Effects of cultivation environment and culture layer on metabolite profiles, microbial safety, and toxic/trace element accumulation in cultivated Gracilaria.

The rapid expansion of tropical seaweed aquaculture has increased the need to understand better how farming environments influence the metabolic quality and biosafety of cultivated macroalgal biomass under field-scale conditions. This study investigated the interactive effects of cultivation environment (pond vs. nearshore) and culture layer (surface vs. on-bottom) on putative metabolite profiles, microbial quality, and toxic/trace element dynamics in cultivated Gracilaria sp. from the southeastern coast of Bangladesh using a fully factorial (2 × 2) experimental design. Gas chromatography-mass spectrometry (GC-MS) identified 45 putatively annotated metabolites, with multivariate analyses revealing clear treatment-specific metabolic differentiation among cultivation environments and culture layers. Pond systems were generally associated with greater abundances of hydrocarbon-derived and lipid-associated metabolites, whereas nearshore cultivation was characterized by relatively higher abundances of long-chain fatty acid-associated, oxygenated, and nitrogen-containing metabolites. Microbial abundance also differed significantly among cultivation systems, with surface nearshore treatments exhibiting higher presumptive Salmonella/Shigella and enteric Gram-negative bacterial loads, whereas on-bottom pond systems showed comparatively greater presumptive Vibrio abundance. Toxic heavy metal and metalloid accumulation varied with cultivation conditions, with on-bottom pond systems accumulating higher concentrations of As, Cr, Hg, and Pb, while Cd concentrations were comparatively higher in nearshore systems. Several essential minerals and trace elements were enriched in near-shore and on-bottom cultivation systems. Overall, the findings demonstrate that cultivation environment and culture layer strongly influence the biochemical quality, microbial safety, and elemental composition of cultivated Gracilaria sp., highlighting the importance of balancing biomass quality, food safety, and environmental conditions when selecting cultivation strategies for sustainable seaweed aquaculture.