Heme containing peroxidase, for example manganese peroxidase (MnP), is easily inactivated by the hydrogen peroxide (H₂O₂) presented in the reaction. Here we extremely increased the H₂O₂ stability of MnP by molecular evolution and immobilization in mesoporous materials. A mutant MnP library containing three randomized amino acid residues located in the entry site of H₂O₂-binding pocket of MnP was evolved on a 384-well plate using SIMPLEX (single-molecule PCR-linked in vitro expression). The screening of more than 10⁴ samples independently expressed for improved H₂O₂ stability led to four positive mutants, the H₂O₂ stability of which was nine times higher than that of the wild-type. Immobilized MnP mutant in mesoporous material (FSM) showed the high H₂O₂ stability, more than 50 folds than wild type MnP. But the stability of immobilized wild type MnP was not improved so much as that for immobilized mutant MnP. [DOI: 10.1380/ejssnt.2005.207]