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Effect of a natural organic acid-icing system on the
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microbiological quality of commercially relevant chilled fish
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species
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Authors: Minia Sanjuás Reya, Bibiana García-Sotoc, José R. Fuertes-Gamundic,
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Santiago Aubourgb and Jorge Barros-Velázqueza,*
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Affiliations: aDepartment of Analytical Chemistry, Nutrition and Food Science, School
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of Veterinary Sciences, University of Santiago de Compostela, E-27002 Lugo, Spain;
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b
Department of Seafood Science and Technology, Institute for Marine Research (IIM-
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CSIC), C/ Eduardo Cabello 6, E-36208 Vigo, Spain; and Cooperativa de Armadores del
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Puerto de Vigo (ARVI), Muelle del Berbés, Vigo, Spain
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Corresponding Author: Professor Jorge Barros Velázquez, Department of Analytical
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Chemistry, Nutrition and Food Science, School of Veterinary Sciences, University of
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Santiago de Compostela, E-27002 Lugo, Spain. Tel.: +34.600.942264; Fax:
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+34.981.825474; E-mail: jorge.barros@usc.es
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Running title: Organic acid-icing system for fish products
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Abstract. Natural preservative organic acids (ascorbic, citric and lactic acids) were used
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to prepare a novel organic acid-flake icing system and evaluated for the preservation of
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three commercially relevant marine species during refrigerated storage: hake
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(Merluccius merluccius), megrim (Lepidorhombus whiffiagonis) and angler (Lophius
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piscatorius). The icing system was prepared with fresh water and two different
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concentrations of a commercial acid mixture-formula containing the three organic acids
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at 800 ppm and 400 ppm (C-800 and C-400 batches, respectively). Aerobic mesophiles,
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psychrotrophs, proteolytic bacteria, trimethylamine and pH levels were evaluated in fish
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muscles throughout storage; these results were then compared with sensory analyses.
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Significantly (P<0.05) lower counts of mesophiles were found for hake and megrim in
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the C-800 and C-400 batches compared with the C-0 control batch. In the case of
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angler, significantly (P<0.05) lower counts of mesophiles, psychrotrophs and
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proteolytic microorganisms (P<0.05) were found for fish stored in the C-800 icing
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conditions. Both C-400 and C-800 megrim batches exhibited significantly (P<0.05)
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lower pH values than the C-0 control batch, and this result was also observed in the C-
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800 angler batch. These results agreed with the sensory analysis, which revealed that the
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shelf lives of the three fish species in the C-800 batch were extended. According to the
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parameters assessed, storage of hake, megrim and angler in the proposed ice system
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containing natural organic acid preservatives better protects their microbial and sensory
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qualities.
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Keywords: fish, storage, organic acids, icing systems, microbial quality, sensory
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quality.
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2
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1. Introduction
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The quality and freshness of marine species rapidly decline post-mortem due to a
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variety of microbial and biochemical degradation mechanisms (Pigott & Tucker, 1990;
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Whittle, Hardy, & Hoobs, 1990). Thus, the sensory quality and nutritional value of
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chilled fish deteriorate as a result of different damage pathways, such as endogenous
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enzymatic activity, microbial development and lipid oxidation mechanisms (Olafsdóttir
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et al., 1997; Whittle, Hardy, & Hoobs, 1990).
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To slow the mechanisms involved in quality loss, the fish specimens should be
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refrigerated immediately after capture. Therefore, fish have traditionally been cooled
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and stored in either flake ice (Nunes, Batista, & Morâo de Campos, 1992), refrigerated
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sea water (Kraus, 1992), or ice slurries (Barros-Velázquez, Gallardo, Calo, & Aubourg,
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2008; Rodríguez, Barros-Velázquez, Piñeiro, Gallardo, & Aubourg, 2006; Rodríguez,
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Losada, Aubourg, & Barros-Velázquez, 2004), or they have been preserved by exposure
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to chemical agents (Hwang & Regenstein, 1995). However, the seafood industry is
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always searching for new preservation strategies to extend the shelf life of fish and
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provide the consumers with fish material with the best quality, both at the sensory and
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nutritional levels.
