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Background: β-alanine, a non-essential amino acid found in the diet and produced through nucleotide catabolism, is significant for muscle performance due to its role in carnosine synthesis. This study aims to assess the impact of a 4-week β-alanine supplementation on neuromuscular fatigue in individuals engaging in High-Intensity Functional Training (HIFT) and its subsequent effect on sports performance, distinguishing between central fatigue from the CNS and peripheral fatigue from the muscular system. Materials and methods: This study (a randomized controlled trial) comprised a total of 27 subjects, who were divided into two groups. Group A (the control group) was administered sucrose powder, while Group B (the experimental group) was given β-alanine powder. The subjects were randomly assigned to either the experimental or control groups. This study lasted four weeks, during which both groups participated in high-intensity interval training (HIFT) on the first day to induce fatigue and work close to their VO2 max. Results: Statistically significant changes were in the sports performance variables, specifically vertical jump and jumping power (p = 0.027). These changes were observed only in the group that had been supplemented with β-alanine. Nevertheless, no alterations were observed in any other variables, including fatigue, metabolic intensity of exercise, or perceived intensity (p > 0.05). Conclusions: A four-week β-alanine intake program demonstrated an improvement in the capacity of subjects, as evidenced by enhanced vertical jump and power performance. Nevertheless, it does result in discernible alterations in performance. [ABSTRACT FROM AUTHOR]