Compared Effect of a Fruit Milk Shake With a Protein-Carbohydrate Supplement on Recovery After Resistance Exercise

The aim of this study is to assess the effect of ingesting a commercial carbohydrate and protein dietary supplement in powder form (P-CHO supplement) or a milk shake with skimmed milk, strawberries and banana (MS), after resistance exercise, in muscle damage, oxidative stress, inflammation and functional recovery. It is hypothesised if the ingestion of a milk shake with skimmed milk and fruit (strawberry and banana) has the same impact on markers of muscle damage, oxidative stress, inflammation and functional recovery induced by resistance exercise, as the intake of a commercial powder with the same CHO and protein amounts.

Fifteen adult athletes from the Portuguese Athletic Federation will complete 2 trials separated at least by 2 weeks. Alternate legs and drinks will be used in each trial and participants will be overnight-fasted. This study will have a single-blind, randomized, crossover, repeated-measures experimental design. In each trial, after warm-up, the eccentric peak torque of the knee joint extensors will be determined using an isokinetic dynamometer. After this, participants will complete an isokinetic exercise until exhaustion at a constant angular velocity of 60° • s-1. After the exhaustion protocol, athletes will perform again the peak torque determination. Immediately after, participants will drink the P-CHO supplement or MS during the first 2 h. Both drinks will contain 0.8-1.2 g carbohydrates • kg-1 • h-1 and 0.2-0.4 g protein • kg-1 • h-1. Twenty four and 48 h after the exhaustion protocol participants will return to the laboratory to repeat the peak torque determination. Blood samples will be collected before warming-up, immediately and 2 h after the last peak torque determination and 24 h and 48 h after. Serum samples will be analyzed for creatine kinase, lactate dehydrogenase, interleukine-6, protein carbonyls and total antioxidant status. The delayed onset muscle soreness, using a visual analogue scale, and girths will be measured at the same moments as blood sampling. Two-way repeated-measures ANOVA will be used for statistical analysis of the data.

Sample:

Fifteen adult athletes from the Portuguese Athletic Federation will be invited to participate in this study. They must be healthy and lactose tolerant. Athletes will be voluntary and should receive an explanatory document with the aims of the study. They must also sign an informed consent to participate. To help ensuring that the metabolic conditions are similar between trials, subjects will be instructed to fulfill a food record and a physical activity record of the 2 days prior to each trial (Shirreffs, Watson, \& Maughan, 2007). It will be asked not to make drastic changes in diet and to avoid strenuous exercise during this period.

Experimental Protocol:

This study will have a single-blind, randomized, crossover, repeated-measures experimental design. As shown in Figure 1, each participant should complete 2 trials, with a wash out period of, at least, 2 weeks. In one trial subjects will randomly drink the P-CHO supplement and in the other, the MS. In order to minimize any repeated bout effect, different legs will be used in each trial (Howatson et al., 2010).

About one week before the exercise protocol, participants will be familiarized with the experimental procedures and measurements. At this time, anthropometric assessments (BW, height and body fat percentage) will also be taken.

All tests should be performed in the morning, in a fasted state. At the beginning of each trial, the participants will warm-up for 5 min using a cycle ergometer at an intensity of 70-100 rpm. After the warm-up, the eccentric peak torque of the knee joint extensors muscles will be determined by isokinetic dynamometry (Biodex®, system IV). Then, the participants will complete concentric/eccentric knee extension/flexion exercise protocol until exhaustion. After exhaustion, eccentric peak torque determination will be repeated. Immediately after finishing the isokinetic test, participants will drink the P-CHO supplement or MS during the first 2 hours post-exercise. The drinks will be taken in 2 bolus per hour, during 2 h. During the 2 days after the exhaustion protocol (24 h and 48 h after), participants will return to the laboratory to repeat eccentric peak torque determination after a similar warm-up of the day before.

Drinks' Composition:

The nutritional formula of both drinks will be equal in terms of CHO and protein content in order to achieve the recommendation values of 0.8-1.2 g CHO • kg-1 BW • h-1 and 0.2-0.4 g protein • kg-1 BW • h-1 (Beelen et al 2010). They will also have similar amounts of vitamin C.

The fruit content of the MS will be 100 g of strawberry and the amount of banana necessary to guaranty 0.8-1.2 g CHO • kg-1 BW • h-1. Regarding the P-CHO supplement, it has to be available in a common local market and meet the recommend amounts of CHO and protein for the recovery period. If necessary, vitamin C will be added to match the MS content.

