CHO and B-PLA trials, participants were allowed 15 min to consume a standardized breakfast (as per preliminary trial) but remained fasted in the F-CHO and F-PLA trials. At 45 min, participants in all trials rested for a further hour. At 1 h 45 min, participants completed a 1-h 30-min submaximal (50% V˙O2max) walking test at a 10% gradient, carrying a 10-kg rucksack, to mimic the demands of high altitude trekking. Within each nutritional subgroup (breakfast omission and breakfast consumption), one trial consumed a carbohydrate and one trial consumed a placebo beverage. Each beverage was consumed preexercise (600 mL) and every 15 min during exercise (150 mL). A total of 1.5 L of carbohydrate or placebo solution was consumed over the course of the trial. After a short rest period, participants then completed a self-paced 3-km time trial at 10% gradient, carrying a 10-kg rucksack. A rolling start was used, in which the speed required to elicit 50% V˙O2max was utilized. Once the time trial had started, participants had full control over speed. Participants were informed of their distance every 500 m but were blinded to their speed and time (24). After exercise, participants rested for a further 30 min before returning to sea level conditions.

Measurements

Heart rate, SpO2, and RPE

Heart rate and SpO2 were measured every 15 min during rest. Heart rate, SpO2, and RPE were measured every 10 min throughout exercise.

Expired breath collection

Expired gas breath samples were collected using an online gas analysis system (Metalyser, Cortex, Germany). These measurements were made intermittently throughout exercise (20–30 min, 50 min–1 h, 1 h 5 min–1 h 15 min, and 1 h 20 min–1 h 30 min). Participants were fitted with a facemask 5 min before the collection period while the participant was seated. In addition, samples of expired gas were collected in duplicate via a mixing chamber in 12-mL Labco Exetainers (SerCon Ltd., Crewe, UK) for the analysis of 13C/12C in expired CO2 during the final 60 s of each gas collection period (preexercise, 60 min, 75 min, and 90 min exercise).

Blood sampling

Venous blood samples were drawn for the analysis of plasma glucose, plasma lactate, and serum FFA at baseline (before entry to the chamber), 30 min (preprandial), and 1 h 45 min (postprandial), as well as during the submaximal walking test at 30, 60, 75, and 90 min and also after exercise at 4 h 30 min (postexercise). Samples for serum insulin were drawn at all time points with the exclusion of 30 and 75 min of the submaximal walking test. Samples for the analysis of 13C plasma glucose enrichment were drawn at 1 h 45 min (postprandial) and at 60, 75, and 90 min during submaximal exercise.

Analyses

Commercially available enzyme-linked immunosorbent assay kits were used to determine serum concentrations of insulin (IBL, Hamburg, Germany). To eliminate interassay variation, all samples from each participant were analyzed on the same plate. Plasma glucose and lactate, and serum FFA were measured photometrically with reagents from Instrumentation Laboratories (Lexington, MA) and Randox Laboratories (Crumlin, UK). The within-batch coefficients of variation were as follows: insulin 6.1%, glucose 1.6%, lactate 2.4%, and FFA 3.8%.

The 13C abundance of stock glucose and 13C enrichment of spiked glucose was determined using LC-IRMS (Isoprime), using L-fucose as an isotopic internal standard as previously described (20). The 13C/12C ratio in expired CO2 was determined using IRMS (AP2003; GVI Instruments Ltd., Manchester, UK). The isotopic ratio 13C/12C is derived against laboratory CO2 (itself calibrated against VPDB) from the ion beam area ratio measurements with correction of the small contribution of 12C16O17O at m/z 45 (25). The 13C/12C ratio in plasma glucose was determined using LC-IRMS as detailed previously (20).

Calculations

Substrate oxidation was calculated using relevant equations for exercise periods (26). The isotopic enrichment of the ingested glucose was expressed in standard δ13C units (‰) relative to VPDB (25). Exogenous carbohydrate oxidation derived from the ingested glucose was calculated using the following equation (27):

exogenous carbohydrate oxidationg·min1=V˙CO2RexpRref/RexoRref/k

where V˙CO2 is represented in liters per minute, Rexp is the measured isotopic composition in expired CO2 during exercise at different time points, Rref is the isotopic composition of expired CO2 during exercise with the ingestion of placebo (background), Rexo is the measured isotopic enrichment of the ingested glucose, and k is the rate adjusted value for the complete oxidation of glucose (27). Changes in the background isotopic composition of expired CO2 with a placebo beverage could not be determined from the experimental trials because of the enrichment of the placebo beverage with U-13C6 D-glucose. Therefore, data from the preliminary trial (visit 3) in which expired gas was collected during exercise before and after breakfast were utilized. The use of Rref from expired CO2 during exercise with placebo is typical of studies in this area (21). The 13C enrichment of exogenous glucose is high, and the use of Rref during exercise cancels the confounding effect of small fluctuations in background enrichment of expired CO2 derived from the Western European diet. Endogenous carbohydrate oxidation was calculated by subtracting exogenous carbohydrate oxidation from total carbohydrate oxidation.

