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Added Protein Maintains Efficacy of a Low-Carbohydrate Sports Drink

Martínez-Lagunas, Vanessa; Ding, Zhenping; Bernard, Jeffrey R; Wang, Bei; Ivy, John L

Journal of Strength and Conditioning Research: January 2010 - Volume 24 - Issue 1 - p 48-59
doi: 10.1519/JSC.0b013e3181c32e20
Original Research
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Martínez-Lagunas, V, Ding, Z, Bernard, JR, Wang, B, and Ivy, JL. Added protein maintains efficacy of a low-carbohydrate sports drink. J Strength Cond Res 24(1): 48-59, 2010-The purpose of the present study was to investigate the aerobic capacity characteristics of an isocaloric carbohydrate (CHO) plus protein (PRO) drink and a low-calorie CHO plus PRO drink against a traditional 6% CHO sports beverage. Twelve male and female trained cyclists exercised on 4 separate occasions at intensities that varied between 55 and 75% o2max for 2.5 hours and then at 80% o2max until fatigued. Supplements (255.4 ± 9.1 mL) were provided every 20 minutes and consisted of a 4.5% carbohydrate plus 1.15% protein complex (CHO/PRO H), a 3% carbohydrate plus 0.75% protein complex (CHO/PRO L), a 6% carbohydrate supplement (CHO), or a placebo (PLA). Time to fatigue at 80% o2max was significantly longer (p < 0.05) during the CHO (26.9 ± 6.1 minutes, mean ± SE), the CHO/PRO H (30.5 ± 5.9 minutes), and the CHO/PRO L (28.9 ± 6.5 minutes) trials compared with the PLA trial (14.7 ± 3.4 minutes), with no significant differences among the CHO, CHO/PRO H, and CHO/PRO L treatments. In general, blood glucose, plasma insulin, and carbohydrate oxidation were elevated above PLA during the CHO, CHO/PRO H, and CHO/PRO L trials, whereas plasma free fatty acids, rating of perceived exertion, and fat oxidation values were lower during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial. Only minor differences in blood parameters occurred among the CHO, CHO/PRO H, and CHO/PRO L treatments. In summary, partially substituting PRO for CHO in a sports drink did not enhance aerobic capacity, but substitution was able to occur without loss of efficacy. Thus, adding PRO to a low-calorie CHO sports drink may be an effective strategy to enhance aerobic capacity while limiting carbohydrate and caloric consumption.

Department of Kinesiology and Health Education, The University of Texas at Austin Exercise Physiology and Metabolism Laboratory, Austin, Texas

Address correspondence to John L. Ivy, johnivy@mail.utexas.edu.

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Introduction

The benefits of carbohydrate (CHO) supplementation during exercise on aerobic capacity or aerobic performance have been widely studied. Investigators have consistently documented improved aerobic capacity or performance in athletes ingesting CHO beverages vs. water or other placebo beverages during prolonged exercise lasting more than 2 hours (8-10,12,13,15,19,22,24,37,38) or during shorter exercise of high intensity lasting approximately 1 hour (3,25).

More recently, it has been suggested that the addition of small amounts of protein (PRO) or amino acids (AA) to CHO supplementation may further improve aerobic capacity or performance, facilitate recovery, and improve subsequent performance compared with traditional CHO-only sports beverages (2,4,7,21,23,29-31,36,39). For example, Ivy et al. (23) and Saunders et al. (30,31), who compared the performance effect of adding PRO to CHO supplementation during endurance exercise, reported that aerobic capacity was significantly improved after the consumption of a CHO/PRO beverage in comparison with an isocarbohydrate CHO beverage. In these studies, the CHO/PRO beverage contained about 20 to 25% more calories than the CHO-only supplement (23,30,31). Thus, the additional performance effect of the CHO/PRO supplement over the CHO-only supplement reported could have been attributed to the higher caloric content of the CHO/PRO beverage rather than a protein-specific physiological mechanism per se.

To comprehend better the benefits of adding PRO to CHO supplementation, some investigators opted to match the total calories of the CHO and CHO/PRO beverages. Recently, 2 studies designed to evaluate the aerobic capacity (32) and aerobic performance (28) effects of consuming isocaloric CHO/PRO and CHO-only drinks during cycling exercise found no aerobic capacity or performance differences between drinks. However, conditions were different from studies previously reporting improvements in aerobic capacity such as calories ingested, exercise protocols, and amounts of CHO or PRO in the experimental beverages. Therefore, further research is warranted to clarify the potential contributions the addition of PRO to a CHO sports drink can provide in the form of enhancing aerobic capacity. Additionally, investigators have yet to investigate whether a CHO/PRO supplement with less CHO and caloric content could be more effective than or as effective as a traditional CHO-only supplement in improving aerobic capacity. This would be particularly beneficial for athletes and individuals who are concerned about caloric intake during training and control of body weight.

