Estuaries exist across a large climatic gradient in the northwestern Gulf of Mexico, capturing a range of hydrologic conditions and estuarine functioning. We examined freshwater inflow, salinity, and stable isotope compositions (δ¹³C, δ¹⁵N) of oysters, suspended particulate organic matter (SPOM), and surface sediment organic matter (SSOM) from five estuaries across the hydrologic gradient. All five estuaries experienced large decreases in freshwater inflow over the last 40 yr, with three estuaries being subject to freshwater inflow reductions of more than 85%. Generally, these freshwater inflow decreases were associated with estuarine salinity increases. Across the spatial gradient, average salinity generally increased from northeast to southwest estuaries. SPOM in the northeastern, lower salinity estuaries generally contained more continental organic matter and was of higher quality (i.e., lower C/chlorophyll a ratio), as compared to southwestern, higher salinity estuaries. Similarly, both SSOM and oyster δ¹³C values were positively correlated with salinity, further highlighting that food webs in lower salinity estuaries are more greatly influenced by continental organic matter than those in higher salinity estuaries. A decrease in the connectivity between continental and coastal habitats may have broad consequences for flows of organic matter, and estuarine function and health. Conducting studies across large-scale hydrologic gradients can provide a useful approach to informing and predicting shifts in estuarine functioning.