Untargeted Metabolomics Reveals Seasonal and Tissue-Specific Metabolic Shifts in Holothuria cinerascens

Sea cucumbers are valuable marine invertebrates known for their biologically active compounds with health-promoting properties. However, research has largely focused on select high-value species, overlooking others with promising bioactive potential. This study presents the first untargeted metabolomic analysis of Holothuria cinerascens from KwaZulu-Natal, South Africa, using ¹H-nuclear magnetic resonance (NMR) and ultra performance liquid chromatography quadruple time-of-flight mass spectrometry (UPLC-QTOF-MS) to assess metabolic and seasonal variability across three tissues: body wall, gonad and gut/mesentery. The body wall exhibited the highest metabolite diversity, with elevated levels of amino acids and potential triterpene glycosides, likely linked to stress or defence responses, whereas the gut/mesentery showed higher levels of sugars (galactose, xylose) and glycerol, possibly reflecting energy metabolism, diet or microbial activity. The gonad showed the lowest overall metabolite abundance but the highest levels of betaine and pyruvate. Seasonal differences were most pronounced in the gonad and gut/mesentery, likely related to reproductive activity and increased food availability. Compound identification was limited by structural isomerism and gaps in holothurian literature and databases, leaving several metabolites unidentified. These findings highlight the untapped potential of H. cinerascens and underscore the need to expand holothurian research. Future studies should prioritise compound characterisation and examine environmental influences on metabolite profiles to inform sustainable cultivation practices and natural product discovery.