The purpose of the present study was to investigate the ability of PRO to reduce the need for CHO in a sports drink without reducing or possibly improving the efficacy of the drink relative to enhancing aerobic capacity. We hypothesized (a) that aerobic capacity will be enhanced with CHO and CHO plus PRO supplementation compared with a non-caloric placebo beverage and (b) that aerobic capacity would not be compromised and possibly improved when reducing the CHO content of a sports beverage from 6 to 4.5% or 3% when combined with a small amount of protein. Cardiorespiratory measurements and venous blood samples were also collected during the experimental trials to investigate other potential differences in physiological responses among the supplements.

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Methods

Experimental Approach to the Problem

To investigate the ability of PRO to reduce the need for CHO in sports drinks and, thus, possibly reduce the need for excess calories while maintaining or enhancing aerobic capacity, we compare the effects of the following experimental beverages on aerobic capacity as measured by time to fatigue while cycling: (a) traditional 6% carbohydrate sports drink (CHO); (b) 4.5% carbohydrate plus 1.15% protein drink (CHO/PRO H) and 3% carbohydrate plus 0.75% protein beverage (CHO/PRO L); and (c) artificially colored, flavored, and sucralose-sweetened water placebo (PLA).

After baseline testing and a familiarization trial, each subject completed 4 randomized and counterbalanced experimental trials, in which they received one of the above-mentioned experimental beverages at regular intervals during prolonged cycling exercise of varying intensity. The experimental drinks were similar in color, taste, and texture and were randomly distributed by a laboratory technician who was not involved in the data collection. Neither the subjects nor the investigators were aware of the treatments the subjects were on. Therefore, our study followed a double-blind, placebo-controlled, and within-subject repeated measures experimental design.

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Subjects

The subjects were 12 trained cyclists (5 women and 7 men) who had a background in either road cycling or triathlon and who were accustomed to cycling for prolonged periods (3-5 hours). While there may be concern for a mixed gender study population, research has demonstrated that men and women respond to CHO/PRO supplementation in a similar manner (31). Table 1 shows their characteristics in more detail. Before volunteering to participate in this study, all subjects were given a detailed explanation of the purpose, procedures, and potential risks of the study. Additionally, all subjects completed a medical screening questionnaire and signed a written informed consent form. The medical screening questionnaire, the written informed consent form, and the experimental protocol were approved by The University of Texas at Austin's Institutional Review Board before the start of the study.

Table 1

Table 1

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Procedures

Pre-Experimental Procedures

Before the experimental trials, all subjects reported to the laboratory for assessment of their o2max and for familiarization with the exercise protocol. The o2max test was performed on the same electrically braked cycle ergometer used in the familiarization and experimental trials (Lode Excaliber Sport 911900; Lode BV, Groningen, The Netherlands). The protocol for establishing o2max consisted of a 4-minute warm-up followed by a 1-minute stage beginning at 50 W, with increasing workload of 25 W every minute until exhaustion. Subjects breathed through a Hans-Rudolph 2-way, non-breathing valve, with expired gases directed to a mixing chamber for analysis of oxygen (O2) and carbon dioxide (CO2). Inspired volumes were measured using a dry gas meter (Max-1; Physio-Dyne Instruments Corp., Quogue, NY, USA). Outputs from these instruments were directed to a laboratory computer for calculation of ventilation, O2 consumption (o2), CO2 production (co2), and respiratory exchange ratio (RER). The criteria used to establish o2max were a plateau in o2 with increasing exercise intensity and RER >1.10. This respiratory gas analysis system was properly calibrated several minutes before the start of the o2max test and all familiarization and experimental trials.

Seven days after their o2max test, the subjects reported to the laboratory to perform a familiarization trial, which simulated the exercise protocol and supplement timing of the experimental trials, with the exception that only water was provided and there were no blood draws. This familiarization trial was also used to adjust and/or verify appropriate workloads for the experimental trials. After the o2max test, all subjects were instructed to maintain a training log (for the 3 days before their familiarization and experimental trials) and a diet record (for the 2 days before their familiarization and experimental trials). Subjects were asked to keep their physical activity, dietary intake, and sleep pattern as constant as possible for the 48 hours before each trial and to avoid intense exercise training during this period. The subjects were required to provide an electronic copy of their training and dietary logs to the investigators to ensure they maintained constant exercise training and diet patterns. The training and food records recorded before the familiarization trial were used to establish the physical activity and dietary pattern of the subjects for the subsequent experimental trials. The training and diet were not standardized among subjects because each subject served as his or her own control.

