This study aimed to investigate the effect of IBA at different concentrations (0, 2000 and  4000 ppm) and putrescine (0, 100 and 200 ppm) on rhizogenesis and the biochemical changes  of leafy hardwood cuttings of Olea europaea L. This experiment was conducted as a factorial  in a complete randomized block design with three replications. The result showed that  rhizogenesis was the highest and callus induction was the lowest in the treatment with 4000  ppm IBA. Furthermore, co-treatment with IBA and putrescine increased rhizogenesis and  callus induction two-times over their individual treatments. Moreover, biochemical changes  of protein, carbohydrate (C), nitrogen (N) percentage, C/N ratio, peroxidase activity and  polyphenol oxidase showed significantly different results in two parts of the leaf (bud) and  stem bark of the cutting (root and callus). Overall, based on the results, it can be concluded  that IBA application at 4000 ppm can be recommended as a suitable treatment for rhizogenesis  of hard-to-root ‘Konservalia’ cultivar.