Theoretical models for the description of microdialysis outflow:inflow (O/I) ratio for 3H2O and [14C]ethanol were developed, taking into account the nutritive fraction of total blood flow in muscle. The models yielded an approximately exponential decay expression for the O/I ratio, dependent on the physical dimensions of a linear probe (length and radius), the flow rate through the probe, muscle blood flow (including the nutritive fraction) and the diffusion coefficients for the tracer in the probe and muscle. The models compared favourably with experimental data from the constant-flow perfused rat hindlimb. Estimates of the nutritive fraction of total blood flow from experimental data were determined by minimizing the error between model and experimental data. The nutritive fraction was found to be 0.22 ± 0.04 under basal perfusion conditions. When 70 nm noradrenaline (norepinephrine) was included in the perfusion medium, the nutritive fraction was 0.91 ± 0.06 (P < 0.05). The inclusion of 300 nm serotonin, decreased the nutritive fraction to 0.05 ± 0.01 (P < 0.05). This model can be applied to the determination of nutritive fraction of skeletal muscle blood flow in physiologically relevant microvascular conditions such as during exercise and in disease states.