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Natural organic acids are preservatives that have been receiving increasing attention
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as minimal processing strategies because they are easily attained, have a low
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commercial cost and can be used at a wide range of permitted concentrations. These
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organic acids are soluble in lipids in their undissociated forms, which allows them to
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cross the microbial membrane into the microbial cytoplasm, where the acids tend to
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dissociate and deliver hydrogen ions and a particular anion (Booth, 1985; Booth &
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Kroll, 1989). As a result, microorganisms are forced to export the excess hydrogen ion
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to maintain a physiological pH inside the cell, which is an energy-depleting process that
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limits bacterial growth. Otherwise, the excess hydrogen ions in the cytoplasm may
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cause the pH to decrease to levels that are incompatible with bacterial growth (Gould,
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1996).
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Among the natural organic acids, ascorbic acid (AA) and citric acid (CA) and their
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salts are well-known chelators and acidulants in biological systems, which are
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especially beneficial for fish oil and emulsions (Kelleher, Silva, Hultin, & Wilhelm,
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1992; Osborn-Barnes & Akoh, 2003), minced fish (Hwang & Regenstein, 1995;
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Stodolnik, Blasiak, & Broszedzka, 1992), fish fillets (Badii & Howell, 2002;
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Pourashouri, Shabanpour, Aubourg, Rohi, & Shabani, 2009) and whole fish (Aubourg,
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Pérez-Alonso, & Gallardo, 2004). Treatment with lactic acid (LA) has also been
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reported to be effective for extending the shelf lives of fish fillets (Kim, Hearnsberger,
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& Eun, 1995; Metin, Erkan, Varlik, & Aran, 2001) and coated fish (Gogus, Bozoglu, &
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Yurdugul, 2006). Interestingly, the effectiveness of LA in inhibiting the growth of
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Gram-negative bacteria, which are responsible of fish spoilage, has also been reported
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(Alakomi et al., 2000). However, when employed above certain concentrations or for
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relatively long exposure times, evidence of fish digestion by the organic acids has been
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observed, which may produce unfavourable sensory and physical properties of the fish
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(Kim et al., 1995; Metin et al., 2001).
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This study provides a novel strategy that employs aqueous solutions of natural
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organic acids as biopreservatives to prepare flake ice for chilling fish specimens. We
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have previously demonstrated the biochemical benefits of employing an organic acid-
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icing system, including AA, CA and LA, on commercially relevant fish species in terms
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of lipid oxidation and hydrolysis (García-Soto, Sanxuás, Barros-Velázquez, Fuertes-
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Gamundi, & Aubourg, 2010). However, the microbial effects of this system have not
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been evaluated to date. Accordingly, this work focused on studying the effects of an
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organic acid-icing system on the microbial development and shelf lives of three
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commercially
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Lepidorhombus whiffiagonis; and angler, Lophius piscatorius) from the Grand Sole
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North Atlantic fishing bank during refrigerated storage.
relevant
fish
species
(hake,
Merluccius
merluccius;
megrim,
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2. Materials and methods
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2.1. Preparation of icing systems including organic acids
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A commercial formula (BPS2) including an organic acid mixture was provided by
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the company Atlantic One S.L. (Vigo, Spain) for use in the present work. This product
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consisted of a water-soluble viscous liquid of AA, CA and LA (1 meq of each acid/120
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mg product) in glycerol, which is regarded as safe (GRAS) for use in foods according to
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the European and American regulatory agencies (Giese, 1996; Madrid, Madrid, &
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Madrid, 1994).
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Preliminary trials were performed to optimise the most convenient product
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concentrations for the icing system. Thus, the effects of a wide range (70-2000 ppm) of
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aqueous concentrations of the organic acid mixture on the sensory qualities of fish
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specimens were initially tested. From this analysis, it was concluded that 800 ppm and
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400 ppm provided the most convenient results (data not shown). Therefore, both
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concentrations were selected and employed in this study as the C-800 and C-400
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batches, respectively. The organic acid-icing mixture was prepared as flake ice with
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water containing the desired concentrations of organic acids in an Icematic F100
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Compact device (Castelmac Spa, Castelfranco, Italy). Both batches were compared to
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fish kept in traditional ice prepared from water (C-0 batch).