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Experimental Procedures

After baseline testing and familiarization, each subject completed 4 randomized experimental trials, separated by 7 days, in which CHO, CHO/PRO H, CHO/PRO L, or PLA was provided during exercise. Possible trial sequence was established, and the trial sequence then randomly selected for each subject. PacificHealth Laboratories, Inc., Matawan, NJ, provided all experimental drinks. The protein consisted of whey protein, and the carbohydrate consisted of dextrose. Detailed ingredients of each supplement are presented in Table 2.

Table 2

Table 2

The amount of fluid ingested during exercise was standardized depending on the body weight of each subject. The rates of carbohydrate delivery for the CHO, CHO/PRO H, and CHO/PRO L supplements were 0.7, 0.52, and 0.35 g·kg1·h1, respectively. Protein was provided at a delivery rate of 0.13 g·kg1·h1 with the CHO/PRO H supplement and 0.09 g·kg1·h1 with the CHO/PRO L supplement. On average, subjects ingested 255.4 ± 9.1 ml (mean ± SE) of fluid every 20 minutes until fatigue, which ensured a carbohydrate delivery rate of 46.0 ± 1.6 g·h1 for the CHO supplement, 34.5 ± 1.2 g·h1 for the CHO/PRO H supplement, and 23.0 ± 0.8 g·h1 for the CHO/PRO L supplement. The protein delivery rate was 8.8 ± 0.3 g·h1 for the CHO/PRO H supplement and 5.7 ± 0.2 g·h1 for the CHO/PRO L supplement. Kilocalories delivered were 184 ± 6.4·h1, 173 ± 6.0·h1, and 114 ± 1.0·h1 for the CHO, CHO/PRO H, and CHO/PRO L supplements, respectively. All subjects performed each trial in a room of 21 to 23°C at the same time of the day (early morning, either 6:30 or 7:00 am) and the same day of the week over a 4-week period. A floor fan was directed toward the exercising subject to reduce thermal stress.

On the day of an experimental trial, the subjects reported to the laboratory 30 minutes before the start of exercise of having fasted for 12 hours. During the fasting time, the subjects were allowed to consume only water. On reporting to the laboratory, the subjects were weighed and fitted with a heart rate (HR) monitor (Polar Beat; Polar Electro Oy, Kempele, Finland) secured in place around their chest. A catheter was also inserted into a forearm vein of the subjects, which was fitted with a 3-way stopcock, extended with a catheter extension, and taped in place. The catheter was kept patent with regular sterile saline flushes every 15 minutes during the exercise protocol. Once the catheter was in place, the subjects mounted the cycle ergometer. After the remaining seated for 2 to 3 minutes, a resting blood sample was drawn, HR was recorded, the first drink was provided, and then the cycling exercise started.

The exercise protocol used was a modification of the one used by Brouns (6) and is illustrated in Figure 1. Cycling started with a 24-minute warm-up at 55% o2max and was followed by twelve 8-minute intervals alternating between 55 and 75% o2max. This sequence was then followed by 10 shorter intervals of 3 minutes each alternating between the same intensities of 55 and 75% o2max. After this sequence, subjects cycled at 80% o2max until fatigue. At this stage of the exercise protocol, subjects were required to stay on the saddle at all times. All subjects received constant verbal encouragement to ride as long as possible while maintaining a pedaling cadence of 70 to 100 rpm. When the subjects could no longer maintain a pedaling cadence of 60 rpm for the first time, one of the investigators manually helped to restore pedal cadence above 60 rpm. Once the subjects dropped their pedaling rate for the second time to less than 60 rpm, they were asked to stop the exercise and this was considered their point of fatigue. During each ride, the subjects were not aware of how long they had ridden, as all timing devices were removed from their sight. No feedback on performance was provided to the subjects until the study was completed.

Figure 1

Figure 1

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Data and Sample Collection and Analyses

Ventilation, o2, co2, and RER were recorded at different times throughout the exercise protocol (see Figure 1) with the respiratory gas analysis system used for o2 assessment to verify that the subjects were working at the proper intensity and to determine energy expenditure and CHO and fat oxidation rates. Average energy expenditure (kcal·min1) was computed from RER and o2 (L·min−1) using the Weir equation (35) with the assumption that PRO oxidation during exercise was negligible. CHO and fat oxidation rates (g·min−1) were calculated from co2 and o2 (L·min1) using the formulas used by Frayn (16).