Exhaustion Protocol:

For inducting exhaustion, a concentric/eccentric knee extension/flexion exercise until exhaustion will be used. At the first day of each trial, the exhaustion protocol will be conducted. Securely fastened into the test chair of the isokinetic dynamometer with the knee joint of the test limb aligned with the axis of rotation of the dynamometer and the distal lower limb secured to the dynamometer's test arm, subjects will initially complete 3 maximal repetitions of knee joint extension and flexion at a constant angular velocity of 60°•s-1 to determine the subject's maximal peak torque of the knee extensors muscles (the best of the 3 repetitions). After this, the exhaustion protocol it will be conducted. The protocol will consist of 3 bouts of a concentric/eccentric knee extension/flexion exercise, at 60°•s-1, with a 200-second rest time between sets. The first and second set will be composed by 100 repetitions; in the third set, subjects will perform n repetitions until the eccentric peak value of 3 consecutive repetitions fell below 25% of the initial knee extensor eccentric peak torque value. The range of motion will be from 50° to 110° flexion in the knee (0º = full knee extension).

Participants will receive verbal encouragement to perform maximally throughout the exercise protocol.

Functional Recovery:

To monitor force recovery, measures of eccentric peak torque of the knee joint extensor muscles will be determined. Athletes will perform the isokinetic strength tests immediately before and after the exhaustion protocol and 24 and 48 h after the exhaustion protocol, for each trial. The exercise will consist of 3 maximal repetitions to determine eccentric knee joint extension peak torque at a constant angular velocity of 60°•s-1. The range of motion will be similar to the one used in the exhaustion protocol.

Blood Sampling and Biomarkers:

For each trial, blood will be taken from an antecubital vein after the participant arrives at the laboratory, in a fasted state. Blood samples will also be collected immediately after the last peak torque determination and 2, 24 and 48 h after.

The following blood markers will be used:

* Muscle damage: CK and LDH;
* Inflammation: IL-6;
* Oxidative stress: protein carbonyls and TAS.

Other Measurements:

The DOMS and circumferential measurements will be measured at the same moments as blood sampling.

A visual analogue scale (VAS), 10 cm in length, will be used to determine DOMS (Bijur, Silver, \& Gallagher, 2001).

The mid-thigh girth measurement will be done according to the anthropometric technique suggested by the International Society for the Advancement of Kinanthropometry (ISAK). Three other girths will be measured: 5, 15 and 25 cm above the superior border of the patella. These measurements will allow monitoring the oedema caused by inflammation.

Body Composition:

Weight and height will be measured according to the methodology proposed by the International Society for the ISAK. It will be calculated the body mass index (BMI) and the percentage of body fat by measuring four skinfolds (biceps, triceps, subscapular and suprailiac) according to the technique proposed by ISAK. The value of the measurements will be converted to body density and using the formula: Durnin and Womersley 1974 (Costa, 2001). Later, the body density value will be converted to percentage of body mass, using the formula of Siri (Costa, 2001).

Dietary Intake:

Athletes will be asked to carry out food records 2 days prior to each trial. Each individual will be informed both orally and in writing, by an expert nutritionist, on their correct completion. It also will be required to register the time and place where food consumption was made. Subsequently, data will be processed through a computer program (Food Processor®) to convert the information into energy and nutrients and calculate mean intakes.

Physical Activity:

Two days prior to each trial, athletes will record the type, intensity and duration of physical activities in registration documents distributed previously.

Statistical Analysis:

Statistical analysis will be performed using the Statistical Package for Social Sciences (SPSS 18 for Windows TM®). The results will be expressed as mean ± standard deviation and it will be considered a statistically significant for p \< 0.05. Normality will be assessed by the Kolmogorov-Smirnov test. Data will be analyzed by two-way repeated-measures ANOVA (treatment \[two levels: MS and P-CHO supplement\] vs time \[five levels\]) to determine whether there are any statistically significant effects of time or treatment (Bowtell, Sumners, Dyer, Fox, \& Mileva, 2011). The Mauchly sphericity test will be used to check homogeneity of covariance for all ANOVA analyses; violations of the assumption of sphericity will be correct using the Greenhouse-Geisser adjustment. Where appropriate, the effect size (ƞ2) will also be calculated.

Eligibility Criteria:
Inclusion Criteria:

* healthy and lactose tolerant

Exclusion Criteria: