Poria cocos is an edible medicinal fungus known as 'Fuling' in China. The switching from traditional cultivation of P. cocos to substitute cultivation has become a new trend. This study was to determine the comparison of the composition and efficacy of traditionally cultivated P. cocos decoction (TCPCD) and substituent-cultivated P. cocos decoction (SCPCD). We studied the pharmacodynamic differences between TCPCD and SCPCD by establishing three animal models. 53 triterpenoids were identified in TCPCD and SCPCD. TCPCD and SCPCD could upregulate the spleen-deficiency indicators and brain-gut peptides and attenuate aquaporin (AQP)1 and AQP2 in the rat model of functional dyspepsia. Both extracts reduced dopamine, 5-hydroxytryptamine, γ-aminobutyric acid levels in rats with anxiety disorder. Moreover, both TCPCD and SCPCD improved xenobiotic resistance and decreased the levels of interleukin-2 and tumour necrosis factor-α of immunocompromised mice. Our findings provide a scientific basis for the promotion of cultivation of P. cocos using substitute materials.