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2.2. Fish material, processing and sampling
Specimens
of
hake
(Merluccius
merluccius;
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individuals),
megrim
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(Lepidorhombus whiffiagonis; 78 individuals) and angler (Lophius piscatorius; 78
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individuals) were caught near the Galician Atlantic coast (North-Western Spain) in
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autumn of 2009 and kept in ice on-board. Upon arrival to the laboratory, the specimens
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of each species were analysed by triplicate on day one. The remaining specimens were
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divided into three batches and stored in different types of ice (C-800, C-400 and C-0
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conditions) at a 1:1 fish-to-ice ratio. All batches were placed in a refrigerated room
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(4ºC). The storage boxes employed allowed draining, and ice was renewed when
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required. Fish samples from different batches were collected for analysis on days one,
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five, eight and 12; day 15 only was considered for hake. On day zero, six individuals of
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each fish species were analysed as starting fish (three groups of two individuals). For
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each fish species and icing condition, three different groups were analysed separately to
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achieve statistically significant results (n = 3).
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2.3. Microbiological analyses
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Samples of 10 g of fish muscle were dissected aseptically from chilled fish
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specimens, mixed with 90 ml of 0.1% peptone water (Merck, Darmstadt, Germany), and
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homogenised in a stomacher (AES, Combourg, France) as previously described (Ben-
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Gigirey, Vieites Baptista de Sousa, Villa, & Barros-Velázquez, 1998; Ben-Gigirey,
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Vieites Baptista de Sousa, Villa, & Barros-Velázquez, 1999). In all cases, serial
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dilutions from the microbial extracts were prepared in 0.1% peptone water. Total
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aerobes were investigated by surface inoculation into plate count agar (PCA, Oxoid
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Ltd., London, UK) after incubation at 30ºC for 48 h. Psychrotrophs were also
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investigated in PCA, but incubation was carried out at 7–8ºC for seven days.
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Microorganisms exhibiting a proteolytic phenotype were investigated in casein-agar
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medium after incubation at 30ºC for 48 h, as previously described (Ben-Gigirey, Vieites
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Baptista de Sousa, Villa, & Barros-Velázquez, 2000).
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2.4. TMA-N analysis
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Trimethylamine-nitrogen (TMA-N) values were determined by the picrate method
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as previously described (Tozawa, Erokibara, & Amano, 1971). This technique involves
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the preparation of a 5% trichloroacetic acid extract of fish muscle (10 g/25 ml). The
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results were expressed as mg TMA-N/100 g muscle.
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2.5. Assessment of pH value
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The evolution of pH in hake, megrim and angler during storage times was
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determined by means of a pH-meter fitted with a 6-mm diameter insertion electrode
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(Crison, Barcelona, Spain). The pH level was measured through penetration of the
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electrode into the fish samples.
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2.6. Sensory analysis
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The sensory analysis was conducted by a sensory panel of five experienced judges,
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according to the guidelines concerning fresh and refrigerated fish (Council Regulation,
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1990). Te panellists had been involved in sensory analysis of different kinds of fish and
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seafood products for the last ten years. Before the present experiment, a special training
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was administered on chilled hake, megrim and angler.
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Four categories were used to rank the samples: highest quality (E), good quality (A),
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fair quality (B) and unacceptable quality (C). Sensory assessment of the fish included
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the following parameters: gills (odour, colour and mucus development), eyes, external
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odour (development of putrid and other off-odour), muscle odour (raw and cooked fish;
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development of putrid and other off-odour) and taste (cooked fish; development of
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putrid and other off-taste). At each sampling time, the fish muscle portions were
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presented to the panellists in individual trays, which were scored individually. The
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panel members shared the samples tested.
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2.7. Statistical analyses
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Bacterial counts were transformed into log CFU/g before undergoing statistical
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analysis. Data corresponding to the three icing batches were subjected to one-way
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analysis of variance to assess significant (P<0.05) differences between batches. The
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PASW Statistics 18 software for Windows (SPSS Inc., Chicago, IL, USA) was used to
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explore the statistical significance of the results, including multivariate contrasts and
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multiple comparisons by Scheffe and Tukey tests. A confidence interval at the 95%
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level was used in all cases.