Respiratory gas collection periods were limited to 3 to 6 minutes, but only the middle 1 to 3 minutes of each collection was used to determine o2 and RER (the first and last minute of each collection were not considered for these calculations) to assure more accurate readings at the different exercise intensity stages. Heart rate and subjective ratings of perceived exertion (RPE) on a Borg Scale ranging from 6 to 20 (5) were recorded at the same time as the respiratory gas collections (see Figure 1).

Five milliliters of venous blood was drawn before the start of exercise, at the end of predetermined exercise stages, and immediately after the cessation of exercise (see Figure 1). One drop of blood (150 μL) was immediately used to measure plasma glucose concentration in duplicate with a glucose meter (One Touch Basic; Lifescan, Inc., Milpitas, CA, USA). The average of the 2 glucose measurements was recorded. Before using the glucose meter in the study, its validity and reliability were verified by comparing values obtained from the glucose meter with those from a YSI23A glucose analyzer (Yellow Spring Instruments, Yellow Springs, OH, USA). The glucose meter was calibrated at the beginning of each experimental trial using standards provided by Lifescan, Inc. The coefficient of variation (CV) for the glucose assay was 8.8%. After blood glucose determination, each blood sample was anticoagulated with 0.2 mL of EDTA (24 mg·mL−1, pH 7.4). Then, 0.5 ml of the anticoagulated blood was transferred to another tube containing 1 mL 10% perchloric acid (PCA). All tubes were centrifuged for 10 minutes at 3,000 rpm with a JS-7.5 rotor in a Beckman J2-21 centrifuge. Plasma and PCA extracts were separated into aliquots for each assay and stored in mircofuge tubes at −80°C until further analysis. The plasma samples were analyzed for insulin and free fatty acids (FFAs). Plasma insulin was assayed using a radioimmunoassay kit (MP Biomedicals, Orangeburg, NY, USA) based on the principles described by Goetz et al. (18) and had a CV of 6.0%. Plasma FFAs were measured using the colorimetric assay procedure of Duncombe (14) but modified by using the extraction reagent of Noma et al. (26) and had a CV of 4.5%. Blood lactate was determined from the PCA extracts by enzymatic analysis according to Hohorst (20). The CV for the lactate assay was 1.5%. All assays were run in duplicate.

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Statistical Analyses

All dependent variables were analyzed using a general linear model for repeated measures. Time to fatigue was analyzed using a 1-way analysis of variance (ANOVA). All the other dependent variables that included multiple measures per trial (HR, RPE, energy expenditure, RER, carbohydrate and fat oxidation, blood glucose and lactate, and plasma insulin and FFAs) were analyzed using a 2-way (treatment × time) ANOVA. The independent variables were the 4 different experimental beverages used: CHO, CHO/PRO H, CHO/PRO L, and PLA. Significant differences among mean were determined using the Fisher Least Square Difference procedure. The level of significance for all analyses was set at p ≤ 0.05. All data are expressed as mean ± SE unless otherwise indicated. SPSS for Windows GradPack version 16.0 (SPSS, Inc., Chicago, IL, USA) was used for all statistical analyses.

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Results

Aerobic Capacity

As expected, the average time to fatigue during exercise at 80% o2max after 150 minutes of varying intensity cycling was significantly longer (p < 0.05) during the CHO, the CHO/PRO H, and the CHO/PRO L trials compared with the PLA trial (Figure 2). The increase in time to fatigue was 83.0% longer during the CHO trial, 107.5% during the CHO/PRO H trial, and 96.6% during CHO/PRO L trial compared with the PLA trial. Comparing the CHO and CHO/PRO H trials, 9 of 12 subjects performed better on the CHO/PRO trial. However, this tendency for the subjects to ride longer duirng the CHO/PRO H trial compared with the CHO trial was not significant (p = 0.073). Time to fatigue for the CHO/PRO L trial did not differ from that of the CHO/PRO H trial or the CHO trial. In addition, there was no difference in the response of the male and female subjects.

Figure 2

Figure 2

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Heart Rate and Perceived Exertion

Heart rate gradually rose during the course of exercise. Average HR (from 5 to 152 minutes of HR collection) was 144.9 ± 3.8 b·min1 during the CHO trial, 146.1 ± 3.4 b·min1 during the CHO/PRO H trial, 144.0 ± 3.6 b·min1 during the CHO/PRO L trial, and 143.5 ± 3.3 b·min1 during the PLA trial. No significant treatment or treatment × time differences in HR were found among the 4 experimental beverages.