Computations were made on the assumption that, in response to exercise, 13C is not lost irreversibly in pools of tricarboxylic acid cycle intermediates and/or bicarbonate, and that 13CO2 recovery in expired gases was complete or almost complete during exercise (28). Such computation has been shown to underestimate exogenous carbohydrate oxidation rates at the beginning of exercise because of the delay between 13CO2 production in tissues and expired 13CO2 at the mouth (29). As such, exogenous carbohydrate oxidation rates are presented from 60 min onward during submaximal exercise, where it is expected that there would be isotopic equilibrium in the tissues and at the mouth.

Plasma glucose oxidation was computed at 60, 75, and 90 min during the submaximal walking test, based on the isotopic composition of plasma glucose (Rglu) using the following equation (30):

plasma glucose oxidationg·min1=V˙CO2RexpRref/RgluRref/k

The oxidation rate of muscle glycogen (g·min−1) either directly or through the lactate shuttle (31) was calculated by subtracting plasma glucose oxidation from total carbohydrate oxidation. Finally, the amount of glucose released from the liver was estimated as the difference between plasma glucose oxidation and exogenous carbohydrate oxidation. Liver glucose oxidation values are representative of contributions from liver glycogen, gluconeogenesis, as well as residual glucose uptake from the gut derived from previous breakfast consumption.

Statistical analysis

Data are expressed as mean ± SD in text and mean ± SE in figures. All data were analyzed using IBM SPSS statistics for Windows (version 24; SPSS Inc., Chicago, IL). The trapezoid method was used to calculate area under the curve (AUC) for substrate oxidation and hormone concentrations. The periods of AUC were defined as preprandial (0–30 min), postprandial (45–1 h 45 min), submaximal exercise (0–60 min and 60–90 min), and postexercise (4 h–4 h 30 min). Two-way repeated-measures ANOVA (time–trial) was used to determine differences between absolute and relative carbohydrate and fat oxidation, hormone concentrations, δ13CO2 in expired gas and plasma glucose, and rates of oxidation of plasma glucose, liver glucose, and muscle glycogen. One-way repeated-measures ANOVA was used to determine differences between trials for absolute (AUC) and relative contributions of endogenous carbohydrate, liver glucose, muscle glycogen, plasma glucose (only absolute) oxidation, energy expenditure, heart rate, SpO2, RPE, and time trial times. Where significant main effects were found, further post hoc analysis was performed using Bonferroni correction for multiple comparisons. Paired sample t-tests were used to determine differences in relative and absolute exogenous carbohydrate oxidation and total absolute exogenous carbohydrate oxidation (AUC). Effect sizes are presented as Cohen’s d and interpreted as ≤0.2 trivial, >0.2 small, >0.6 moderate, >1.2 large, >2 very large, and >4 extremely large (32).

RESULTS

Maximal oxygen uptake and walking speeds

V˙O2max in hypoxia was 40.6 ± 4.3 mL·kg·min−1, and this elicited a walking speed of 2.9 ± 0.5 km·h−1 in the experimental trials (B-CHO, 50.0% ± 8.4% V˙O2max; B-PLA, 49.0 ± 8.1 V˙O2max; F-CHO, 49.3 ± 8.3 V˙O2max; F-PLA, 49.0 ± 8.1 V˙O2max; P = 0.99).

Energy expenditure

Energy expenditure was not significantly different between trials (B-CHO, 4003 ± 671 kJ; B-PLA, 3648 ± 726 kJ; F-CHO, 3768 ± 598 kJ; F-PLA, 3563 ± 621 kJ; all P = 0.99, d ≤ 0.32).

