Aquaculture Europe 2023

September 18 - 21, 2023


Add To Calendar 20/09/2023 14:30:0020/09/2023 14:45:00Europe/ViennaAquaculture Europe 2023ELECTRO-STUNNING PARAMETERS AND WATER TEMPERATURE SIGNIFICANTLY AFFECT FLESH QUALITY IN MEDITERRANEAN MARINE FISH SPECIESSchubert 3The European Aquaculture Societywebmaster@aquaeas.orgfalseDD/MM/YYYYaaVZHLXMfzTRLzDrHmAi181982


 Rafael Angelakopoulos1 , Alexia Fitsili1, Arkadios Dimitroglou2, Leonidas Papaharisis3, Katerina A. Moutou1

1 Department of Biochemistry and Biotechnology, University of Thessaly, Biopolis, 41500 Larissa, Greece

2 Department of Animal Science, Agricultural University of Athens, Iera Odos 75, 11855, Athens, Greece

3Department of Research & Development, Avramar Aquaculture SA, 341 00 Chalkida, Greece




 Fish fillet proteolysis is a complex process that involves various biochemical and structural changes post-mortem, along with microbial activity. These changes can result in a decline in fish fillet quality and shelf life,  often manifested as muscle softening and the creation of gaps in the myocommata

. Proteases responsible for muscle deterioration, namely, calpains, cathepsins and metalloproteases, can originate from both muscle tissue and the digestive system, and their activity is influenced by factors such as stress, temperature, and handling during harvesting and transportation

 . In Mediterranean aquaculture, species such as  Sparus aurata , Dicentrarchus labrax , and Pagrus major are harvested using ice slurry that can c ause  stress on fish. This stress can exacerbate proteolytic cleavage and muscle softening, leading to further degradation of fillet quality

 . The need for a more humane method of fish harvesting has led to the application of electro-stunning , suggested  by both EFSA and OIE. However, before proceeding with the wide application of e lectro-stunning  in Mediterranean marine fish farming,  it is necessary to ensure that  fish fillet quality is not compromised

. In this direction, the effects of electro-stunning on  fish fillet proteolysis and flesh quality  are investigated in the above Mediterranean farmed fish species , the gilthead sea bream, European seabass and the red seabream.

Materials and methods

 The fish were harvested at the same fish farm in Astakos , Aitoloakarnania , Greece, during three different temperature periods: Warm (August, 25°C), Moderate (June, 21°C), and Cold (February/March, 15.5°C). Three harvest methods  were applied: the conventional method of ice slurry and two electro-stunning settings of higher and lower voltage , with the same flow rate. White muscle samples were excised  from each fish on slaughter day (Day 0) and on days 1, 2, 5, 7, and 13 post-harvest and were snap-frozen in liquid nitrogen. The activity of Calpain, Collagenase, Cathepsin B, and L was assayed using the Barrett and Kirschke method with minor refinements, and protein content was quantified using the Bradford method. Activity was expressed as fluorescence units change per minute per mg protein. Additionally, white muscle  samples at harvest day (Day 0) and on days 7 and 13 post-harvest underwent  histological analysis to assess flesh quality



 The post-mortem activity of proteolytic enzymes in European seabass, gilthead seabream, and red seabream at different water temperatures and harvest methods was investigated. The results showed that calpain and collagenase activities were activated early post-mortem, and species-specific variations were observed in enzyme activity levels. Cytoplasmic calpains had the highest average activity among proteolytic enzymes, while collagenase activity shared a similar temporal pattern with calpain. Cathepsin B and L also showed a positively correlated activation regardless of the harvest method.  Changes in muscle histology caused by the action of proteolytic enzymes can lead to flesh softening and loss of texture. The myofibrils, which make up most of the muscle fiber volume, are  particularly vulnerable to degradation by endogenous proteases

. In our study, we observed an increase in the average single fiber volume density between days 0 and 7, irrespective of the harvest method used, though this increase was significant in electro-stunned groups. The temperature had a significant effect on enzyme activity and the histological phenotype, with the highest activities observed in the moderate temperature period for all species. Harvest method had a milder effect  than the water temperature  on enzymatic activities and the histological phenotype.

Discussion and conclusion

 In the present study the effect of electro-stunning as a harvest method  was  investigated  on the proteolytic activity and flesh quality of three Mediterranean farmed fish  species. It is known that electro-stunning can have an impact on pre-slaughter stress and pH levels, which in turn, may affect the activation of proteolytic enzymes

. The observed variations in enzyme activity among different species of fish, water temperatures, and harvest methods can be attributed to differences in muscle composition, physiological status, and post-mortem metabolism.  Previous studies have suggested that this increase may be due to osmotic phenomena resulting from changes in intracellular membrane permeability and disruptions in ion balance in the cytoplasm


 These findings suggest that  water temperature, species and harvest method have an impact in the histological phenotype due to protease activity  and they highlight the importance of developing species-specific humane post-harvest strategies to effectively preserve fish fillet quality ensuring animal welfare.


 This research was co-financed by Greece and the European Union, the European Maritime and Fisheries Fund, in the context of the implementation of the Greek Operational Programme for Fisheries, Priority Axis “Innovation in Aquaculture”, project title “Development and industrial scale evaluation of an innovative humane slaughter system and assessment of welfare in aquaculture marine fish species” MIS 5010690.