Rate of perceived exertion increased gradually during the course of exercise. Average RPE was significantly lower (p < 0.05) during the CHO (12.8 ± 0.4), CHO/PRO H (12.3 ± 0.4), and CHO/PRO L (12.6 ± 0.4) trials compared with the PLA trial (13.1 ± 0.4). Additionally, RPE during the CHO/PRO H trial was significantly lower than during the CHO trial.

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Respiratory Exchange Ratio, Energy Expenditure, and Substrate Utilization

The average RER during the CHO (0.899 ± 0.005), the CHO/PRO H (0.893 ± 0.006), and the CHO/PRO L (0.888 ± 0.008) trials was significantly higher (p < 0.05) than during the PLA trial (0.872 ± 0.007). Average RER was not significantly different among the CHO, CHO/PRO H, and CHO/PRO L trials (Table 3). Additionally, some significant RER differences (treatment × time) were found from 40 to 152 minutes of exercise. Generally, RER was significantly higher during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial, but no significant differences were identified among the CHO, CHO/PRO H, and CHO/PRO L treatments.

Table 3

Table 3

Average energy expenditure was 12.24 ± 0.87 kcal·min1 during the CHO trial, 12.16 ± 0.88 kcal·min1 during the CHO/PRO H trial, 12.26 ± 0.94 kcal·min1 during the CHO/PRO L trial, and 12.07 ± 0.83 kcal·min1 during the PLA trial. No significant treatment or treatment × time differences in energy expenditure were found among the 4 experimental beverages.

Average CHO oxidation during the CHO (2.23 ± 0.17 g·min1), CHO/PRO H (2.14 ± 0.15 g·min1), and CHO/PRO L (2.11 ± 0.17 g·min1) trials was significantly higher (p < 0.05) than during the PLA trial (1.90 ± 0.15 g·min1). There were no significant differences in average CHO oxidation among the CHO, CHO/PRO H, and CHO/PRO L trials (Table 3). Several significant differences (treatment × time) were found in CHO oxidation from 40 to 152 minutes of exercise. In general, CHO oxidation was significantly higher during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial, but no significant differences were found among the CHO, CHO/PRO H, and CHO/PRO L treatments.

Average fat oxidation during the CHO (0.41 ± 0.04 g·min1), CHO/PRO H (0.44 ± 0.05 g·min1), and CHO/PRO L (0.46 ± 0.05 g·min1) trials was significantly lower (p < 0.05) than during the PLA trial (0.53 ± 0.05 g·min1). Average fat oxidation was not significantly different among the CHO, CHO/PRO H, and CHO/PRO L treatments (Table 3). Several significant differences (treatment × time) were found in fat oxidation from 40 to 135 minutes of exercise. Overall, fat oxidation was significantly lower during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial, but no significant differences were identified among the CHO, CHO/PRO H, and CHO/PRO L treatments.

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Blood Analyses

Average blood glucose was significantly higher (p < 0.05) during the CHO (4.11 ± 0.11 mmol·L1), CHO/PRO H (4.10 ± 0.11 mmol·L1), and CHO/PRO L (3.94 ± 0.10 mmol·L1) trials compared with the PLA trial (3.61 ± 0.10 mmol·L1). No significant differences in average blood glucose were found among the CHO, CHO/PRO H, and CHO/PRO L trials (Figure 3). Additionally, treatment × time differences occurred from 32 minutes of exercise to fatigue. Overall, blood glucose was significantly higher during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial, and also some significant differences among the CHO, CHO/PRO H, and CHO/PRO L occurred at 32 minutes, 144 minutes, and at fatigue. At 32 minutes of exercise, blood glucose was significantly higher during the CHO trial than during the CHO/PRO H and CHO/PRO L trials. At 144 minutes, blood glucose was significantly higher during the CHO and CHO/PRO H trials compared with the CHO/PRO L trial. At fatigue, blood glucose was significantly higher during the CHO/PRO H trial compared with the CHO, CHO/PRO L, and PLA trials.