Total carbohydrate and fat oxidation

Results for total carbohydrate and fat oxidation for the full exercise duration are described in supplemental digital content (see text document, Supplemental Digital Content 1, Results for total carbohydrate and fat oxidation and δ13C in expired gas and plasma glucose at rest and exercise, https://links.lww.com/MSS/C170). Because of the necessary partitioning of exercise (i.e., 0–60 min and 60–90 min) as a result of 13C ingestion, exercise will be discussed in relation to the first 60 min, and last 30 min herein.

During the first hour of exercise, absolute (Table 1) and relative contributions of carbohydrate oxidation to energy expenditure were significantly higher after breakfast consumption compared with omission in the carbohydrate (absolute, P < 0.01, d = 1.10; relative, 51.0% ± 10.4% vs 38.1% ± 7.8%, P < 0.01, d = 1.42) and placebo trials (absolute, P < 0.01, d = 1.07; relative, 47.8 ± 10.0 vs 32.1 ± 12.3, P < 0.01, d = 1.41). In addition, absolute and relative contributions of carbohydrate oxidation to energy expenditure were not significantly different between carbohydrate and placebo trials after breakfast consumption (absolute, P = 0.86, d = 0.35; relative, P = 0.99, d = 0.32) or omission (absolute, P = 0.99, d = 0.39; relative, P = 0.99, d = 0.60). In the same period, absolute fat oxidation was significantly higher after breakfast omission compared with consumption in the placebo trials (P < 0.01, d = 0.77), but not in the carbohydrate trial (P = 0.15, d = 0.65). The relative contribution of fat oxidation to energy expenditure was significantly higher after breakfast omission compared with consumption in the carbohydrate (61.9% ± 7.8% vs 49.0% ± 10.4%, P < 0.01, d = 1.42) and placebo trials (67.9% ± 12.3% vs 52.2% ± 10.0%, P < 0.01, d = 1.41). In addition, absolute and relative contributions of fat oxidation were not significantly different between carbohydrate and placebo trials after breakfast consumption (absolute, P = 0.99, d = 0.13; relative, P = 0.99, d = 0.32) or omission (absolute, P = 0.99, d = 0.30; relative, P = 0.99, d = 0.60).

TABLE 1 - AUC values for absolute carbohydrate and fat oxidation during exercise in all trials.
Carbohydrate Oxidation (g) Fat Oxidation (g)
0–90 min 0–60 min 60–90 min 0–90 min 0–60 min 60–90 min
B-CHO 76.72 ± 22.11* 48.27 ± 13.54* 28.45 ± 8.84 31.52 ± 9.28 20.09 ± 5.73 11.43 ± 3.68
B-PLA 67.17 ± 21.20* 43.57 ± 13.39* 23.60 ± 7.96* 33.91 ± 10.22* 20.84 ± 6.20* 13.07 ± 4.04*
F-CHO 57.18 ± 16.21 33.38 ± 9.40 23.80 ± 6.92† 37.12 ± 9.24 23.81 ± 5.66 13.31 ± 3.64†
F-PLA 44.14 ± 22.81 28.71 ± 14.50 15.43 ± 8.48 42.29 ± 10.00 25.60 ± 6.23 16.69 ± 3.91
*Significant difference to alternative nutritional status with the same supplement.
†Significant difference to alternative supplement within the same nutritional status.
B-CHO, breakfast consumption and carbohydrate supplementation; B-PLA, breakfast consumption and placebo; F-CHO, breakfast omission and carbohydrate supplementation; F-PLA, breakfast omission and placebo.

During the last 30 min of exercise, absolute (Table 1) and relative (Fig. 2) contributions of carbohydrate oxidation were higher after breakfast consumption compared with omission in the placebo (absolute, P = 0.02, d = 0.99; relative, P < 0.01, d = 1.43) but not carbohydrate trials (absolute, P = 0.20, d = 0.59; relative, P = 0.20, d = 0.73). In addition, absolute and relative contributions of carbohydrate oxidation were significantly higher in the carbohydrate compared with placebo trial after breakfast omission (absolute, P = 0.02, d = 1.08; relative, P < 0.01, d = 1.52) but not consumption (absolute, P = 0.39, d = 0.58; relative, P = 0.38, d = 0.66). In the same period, absolute and relative contributions of fat oxidation to energy expenditure were higher after breakfast omission compared with consumption in the placebo (absolute, P < 0.01, d = 0.91; relative, P < 0.01, d = 1.43) but not carbohydrate trials (absolute, P = 0.57, d = 0.51; relative, P = 0.20, d = 0.73). In addition, absolute and relative contributions of fat oxidation were significantly higher in the placebo compared with carbohydrate trials after breakfast omission (absolute, P = 0.02, d = 0.90; relative, P < 0.01, d = 1.52) but not consumption (absolute, P = 0.77, d = 0.42; relative, P = 0.38, d = 0.66).

