The Brazilian biomes Atlantic Forest, Cerrado, and Caatinga are considered biodiversity hotspots in the world and are fundamental to the global and regional carbon balance. However, deforestation and/or burning and changes in land use and land cover have contributed to the increase of carbon dioxide (CO2) in the atmosphere as well as the consequent greenhouse effect and global warming. Net photosynthesis is a natural CO2 levels regulation process in the atmosphere and it is influenced by vegetation composition, climatic and anthropogenic factors. In this sense, this study aimed to evaluate the Net Photosynthesis (PSN) in 20 years in the Brazilian biomes Atlantic Forest, Cerrado, and Caatinga, to verify the monthly correlation between PSN and intervening factors, such as climatic variables and burned areas, as well as to determine trends of increase and decrease over the years of all variables studied. The study was based on eight-day MOD17 Net Photosynthesis images and the results showed that monthly PSN in the three biomes is more positively correlated (p-value < 0.05) with actual evapotranspiration together with water availability index and has more negative correlation (p-value < 0.05) with land surface temperature. Spatially, during the following 20 years, the Mann-Kendal test showed an PSN decreasing trend in the Atlantic Forest biome and an increasing trend in the Cerrado biome. These trends observed in the mentioned biomes are related to the advance of agriculture and cattle ranching. Finally, this study provides relevant information on net photosynthesis and interference of climatic and anthropogenic factors, being essential for monitoring and formulation of policies that stimulate carbon absorption in biomes.