NAP: Recovery (2014)


Rowlands DS, Rossier K, Thorp RM, Graham DF, Timmons BW, Stannard SR, Tarnopolsky MA. Effect of dietary protein content during recovery from high-intensity cycling on subsequent performance and markers of stress, inflammation, and muscle damage in well-trained men. Appl Physiol Nutr Metab. 2008; 33: 39-51.

Study Design:
Randomized Crossover Trial
A - Click here for explanation of classification scheme.
Quality Rating:
Positive POSITIVE: See Quality Criteria Checklist below.
Research Purpose:

To examine the effect of a protein-enriched, high carbohydrate, post-exercise recovery diet on next-day high-intensity cycling performance. 

Inclusion Criteria:

Male endurance-trained cyclists.

Exclusion Criteria:

Precluding health conditions.

Description of Study Protocol:


Not described.


Randomized, cross-over trial:

  • Day One, consumed low-PRO diet with final meal consumed four hours prior to exercise. In the lab, subjects completed 2.5-hour cycling intervals:
    • 12 minutes warm-up at 30% Wmax, five minutes at 40% and five minutes at 50% Wmax
    • 10 two-minute periods at 90% Wmax; 12 two-minute periods at 80% Wmax, alternated with two-minute recovery periods at 50% Wmax, finishing with three five-minute periods at 70%, interspersed with three five-minute periods at 50%. 
  • On Day Two, a small breakfast was consumed (50g cereal bar), 15 minutes later, cycling began
    • Warm-up and six two-minute intervals at 80% Wmax, alternated with two-minute recovery periods. Performances test consisted of 10 sprints alternated with 10 recovery periods at 40% Wmax.

Blinding Used



  • Protein-rich recovery feeding intervention (high-protein) and an isocaloric low-protein control condition (control)
  • High-protein was a chocolate-coated bar and milk-like drink formulation (chocolate or vanilla-flavored)
  • Low-protein was an isocaloric bar and milk-like drink formulated to resemble high-protein
  • High-protein provided 1.6g CHO, 0.8g protein and 0.29g fat per kg of fat free mass (FFM) per hour and the control 2.35g CHO, 0.12g protein and 0.29g fat
  • Bar and drink servings were given every 30 minutes, starting immediately after exercise on Day One, for four hours, totaling eight bars and drinks during this period. 

Statistical Analysis

The effect of protein-enriched intervention relative to control on dependent outcomes was estimated from mixed modeling. Most dependent outcomes were analyzed after log transformation to reduce or eliminate the heteroscedastic error apparent after preliminary visual assessment of the residuals; the exception was perceptual data and data sets with negative values where log-transformation is not appropriate. Appropriate mixed (fixed and random effects) linear models were applied to all data sets. 

For the analysis of sprint performance outcome was determined from the interaction between the following fixed effects: Treatment, order of treatment (treatment one or two), day (Day Two or Four). For the analysis of sprint performance, lactate and glucose and psychometric responses, sprint number or time was coded as a quantitative number predictor (as in linear regression) to model the effect of time on outcomes (slope and fatigue effects). For the analysis of creatine kinase, cytokinines, malonyl dealdehyde (MDA) and nitrogen balance, estimates were derived from repeated-measures models (as in traditional ANOVA).

Data Collection Summary:

Timing of Measurements

Blood sample five minutes after exercise and every 30 minutes for the following 120-minute recovery, with a final sample at 180 minutes on Days One and Four. On Day Two, blood at baseline, 15 minutes after small breakfast and every 30 minutes for the first 90 minutes into recovery. Urine collection from completion of exercise on Day One, until the beginning of exercise on Day Four.

Dependent Variables

  • Ratings of fatigue and exertion: Visual analog scales immediately after priming intervals two and five and sprints one, four,seven and 10 on Days Two and Four. Measured categories were sensations of tiredness, leg soreness, ability to sprint, level of exertion and nausea using linear interval Likert scales from one (nothing) to nine (maximal).  
  • Plasma lactate and glucose: Automated analyzer
  • Skeletal muscle membrane damage: Creatine kinase (CK) by enzymatic assay
  • Systemic stress and inflammation: Cortisol, interleukin-6 (IL-6), tumor necrosis factor a (TNT-alpha), and C-reactive protein (CRP) on Day One at 90 minutes recovery, Day Two after fasting and 90 minutes of recovery and Day Four after fasting
  • Oxidative stress: Lipid peroxide MDA
  • Urinary urea: Spectrophotometry
  • Urinary creatine and creatinine: High-performance liquid chromatography
  • Nitrogen balance: Calculated during three periods to total 60 hours
    1. From completion of cycling on Day One to final urine collection shortly before the pre-exercise meal on Day Two
    2. During exercise on Day Two
    3. From the end of exercise on Day Two to the final urine collection on morning of Day Four preceding the pre-exercise meal
  • Nitrogen outputs were urinary urea, creatine and creatinine (measured). Additional nitrogen losses were estimated from sweat at rest, sweat during exercise on Day Two and from feces and miscellaneous loss throughout the 60-hour collection.

