We investigated the relationship between muscle microvascular responses during reactive hyperemia as assessed using near-infrared spectroscopy (NIRS) with changes in skeletal muscle oxygen saturation during exercise. Thirty young untrained adults (M/W: 20/10; 23 ± 5 years) completed a maximal cycling exercise test to determine exercise intensities performed on a subsequent visit separated by seven days. At the second visit, post-occlusive reactive hyperemia was measured as changes in NIRS-derived tissue saturation index (TSI) at the left vastus lateralis muscle. Variables of interest included desaturation magnitude, resaturation rate, resaturation half-time, and hyperemic area under the curve. Afterwards, two 4-minute bouts of moderate intensity cycling followed by one bout of severe intensity cycling to fatigue took place while TSI was measured at the vastus lateralis muscle. TSI was averaged across the last 60-s of each moderate intensity bout then averaged together for analysis, and at 60-s into severe exercise. The change in TSI (∆TSI) during exercise is expressed relative to a 20 W cycling baseline. On average, the ΔTSI was −3.4 ± 2.4 % and −7.2 ± 2.8 % during moderate and severe intensity cycling, respectively. Resaturation half-time was correlated with the ΔTSI during moderate (r = −0.42, P = 0.01) and severe (r = −0.53, P = 0.002) intensity exercise. No other reactive hyperemia variable was found to correlate with ΔTSI. These results indicate that resaturation half-time during reactive hyperemia represents a resting muscle microvascular measure that associates with the degree of skeletal muscle desaturation during exercise in young adults.