A Sustainable Workflow for the Recovery of Glucosylceramides from Apple Pomace

Apple pomace, a major agro-industrial byproduct of the juice industry, remains an underexploited renewable source of high-value sphingolipids. In this study, we developed an integrated, more sustainable workflow for the enrichment and recovery of phytoceramides from three apple pomace samples differing in geographical origin, cultivation practice, and processing history using chlorinated and n-hexane-free solvents. Nine hydroalcoholic solvent systems were evaluated by ultrasound-assisted extraction to balance extraction efficiency and environmental performance. UHPLC-VIP-HESI-MS/MS lipidomics of the phytoceramide-enriched fractions enabled a putative annotation of 64 CER and GlcCER molecular species, with phytosphingoid-based species predominating. Subsequent purification of these fractions by semi-preparative HPLC yielded four major GlcCERs, whose structures were elucidated by high-resolution MS and 1D/2D NMR spectroscopy. Total GlcCER content was 391 mg/Kg of dry pomace for Bodensee organic pomace (E1), 264 mg/Kg, for Saale-Unstrut organic pomace (E2), and 402 mg/Kg for Saale-Unstrut conventional pomace (E3). Overall, the workflow integrates green extraction, sustainable downstream processing, and advanced molecular characterization, demonstrating the potential of apple pomace for high-value sphingolipid valorization within a circular bioeconomy. Furthermore, this streamlined three-step process demonstrates feasibility at kilogram scale and provides a practical basis for further scale-up studies toward potential industrial application.