Figure 3

Figure 3

Average plasma insulin was significantly higher (p < 0.05) during the CHO (10.17 ± 0.77 mlU·L1), CHO/PRO H (10.83 ± 0.46 mlU·L1), and CHO/PRO L (9.79 ± 0.43 mlU·L1) trials compared with the PLA trial (7.83 ± 0.40 mlU·L1). Additionally, average plasma insulin during the CHO/PRO H trial was significantly higher than during the CHO/PRO L trial. Plasma insulin concentration decreased during exercise in all trials, and several significant differences (treatment × time) occurred from rest to fatigue (Figure 4). Overall, plasma insulin was significantly higher during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial, and also some significant differences among the CHO, CHO/PRO H, and CHO/PRO L occurred at 104, 144, and 147 minutes of exercise. At 104 and 144 minutes of exercise, plasma insulin was significantly higher during the CHO/PRO H trial compared with the CHO/PRO L trial. At 147 minutes, plasma insulin was significantly higher during the CHO trial and the CHO/PRO H trial vs. the CHO/PRO L trial. At fatigue, plasma insulin was significantly higher during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial.

Figure 4

Figure 4

Blood lactate concentration rose and fell during the course of exercise in relationship to the changes in exercise intensity but was significantly higher (p < 0.05) at fatigue compared with at rest in all 4 trials (Figure 5). However, no significant differences in blood lactate concentration were identified during the course of exercise among the trials.

Figure 5

Figure 5

Average plasma FFA concentration was significantly lower during (p < 0.05) the CHO (0.21 ± 0.02 mmol·L1), CHO/PRO H (0.26 ± 0.03 mmol·L1), and CHO/PRO L (0.27 ± 0.03 mmol·L1) trials compared with the PLA trial (0.43 ± 0.05 mmol·L1). Additionally, average plasma FFA concentration during the CHO trial was significantly lower than during the CHO/PRO L trial. Plasma FFA concentration progressively increased during the course of exercise in all trials, and several significant differences (treatment × time) among trials occurred from 32 minutes of exercise to fatigue (Figure 6). Overall, plasma FFA concentration was significantly higher during the PLA trial compared with the CHO, CHO/PRO H, and CHO/PRO L trials, and also significant differences among the CHO, CHO/PRO H, and CHO/PRO L occurred at 32 minutes of exercise and at fatigue. At 32 minutes of exercise, plasma FFA concentration was significantly lower during the CHO trial compared with the CHO/PRO H trial. At fatigue, plasma FFA concentration was significantly lower during the CHO compared with the CHO/PRO L trial.

Figure 6

Figure 6

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Discussion

The primary objective of the present study was to test the ability of protein to reduce the need for carbohydrate in sports drinks and thus possibly reduce the need for excess calories while maintaining or enhancing aerobic capacity. To achieve this objective, we compared the effects of 4 different experimental supplements (CHO, CHO/PRO H, CHO/PRO L, and PLA) on time to fatigue during prolonged cycling of varying intensity.

Our study results show that time to fatigue during the CHO, CHO/PRO H, or CHO/PRO L trials were indeed significantly longer compared with the PLA trial. This finding confirms our first research hypothesis and it is in agreement with previous studies that have shown that CHO and CHO plus PRO supplementation during exercise enhances aerobic capacity or aerobic performance compared with water or other placebo beverages (3,4,7-10,13,15,19,21-23,30,36-39). For the most part, investigators agree that CHO supplementation during exercise improves endurance capacity or performance by maintaining euglycemia and a higher rate of CHO oxidation, especially during the latter stages of exercise (8-10,19,22,37). In the present study, both blood glucose (from 32 minutes of exercise to fatigue) and CHO oxidation (from 40 to 152 minutes of exercise) were significantly higher during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial. Thus, it seems that the improved performance after the CHO-supplemented trials in this study was indeed related to these 2 commonly proposed mechanisms. Our RPE results also confirm the additional aerobic capacity benefit of supplementing with CHO, CHO/PRO H, or CHO/PRO L compared with PLA. Rate of perceived exertion was significantly lower during the CHO, CHO/PRO H, or CHO/PRO L trials compared with the PLA trial from 48 to 152 minutes of exercise.

Time to fatigue was not significantly different between the CHO, and isocaloric CHO/PRO H treatment or the hypocaloric CHO/PRO L treatment. This finding supports the first part of our second hypothesis, which stated that aerobic capacity would not be compromised when the amount of CHO in a sports drink was reduced from 6 to 4.5% or 3% if replaced or partially replaced with a small amount of PRO. However, the study findings did not support the second part of this hypothesis because aerobic capacity was not enhanced after the ingestion of either the CHO/PRO H or the CHO/PRO L treatment beyond that produced by the CHO-only treatment.