F2
FIGURE 2:
The relative (% energy yield) contribution to energy expenditure during the last 30 min of exercise in all trials. B-CHO, breakfast consumption and carbohydrate supplementation; B-PLA, breakfast consumption and placebo supplementation; F-CHO, breakfast omission and carbohydrate supplementation; F-PLA, breakfast omission and placebo supplementation. *Significant difference in relative carbohydrate oxidation. †Significant difference in relative liver glucose oxidation. ‡Significant difference in relative muscle glycogen oxidation (P < 0.05).

Expired gas and plasma glucose

Results for δ13CO2 enrichment in expired gas and plasma glucose are presented in supplemental digital content (see text document, Supplemental Digital Content 1, Results for total carbohydrate and fat oxidation and δ13C in expired gas and plasma glucose at rest and exercise, https://links.lww.com/MSS/C170; see Figure, Supplemental Digital Content 2, δ13C in expired gas and plasma glucose at rest and exercise, https://links.lww.com/MSS/C171).

Exogenous and endogenous carbohydrate oxidation

There was no significant difference in exogenous carbohydrate oxidation rates (Fig. 3A) after breakfast consumption compared with omission in the carbohydrate trials at 60 min (0.27 ± 0.12 vs 0.31 ± 0.07 g·min−1), 75 min (0.34 ± 0.10 vs 0.38 ± 0.10 g·min−1), or 90 min (0.43 ± 0.13 vs 0.43 ± 0.12 g·min−1) (P = 0.30). The contribution of exogenous carbohydrate oxidation in the placebo trials was considered negligible (<0.001 g·min−1). The relative contribution of exogenous carbohydrate oxidation in the last 30 min of exercise was not significantly different between breakfast consumption and omission in the carbohydrate trials (18.6% ± 3.5% vs 20.8% ± 3.4%, P = 0.14, d = 0.59; Fig. 3A). The total absolute exogenous oxidation in the last 30 min of exercise was also not significantly different between breakfast consumption and omission in the carbohydrate trials (P = 0.23, d = 0.28; Table 2).

F3
FIGURE 3:
Oxidation rates of exogenous carbohydrate, liver glucose, plasma glucose, and muscle glycogen during the final 30 min of exercise. Values are presented as mean ± SE. B-CHO, breakfast consumption and carbohydrate supplementation; B-PLA, breakfast consumption and placebo supplementation; F-CHO, breakfast omission and carbohydrate supplementation; F-PLA, breakfast omission and placebo supplementation. A, Significant difference between breakfast consumption and omission in the carbohydrate trials. B, Significant difference between breakfast consumption and omission in the placebo trials. C, Significant difference between the carbohydrate and the placebo trials after breakfast consumption. D, Significant difference between the carbohydrate and the placebo trials after breakfast omission.
TABLE 2 - Carbohydrate oxidation from various sources during the last 30 min of exercise in all trials.
Exogenous Oxidation (g) Endogenous Oxidation (g) Muscle Glycogen (g) Glucose from the Liver (g) Plasma Glucose (g)
B-CHO 10.35 ± 3.22 18.10 ± 6.21* 13.83 ± 5.43 4.27 ± 1.42*† 14.62 ± 4.48†
B-PLA 0.02 ± 0.01 23.58 ± 7.96* 15.77 ± 7.61 7.81 ± 1.65* 7.83 ± 1.65*
F-CHO 11.22 ± 2.95 12.58 ± 5.34 10.51 ± 4.91 2.07 ± 0.73† 13.29 ± 3.33†
F-PLA 0.02 ± 0.01 15.41 ± 8.48 10.70 ± 8.04 4.71 ± 1.69 4.73 ± 1.69
*Significant difference to alternative nutritional status with the same supplement.
†Significant difference to alternative supplement within the same nutritional status.
B-CHO, breakfast consumption and carbohydrate supplementation; B-PLA, breakfast consumption and placebo supplementation; F-CHO, breakfast omission and carbohydrate supplementation; F-PLA, breakfast omission and placebo supplementation.

