Background Tuna muscle greening is a problem that occurs after heating. A hypothesis has been postulated to address this problem, involving a conserved Cys residue at position 10 (Cys-10) present on tuna myoglobin (Mb) that is exposed during the thermic treatment, forming a disulfide bond with free cysteine (Cys) in the presence of trimethylamine oxide (TMAO), resulting in the greening of the tuna Mb. Methods We present a study using skipjack tuna (Katsuwonus pelamis) metmyoglobin (MbFe(III)-H2O) where the effect of free Cys (1–6 mM), TMAO (1.33 mM), and catalase on the greening reaction (GR) was monitored by UV-vis spectrometry during thermal treatment at 60 °C for 30 min. Moreover, the participation of Cys-10 on the GR was evaluated after its blocking with N-ethymaleimide. Results The GR occurred in tuna MbFe(III)-H2O after heat treatment with free Cys, forming sulfmyoglobin (MbFe(II)-S) as the responsible pigment for the tuna greening. However, the rate constants of MbFe(II)-S production depended on Cys concentration (up to 4 mM) and occurred regardless of the TMAO presence. We postulate that two consecutive reactions involve an intermediate ferrylmyoglobin (promoted by H2O2) species with a subsequent MbFe(II)-S formation since the presence of catalase fosters the reduction of the rate reaction. Moreover, GR occurred even with blocked Cys-10 residues in tuna Mb and horse Mb (without Cys in its sequence). Discussion We found that GR is not exclusive to tuna Mb´s, and it can be promoted in other muscle systems. Moreover, Cys and thermal treatment are indispensable for promoting this pigmentation anomaly.

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Jumbo squid () muscle proteins show low functionality with limited use in gel products. This work aims to assess the influence of adding the natural and commercially available fibre, amidated low-methoxyl pectin (at 0.5, 1.0, 1.5, 2.0 and 3.0%), on the physicochemical and functional characteristics of jumbo squid () mantle muscle gels. The addition of 0.5% fibre showed an immediate effect on the gel texture profile analysis, improving hardness (p<0.05) from (3.4±0.7) N of the control (no added fibre) to (5.2±0.9) N, and increasing elasticity (p≥0.05). Shear force was significant only at 3.0% fibre addition. Water holding capacity also improved (p<0.05) with fibre addition (from 75% in the control to 90-95% after the treatments). Whiteness was affected (p<0.05) when 3.0% fibre was added. Differential scanning calorimetry showed two endothermic transition peaks in the gels. The second peak (actin) increased (p<0.05) by 1-2 °C with fibre addition. Therefore, the present study demonstrates that amidated low-methoxyl pectin (0.5-3.0%) is an excellent ingredient to improve jumbo squid mantle muscle protein functionality, increasing the gel texture and water retention characteristics.

Freezing conditions affect fish muscle protein functionality due to its denaturation/aggregation. However, jumbo squid (Dosidicus gigas) muscle protein functionality remains stable even after freezing, probably due to the presence of low-molecular-mass compounds (LMMC) as cryoprotectants. Thus, water-soluble LMMC (<1 kDa) fraction obtained from jumbo squid muscle was evaluated by Fourier transform infrared spectrometry. From its spectra, total carbohydrates, free monosaccharides, free amino acids and ammonium chloride were determined. Cryoprotectant capacity and protein cryostability conferred by LMMC were investigated by differential scanning calorimetry. Fraction partial characterization showed that the main components are free amino acids (18.84 mg/g), carbohydrates (67.1 µg/mg) such as monosaccharides (51.1 µg/mg of glucose, fucose and arabinose in total) and ammonium chloride (220.4 µg/mg). Arginine, sarcosine and taurine were the main amino acids in the fraction. LMMC, at the mass fraction present in jumbo squid muscle, lowered the water freezing point to –1.2 °C, inhibiting recrystallization at 0.66 °C. Significant myofibrillar protein stabilization by LMMC was observed after a freeze-thaw cycle compared to control (muscle after extraction of LMMC), proving the effectiveness on jumbo squid protein muscle cryo- stability. Osmolytes in LMMC fraction inhibited protein denaturation/aggregation and ice recrystallization, maintaining the muscle structure stable under freezing conditions. LMMC conferred protein cryostability even at the very low mass fraction in the muscle.