Background: Recent research suggests that omega-3 fatty acids may play a role in bone metabolism through their influence on bone mineral density (BMD) and the regulation of bone turnover markers. However, epidemiological evidence linking omega-3 intake to the risk of developing osteoporosis is still emerging and remains inconclusive. This study aims to clarify the role of dietary omega-3 fatty acids in the prevention of osteoporosis. Methods: We analyzed data from 8,889 participants categorized into normal, osteopenia, and osteoporosis groups based on their BMD scores from the National Health and Nutrition Examination Survey (NHANES). We measured dietary omega-3 intake using two 24-h dietary recall interviews. Dietary omega-3 intake was quantified and divided into quartiles. Multivariate logistic regression and subgroup analysis were used to explore the correlation between dietary omega-3 intake and osteoporosis. The dose–response relationship between the two was analyzed with a restricted cubic spline (RCS). Results: Higher dietary intake of omega-3 fatty acids was inversely associated with the risk of osteoporosis. Participants in the highest quartile of omega-3 intake had a significantly lower risk (OR = 0.71, 95% CI: 0.53–0.93) compared to those in the lowest quartile, with a consistent trend across all adjusted models (p for trend <0.05). Subgroup analyses indicated stronger associations in individuals under 60 years of age, female and non- smokers. In individuals aged under 60, higher omega-3 intake was associated with significantly reduced osteoporosis risk (OR = 0.51, 95%CI: 0.26–0.95), females showed a protective effect of high omega-3 intake against osteoporosis (OR = 0.65, 95% CI: 0.49–0.87). Among non- smokers, higher omega-3 intake was associated with a lower risk of osteoporosis (OR = 0.64, 95% CI: 0.45–0.90), whereas in smokers, the association was not evident (OR = 0.91, 95%CI: 0.55–1.52). No significant associations were found in older participants or smokers. Intake of omega-3 and osteoporosis were linearly related (p for nonlinear = 0.366). Conclusion: This study demonstrates a significant inverse relationship between dietary omega-3 fatty acid intake and osteoporosis risk, suggesting omega-3 s play a crucial role in bone health. However, further longitudinal studies are needed to confirm these findings and refine dietary recommendations for osteoporosis prevention.