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3. Results and discussion
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3.1. Microbiological analyses
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The comparative evolution of aerobic mesophiles during storage for the three fish
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species can be seen in Figures 1, 2 and 3. Statistically significantly (P<0.05) lower
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mesophile counts were found for hake (Figure 1) and megrim (Figure 2) stored in the C-
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192
800 and C-400 systems compared with the C-0 control batches. On the other hand, the
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angler (Figure 3A) C-800 batch exhibited statistically significantly (P<0.05) lower
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mesophiles counts than the C-400 and C-0 batches, and there were no statistically
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significant differences between two latter batches. The aerobic mesophile numbers for
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hake and megrim C-800 and C-400 batches were below 7 log CFU/g and for angler was
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below 6 log CFU/g at the end of storage. These values obtained for the fish specimens
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stored in the proposed organic icing system were below those generally considered
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necessary to induce fish spoilage (Gram & Huss, 1996). In contrast, the mesophile
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counts were as high as 7.15 log CFU/g and 7.26 log CFU/g for the hake and megrim C-
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0 batches, respectively.
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Our results agreed well with other authors that reported the effectiveness of organic
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acids for inhibition of aerobic bacteria evolution in spiny dogfish (Squalus acanthias)
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(Sengoer, Mol, & Uecok, 2007). In addition, other authors found that fish products
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treated with ellagic acid in combination with ascorbic acid and sodium ascorbate
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exhibited a significant delay in the proliferation of aerobic mesophiles (Zambuchini,
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Fiorini, Verdenelli, Orpianesi, & Ballini, 2008)
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With respect to the psychrotrophs, statistically significantly (P<0.05) lower counts
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were determined for angler (Figure 3B) stored in the C-800 system compared with the
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C-400 and C-0 batches. Nevertheless, the psychrotroph counts on the last day of storage
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(12 days) were lower than 7 log CFU/g for any of the batches tested. These results also
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agree with a previous study in which other fish species, such as spiny dogfish, were
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treated with salts, ascorbic acid and citrate (Sengoer, Mol, & Uecok, 2007; Yang,
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Jhaveri, & Constantinides, 1981). No significant differences for psychrotrophs were
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found for megrim or hake.
9
216
Finally, the counts of proteolytic microorganisms were also significantly (P<0.05)
217
lower in the angler C-800 batch than in the other two batches (Figure 3C). The role of
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proteolytic microorganisms in the degradation of fish muscle has been highlighted
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(Rodríguez, Barros-Velazquez, Ojea, Pineiro, & Aubourg, 2003). Accordingly, the
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partial inhibition of this microbial group by the organic acids tested in this study is
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another remarkable result in terms of fish quality. As in the case of the psychrotrophs,
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no significant differences for proteolytic microorganisms were found in megrim or
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hake.
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According to the results of the microbial analysis, the presence of organic acids in
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the icing system significantly slowed down the growth of certain microbes in fish
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muscles; this result was especially relevant for angler, and to a lesser extent, for hake
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and megrim. Other studies of flow ice systems prepared from marine water indicated
228
that when the ice melts, the salt solution may exert a washing effect of the fish surface,
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which reduces the surface microbial load and thus limits microbial diffusion into the
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muscle through the skin (Campos, Rodríguez, Losada, Aubourg, & Barros-Velázquez,
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2005; Rodríguez, Barros-Velázquez, Piñeiro, Gallardo & Aubourg, 2006; Rodríguez,
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Losada, Aubourg, & Barros-Velázquez, 2004). Likewise, in this study, melting of the
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ice flakes prepared with organic acids may exert a similar washing effect, which also
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prevents the formation of surface microbial biofilms that may induce fish spoilage.
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3.2. TMA-N evolution
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The evolution of TMA-N in fish muscles during refrigerated storage is presented in
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Table 1. In the case of hake, the C-800 batch exhibited lower levels of TMA-N
239
formation throughout storage compared with the other two batches, which indicates that
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the combination of organic acids at this proportion is beneficial for decreasing the
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production of these amines by microbes. In the case of megrim, the inclusion of organic
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acids in the icing system did not produce any significant improvement in terms of
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TMA-N formation. Finally, for the angler, the incorporation of CA, AA and LA in the
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C-800 batch provided significant protection against TMA-N formation compared with
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the C-0 batch at all sampling times. Moreover, the effect of the C-400 batch, which
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included intermediate concentrations of the natural organic acids, was also favourable
247
compared with the C-0 batch.