Total absolute endogenous carbohydrate oxidation in the last 30 min of exercise (Table 2) was significantly higher after breakfast consumption compared with omission in the carbohydrate (P = 0.03, d = 0.95) and placebo trials (P = 0.02, d = 0.99). There was no significant difference in endogenous carbohydrate oxidation in the last 30 min of exercise between the carbohydrate and the placebo trials after breakfast consumption (P = 0.18, d = 0.77) or omission (P = 0.99, d = 0.41).

Oxidation of plasma glucose, liver glucose, and muscle glycogen

Plasma glucose oxidation rates (Fig. 3B) were significantly higher after breakfast consumption compared with omission in the placebo trials during exercise at 60, 75, and 90 min (all P < 0.01, d > 1.51) but not carbohydrate trials (P > 0.31, d < 0.55). Plasma glucose oxidation rate was higher in the carbohydrate compared with placebo trials after breakfast consumption at 75 and 90 min (P < 0.01, d > 2.25) but not 60 min (P = 0.09, d = 1.23). Plasma glucose oxidation rate was higher in the carbohydrate compared with placebo trials after breakfast omission at 60, 75, and 90 min (P < 0.01, d > 3.20). Total absolute plasma glucose oxidation during the last 30 min of exercise (Table 2) was significantly higher after breakfast consumption compared with omission in the placebo (P < 0.01, d = 1.86) but not carbohydrate trials (P = 0.99, d = 0.34). Total absolute plasma glucose oxidation was also significantly higher in the carbohydrate compared with placebo trials after breakfast consumption (P < 0.01, d = 2.21) and omission (P < 0.01, d = 3.42).

Liver glucose oxidation rates (Fig. 3C) were significantly higher after breakfast consumption compared with omission at 60, 75, and 90 min in the carbohydrate (P < 0.049, d > 1.55) and placebo trials (all P < 0.01, d > 1.51). Liver glucose oxidation rates were also significantly higher in the placebo compared with carbohydrate trials at 60, 75, and 90 min after breakfast consumption (P < 0.02, d > 1.83) and omission (P < 0.01, d > 1.83). Relative and total absolute (Table 2) liver glucose oxidation was significantly higher after breakfast consumption compared with omission during the last 30 min of exercise in the carbohydrate (absolute, P < 0.01, d = 2.03; relative, 7.7% ± 1.6% vs 3.8% ± 0.8%, P < 0.01, d = 3.20) and placebo trials (absolute, P < 0.01, d = 1.86; relative, 14.8% ± 2.3% vs 8.7% ± 2.8%, P < 0.01, d = 2.42). Relative and total absolute liver glucose oxidation was significantly lower in the carbohydrate compared with placebo trials after breakfast consumption (relative, P < 0.01, d = 3.67; absolute, P < 0.01, d = 2.31) and omission (relative, P < 0.01, d = 2.77; absolute, P < 0.01, d = 2.18).

Muscle glycogen oxidation rates (Fig. 3D) were significantly higher after breakfast consumption compared with omission at 60 min in the carbohydrate trials (P = 0.04, d = 0.98) but not placebo trials (P = 0.13, d = 0.69). Muscle glycogen oxidation rates were not significantly different at 60 min between carbohydrate and placebo trials after breakfast consumption (P = 0.99, d = 0.03) or omission (P = 0.99, d = 0.06). There was no significant difference between trials in muscle glycogen oxidation rates during exercise at 75 min (P > 0.12, d < 0.64) or 90 min (P > 0.19, d < 0.57). The relative but not absolute (Table 2) contribution of muscle glycogen oxidation to energy expenditure during the last 30 min of exercise was higher after breakfast consumption compared with omission in the placebo trials (relative, 29.4% ± 11.1% vs 19.2% ± 12.2%, P = 0.04, d = 0.87; absolute, P = 0.14, d = 0.65) and approached significance in the carbohydrate trials (relative, 25.4.0% ± 9.4% vs 19.4% ± 7.5%, P = 0.09, d = 0.71; absolute, P = 0.14, d = 0.64). There was no significant difference in the relative or absolute contribution of muscle glycogen oxidation to energy expenditure during the last 30 min of exercise in the carbohydrate compared with placebo trials after breakfast consumption (relative, P = 0.99, d = 0.38; absolute, P = 0.99, d = 0.30) or omission (relative, P = 0.99, d = 0.02; absolute, P = 0.99, d = 0.03).