Ivy et al. (23) and Saunders et al. (30,31) reported improved aerobic capacity with adding PRO to CHO supplementation during exercise. Ivy et al. (23) reported a 36% performance improvement during prolonged exercise of varying intensity, while Saunders et al. (30) reported a 29 and 40% performance improvement for subjects cycling at 75% and again at 85% o2peak to fatigue, respectively. In a subsequent study, Saunders et al. (31) reported a performance benefit of 13% during a ride to exhaustion at 75% o2peak when subjects consumed a CHO plus PRO gel vs. a CHO-only gel. The experimental supplements in these 3 studies were matched for total CHO content but not for total caloric content (23,30,31). Thus, the CHO/PRO supplements in these studies contained 20 to 25% more calories than their CHO-only supplements. In this regard, several recent studies found that exercise performance was not improved by the addition of protein to a CHO supplement when the CHO/PRO and CHO supplements tested were isocaloric (28,32). Consequently, the additional performance effect of the CHO/PRO supplement over the CHO-only supplement reported in the studies by Ivy et al. (23) and Saunders et al. (30,31) could be attributed to the higher caloric content of the CHO/PRO supplement rather than a protein-mediated physiological mechanism per se. While our present results support this line of reasoning, it should be noted that there was a tendency for the CHO/PRO H supplement to perform better than the CHO supplement as 9 of 12 subjects cycled longer when consuming the CHO/PRO H supplement. Furthermore, subjects performed as well with the hypocaloric CHO/PRO L supplement as with the CHO supplement.

To our knowledge, this is the first study that compared the performance effects of a low-calorie CHO plus PRO beverage vs. a CHO-only supplement. The finding that the CHO/PRO L supplement was as effective in improving endurance performance as the traditional 6% CHO sports drink despite containing 50% less carbohydrate and approximately 38% fewer calories is novel. This finding and the tendency for improved aerobic capacity with the CHO/PRO H supplement suggest that the added protein may benefit aerobic capacity other than by simply providing an additional fuel source. A limitation to the study design, however, is the absence of a low CHO-only trial. Only a performance reduction with a low CHO supplement can validate the necessity for adding protein to a low CHO supplement. However, the maximal rate of carbohydrate oxidation when consuming a liquid supplement of 6 to 8% glucose is 60-70 g·h1 (11,24), and recommendations for CHO ingestion during prolonged exercise range between 30 and 60 g·h1 (1,24). In the present study, 46.0 ± 1.6, 34.5 ± 1.2, and 23.0 ± 0.8 g of carbohydrate were delivered per hour for the CHO, CHO/PRO H, and CHO/PRO L treatments, respectively. Clearly, the carbohydrate deliver rate during the CHO/PRO L treatment is well below that recommended, and while a low amount of carbohydrate may have an ergogenic effect, it is likely not to have the efficacy of a high-carbohydrate supplement or a low-carbohydrate supplement with added protein.

Van Essen and Gibala (33) found no difference in 80-km cycling time trial performance when comparing a 2% protein, 6% carbohydrate drink (CHO-PRO) with a 6% carbohydrate (CHO) drink. Carbohydrate was provided at a rate of 60 g·h1, a rate substantially higher than used in studies that found a significant benefit in performance with the CHO/PRO supplement (23,30,31). Van Essen and Gibala (33) suggested that when CHO is ingested at levels that approach the optimal rate of CHO oxidation of approximately 60-70 g·h1 (24), the addition of PRO to a CHO supplement does not further enhance performance. Their argument is supported by the findings of Osterberg et al. (27). In the study by Osterberg et al. (27), subjects completed 120 minutes of constant-load ergometer cycling followed by a time trial in which a set amount of work (7 kJ·kg−1) was completed as quickly as possible. Subjects received 250 ml at 15-minute intervals of a 6% CHO, 7.5% CHO/1.6% PRO, or placebo supplement. No difference in performance was noted between the CHO and CHO/PRO treatments. Therefore, the beneficial effects of added protein to a sports drink may only occur when suboptimal amounts of carbohydrate are provided.

To determine the mechanism(s) by which partial replacement of CHO with PRO maintains aerobic capacity, we investigated some potential physiological response differences in cardiorespiratory measurements and blood metabolites among our treatments. We identified only minor differences among the CHO, CHO/PRO H, and CHO/PRO L treatments in blood glucose, plasma insulin, plasma FFA, and RPE. In addition, we found no significant differences in HR, energy expenditure, RER, carbohydrate and fat oxidation, or blood lactate among these treatments.

