Eco-friendly extraction, optimization, and characterization of carotenoprotein from shrimp waste biomass using a novel alkaline protease, Ecoenzyme (ECOENZYME–ALKP)

Shrimp processing generates a substantial amount of waste, rich in valuable compounds like carotenoproteins. Traditional extraction methods often rely on harsh chemicals and consume significant energy, raising environmental concerns. This study introduces a sustainable alternative by using ecoenzyme (ECOENZYME–ALKP), a commercial alkaline protease from Bacillus subtilis with very high activity (200,000 U/g), to extract carotenoproteins from Pacific white shrimp waste for the first time. The quality of the extracted carotenoproteins was evaluated based on their chemical composition, antioxidant activities (DPPH, ABTS, FRAP), color, microstructure (SEM), and spectroscopic (FTIR) properties. Response surface methodology (RSM) was employed to optimize the hydrolysis conditions, identifying the optimal parameters as 36.58 °C, 2.99 h, pH 7.94, and 158.17 μl/100 g enzyme concentration. These conditions yielded a maximum degree of deproteinization (DDP) of 94.79% and a degree of hydrolysis (DH) of 51.28%. The results were compared to traditional NaOH methods, with the carotenoprotein powder produced by ecoenzyme (CPE) showing superior protein content, higher whiteness index, enhanced antioxidant activities, and stronger beta-sheet intensity compared to the carotenoprotein powder produced by chemical method (CPC). Both CPE and CPC exhibited increased DPPH radical scavenging, ABTS, and FRAP activities as concentrations increased up to 9 mg/ml (p < 0.05). The ecoenzyme proved to be both efficient and eco-friendly in producing high-quality carotenoproteins, making the resulting protein powder a viable option for use as a functional food ingredient for humans or animal feed. Notably, the ecoenzyme required significantly less enzyme (158.97 μl/100 g) compared to previous studies, highlighting its potent hydrolyzing ability.