Blood biochemistry

A significant effect of time (all P < 0.01) was observed for all analytes (Fig. 4). A significant effect of trial was observed for all analytes (P < 0.01), except lactate (P = 0.17). Further, a significant interaction effect of time–trial was also observed for all analytes (all P < 0.01). All significant pairwise statistical comparisons are presented in Figure 4.

F4
FIGURE 4:
Plasma glucose, serum FFA, plasma lactate, and serum insulin concentrations over the full experimental trial. Values are presented as mean ± SE. The thin arrow represents the timing of breakfast in the breakfast consumption trials. The black rectangle represents the exercise period. B-CHO, breakfast consumption and carbohydrate supplementation; B-PLA, breakfast consumption and placebo supplementation; F-CHO, breakfast omission and carbohydrate supplementation; F-PLA, breakfast omission and placebo supplementation. a, Significant difference between breakfast consumption and omission in the carbohydrate trials; b, significant difference between breakfast consumption and omission in the placebo trials; c, significant difference between the carbohydrate and the placebo trials after breakfast consumption; d, significant difference between the carbohydrate and the placebo trials after breakfast omission. Significance P < 0.05.

Three-kilometer time trial performance

Time to completion (see Figure, Supplemental Digital Content 3, Time trial performance, https://links.lww.com/MSS/C172) was not significantly different between trials (B-CHO, 2121 ± 230 s; B-PLA, 2154 ± 284 s; F-CHO, 2134 ± 289 s; F-PLA, 2209 ± 213 s; P = 0.99, d ≤ 0.30).

Heart rate, SpO2, and RPE

There were no significant differences between trials for SpO2 (P ≥ 0.45, d ≤ 0.51), heart rate (all P = 0.99, d ≤ 0.36), and RPE (all P = 0.99, d ≤ 0.36) (see Table, Supplemental Digital Content 4, Heart rate, SpO2 and RPE data, https://links.lww.com/MSS/C173).

DISCUSSION

This study investigated the effect of carbohydrate supplementation on substrate oxidation and time trial performance after both breakfast consumption and omission. In the first 60 min of exercise, carbohydrate supplementation had no effect on substrate oxidation after breakfast consumption or omission. However, in the final 30 min of exercise, carbohydrate supplementation increased the relative carbohydrate contribution to energy expenditure after breakfast omission, but not consumption. This was likely explained by an increased contribution of exogenous carbohydrate and a concomitant reduction in the relative contribution of muscle and liver glycogen oxidation to energy expenditure during exercise after breakfast omission compared with consumption in hypoxia. These findings suggest that suboptimal concentrations of endogenous carbohydrate stores during exercise >60 min may have been observed in the breakfast omission, but not consumption trials. Interestingly, a reduction in liver glucose oxidation suggests a liver glycogen-sparing effect of carbohydrate supplementation during submaximal exercise; however, there was no effect of carbohydrate supplementation on time trial performance in hypoxia.

The finding that the relative contribution of carbohydrate oxidation to energy expenditure was increased after breakfast consumption compared with omission in hypoxia is consistent with the well-established response to feeding in normoxia (33). The reduction in plasma glucose concentrations in the postprandial state typically observed at the onset of exercise in normoxia was also replicated in hypoxic conditions in the present study, as observed previously (8). A concomitant reduction in FFA availability and oxidation was also observed in the present study after breakfast consumption compared with omission, likely due to the inhibitory effect of insulin on lipolysis (34). Interestingly, there was no effect of carbohydrate supplementation on whole-body substrate oxidation after breakfast consumption or omission in the first hour of exercise. This finding is likely explained by the non–glycogen-limiting duration of exercise (35).

Carbohydrate supplementation induced a higher relative contribution of carbohydrate oxidation to energy expenditure during the last 30 min of exercise in hypoxia after breakfast omission but not consumption. This increased reliance on carbohydrate oxidation was associated with greater plasma glucose concentration and oxidation, derived from the exogenous carbohydrate source. The findings observed in fasted participants are in agreement with the normoxic (4) and hypoxic literature (7). This increased contribution of carbohydrate oxidation likely only occurred in the final 30 min of exercise because of the depletion of endogenous carbohydrate stores in such conditions, as discussed previously (35). Data from the final 30 min of exercise in present study support this hypothesis by demonstrating significant reductions in total endogenous carbohydrate oxidation after breakfast omission compared with consumption in both the carbohydrate and placebo trials. These reduced contributions were derived from significant, moderate reductions in muscle glycogen utilization in the placebo trial and nonsignificant (P = 0.09), moderate reductions in the carbohydrate trials. In addition, significant, large/very large reductions in liver glucose oxidation were also observed after breakfast omission compared with consumption in the carbohydrate and placebo trials. Further, the absence of feeding both before and during exercise in the placebo trial after breakfast omission resulted in a low insulin concentration and likely facilitated an increased reliance on fat oxidation during exercise in this trial (34).