During variable intensity exercise, CHO supplementation has been found to spare muscle glycogen utilization (38). Moreover, postexercise muscle glycogen resynthesis seems to be enhanced when supplemented with a CHO/PRO supplement as compared with a CHO-only supplement (21,39). It is therefore possible that the efficacy of the CHO/PRO supplementation was maintained by eliciting a greater sparing of muscle glycogen and possibly liver glycogen. In the present study, we did not directly assess glycogen stores, and therefore, further investigation is required to determine whether adding PRO to CHO supplementation produces a greater sparing of glycogen during variable intensity exercise than previously demonstrated with a CHO supplement. However, it should be noted that subjects did not become hypoglycemic during the CHO/PRO trials, and in fact, during the CHO/PRO H trial, blood glucose was significantly higher at fatigue than all other trials.

Another possibility for the effectiveness of the CHO/PRO supplements is a reduction in exercise-induced muscle damage. Both Saunders et al. (30) and Romano-Ely et al. (28) reported that CHO plus PRO supplementation significantly attenuated postexercise muscle damage (indirectly assessed by postexercise levels of plasma creatine kinase [CPK] and lactate dehydrogenase [LDH]) compared with CHO-only supplementation. Saunders et al. (30) compared isocarbohydrate treatments and reported that CPK values at 12-15 hours post exercise were 83% lower after the CHO plus PRO trial than during the CHO-only trial. Romano-Ely et al. (28) compared isocaloric treatments and reported that CPK (53%) and LDH (9%) postexercise levels at 24 hours were significantly lower in the CHO plus PRO trial compared with the CHO trial. More recently, Valentine et al. (32) demonstrated that a CHO/PRO supplement was not only more effective than an isocaloric or isocarbohydrate supplement in attenuating postexercise muscle damage (indirectly assessed by CPK and myoglobin levels) but also in improving muscle function. The latter because the number of knee extensions performed at 70% 1-repetition maximum 24 hours after the experimental trial was significantly higher after the CHO/PRO supplementation compared with their other treatments. In addition, other investigators have found that CHO plus PRO supplementation could reduce muscle damage during resistance exercise (2). Thus, the physiological mechanism(s) by which the replacement of PRO for CHO maintains the efficacy of a performance-enhancing supplement could be related to maintaining the integrity of the muscle tissue.

Finally, the addition of protein could provide precursors for the anaplerotic reactions required to maintain Krebs cycle intermediates in skeletal muscle. At the onset of exercise, there is a rapid expansion of most but not all Krebs cycle intermediates. As exercise persists, the concentration of oxaloacetate and α-ketoglutarate can reach critically low levels (17). It has been proposed that fatigue during prolonged exercise may result from the inability of mitochondria to sustain aerobic energy production due to the depletion of 1 or more Krebs cycle intermediates (34). Although carbohydrate supplementation is thought to provide some assistance with this process, it may not be as effective as protein supplementation in providing the AA that directly enter into the anaplerotic reactions required for the replacement of essential Krebs cycle intermediates.

In summary, the CHO/PRO H supplement was not found to be anymore beneficial in improving aerobic capacity than a traditional 6% isocaloric CHO supplement. However, it was found that with the addition of protein, the carbohydrate and caloric content of a sports drink could be substantially reduced without loss of efficacy. While the mechanism by which the protein works is unknown, the finding has significant implications for endurance athletes and individuals who are concerned about caloric intake during training and control of body weight. Further investigation regarding the specific physiological mechanisms by which added protein maintains the efficacy of a sports drink when it contains reduced levels of carbohydrate, or when suboptimal levels of carbohydrate are provided, is warranted.

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Practical Applications

Nutritional supplementation during exercise is important for maintaining a quality workout regimen and performing optimally during competition. However, many athletes compete in sports in which weight control is essential and nutritional supplementation is avoided for fear of unwanted weight gain. Moreover, individuals who exercise regularly to maintain optimal cardiovascular fitness and body composition are reticent about consuming nutritional supplements during exercise or post exercise for fear of increasing their caloric intake and limiting improvements in body composition they are striving to obtain. The present study demonstrates that the carbohydrate and caloric content of a sport drink can be significantly reduced and that the efficacy of the drink maintained with the addition of a small amount of protein. A low-carbohydrate beverage containing a small amount of protein thus provides the ergogenic properties of more traditional sports drinks, while alleviating the concerns of the athlete or regular exerciser consuming unnecessary calories or simple carbohydrates.

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Acknowledgments

The authors would like to sincerely thank the subjects for their outstanding effort and dedication throughout the course of the study. This research project was funded by Pacific Health Laboratories, Inc., Matawan, NJ. No limitations or restrictions were placed on publication of data by the funding source. The results of this study do not constitute endorsement of any products by the authors or National Strength and Conditioning Association.

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Keywords:

aerobic capacity; endurance; exercise metabolism; fatigue

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