A series of experiments was performed in an aerobic chemostat reactor using a multi-substrate system consisting of acetate, phenol, and 2,4-dichlorophenol (DCP). The phenolic compounds require initial oxygenation reactions, while acetate is oxidized without oxygenations. The biomass completely dechlorinated DCP and utilized all of the substrates simultaneously as electron donors and carbon sources. However, DCP removal was less than for phenol and depended on the solids retention time. A novel substrate-specific yield analysis indicated that true yield values were approximated well by the number of electrons removed in non-oxygenation reactions. Experiments for estimating the kinetic parameters for utilization of the phenolic compounds were designed to eliminate the effects of the key cosubstrates of oxygenation reactions, O-2 and the reduced intracellular electron carrier, NADH + H+. The maximum specific rate of substrate utilization, q(max), and the half-maximum rate concentration, K,for phenol and DCP were estimated. The kinetics for DCP were much slower than those for phenol, and the largest effect was a half-maximum rate concentration, which was 19 times larger for DCP. The larger K for DCP explains why DCP removal was low and sensitive to the solids retention time.