Nine fish per treatment were filleted, and the right fillets were weighed, packaged in a plastic bag and then refrigerated at + 4 °C. After 24 h at + 4 °C, the right fillets were gently dried with paper to remove excess moisture, and then weighed. Subsequently, the muscle pH (pH24 h) and flesh color were assessed on the inside portion of the cranial, medial and caudal region of each fillet. The pH24 measurement was performed using a Crison MicropH 2001 (Crison Instruments, Barcelona, Spain) equipped with a combined electrode and an automatic temperature compensator. The flesh color was analysed using a bench colorimeter Chroma Meter CR-400 (Konica Minolta Sensing Inc., Osaka, Japan). The results were expressed in terms of lightness (L*), redness (a*) and yellowness (b*) in the CIELAB color space model [28].

The water holding capacity was calculated as follows:

Drip loss (DL; %) = [(raw fillet weight (g) − raw fillet weight after 24h (g)) / raw fillet weight (g)] × 100

The fillets were then individually vacuum-packaged in a plastic bag and stored at − 20 °C. After total freezing, the fillets were thawed at + 4 °C, removed from the bags, dried with paper, and weighed to calculate the thawed loss (TL) as follows:

Thawing loss (TL; %) = [(raw fillet weight (g) − thawed fillet weight (g)) / raw fillet weight (g)] × 100

The same fillets were then vacuum-packaged in a plastic bag and cooked in a fish kettle for 10 min at 80 °C (core temperature of the fillets: 75 °C). After cooking, the bags were removed from the fish kettle and cooled in fresh water for 15 min to stop the cooking process. Then, the fillets were removed from the bags, dried with paper and weighed again to calculate the cooking loss (CL), as follows:

Cooking loss (CL; %) = [(raw fillet weight (g) − cooked fillet weight (g)) / raw fillet weight (g)] × 100

Following cooking loss determination, a cooked fish sample (1.5 cm × 1.5 cm) from each fillet was sheared perpendicular to the fibre direction using the Instron 5543 Universal Testing Machine (Instron Corporation, Canton, Massachusetts, USA) equipped with a straight edged shear blade (crosshead speed of 30 mm/min). The maximum peak force recorded during the analysis was reported as Newton (N) shear force.

The nine left fillets per treatment were frozen, finely ground with a knife mill (Grindomix GM200; Retsch GmbH, Haan, Germany) and freeze-dried (Edwards MF 1000, Milan, Italy) to determine their proximate composition (DM, CP, EE, and ash), according to the same procedures implemented for feed analyses [23, 24]. The freeze-dried and ground samples of the fish fillet were also used to assess their FA composition. After dichloromethane-methanol extraction of total lipids from fillets, a basic saponification and a BF3 esterification were used for the determination of the fatty acid composition, adding tridecanoic acid as internal standard, as reported by Renna et al. [29]. FAME were separated using the same analytical instruments and temperature program previously reported for the FA analysis of feeds. Peaks were identified by injecting pure FAME standards as reported in Renna et al. [30]. The results were expressed as mg/100 g wet weight (ww). All chemical analyses were performed in duplicate.

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