Soccer is one of the most widely practiced sports in the world and enjoys broad participation in Brazil,1 many professional teams invest in identifying and selecting potential new players, fostering the development of youth with the potential to join professional squads2 in this context, sexual maturation plays a major role in the physical performance of young athletes, particularly in terms of strength and speed. As individuals progress through puberty, muscle development tends to accelerate, contributing to improvements in the body's functional capacities.2

In the current landscape, physical fitness is considered a key determinant of athletic performance, especially in sports with high physiological demands such as soccer. Achieving this critical level of performance requires properly structured and appropriately intense training.3

Given the high physical intensity involved in playing soccer, especially among children, it is essential to understand its effects on cardiovascular health. Differences in cardiovascular health can already be present in childhood, with potential long-term consequences. Identifying and monitoring possible risk factors at this stage of life is crucial for the prevention of future cardiovascular diseases.4

There are several ways to assess cardiac autonomic modulation, with heart rate variability (HRV) standing out as a non-invasive method used to evaluate the autonomic nervous system's modulation at the cardiac sinus node. It describes the oscillations between consecutive RR intervals recorded by the electrocardiogram.5

To analyze HRV, parameters can be obtained through linear methods—in both the time and frequency domains—as well as through nonlinear methods.6 Among the many factors that influence cardiac autonomic modulation, the athlete’s emotional state is particularly relevant. An anxious state increases sympathetic modulation, which negatively affects athletic performance in terms of motor skills and decision-making.7

Regarding anxiety, it refers to an emotional state of fear or worry.8 Anxiety rates in children and adolescents increased following the COVID-19 pandemic.9 However, it is necessary to distinguish between state anxiety and trait anxiety. State anxiety refers to a temporarily elevated level of anxiety, while trait anxiety is considered a personality characteristic or individual profile.10 Anxiety becomes a key topic in sports due to its direct impact on athletic performance, especially among children and adolescents. When these individuals exhibit high levels of state anxiety, they are unable to reach their full potential in sports, which compromises their physical fitness.11

In this context, it is observed that physical fitness among children and adolescents participating in youth academies or soccer schools is progressively developed over their athletic journey toward the professional level. This development is influenced by factors such as training quality, socioeconomic conditions, and the stage of biological and sexual maturation.12

Although previous studies have separately investigated cardiac autonomic modulation, anxiety, or physical fitness in young athletes, evidence remains scarce regarding the integrated analysis of these variables while considering different stages of biological maturation.

Based on this, this study aimed to identify cardiac autonomic modulation, anxiety levels, and physical fitness among players at different stages of sexual maturation.

The findings presented in Table 1 showed statistically significant differences between the groups in anthropometric and physical performance variables. Prepubescent boys had significantly lower weight and height compared to pubescent boys (p = 0.01 for both variables). In the aerobic capacity test, the prepubescent group covered a shorter distance than the pubescent group (p < 0.01), indicating lower cardiorespiratory performance. Similar differences were observed in the standing long jump (p = 0.01) and square test (p = 0.04), with poorer performance among prepubescent participants. On the other hand, no significant differences were found in BMI percentile (p = 0.39) or the 20-meter sprint test (p = 0.55).

Regarding cardiac autonomic modulation (Table 2), heart rate variability analyses showed that prepubescent boys exhibited reduced values in time-domain indices such as SDNN (p = 0.007) and RMSSD (p = 0.003), indicating lower parasympathetic activity. In the frequency domain, significant differences were found between groups in both absolute and normalized HF values (p = 0.003 and p = 0.001, respectively), with lower values among the prepubescent group. Additionally, a higher LF/HF ratio was observed (p = 0.002), suggesting greater sympathetic dominance in this group. Nonlinear indices also reflected lower autonomic modulation in prepubescent participants, with reduced values of SD1 (p = 0.01), SD2 (p = 0.005), and the SD2/SD1 ratio (p = 0.002). These findings support the hypothesis that puberty is associated with changes in cardiac autonomic balance, with greater parasympathetic involvement observed in pubescent boys.

The analysis of anxiety classification based on the MASC scale scores (Table 3) revealed a trend toward a difference between the groups, with a p-value of 0.06. It was observed that 96.8 % of pubertal boys presented minimal levels of anxiety (score ≤ 54), while among prepubertal boys this percentage was 77.3 %. Additionally, only the prepubertal group presented cases with higher levels of anxiety: 13.6 % were classified as having mild anxiety (55–59), 4.5 % with moderate anxiety (60–64), and 4.5 % with severe anxiety (≥ 65), whereas no pubertal participants fell into these categories.