Independent Variables

  • Protein-rich recovery feeding intervention (high-protein) and an isocaloric low-protein control condition (control)
  • High-protein was a chocolate-coated bar and milk-like drink formulation (chocolate or vanilla-flavored).

Control Variables

Two four-day experimental blocks separated by 14 days.

  • On Day One: Standardized animal flesh-free diet four hours before testing. Fatigue inducing ride of 2.5-hour intervals with an average estimated total work performed of 10.5mJ. Recovery food was consumed in units immediately after the first blood draw and in 30-minute intervals for the first three hours, with the last unit consumed one hour after leaving the lab.
  • On Day Two: After a small breakfast a repeated-sprint performance (7.7mJ). Ingestion of recovery food same as on Day One. Standardized CHO-rich diet that was provided was then started and followed until late evening of Day Three.
  • Day Four was identical to Day Two.
Description of Actual Data Sample:
  • Initial N: 12 males
  • Attrition (final N): 12
  • Age: 34±10 years
  • Ethnicity: Not described
Other Relevant Demographics
  • VO2max4.9±0.6L per minute
  • Peak power output: 362±29W
  • Training for 7.5±3.5 years, with a weekly average training of 12.0±3.5 hours during the previous six months.
  • Height: 179±4cm
  • Body mass: 75.9±4.3kg
  • Fat free mass (FFM): 68.2±2.3kg
  • Body fat mass: 10±4.0%.

New Zealand.
Summary of Results:


  • On Day Two, there was no clear effect of protein dose on overall sprint mean power. However, on Day Four, mean power was substantially higher in the protein-enriched condition, relative to control. The decline in mean power from sprint one to 10 (fatigue effect) was greater in control on Day Two vs. protein-enriched; on Day Four the difference in fatigue was unclear.
  • With the treatment effect factored out, overall sprint mean power on Day Four compared with Day Two was 8.7% higher (95% CI, 4.7% to 13%; P<0.0001) during the first and 7.9% (±3.5% to 12.4%; P<0.0001) during the second experimental block, respectively. The second order of trial increased overall sprint mean power on Days Two and Four, relative to the first by an insubstantial 1.9% (-2.1% to 6.0%; 0.36) and 1.0% (-3.0% to 5.2%; 0.62), respectively.
  • There were likely trivial effects of the protein-enriched condition on psychometric measures on Day Two, except for a likely decrease in the perception of tiredness. On Day Four, possible small increases in strength, decreases in nausea and a very likely increase in perceived effort were observed in the protein-enriched condition, relative to control. The change in perception ratings from sprint one to 10 (slope) were unclear, with the exception in Day Two of the rating of leg soreness increasing by 0.14 units (95% CI, 0.04 to 0.22; P=0.01) more per sprint and the rating of leg strength decreasing 0.08 units (0.00 to 0.15; P=0.05) more per sprint in the protein-enriched vs. control. 
  • Glucose was 15% (95% CI, 9% to 21%; P=0.0001) and 24% (15% to 33%; P=0.0001) lower in protein-enriched on Day One and Two recoveries, respectively. The rate of decline in glucose in protein-enriched was 23% (7% to 40%; P=0.015) and 75% (37% to 121%; P=0.0003) greater on Days One and Two than in control. On Day Two, there was no difference in glucose following sprints; on Day Four, glucose was 8% (5% to 11%; P=0.0001) higher in protein-enriched. 
  • Plasma lactate, during sprints was 15% higher (-2% to 35%; P=0.15) on Day Two in control. On Day Four, lactate was 13% (4% to 22%; P=0.013) higher during protein-enriched.
  • No clear effect of diet on oxidative stress markers
  • Nitrogen balance was positive in the protein-enriched and negative in control from Day One post-exercise to the morning of Day Two; the net difference between conditions was 207mg N per kg FFM (95% CI, 207 to 386). There was no difference in nitrogen balance between conditions for the remaining time. The overall 60-hour net gain in protein-enriched relative to the net loss in control was 349mg N per kg FFM (191 to 507).
Author Conclusion:
  • The ingestion of a protein-enriched, high-carbohydrate, mixed diet for two days during a four-hour recovery period after high-intensity cycling provided a substantial enhancement to subsequent performance 60 hours, but not 15 hours after the first bout of interval cycling
  • The ergogenic effect was linked to a positive nitrogen balance during recovery on Day One, but the specific nutrient-mediated effect appears delayed and the mechanisms to be determined.
Funding Source:
Fonterra, New Zealand; Bronston and Jacobs, Auckland; Nice and Natural, Auckland (Product support)
Food Company:
Pharmaceutical/Dietary Supplement Company:
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? N/A
  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.) 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? Yes
  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