NAP: Recovery (2014)


Beelen M, Van Kranenburg J, Senden JM, Kuipers H, Van Loon LJC. Impact of caffeine and protein on postexercise muscle glycogen synthesis. Med Sci Sports Exerc. 2012; 44 (4): 692-700.

Study Design:
Randomized Crossover Trial
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Quality Rating:
Positive POSITIVE: See Quality Criteria Checklist below.
Research Purpose:

To assess the effect of protein or caffeine co-ingestion on post-exercise muscle glycogen synthesis rates when optimal amounts of CHO (1.2g per kg-1 per hour-1) is consumed. 

Inclusion Criteria:

Male cyclist who cycled at least 100km per week-1 and had a training history of more than three years.

Exclusion Criteria:
  • Females
  • Cycling less than 100km per week-1.
Description of Study Protocol:
  • Recruitment: Not described
  • Design: Randomized crossover trial; three tests separated by at least one week 
  • Blinding used: Double-blind.


  • One of three beverages given in amounts of three ml per kg-1 every 30 minutes during recovery (randomized order):
    • CHO only (CHO): 1.2g per kg-1per hour-1 CHO
    • CHO+protein+leucine mixture (CHO+PRO): 1.2g per kg-1per hour-1 CHO with 0.2g per kg-1per hour-1 casein protein hydrolysate and 0.1g per kg-1per hour-1 leucine
    • CHO+caffeine (CHO+CAF): 1.2g per kg-1per hour-1 CHO with 1.7g per kg-1per hour-1 caffeine.
  • All drinks were flavored with 0.05g per L-1 sodium saccharinate, 0.9g per L-1 citric acid and 5.0g per L-1 cream vanilla flavor. CHO source was 50% glucose and 50% maltodextrin. Each drink was labeled with 0.32g per L-1 (U-13C6) glucose.

Statistical Analysis

  • Plasma insulin and glucose responses were calculated as area under the curve
  • A two-factor repeated-measures ANOVA with time and treatment as factors was used to compare differences between treatments over time
  • In case of significant F-ratios, Bonferroni post-hoc tests were applied to locate the differences
  • For non-time-dependent variables, a paired Student's T-test was used to compare differences between treatment and control
  • The results from the questionnaires were analyzed by the Friedman non-parametric test. 
Data Collection Summary:

Timing of Measurements

  • Questionnaire at five, 175 and 355 minutes
  • Blood samples at 15-minute intervals for the first 90 minutes of recovery and every 30 minutes after until 360 minutes
  • Muscle biopsy at cessation of exercise and after the final blood sample.

Dependent Variables

  • Gi symptoms and taste of beverage: Questionnaire using a 10-point scale (1=not al all, 10=very, very much; or 1=horrible, 10=very tasty, respectively)
  • Muscle glycogen concentration and synthesis rates: Muscle biopsy from the middle region of the vastus lateralis muscle
  • Plasma insulin, glucose, lactate, FFA, adrenalin and noradrenalin and caffeine: Blood samples.

Independent Variables

  • CHO only (CHO)
  • CHO+protein+leucine mixture (CHO+PRO)
    CHO+caffeine (CHO+CAF) beverage.

Control Variables

  • Standardized dinner the evening prior to testing
  • No physical exhaustive labor or exercise and diet as constant as possible the two days prior to testing
  • A food and activity diary kept during these two days was used to standardize food intake and physical activity before the second and third tests
  • Abstinence from caffeine containing food and beverages for two days prior to tests
  • Muscle glycogen depletion by intense exercise protocol on a cycle ergometer until pedaling speed could not be maintained at 70% Wmax after pedaling bouts at 90% and 80% Wmax
  • Recovery of six hours.
Description of Actual Data Sample:
  • Initial N: 14 males
  • Attrition (final N): 14
  • Age: 24±1 years
  • Ethnicity: Not described
  • Other relevant demographics: Maximal workload capacity (Wmax), 387±11W; VO2max, 61.5±1.2ml per kg per minute
  • Anthropometrics
    • BW: 71.6±2.5kg
    • Body mass index: 21.7±0.4km/m2.
  • Location: The Netherlands.
Summary of Results:


  • Total cycling time did not differ between experiments
  • All drinks were well tolerated; 95±2% of the total volume of test drink (2.6±0.9 L) was ingested, with no differences between treatments.  The main complaints of bloated feeling, belching and urge to urinate were no different between experiments, although the taste of the CHO+PRO was rated significantly lower than the the other two drinks (P<0.01).
  • Plasma insulin was higher in CHO+PRO, compared with CHO (P<0.05). Plasma glucose did not differ between experiments. No differences in plasma [U-13C6] glucose enrichments were observed between treatments.
  • Plasma lactate over time was lower in CHO+PRO vs. CHO (P<0.012). There were no differences between CHO and CHO+CAF. 
  • Plasma FF over time was higher in CHO+CAF vs. CHO (P<0.01).  There were no differences between CHO and CHO+PRO. 
  • There were no differences between trials over time in plasma adrenalin. Plasma noradrenaline over time was lower in CHO+PRO vs CHO. There were no differences between CHO and CHO+CAF.
  • Plasma caffeine increased significantly (P<0.01) in CHO+CAF and was below detection (P<0.05mg per L-1) in the other two trials
  • Post-exercise muscle glycogen did not differ between experiments. After six hours of post-exercise recovery, muscle glycogen increased, but the concentrations were not significantly different between trials.
  • Histochemical analyses of muscle biopsies revealed no differences in muscle glycogen content between trials or between Type I and Type II fibers. There was a significant correlation between mixed muscle glycogen, determined by PAS staining and muscle glycogen measured using biochemical assay, with a Pearson correlation coefficient of 0.24 (P<0.01).
Author Conclusion:

Coingestion of an insulinotropic amino acid-protein mixture of caffeine does not further accelerate post-exercise muscle glycogen synthesis when an optimal amount of CHO (1.2g per kg-1 per h-1) is already ingested.

Funding Source:
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? N/A
  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? N/A
  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? Yes
  5.1. In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate? Yes
  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.) Yes
  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? Yes
  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