The analysis of the stress index by sexual maturation (Table 4) revealed a statistically significant difference between the prepubertal and pubertal groups (p = 0.009). Pubertal children showed a higher prevalence of low (35.5 %) and normal (64.5 %) stress levels, whereas in the prepubertal group, these values were 18.2 % and 50 %, respectively. On the other hand, 27.3 % of prepubertal individuals presented elevated stress levels (score between 13 and 22), compared to only 5 % in the pubertal group. Additionally, only the prepubertal group had one case of high stress (4.5 %), and no child in either group was classified as having very high stress (score >30). These data indicate that children in the prepubertal phase tend to exhibit higher levels of stress, suggesting possible emotional vulnerability prior to the onset of sexual maturation.

This study aimed to analyze cardiac autonomic modulation, anxiety levels, and physical fitness in players at different stages of sexual maturation. The main results of the present study indicate that pubertal boys exhibit greater vagal autonomic modulation and lower sympathetic cardiac autonomic modulation. Sexual maturation is associated with changes in cardiac autonomic modulation and stress levels, corroborating the study by Leonard et al.17,which showed improved vagal autonomic indices in adolescents, reinforcing that sexual maturation influences cardiac autonomic function.

Additionally, it is extremely important to consider other factors that modulate HRV, such as physical activity level. According to Sharma et al. ,18 regular physical exercise positively influences heart rate variability by increasing parasympathetic activity. Furthermore, global variability indices were higher in this group, indicating a better adaptive capacity of the cardiac autonomic nervous system (ANS), similar to the findings of Chen 2022. ,19 who reported improved global HRV indices in adolescents.

These findings are consistent with other studies that have demonstrated differences in vagal HRV in pubertal individuals. As observed by Chen at al. ,20 sexual maturation, combined with a high level of physical activity, influences cardiac autonomic control. Similarly, Oliveira at al.21 reported greater parasympathetic predominance in pubertal boys. These findings reinforce the hypothesis that sexual maturation promotes more efficient autonomic modulation. In the same logic, according to Sisakova,22 although children already have cardiac autonomic regulation, this balance only becomes truly effective as they grow, providing greater maturation of the cardiac ANS in this population.

In this sense, these results can be explained from a physiological standpoint. In a study by Dogru et al.23 conducted on men, a positive correlation between total testosterone and parasympathetic activity and a negative correlation with sympathetic activity was evidenced. Testosterone plays important roles in neural maturation and functioning. Similarly, Oskui et al.24 state that testosterone causes vasodilation in coronary arteries and can reduce inflammatory markers. Therefore, normal testosterone levels play an important role in maintaining cardiovascular health. Furthermore, Butler at al.25 confirmed that testosterone increases as puberty progresses, with differences across all levels of sexual maturation. Thus, the hypothesis raised regarding our results is that pubertal boys have higher testosterone levels compared to prepubertal boys, suggesting better cardiovascular adaptation, as well as a significant influence on performance tests.

Moreover, this study identified that pubertal boys showed superior performance in aerobic capacity, lower limb power, and agility tests compared to prepubertal boys. Manzano et al.26 found similar results, stating that the transition process to the pubertal phase leads to body improvement and development of physical capacities.

Our study compared the sexual maturation of boys; however, as shown by Mandigout et al. ,27 who studied the effects of a training program in preadolescents classified in stage 1 of the Tanner scale, their study found increased global HRV variables in healthy prepubertal children. Nevertheless, as analyzed in our study, when compared to boys in the pubertal phase, there are differences in HRV indices when compared to pubertal boys.

Beyond the difference observed in cardiac autonomic modulation, we found that prepubertal individuals show higher cardiac stress indices according to Baevsky's stress index, as well as higher anxiety scores. These findings align with Dieleman et al.28 who highlight the influence of anxiety on an individual’s physiological stress, which can serve as an indicator of chronic stress in children. Similarly, Sharma et al.18 showed that children classified as anxious had parasympathetic reactivity outside the normal range. These findings reinforce the importance of maturational stage and emotional aspects, such as anxiety, in interpreting HRV indices in pediatric populations, suggesting that prepubertal boys have poor cardiac autonomic nervous system (ANS) adaptation, indicating that the onset of puberty may represent a phase of greater psychophysiological vulnerability.

