Regional ecosystem health assessment is an important method for obtaining information on ecosystem condition, which has practical guiding significance for the ecological restoration and management. This study established an ecosystem health assessment framework by integrating ecosystem integrity and human demand for ecosystem services from the perspective of coupled human and ecosystems, which integrated ecosystem services supply-demand into the framework of "Vigor-Organization-Resilience". Taking the Guangdong-Hong Kong-Macao Greater Bay Area as a case study, this study analyzed the spatio-temporal variation characteristics of ecosystem health, and explored the relationship between urbanization and ecosystem health from 2005 to 2018. The results indicated that:(1) The level of ecosystem health was low in the central region of the study area and high in the surrounding areas from 2005 to 2018. The well, relatively well and ordinary health levels dominated throughout the study area in 2005 and 2018. From the perspective of temporal changes, the ecosystem health in the surrounding mountains of the study area was improved, while the ecosystem health in the central region was gradually deteriorated. (2) The ecosystem vigor in the study area played an important role in improving the regional ecosystem health. In addition, the ecosystem services supply-demand could appropriately modify the results of the traditional ecosystem health assessment, and the deficit of ecosystem services supply-demand in the central region aggravated the deterioration of ecosystem health. (3) Ecosystem health had significantly negative spatial correlations with land urbanization and population urbanization in the study area, and the negative impact of land urbanization on ecosystem health was increasingly significant, especially in the central region. This study can provide a new idea of ecosystem health assessment, and provide scientific references for ecological protection and management in the study area at the same time.