The relation between the oxidative burst and phenylpropanoid pathways has been studied using the sugarcane cultivar C86-56, which does not release phenolics in agar-base micropropagation systems. In stationary liquid culture, a significant production of phenolic compounds and plant survival were determined in sugarcane plants treated with 5 mM H2O2. The spectrophotometer determinations and the gene expression analysis corroborated that releasing of phenolics and soluble -quinones was induced during the first 24 h of treatment. In comparison with the control treatments, sugarcane plants treated with H2O2 demonstrated differences in the micropropagation-related variables when multiplied in Temporary Immersion Bioreactors (TIBs) supplemented with polyethyleneglycol (PEG 20%). Expression of selected genes related to photosynthesis, ethylene, auxins, oxidative burst, and defense pathways were confirmed during the entire PEG 20% stress in the plants coming from the 5 mM H2O2 treatment; whereas, much more heterogeneous expression patterns were evidenced in plants stressed with PEG but not previously treated with H2O2. RT-PCR expression analysis supports the hypothesis that while H2O2 induces the oxidative burst, the phenylpropanoids pathways elicit and maintain the defensive response mechanism in micropropagated sugarcane plants.