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The amount of time spent in sedentary behaviors (SB) progressively increases with age, while reducing time spent in light-intensity physical activity (LPA) and moderate- to vigorous-intensity physical activity (MVPA). These trajectories in PA and SB are linked to accelerated reductions in physical functioning.This study aimed to examine the association of substituting SB time with LPA and MVPA on physical function in older adults.Ninety-one older adults (mean age, 70.7 ± 10.2 yr) wore a hip-mounted accelerometer to measure SB, LPA, and MVPA time. Measures of physical function included a 400-m walk test (400W), the usual gait speed (UGS), the five times sit-to-stand (5xSTS) test, and the short physical performance battery (SPPB). Isotemporal substitution regression modeling was performed to assess the relationship of replacing the amount of time spent in one activity for another.Replacing 30 min·d−1 of SB with LPA was associated with a significant improvement in 400W (P = 0.0497), whereas MVPA resulted in a significant improvement (P < 0.01) in 400W, UGS, 5xSTS, and SPPB. Replacing 60 min·d−1 of SB with 10 min·d−1 of MVPA and 50 min·d−1 of LPA was associated with significant improvements in the 400W, UGS, and 5xSTS (P < 0.05). Meanwhile, as little as 5 min·d−1 of MVPA and 55 min·d−1 of LPA were linked to a 78% increased odds of scoring with good function in the SPPB (P = 0.0247).Replacing SB with LPA was linked to a significant improvement in the 400W, but not the other brief functional measures. Mixed doses of LPA and MVPA may add flexibility to interventions targeting reductions of SB in older adults for clinically relevant improvements in physical function.