Hypolimnetic oxygen injection is a management tool used to improve water quality by preventing anoxia and associated phosphorus release from the sediments. An additional benefit would be formation of a low‐oxygen refuge for large‐bodied herbivorous zooplankton in the metalimnion. The magnitude and timing of hypolimnetic oxygenation was deliberately manipulated during summer 1997 in Irondequoit Bay, a eutrophic Lake Ontario embayment (New York, USA), to maintain metalimnetic dissolved oxygen (DO) concentrations in the range of 1–2 mg/L. We assessed the vertical distributions of fish and zooplankton in 1997, a year with controlled oxygenation, and 1996, a year with oxygenation but without a deliberate attempt to create amenable refuge conditions. After initiation of oxygenation, June through early August metalimnetic DO in 1997 was 1.8 ± 0.5 mg/L (means ± 2 se), whereas metalimnetic DO in 1996 was 3.8 ± 1.0 mg/L. Hydroacoustic surveys in 1997 and gillnet catches in both years indicated that >96% of planktivorous fish, alewives (Alosa pseudoharengus) and emerald shiners (Notropis atherinoides), were restricted to depths with DO > 4.0 mg/L. A refuge effect in 1996 was slight and evidenced only by significantly larger zooplankton in the metalimnion while densities of Daphnia galeata medotae and D. retrocurva were similar or significantly greater in the epilimnion. In 1997, the refuge effect was strong despite an almost twofold increase in planktivorous fish abundance compared to 1996. Overall zooplankton densities were similar in the epilimnion and metalimnion while zooplankton size and densities of both daphnids were significantly greater in the metalimnion. Densities of an invertebrate predator, Mesocyclops edax, were also significantly higher in the metalimnion compared to the epilimnion. Despite high fish abundance in 1997, mean summer (late May through August) chlorophyll a decreased 29% compared to 1996, consistent with expectations from grazing by large Daphnia protected in the refuge. Presence of large‐bodied zooplankton in 1997 despite high planktivore abundance demonstrated that successful creation of a low‐oxygen refuge from fish predation was possible in an open Great Lakes embayment where traditional biomanipulation via fish removal was not practical.