Towards standardization of carotenoid analysis in crustaceans: effects of storage, extraction time, and calculations

Carotenoids play an essential role in aquaculture by supporting growth, survival, and stress tolerance in cultured species, while also contributing to product coloration, an important indicator of its quality and market value. Reliable monitoring of carotenoid levels in cultured organisms and feeds, as well as the development of effective feed enrichment strategies, requires standardized analytical procedures. This study provides a comparative evaluation of sample preparation methods and calculation approaches used for spectrophotometric determination of total carotenoids, using Artemia as a model organism. The results show that extending acetone extraction from one to three days increases the measured carotenoid concentration in fresh samples, whereas the opposite trend is observed in frozen samples, where prolonged extraction leads to pigment degradation. Additionally, the use of different calculation formulas can produce multi‑fold discrepancies in carotenoid concentrations derived from identical absorbance data. These findings highlight the need for unified protocols for sample preparation, extraction, and data processing to ensure consistency and comparability of carotenoid measurements in aquaculture research and practice.