Orientador: Prof. Dr. Ricardo Pereira Ribeiro

 Data da Defesa: 11/06/2015



INTRODUCTION: Fish management procedures in aquiculture, such as capture, handling and transport, are often traumatic and cause serious physiological and biochemical reactions. The importance of the reduction of pre-slaughter stress is due to the fact that vigorous swimming intensely employs the white muscle, increases the anaerobic glycolysis and the production of lactic acid, and decreases muscle´s pH with the structural degradation of muscular proteins. The above diminishes the meat´s water-holding capacity (WHC). The genetic factors bonded to changes in the muscle´s WHC are related to alterations in the activity of ryanodine receptors which may decrease the ability of this channel to control the release of calcium to the cytoplasm of the muscular cell, especially in periods of physical stress. Calcium excess in the cytosol causes fast muscular contraction, with an acceleration of anaerobic metabolism and post-mortem glycolysis. The muscle´s pH is drastically decreased, affecting the muscle´s capacity to retain water in the intracellular stores. Changes in muscle quality derived from pre-slaughter stress diminish shelf life and bring economic liabilities for the fish industry. AIMS: Current analysis assesses the pre-slaughter stress caused by transport and its influence on stress indicators, fillets´ quality and sensorial factors, and genetic expression of the ryanodine receptor protein (RyR) in Nile tilapias. MATERIALS AND METHODS: The establishment of pre-slaughter management situations characterizing acute stress ante-mortem levels in the tilapias was conducted by an assay with different densities and periods of transport. The assay had a 3 x 4 factorial design, with three densities (100, 200 and 400 kg live weight/m³) at four transport periods (60, 120, 180 and 240 minutes), coupled to a control treatment (fish retrieved from the tank and immediately euthanized), with a total of 13 treatments and 10 replications per treatment (the fish was the experimental unit), with 130 fish (average weight 866.86±143.98 g and total length 33.80±1.63 cm). Levels of serum cortisol and plasma glucose as stress indicators, the gene expression of two isoforms of the ryanodine receptor protein (RyR1 and RyR3), coupled to the following quality parameters of the fillets: pH, colorimetry (luminosity L* and yellow-blue component b*), water-holding capacity, loss of water due to pressure and loss of water due to cooking, were analyzed. Different treatments of the factorial design were selected for the sensorial analysis, divided into two experiments. Sensorial Experiment 1: fish at average density 200 kg/m³ at 60 and 240 min (treatments 200 kg/m³ and 60 min; 200 kg/m³ and 240 min), and compared to fish of control treatment. Sensorial Experiment 2: fish transported during 180 min, at densities 100 and 400 kg/m³ (treatments 100 kg/m³ + 180 min; 400 kg/m³ + 180 min), compared to fish of control treatment. Sensorial analysis was undertaken with 90 non-trained testers, with two tests (acceptance and purchase intention). Tasters had to answer questions on the physical aspect, color, softness, juiciness and taste characteristic of the fish, through a 9-point hedonic scale, between 1 (I disliked it very much) and 9 (I like it very much). The purchase intention test was undertaken by a 5-point hedonic scale, between 1 (I would not buy it) and 5 (I would surely buy it). RESULTS AND DISCUSSION: There was no effect by density x time interaction on all the parameters evaluated. Transport time alone reduced cortisol levels up to 180 minutes, with increase of WHC and mRNA RyR1 and RyR3 (density 200 kg/m³). Isoforms RyR1 and RyR3 affected differently the water-holding capacity in the tilapia. Defect in calcium regulation mainly occurred in the gene that codified the RyR1. The pre-slaughter stress reduced the RyR1 gene expression and the decrease of preslaughter stress raised the fillets´ water-holding capacity. When treatments in the sensorial analysis only are assessed (Sensorials Experiments 1 and 2), the 60-min period in Sensorial Experiment 1 caused a higher concentration of serum cortisol (27.8 µg/dL) and plasma glucose (167.2 mg/dL) when compared to that at 240 min and to control. Stress caused WHC and a greater water loss by pressure at 60-min period when compared to fish transported during 240 min. The fillets had a higher score for juiciness. Moreover, stress did not affect pH, color and water loss by cooking. Transport densities 100 and 400 kg/m³ in Sensorial Experiment 2 did not affect stress indexes and instrumental quality parameters of the fillets, even though fish transported at 400 kg/m³ had a better acceptance by tasters. CONCLUSIONS: Increase in transport time up to 180 min decreased stress expressed by serum cortisol rates. More stressed fish had a lower RyR1 and RyR3 gene expression and a lower water-holding capacity in the meat. Stress reduction caused a linear increase up to 180 min of the parameters. Fish in treatment 200 kg/m³ and 240 min transport had lower stress rate, higher WHC and lower water loss by pressure, coupled to better juiciness when compared to fish transported at 200 kg/m³ for 60 min. Results give first evidence that pre-finish stress affects genes that codify ryanodine receptors and, consequently, the fillets´ water-holding capacity due to an increase in Ca2+ availability in cytosol. Following the example of other animal species, future perspectives derived from current analysis will provide the early identification of stressprone specimens and the development of meat with lower water-holding capacity. Keywords: water-holding capacity; colorimetry; cortisol; hypermetabolism; Oreochromis niloticus, pH


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