Fish growth and survival are largely determined by the nutritional quality of their food, and the fish that grow quickly during early life stages are more likely to reproduce. To adequately estimate the quality of the prey for fish, it is necessary to understand the trophic links at the base of the food-web. Trophic biomarkers (e.g., stable isotopes and fatty acids) are particularly useful to discriminate and quantify food-web relationships. We explored the connections between plankton food-web components, and the seasonal and spatial dynamics of the trophic biomarkers and how this determines the availability of high-quality prey for juvenile Pacific salmon and Pacific herring in the Strait of Georgia, Canada. We demonstrate that the plankton food-web in the region is largely supported by diatom and flagellate production. We also show that spatial differences in terms of energy transfer efficiency exist in the region. Further, we found that the fatty acid composition of the zooplankton varied seasonally, matching a shift from diatom dominated production in the spring to flagellate dominated production in the summer. This seasonal shift conferred a higher nutritional value to zooplankton in the summer, indicating better quality prey for juvenile salmon and herring during this period.