Pubertal maturation is characterized by significant biological and neuroendocrine changes that can influence both emotional regulation and the functioning of the autonomic nervous system. These developmental changes have been associated with increased emotional sensitivity and a higher risk of anxiety symptoms during early adolescence.29 Additionally, puberty is also associated with modifications in cardiovascular autonomic regulation. Lower heart rate variability (HRV) values have been linked to reduced parasympathetic activity and impaired emotional regulation, which may increase vulnerability to symptoms of anxiety and stress.30 Some studies have already demonstrated associations in which adolescents presenting anxiety symptoms show alterations in HRV indices compared with healthy peers, suggesting dysregulation of cardiac autonomic control.31

Although some studies suggest that anxiety symptoms increase during adolescence, other longitudinal studies have reported a decrease in anxiety symptoms during the early stages of adolescence.32 These findings suggest that the developmental trajectory of anxiety during the transition from childhood to adolescence is heterogeneous and influenced by multiple biological, psychological, and environmental factors. However, changes in body perception during adolescence may also influence psychological outcomes, including anxiety and depressive symptoms.9 Therefore, differences in maturational stage may partially explain variations in anxiety levels observed among young athletes.

The differences in physical fitness observed between pre-pubertal and pubertal players may also be influenced by differences in chronological age. In the present study, pubertal athletes were older, which may reflect greater exposure to training, motor learning, and accumulated practice time. Therefore, the observed differences may not be exclusively related to biological maturation but also to age-related adaptations and training experience. Latorre-Román, et al.,33 found that developmental changes occurring during childhood may influence physical fitness and autonomic cardiac modulation, indicating that age-related factors may also contribute to the differences observed between groups.

The present study has some limitations that should be mentioned. First, we used a cross-sectional design, which does not allow the establishment of causal relationships between the investigated variables. Therefore, the findings should be interpreted as associations between sexual maturation, cardiac autonomic modulation, anxiety levels, and physical fitness in young soccer players. Second, although biological maturation provides important information about developmental status, differences in chronological age within the same maturation stage may influence physiological and psychological variables such as cardiac autonomic modulation, anxiety levels, and physical fitness. Consequently, chronological age should be considered a potential confounding factor when interpreting the results.

Third, sociodemographic variables such as socioeconomic status, family environment, and educational background were not assessed in the present study. These factors may influence psychological outcomes, particularly anxiety levels, represent a potential source of uncontrolled variability. Fourth, although Tanner staging is widely used to assess sexual maturation, some limitations should be acknowledged. Previous studies have reported potential misclassification, particularly when self-assessment methods are used, as children and adolescents may overestimate or underestimate their pubertal stage.34

Fifth, the relatively small sample size (n = 53) may have reduced the study's statistical power and does not allow the generalization of the findings. Sixth, another sample limitation was the inclusion of only boys, limiting the findings to a single gender. Seventh, in the present study, five participants classified as Tanner stage P2 were included in the pre-pubertal group. Although Tanner stage P2 is often considered an early stage of pubertal development, it represents the transition from pre-puberty to puberty and may already reflect initial hormonal changes.

This transitional characteristic may introduce some variability in physiological and psychological responses, potentially influencing variables such as autonomic cardiac modulation, anxiety levels, and physical fitness. Therefore, the inclusion of participants at this stage may represent a potential source of classification bias and should be considered when interpreting the findings.

Despite these limitations, a cross-sectional study was conducted on a subject rarely explored in the literature. The analysis of cardiac autonomic modulation, anxiety levels, and physical fitness in active and healthy children is rarely studied. Future studies should adopt a longitudinal research methodology to observe the evolution of performance, emotional, and physiological variables, providing a better understanding of the effects of sexual maturation on these variables. Additionally, hormone testing should be considered to determine total testosterone levels and understand the effects of this hormone in the maturation process of children and adolescents who play soccer. Therefore, this research becomes an important reference for future studies, providing relevant parameters in this field.

The findings of this study have important practical implications for professionals working with youth athletes. Understanding the relationship between pubertal maturation, cardiac autonomic modulation, anxiety levels, and physical fitness may help coaches, physical trainers, and health professionals better monitor the physiological and psychological development of young soccer players. In addition, these results may contribute to the development of training and monitoring strategies that consider maturational status, promoting athlete well-being and optimizing performance during adolescence.

We conclude that both vagal and global autonomic modulation are greater in pubescent athletes compared to prepubescent athletes. Additionally, we observed higher performance variable values and lower cardiac stress levels in this group. Regarding anxiety, the results indicated differences between maturation stages, highlighting the importance of considering sexual maturation when analyzing psychological and physiological variables in young athletes. These findings suggest that the pubertal stage promotes greater maturation of the cardiac autonomic nervous system and enhanced development of physical capacities.