B-alanine is a non-essential, non-proteinogenic amino acid found in deficient skeletal muscle
concentrations. This deficiency makes B-alanine a rate-limiting factor for carnosine
synthesis. B-alanine supplementation is highly efficient in increasing intramuscular
carnosine content, leading to improved physical performance, especially in high-intensity
exercises (HIIE). It seems that exercise per se can modulate carnosine content; however, it
remains uncertain whether physical training or training status can influence B-alanine
supplementation responses. Thus, this work aims to assess whether HIIE can increase B-alanine
uptake by peripheral tissues and, more specifically, skeletal muscle and increase
intramuscular carnosine synthesis. A crossover counterbalanced study will be conducted. The
volunteers will be evaluated in two conditions: intake of B-alanine + exercise (B-EX) and
B-alanine intake only (B-Ala). The dosage will be 20 mg · kg-1 (powder diluted in water).
This process will be divided into two blocks of six days (Thursday to Thursday - without the
weekend) with a 4-6 weeks washout. In the B-EX block, a 20-minute HIIE session will be held.
Next, B-alanine supplementation will be administered. Immediately after exercise, in a
150-minute rest, collections of venous blood samples will be performed, and the muscle biopsy
and urine collection will be collected pre-test and post-test. In the next three days
(Thursday to Wednesday), this same protocol will be repeated without blood, muscle, or urine
collections. On these days, the B-alanine supplementation will be administrated before and
after the protocol (B-EX and B-Ala), in the same dosage of 20mg.kg-1. On the 6th day
(Thursday), the complete protocol will be carried out, including the collection of biological
materials. In the B-Ala block, the same procedures will be adopted for the B-EX block, with
the replacement of the HIIE for 20 minutes of rest sitting on the cycle ergometer. The
nutritional parameters will be evaluated during the two blocks by dietary diaries in software
(Dietbox) that includes images and the description of all the volunteer food in the day. It
will be determined that during the blocks (B-EX and B-ala), the Beta-alanine intake by food
is limited to 300mg per day. The analysis of the determination of muscle B-alanine, plasma,
and urine, muscle carnosine will be performed by HPLC-ESI + -MS / MS. The gene expression of
carnosine-related enzymes and transporters will be analyzed using the real-time PCR
technique. The enzymes CARNS1, CN2, TauT, PAT1, and phosphorylated Na + / K + / ATPase will
be analyzed using the Western Blot technique. Carnosine synthase activity will be carried out
by incorporating [3H] B-alanine acid in the corresponding dipeptide. The analysis of
paresthesia's presence and intensity will be through a scale adapted from LINGJÆRDE et al.,
1987. The statistical analysis will be performed in the RStudio software and SAS software,
using the mixed model to compare muscle, plasma, and urinary variables. Condition (B-Ex vs.
B-ala) and time of collection (time) will be fixed factors, while the subjects will be random
factors. Data sets containing values obtained at only 2 points in time (e.g., area below the
plasma B-alanine curve) will be analyzed using Student's t-test for dependent samples if the
data distribution is normal. If the data distribution is not normal, we will use the Wilcoxon
sign test for pairs. The normality of data distribution will be assessed qualitatively using
Q-Q Plot. Four different covariance matrix structures will be tested for each data set. The
Schwarz-Bayesian criterion (lowest BIC value) will be used to select the matrix that best
fits each data set. Hypothesis-guided contrast analysis of a single degree of freedom will
test specific effects within and between conditions. The size of the Cohen effect (D) will be
calculated for all variables. Significance will be accepted if p<0.05.