Objective To explore whether Antrodia cinnamomea can prolong the survival time of stroke-prone spontaneously hypertensive rats, and to investigate the underlying mechanisms from the proteomics perspective. Methods A total of 80 male stroke-prone spontaneously hypertensive rats were randomly divided into two groups (control group and Antrodia cinnamomea treated group, n=40). The animals were intragastrically given Antrodia cinnamomea (150 mg/kg). Then the death of the animals in the two groups was observed. Another 6 stroke-prone spontaneously hypertensive rats were randomly divided into drug treatment group and control group, with the drug treatment group intragastrically given Antrodia cinnamomea (150 mg/kg), and the brain proteomics were compared between the two groups 90 days later, with WKY rats used as normal controls. The differentially expressed proteins before and after drug treatment were subjected to mass spectrometry analysis, and the results of mass spectrometry analysis were further confirmed by Western blotting analysis. Results Antrodia cinnamomea significantly prolonged the survival time of stroke-prone spontaneously hypertensive rats (P<0.05), and proteomics results showed that Antrodia cinnamomea increased the expression of glutathione S-transferase (GST) and superoxide dismutase (SOD), which were also confirmed by Western blotting analysis. Further study revealed that the total anti-oxidative capability (T-AOC) of the animals was increased (T-AOC: [66.48±16.17] U/g vs [124.75±28.43] U/g, P<0.05), which was manifested by the increased activity of GST and SOD (GST: [40.33±5.24] U/mg vs [70.50±6.24] U/mg, P<0.05; SOD: [109.25±23.61] U/mg vs [192.60±23.95] U/mg, P<0.05), and decreased level of malondialdehyde ([3.96±0.45] nmol/mg vs [2.04±0.31] nmol/mg, P<0.05). Conclusion Long-term treatment with Antrodia cinnamomea can prolong the lifespan of stroke-prone spontaneously hypertensive rats, which might be related to the increased activity of anti-oxidative enzymes and the decreased oxidative damage.