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The crystal structures would directly affect the physical and chemical properties of the surface of the material, and would thus influence the catalytic activity of the material. α-MnO2, β-MnO2 and γ-MnO2 nanorods with the same morphology yet different crystal structures were prepared and tested as oxidase mimics using 3,3',5,5'-tetramethylbenzidine (TMB) as the substrate. β-MnO2 that exhibited the highest activity had a catalytic constant of 83.75 μmol·m-2·s-1, 2.7 and 19.0 times of those of α-MnO2 and γ-MnO2 (30.91 and 4.41 μmol·m-2·s-1), respectively. The characterization results showed that there were more surface hydroxyls as well as more Mn4+ on the surface of the β-MnO2 nanorods. The surface hydroxyls were conducive to the oxidation reaction, while Mn4+ was conducive to the regeneration of surface hydroxyls. The synergistic effect of the two factors significantly improved the activity of β-MnO2 oxidase mimic. Using β-MnO2, a β-MnO2-TMB-GSH system was established to detect the content of glutathione (GSH) rapidly and sensitively by colorimetry. This method had a wide detection range (0.11-45 μM) and a low detection limit (0.1 μM), and had been successfully applied to GSH quantification in human serum samples.