Aquatic-to-terrestrial subsidies play crucial roles in shaping the dietary niches of riparian predators. These subsidies consist mainly of emerging aquatic insects that facilitate the transfer of nutrients from aquatic to terrestrial ecosystems. Riparian web-building spiders are key in linking aquatic and terrestrial food webs, as they are among the most common and abundant riparian secondary consumers. However, we still lack a comprehensive understanding of how variations in aquatic subsidies affect interactions among riparian predators such as spiders. We examined two broad guilds of riparian spiders, horizontal web-builders and vertical web-builders, using stable isotope and fatty acid analyses to explore the distinct dietary patterns of each guild. We hypothesized that these guilds exhibit distinct dietary patterns driven by their prey capture strategies (web orientation) and the varying contributions of aquatic subsidies across ecological gradients. In fact, horizontal web-builders, which primarily target aquatic-emerging insects, relied more on aquatic resources during high-emergence seasons, particularly in downstream areas where insect emergence was higher than upstream. In contrast, vertical web-builders that capture larger, fast-flying terrestrial insects relied less on aquatic subsidies. Furthermore, the two guilds' isotopic and fatty acid niches were more distinct when insect emergence was higher, while overlap increased when emergence was lower. The fatty acid and stable isotope results varied moderately, but the outcomes were largely consistent, supporting the conclusion that the realized niches of these two guilds are shaped by prey availability. These findings highlight the importance of aquatic-to-terrestrial nutrient flows in structuring riparian predators’ diets and underscore how resource availability mediates inter-guild niche differentiation. Broadly, our study provides insights into how cross-boundary nutrient dynamics influence dietary niche partitioning and species interactions within riparian food webs.