IntroductionSprat (Sprattus sprattus) is an underutilized fish species that may act as an economic and sustainable alternative source of protein due to its good amino acid (AA) profile along with its potential to act as a source of multiple bioactive peptide sequences.Method and resultsThis study characterized the physicochemical, technofunctional, andin vitroantioxidant properties along with the AA profile and score of a sprat protein enzymatic hydrolysate (SPH). Furthermore, the impact of the SPH on the growth, proliferation, and muscle protein synthesis (MPS) in skeletal muscle (C2C12) myotubes was examined. The SPH displayed good solubility and emulsion stabilization properties containing all essential and non-essential AAs. Limited additional hydrolysis was observed followingin vitro-simulated gastrointestinal digestion (SGID) of the SPH. The SGID-treated SPH (SPH-SGID) displayedin vitrooxygen radical antioxidant capacity (ORAC) activity (549.42 μmol TE/g sample) and the ability to reduce (68%) reactive oxygen species (ROS) production in C2C12 myotubes. Muscle growth and myotube thickness were analyzed using an xCELLigence™ platform in C2C12 myotubes treated with 1 mg protein equivalent.mL−1of SPH-SGID for 4 h. Anabolic signaling (phosphorylation of mTOR, rpS6, and 4E-BP1) and MPS (measured by puromycin incorporation) were assessed using immunoblotting. SPH-SGID significantly increased myotube thickness (p< 0.0001) compared to the negative control (cells grown in AA and serum-free medium). MPS was also significantly higher after incubation with SPH-SGID compared with the negative control (p< 0.05).ConclusionsThese preliminaryin situresults indicate that SPH may have the ability to promote muscle enhancement.In vivohuman studies are required to verify these findings.