We provide the first assessment of regional water chemistry and plankton (phytoplankton and crustacean zooplankton) for a suite of lakes near the Sutton River region of the north-central Hudson Bay Lowlands (HBL). We use ordination analyses to examine the spatial variation in water chemistry and plankton across lakes, and to explore the factors that may explain this variation. Based on data collected during summer from 2009 to 2011, we found that in addition to geology, water chemistry was strongly influenced by a lake’s proximity to salt water and the degree of permafrost development within its catchment. Phytoplankton composition varied across lakes based on differences in water depth and nutrient concentrations, with non-filamentous cyanobacteria and chlorophytes more common in shallow lakes, and deeper lakes dominated by planktonic diatoms or filamentous cyanophytes. Crustacean zooplankton community composition and richness in the HBL lakes was similar to communities found in Ontario lakes in more temperate regions within the Precambrian Shield. These baseline data provide a foundation upon which future surveys in this climatically sensitive region may be compared.