Purpose <p>Recently, muscle imbalances have been studied using muscle oxygen saturation (SmO₂) in both legs, from which the variable muscle oxygen saturation asymmetry (MO₂Asy) was derived. The study aimed to measure the predictive capacity of MO₂Asy across different stages of a graded exercise test (GXT).</p> Methods <p>This was a cross-sectional study. Thirty male players (years: 24 ± 6 and 8 ± 3&#xa0;years of experience in professional soccer) performed a GXT, and the following parameters were measured: maximum oxygen consumption (<i>V</i>O₂max), respiratory quotient (RQ), ventilation, heart rate, and rating of perceived exertion (RPE). Near-infrared spectroscopy (NIRS) technology was also used to assess MO₂Asy in the gastrocnemius muscle (GM). A Bayesian ANOVA, Pearson correlation with linear regressions, and a predictive test of the receiver operating characteristic curves were applied.</p> Results <p>MO₂Asy analysis revealed changes from Fatmax and VT1—low-intensity domain to VT2 and <i>V</i>O₂max—high-intensity domain (BF¹⁰ = 7.988; <i>p</i> = 0.096; ES = 0.054). The MO₂Asy significantly influenced RQ (<i>r</i>^<i>2</i> = 71.2%; <i>p</i> &lt; 0.001) and VO₂max (<i>r</i>^<i>2</i> = 75.6%; <i>p</i> &lt; 0.001). Exercise intensity thresholds were established: VT1 (MO₂Asy = 28% to 16%; <i>p</i> = 0.040) and VT2 (MO₂Asy =  ≤ 15%; <i>p</i> = 0.012).</p> Conclusion <p>MO₂Asy decreases with increasing exercise intensity. It is proposed that high-intensity exercise would be characterized by values around ≤ 15% of MO₂Asy in the GM muscle. Although the vastus lateralis has been the preferred site in most studies using NIRS, the GM offers practical advantages that make it more accessible and easier to evaluate, especially in clinical settings and in the field with professional soccer players.</p>