Food availability is a key regulator of the distribution, metabolism, and success of benthic populations. In deep sea ecosystems, hydrodynamics and depth play fundamental roles in determining benthic food resources. Recent studies suggest that the Southern Ocean sub-Antarctic front is shifting southward, with implications for primary production and food availability around the sub-Antarctic Islands embedded in the Antarctic Circumpolar Current. We used fatty acid (FA) and stable isotope (SI) analyses to investigate the trophic pattern of benthic invertebrates and suspended particulate matter (SPM) at three Depths (shallow: 100 m, middle: 300 m, and deep: 600 m) in three hydrographic regions with different flow and productivity regimes around the Prince Edward Islands. Both region and depth affected the SI values of SPM, while feeding guild was the key factor influencing consumer trophic values. Depth affected the delta N-15 of all trophic groups and the FA compositions of suspension feeders. Deeper samples had a higher delta N-15 and showed a greater proportion of mono- and saturated fatty acids, reflecting greater remineralization of SPM and of food reaching the seafloor. Region affected the delta C-13 and FA values of SPM, suspension feeders and deposit feeder/scavengers, with differences between the interisland and open ocean regions. This was probably linked to the retention of nutrients and phytoplankton between the islands. Critically, the effects of depth and hydrographic region were taxon specific, indicating that long-term responses to environmental change may have complex consequences for the feeding ecology and viability of benthic populations, with implications for the higher trophic levels that these populations support.