Although the effect of hypoxia on substrate oxidation during exercise with carbohydrate supplementation has been investigated in fed participants previously (6), the effect of carbohydrate supplementation on substrate oxidation during exercise in hypoxia in a placebo-controlled fashion has not. Our data demonstrate that carbohydrate supplementation has no effect on substrate oxidation during exercise (≤90 min) after breakfast consumption. Similar relative contributions of exogenous carbohydrate oxidation to energy expenditure were observed in the breakfast consumption compared with omission trial. As such, the absence of change in the relative contribution of carbohydrate oxidation between the carbohydrate and the placebo trials after breakfast consumption is likely due to maintained contributions of endogenous carbohydrate oxidation in the fed state (36). In support of this statement, the relative contribution of liver glucose and muscle glycogen oxidation was significantly higher after breakfast consumption compared with omission in the placebo trials, subsequently negating the effects of carbohydrate supplementation.

Findings from Young et al. (7) suggest that the oxidation of exogenous carbohydrate may be suppressed during exercise in hypoxia compared with normoxia. Although unable to confirm the suppression of exogenous carbohydrate oxidation, the rates observed in the present study (breakfast consumption and omission means [60–90 min]: 0.35 and 0.37 g·min−1) are lower than that observed by O’Hara et al. (5) (~0.92 g·min−1), O’Hara et al. (11) (0.82 g·min−1), and Péronnet et al. (6) (0.43 g·min1) but higher than that observed by Young et al. (7) (~0.19 g·min−1). The lower exogenous carbohydrate oxidation rates in the present study compared with O’Hara et al. (5), O’Hara et al. (11), and Péronnet et al. (6) are likely due to reduced exercise intensity (50% vs 74% and 77% V˙O2max respectively). Maximal exogenous carbohydrate oxidation rates typically occur during exercise at up to 64% V˙O2max (37); therefore, it is possible that the exercise intensity used in the present study (50% V˙O2max) did not elicit maximal exogenous carbohydrate oxidation rates. By contrast, the higher exercise intensities used by others were likely sufficient to elicit maximal rates. In addition, the carbohydrate supplement used by some (5,11) included both glucose and fructose, which has been demonstrated to increase exogenous oxidation compared with glucose alone because of the use of distinct intestinal transporters (38). Young et al. (7) supplemented participants with a glucose and fructose solution; however, only the glucose was labeled with a 13C isotopic tracer, and thus fructose oxidation could not be quantified. As such, the low exogenous glucose oxidation rates may be expected, as ingestion rates of glucose alone were just ~1.0 g·min−1 compared with 1.2 g·min−1 in the present study, despite a slightly higher exercise intensity used by Young et al. (7) (~60% V˙O2max vs 50% V˙O2max).

Young et al. (7) suggested that the exogenous carbohydrate oxidation suppression observed in acute hypoxia compared with normoxia infers that exogenous carbohydrate supplementation does not spare endogenous glucose stores to the same degree as in normoxia and therefore provides less of an advantage in hypoxic than normoxic conditions. Although the present study cannot determine this response in comparison with normoxic conditions, these data suggest that a sparing of endogenous carbohydrate may occur in hypoxia. In this regard, significant reductions in liver glucose oxidation were observed in the carbohydrate compared with placebo trials after both breakfast consumption and omission. This indication of an endogenous glucose-sparing effect seems exclusive to liver glycogen stores, as no significant differences in muscle glycogen oxidation were observed between the carbohydrate and the placebo trials after breakfast consumption or omission. A liver glycogen-sparing effect is consistent with some (4,39) but not all (3) normoxic literature. These findings are likely associated with partial attenuation of liver glycogenolysis and gluconeogenesis (4); however, these measurements were beyond the scope of the present study.

