Alzheimer's disease (AD) is a chronic neurodegenerative disease. G protein coupled receptor 50 (GPR50) is a candidate gene for AD. The present study was designed to determine the association between GPR50 methylation and AD. The methylation levels of the GPR50 promoter in 51 patients with AD and 61 healthy controls were determined by bisulfite pyrophosphate sequencing. All participants were Han Chinese, living in Ningbo. It was identified that the GPR50 promoter methylation level was significantly decreased in the male AD group compared with the male control group (9.15 vs. 16.67%, P=0.002). In addition, it was observed that the GPR50 methylation levels of the females was significantly increased compared with that of males in both the patients with AD and the healthy control group (AD patient group: 33.00 vs. 9.15%, P<0.0001; healthy control group: 29.41 vs. 16.67%, P<0.0001). This may be explained by the fact that GPR50 is located on the X chromosome. In addition, GPR50 methylation was positively correlated with plasma cholinesterase levels in female patients with AD (r=0.489, P=0.039). The present study demonstrated that hypomethylation of the GPR50 promoter in peripheral blood may be a potential biomarker for the diagnosis of AD in Chinese Han males.