Indole-3-butyric acid (IBA) and 2,4-dichlorophenoxybutyricacid (2,4-DB) are metabolised byperoxisomal b-oxidation to active auxins that inhibit rootgrowth. We screened Arabidopsis mutants for resistance toIBA and 2,4-DB and identified two new 2,4-DB resistantmutants. The mutant genes encode a putative oxidoreductase(SDRa) and a putative acyl-activating enzyme(AAE18). Both proteins are localised to peroxisomes.SDRa is coexpressed with core b-oxidation genes, butgermination, seedling growth and the fatty acid profile ofsdra seedlings are indistinguishable from wild type. Thesdra mutant is also resistant to IBA, but aae18 is not.AAE18 is the first example of a gene required for responseto 2,4-DB but not IBA. The closest relative of AAE18 isAAE17. AAE17 is predicted to be peroxisomal, but anaae17 aae18 double mutant responded similarly to aae18for all assays. We propose that AAE18 is capable of activating2,4-DB but IBA activating enzymes remain to bediscovered. We present an updated model for peroxisomalpro-auxin metabolism in Arabidopsis that includes SDRaand AAE18.