NAP: Recovery (2014)


Howath KR, Moreau NA, Phillips SM, Gibala MJ. Coingestion of protein with carbohydrate during recovery from endurance exercise stimulates skeletal muscle protein synthesis in humans. J Appl Physiol. 2009; 106: 1,394-1,401.

Study Design:
Randomized Crossover Trial
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Quality Rating:
Positive POSITIVE: See Quality Criteria Checklist below.
Research Purpose:
  • To determine whether ingesting protein with CHO during recovery from prolonged exercise would increase mixed skeletal muscle protein fractional synthetic rate (FSR) and improve whole body protein balance compared with CHO alone
  • A secondary purpose was to determine whether adding protein or additional CHO to a feeding strategy that provided 1.2g CHO per kg-1 per hour-1, the rate generally recommended to maximize post-exercise glycogen synthesis, would further augment this process during recovery. 
Inclusion Criteria:

Recreationally active and habitually engaged in a variety of activities that included running, cycling, weightlifting and intramural sports several times per week.

Exclusion Criteria:
  • Specifically training for a particular sport or event
  • Cardiovascular, pulmonary or metabolic disease.
Description of Study Protocol:
  • Recruitment: Not described 
  • Design: Randomized crossover trial
  • Blinding used: Not mentioned.


Three experimental beverages ingested in random order at a rate of 750ml per hour, in 15-minute intervals during the first three hours of each recovery period:
  • 1.2g CHO per kg body mass-1 per hour-1 (L-CHO), 1.2g CHO+0.4g protein per kg body mass-1 per hour-1 (PRO-CHO; matched to L-CHO for total CHO ingested) or 1.6g CHO per kg body mass-1 per hour-1 (H-CHO; matched to PRO-CHO for total energy ingested)
  • The CHO source was maltodextrin and the protein hydrolyzed whey protein concentrate. To make the drinks comparable in taste, 5g sucralose and orange powder flavoring were added to 750ml of each beverage. To maintain constant infusion of L-(ring-2H5)Phe, an amount of isotope equivalent to 9% of the Phe present in the whey protein was added to the PRO-CHO drinks. 

Statistical Analysis

  • Muscle and blood data using a two-factor (treatment x time) repeated-measures ANOVA, except for muscle FSR and glycogen synthesis rates, which were analyzed using a one-factor (treatment) repeated-measures ANOVA
  • When a significant main effect or interaction was identified, data were subsequently analyzed using Tukey honestly significant difference post-hoc test
  • Integrated AUC calculations were performed and analyzed using a one-factor (treatment) ANOVA. 
Data Collection Summary:

Timing of Measurements

Blood sample at baseline and at cessation of exercise and then at 15-minute intervals for the initial hour of recovery and 30 minutes thereafter; breath sample during the final 15 minutes of the first and fourth hours of recovery; muscle biopsy post- two hours exercise and post- four hours recovery at each of three trials separated by seven days.

Dependent Variables

  • Gastrointestinal distress: Questionnaire after exercise and each hour of recovery to assess 11 parameters ranked on a scale from one to 10, with one being "the absence of any distress" and 10 being "distress strongly present"
  • Mixed muscle fractional synthetic rates (FSR): Gas chromatography mass spectrometry of freeze-dried, powdered muscle biopsy. Based on L-(ring-2H5)phenylalanine infusion
  • Glycogen: Enzymatic assay adapted for fluorometry of freeze-dried muscle
  • Amino acids: Freeze-dried muscle extract using HPLC
  • Glucose, insulin and AUC for insulin: Blood samples
  • Ratio of 13CO2 to 12CO2: Automated breath analysis system
  • Whole body protein kinetics: (l-13C)leucine tracer using equations of Matthews, et al.

Independent Variables

Low CHO vs. high CHO vs. CHO+protein beverage.

Control Variables

  • Standardized meal three hours prior to exercise, two-hour bout of standardized variable-intensity cycling
  • Subjects were asked to keep their habitual exercise pattern and dietary intake as constant as possible over the course of the experiment
  • No physical activity was allowed for 48 hours before each trial and subjects kept a food record for 48 hours before each trial; results showed no difference in total energy intake or macronutrient composition between trials
  • In addition to general nutrition controls, all subjects were provided with a standardized, pre-packaged meal on the day before each experimental trial. Subjects were instructed to ingest the meal as breakfast at 7:00 a.m. on the day of the trial, after having fasted overnight. The meal provided 700kcal and was derived from 82% carbohydrate, 10% fat, 8% protein. Subjects were instructed not to consume any other food or drink, except for water, before reporting to the laboratory.
Description of Actual Data Sample:
  • Initial N: Six males
  • Attrition (final N): Six
  • Age: 22±1 years
  • Ethnicity: Not described
  • Other relevant demographics: VO2peak, 4.4±0.3 L per minute
  • Body mass: 90±5kg 
  • Height: 184±2cm 
  • BMI: 26.4±0.8kg/m2.
Hamilton, Ontario, Canada.
Summary of Results:


  • Leucine flux and oxidation were higher during PRO-CHO compared with H-CHO and L-CHO (main effect treatment, P<0.05) with both variables being higher at four hours vs. one hour (P<0.05)
    • Non-oxidative Leu disposal (NOLD) was lower after four hours vs. one hour (main effect time, P<0.05), with no difference between trials
    • Whole body protein breakdown was lower at one and four hours of recovery during PRO-CHO vs. the other trials (P<0.05)
    • Leucine net balance was highest at one hour of recovery during the PRO-CHO trial (P<0.05)
    • While leucine net balance decreased from one to four hours in all trials (main effect time, P<0.05), it remained positive in the PRO-CHO trial while it was negative in the other two trials (main effect, P<0.05).
  • Muscle FSR was higher (P<0.05) during recovery in PRO-CHO, compared with the other two trials
  • Muscle glycogen content was similar between trials after exercise and increased to a similar extent in all trials over the four-hour period of recovery (main effect for time, P<0.05)
  • There was no difference between trials in muscle glycogen synthesis rate during recovery
  • Total muscle branched-chain amino acid (BCAA) content was higher (P<0.04) after four hours of recovery during PRO-CHO vs. the other two trials, mainly due to changes in leucine and isoleucine
  • After one hour of recovery, the blood concentration of alanine, leucine, isoleucine, valine, arginine, lysine, phenylalanine, proline, threonine and tyrosine were higher in PRO-CHO vs. the other trials and remained higher for the rest of the recovery (P<0.05)
    • Total amino acids were higher during PRO-CHO compared with L-CHO by one hour of recovery and were higher than both CHO trials by 2.5 hours of recovery (P<0.05)
    • Total BCAA concentrations increased during the first hour of recovery during PRO-CHO, compared with the CHO trials and were significantly higher by one hour of recovery and remained higher throughout the remainder of recovery (P<0.05).
  • There was no difference between trials in glucose, insulin, AUC for insulin or total GI distress scores.
Author Conclusion:
  • Ingesting protein with CHO during recovery from prolonged endurance exercise increased muscle FSR and improved whole body net protein balance, compared with CHO alone
  • However, adding protein or additional CHO to a drinking strategy that provided 1.2g CHO per kg-1 per hour-1 did not further augment glycogen synthesis. 
Funding Source:
Natural Sciences and Engineering Research Council of Canada
Reviewer Comments:
Quality Criteria Checklist: Primary Research
Relevance Questions
  1. Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies) Yes
  2. Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about? Yes
  3. Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice? Yes
  4. Is the intervention or procedure feasible? (NA for some epidemiological studies) Yes
Validity Questions
1. Was the research question clearly stated? Yes
  1.1. Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified? Yes
  1.2. Was (were) the outcome(s) [dependent variable(s)] clearly indicated? Yes
  1.3. Were the target population and setting specified? Yes
2. Was the selection of study subjects/patients free from bias? Yes
  2.1. Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study? Yes
  2.2. Were criteria applied equally to all study groups? Yes
  2.3. Were health, demographics, and other characteristics of subjects described? Yes
  2.4. Were the subjects/patients a representative sample of the relevant population? Yes
3. Were study groups comparable? Yes
  3.1. Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT) Yes
  3.2. Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline? Yes
  3.3. Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.) Yes
  3.4. If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis? N/A
  3.5. If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.) N/A
  3.6. If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")? N/A
4. Was method of handling withdrawals described? Yes
  4.1. Were follow-up methods described and the same for all groups? Yes
  4.2. Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.) N/A
  4.3. Were all enrolled subjects/patients (in the original sample) accounted for? Yes
  4.4. Were reasons for withdrawals similar across groups? N/A
  4.5. If diagnostic test, was decision to perform reference test not dependent on results of test under study? N/A
5. Was blinding used to prevent introduction of bias? No
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? No
  5.2. Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.) N/A
  5.3. In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded? N/A
  5.4. In case control study, was case definition explicit and case ascertainment not influenced by exposure status? N/A
  5.5. In diagnostic study, were test results blinded to patient history and other test results? N/A
6. Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described? Yes
  6.1. In RCT or other intervention trial, were protocols described for all regimens studied? Yes
  6.2. In observational study, were interventions, study settings, and clinicians/provider described? N/A
  6.3. Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect? Yes
  6.4. Was the amount of exposure and, if relevant, subject/patient compliance measured? Yes
  6.5. Were co-interventions (e.g., ancillary treatments, other therapies) described? N/A
  6.6. Were extra or unplanned treatments described? N/A
  6.7. Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups? N/A
  6.8. In diagnostic study, were details of test administration and replication sufficient? N/A
7. Were outcomes clearly defined and the measurements valid and reliable? Yes
  7.1. Were primary and secondary endpoints described and relevant to the question? Yes
  7.2. Were nutrition measures appropriate to question and outcomes of concern? Yes
  7.3. Was the period of follow-up long enough for important outcome(s) to occur? Yes
  7.4. Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures? Yes
  7.5. Was the measurement of effect at an appropriate level of precision? Yes
  7.6. Were other factors accounted for (measured) that could affect outcomes? N/A
  7.7. Were the measurements conducted consistently across groups? Yes
8. Was the statistical analysis appropriate for the study design and type of outcome indicators? Yes
  8.1. Were statistical analyses adequately described and the results reported appropriately? Yes
  8.2. Were correct statistical tests used and assumptions of test not violated? Yes
  8.3. Were statistics reported with levels of significance and/or confidence intervals? Yes
  8.4. Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)? N/A
  8.5. Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)? N/A
  8.6. Was clinical significance as well as statistical significance reported? Yes
  8.7. If negative findings, was a power calculation reported to address type 2 error? N/A
9. Are conclusions supported by results with biases and limitations taken into consideration? Yes
  9.1. Is there a discussion of findings? Yes
  9.2. Are biases and study limitations identified and discussed? Yes
10. Is bias due to study's funding or sponsorship unlikely? Yes
  10.1. Were sources of funding and investigators' affiliations described? Yes
  10.2. Was the study free from apparent conflict of interest? Yes