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249
3.3. pH analyses
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Increases in the pH of the fish muscle indicate the accumulation of alkaline
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compounds, such as ammonia compounds and TMA, which are principally derived
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from microbes (Hebard, Flick, & Martin, 1982). In this sense, it has been suggested that
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pH values above 7 may limit the shelf lives of certain fish species (Moral, 1987; Ruiz
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Capillas & Moral, 2001).
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In our study, no significant effects of the organic acids on pH evolution of hake
256
muscle were observed. With respect to the muscles of megrim stored in the C-800 and
257
C-400 systems, the pH values were significantly (P<0.05) lower compared with the C-0
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control batch (Figure 4A). Thus, the pH differences at the end of the storage period
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were more than 0.50 pH units, which indicates that the incorporation of natural
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preservatives to the icing system, at either of the two concentrations tested, significantly
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reduced the microbial and endogenous alkalinising mechanisms that may limit fish shelf
262
life. Likewise, in the case of angler stored in the C-800 icing system, the pH value of
11
263
the muscle was significantly (P<0.05) lower compared with the C-400 and C-0 control
264
batches (Figure 3D).
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These results agree with other studies that reported better pH value maintenance in
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catfish fillets treated with lactic acid (Kim, Hearnsberger, & Eun, 1995) and in sole
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specimens treated with ellagic acid, ascorbic acid and sodium ascorbate (Zambuchini,
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Fiorini, Verdenelli, Orpianesi, & Ballini, 2008). However, and similar to previous
269
analyses, our study includes a novel mixture of organic acids that has not been tested
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before for fish preservation purposes.
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3.4. Sensory analyses
273
The results of the sensory evaluation of the three species tested with the different
274
icing systems are presented in Tables 2, 3 and 4. In the case of hake, the C-800 batch,
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which incorporated the highest concentration of organic acids, showed the best
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behaviour and retained an acceptable quality even after 12 days of storage; however,
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this result was not observed for the other two batches. For hake, storage in the C-800
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organic acid-icing system provided better preservation of all sensory quality parameters,
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especially muscle odour and taste.
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In the case of megrim, similar results were obtained. Thus, only the C-800 batch
281
retained acceptable quality after eight days of storage, as compared with the other two
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icing batches. As was the case for hake, muscle odour and taste was the sensory
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parameters for which the C-800 organic acids-icing system provided the best results.
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Finally, angler specimens exhibited a better behaviour in all batches compared with
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hake and megrim. However, the C-800 batch maintained very good sensory quality up
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until day eight, and the other batches were also considered to be acceptable at that time.
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Moreover, the angler specimens stored in the C-800 organic acid-icing system were
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acceptable on day 12, which was not observed for the other two batches.
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In general and according to these results, no significant beneficial effect of the C-400
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organic acids-icing system was found on fish quality and shelf life compared to the
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control fish. In contrast, the incorporation of AA, CA and LA at the concentrations in
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the C-800 batch resulted in better maintenance of the quality and extension of the shelf
293
life compared with the other two batches. This result agrees with the microbiological
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numbers determined in this study, which showed a significant slowing of microbial
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growth for hake, megrim and especially angler with the combination of the organic
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acids in the C-800 batch.
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4. Conclusions
299
The proposed C-800 organic acid-icing system evaluated in this work, which
300
contained AA, CA and LA, proved to be an effective mixture for fish preservation due
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to its antimicrobial effect. The microbial and sensory results obtained in this study
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allowed us to conclude that this novel icing system slowed microbial growth in hake,
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megrim and especially angler; these results agree with a significant extension of the fish
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shelf life. These results may be of practical interest for improving the preservation of
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fish quality not only on land but also for on-board strategies. Based on the results
306
presented in this work, a strategy for the on-board application of the C-800 organic
307
acid-icing system is currently being applied to these and other commercially relevant
308
fish species from the Grand Sole Bank. Moreover, this study provides a way to optimise
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the concentrations of CA, AA and LA for fish preservation purposes. This process is
310
currently under development in our laboratories, and the potential synergistic effects
13
311
among the organic acids and the diffusion of organic acids from the icing medium to the
312
fish muscle after melting are being considered.
