Introduction: Elevated plasma homocysteine is an established risk factor for cardiovascular and metabolic diseases. Its regulation is influenced by genetic polymorphisms, gender, hormonal status, nutritional intake, and lifestyle habits. While women generally present lower homocysteine levels compared to men, particularly before menopause, sedentary behavior and inadequate intake of B vitamins and folate may increase concentrations. Regular physical activity and supplementation with vitamin B6, vitamin B12, and folic acid are considered effective strategies to reduce homocysteine, but evidence on their combined effects across gender and activity levels remains inconsistent.

Methods: A systematic literature search was conducted in PubMed, Scopus, and Web of Science up to August 2025, following the PRISMA guidelines. Studies were included if they reported plasma homocysteine levels stratified by gender or physical activity status, with or without supplementation of vitamin B6, vitamin B12, and/or folate. Both observational and interventional studies were considered. Data were extracted on study design, population, supplementation regimens, physical activity level, and outcomes.

Results: Across the included studies, women showed consistently lower baseline homocysteine levels than men, with differences narrowing after menopause. Regular physical activity was associated with modest reductions in homocysteine, although the effect varied according to intensity, duration, and training status. Supplementation with vitamin B6, B12, and folate produced significant reductions in plasma homocysteine across all groups, with the largest absolute decreases observed in sedentary individuals and in men. Evidence on the interaction between gender, physical activity, and supplementation remains limited, and heterogeneity across studies prevented meta-analysis.

Conclusions: This systematic review highlights that homocysteine levels are modulated by gender, lifestyle, and nutritional supplementation. B vitamin and folate supplementation is effective in lowering homocysteine, with potentially greater benefits in high-risk groups such as men and sedentary individuals. Further high-quality trials are needed to clarify the synergistic role of gender and physical activity in homocysteine metabolism.