Interestingly, carbohydrate supplementation had no effect on the 3-km time trial performance. This is in agreement with Bradbury et al. (17), who observed no difference in time trial performance (2-mile walk) with a carbohydrate compared with placebo beverage in acute and chronic hypoxia (4300 m). However, these studies are in contrast to Fulco et al. (15), who observed an increase in endurance performance with carbohydrate supplementation after 3 d hypoxic exposure (4300 m) (CHO, 80.1 ± 7 min vs PLA, 104.9 ± 9.0 min). Differences between findings are likely due to variance in experimental design. In this regard, the intensity of the exercise used in the present study may not have been sufficient to deplete endogenous substrate stores to critical levels, therefore suppressing the effect of the supplement. Assuming a rate of glycogenolysis of ~0.7 mmol·kg·min−1 during exercise at 50% V˙O2max (40), a muscle glycogen depletion of ~63 mmol·kg·min−1 likely occurred in the present study (expected concentration at the end of submaximal exercise ~87 mmol·kg−1). As such, it seems likely that muscle glycogen concentration did not reach a critical threshold (<70 mmol·kg−1 wet weight) in which it may be expected calcium release from the sarcoplasmic reticulum is impaired, and subsequently peak power output reduced (41). This hypothesis may also explain the null findings observed by Bradbury et al. (17), as they used similar experimental design. Bradbury et al. (17) also demonstrated no effect of carbohydrate supplementation on endurance performance in sea level conditions, contradicting their hypothesis that hypoxia per se confounds the ergogenic effect of carbohydrate supplementation. By contrast, although Fulco et al. (15) used a shorter submaximal exercise period (45 min), the time trial used was longer in duration; therefore, participants likely spent more time exercising at higher intensities, thus depleting endogenous substrate stores. Further research is required to elucidate the effects of carbohydrate supplementation in conditions directly applicable to real-world scenarios (i.e., day-long mountaineering treks).

Despite the novel findings reported in this study, several limitations must be acknowledged. First, participants in this study were young males, and these findings remain to be elucidated in other populations. For example, women have shown differing metabolic responses in hypoxia (42) and normoxia (43), as well as a varied response to carbohydrate supplementation (11). In addition, this study was conducted in simulated normobaric hypoxia, and caution should be applied when considering the practical application of these findings to terrestrial altitude. Finally, calculations estimating liver glucose oxidation may also comprise residual gut uptake of glucose derived from preexercise breakfast consumption, albeit in small quantities. Future research should investigate the effects of carbohydrate supplementation on endurance performance of a longer duration (≥60 min) after breakfast consumption and omission. In addition, both optimal dose and composition should be elucidated in hypoxia, in both males and females.

In conclusion, breakfast consumption increased carbohydrate oxidation during the first 60 min of exercise regardless of carbohydrate supplementation. However, carbohydrate supplementation matched the effects of preexercise breakfast consumption during the final 30 min of exercise by increasing carbohydrate oxidation after breakfast omission, but not consumption. The reduction in liver glucose oxidation after carbohydrate supplementation suggested that a liver glycogen-sparing effect was present; however, no difference was observed in muscle glycogen oxidation. No effect of carbohydrate supplementation on the 3-km time trial performance was observed after breakfast consumption or omission. These data provide novel information regarding the use of carbohydrate supplementation in hypoxia for populations in differing states of energy balance (i.e., fasted or fed). These findings should be considered in the design of nutritional interventions for mountaineers, military personnel, and athletes exposed to high altitude. Specifically, the findings observed after breakfast omission in the present study may be applicable to individuals experiencing attenuated energy intake as a result of hypoxia-induced appetite suppression (44,45). Carbohydrate supplementation may be a useful nutritional strategy to induce alterations in substrate oxidation for these individuals. Although no changes in endurance performance were observed in the present study, this requires further investigation in chronic hypoxia, in which exercise duration and subsequent glycogen depletion are potentiated.

The authors thank Leeds Beckett University for funding this study, as well as all participants for their time and effort related to participation in this study. They also thank Mrs. Eleanor McKay and Ms. Sandra Small for technical assistance in conducting the plasma 13C glucose and breath 13C analysis at SUERC.

The authors report no competing interests. The results of the present study do not constitute endorsement by the American College of Sports Medicine. The results of the study are presented clearly, honestly, and without fabrication, falsification, or inappropriate data manipulation.

A. G., K. D., R. K., and J. O. H. conceived and designed the study. A. G. and J. R. collected the data. A. G., D. M., and T. P. analyzed the data. A.G. analyzed the data and wrote the manuscript. All authors provided critical feedback on the manuscript before submission.

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Keywords:

ALTITUDE; ENDOGENOUS; EXOGENOUS; UTILIZATION; ENDURANCE

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