The interactions between heterotrophic microbes and high‐molecular‐weight (HMW) dissolved organic carbon in estuaries are complex and poorly understood. This study examined the coupling between hydrolysis of HMW carbohydrates (polysaccharides) and uptake of monosaccharides by bacterioplankton along a salinity gradient in the Chesapeake Bay water column and nearby coastal waters in order to evaluate the potential importance of polysaccharides as a carbon source for the estuarine microbial loop. We measured the rates of enzymatic hydrolysis of six polysaccharides (arabinogalactan, chondroitin sulfate, fucoidin, laminarin, pullulan, and xylan) as well as total carbohydrate and monosaccharide concentrations, bacterioplankton abundance, and monosaccharide assimilation rates. Enzymatic hydrolysis rates were sufficiently rapid to produce on a daily basis 40‐62% of the monosaccharides present in Chesapeake Bay surface waters but a lower percentage (23%) of monosaccharides present in surface water on the continental shelf. Rates of both monosaccharide assimilation and polysaccharide hydrolysis were markedly lower on the continental shelf than in the Chesapeake Bay. These measurements suggest that at the time of sampling, polysaccharides in the Chesapeake Bay were rapidly recycled, while rates of cycling were considerably slower on the nearby continental shelf. In contrast to the apparently rapid turnover of bulk polysaccharides, hydrolysis of two polysaccharides, pullulan and chondroitin sulfate, was essentially undetectable, implying that those substrates would be unavailable to the microbial communities sampled on a timescale of 10 d.