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314
Acknowledgements
315
The authors thank the owner and crew of the CACHACHO and CHANS ships for
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their participation in the present study, and the Vigo harbour fishing fleet in general for
317
kindly providing us with the three fish species. The authors also thank Mr. Rodrigo
318
García Castro for his technical assistance and Atlantic One S.L. (Vigo, Spain) for
319
providing the commercial organic acid mixture BPS2. Mrs. Gabriela Medina is widely
320
acknowledged for her valuable preliminary studies of applying the BPS2 product to
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fish. This work was supported by the Secretaría Xeral de I+D from the Xunta de
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Galicia (Galicia, Spain) through the Research Project PGIDIT 08TAL038E.
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450
19
451
452
453
454
TABLE 1
Trimethylamine (TMA) assessment (mg TMA-N/100 g muscle)* in fish kept in different kinds of ice**
Chilling
Time (days)
1
455
456
457
Hake
Megrim
Angler
C-0
C-400
C-800
C-0
C-400
C-800
C-0
C-400
C-800
0.17
0.10
0.15
2.60
2.16
1.62
2.62 b
1.08 b
0.22 a
(0.05)
(0.03)
(0.03)
(0.35)
(0.20)
(0.77)
(1.48)
(0.57)
(0.18)
5
1.00 b
0.81 ab
0.29 a
5.89
5.98
7.48
4.80
6.15
2.72
(0.49)
(0.19)
(0.06)
(0.21)
(2.97)
(2.51)
(1.57)
(2.18)
(2.43)
8
6.18 b
3.76 ab
1.99 a
12.07
10.42
13.70
9.83 c
7.25 b
5.24 a
(2.46)
(1.17)
(0.36)
(3.85)
(3.97)
(0.93)
(1.37)
(0.91)
(1.84)
12
11.12 b
11.98 b
8.26 a
15.66 ab
12.04 a
18.19 b
11.76 b
8.48 ab
7.58 a
(1.50)
(1.30)
(1.09)
(4.19)
(3.79)
(1.61)
(1.95)
(1.27)
(1.73)
15
17.12 b
16.79 b
12.84 a
(2.18)
(1.56)
(1.04)
* Mean values (n=3) are indicated; standard deviations are expressed in parentheses. Starting TMA values: 0.11±0.02 (hake), 1.43±0.61
(megrim) and 1.87±1.56 (angler).
** Ice conditions: C-0, C-400 and C-800, as detailed in the Materials and methods section.
20
458
459
460
461
TABLE 2
Sensory acceptance* of hake stored under different icing conditions**
Attribute
Chilling Time (days)
Gills
C-0
C-400
C-800
5
B
A
A
8
B
B
B
12
C
B
B
15
C
C
C
Eyes
C-0
C-400
C-800
A
A
A
B
B
B
C
B
B
C
C
C
C-0
C-400
C-800
A
A
A
B
B
B
C
B
B
C
C
C
External
odour
462
463
464
465
Icing
condition
Muscle
C-0
A
B
C
C
odour and
C-400
A
B
C
C
taste
C-800
A
A
B
C
* Starting fish were category E for all attributes considered. Fish on day one were
category A for all attributes considered.
** Icing conditions are expressed as in Table 1.
21
466
467
468
469
TABLE 3
Sensory acceptance* of megrim stored under different icing conditions**
Attribute
Chilling Time (days)
Gills
C-0
C-400
C-800
1
A
A
A
5
B
A
A
8
C
B
B
12
C
C
C
Eyes
C-0
C-400
C-800
A
A
A
A
A
A
B
B
B
B
B
B
C-0
C-400
C-800
A
A
A
B
B
B
B
B
B
C
C
C
C
C
B
C
C
C
External
odour
470
471
472
Icing
condition
Muscle
C-0
A
B
odour and
C-400
A
A
taste
C-800
A
A
* Starting fish were category A for all attributes considered.
** Icing conditions are expressed as in Table 1.
22
473
474
475
476
TABLE 4
Sensory acceptance* of angler stored under different icing conditions**
Attribute
Chilling Time (days)
Gills
C-0
C-400
C-800
1
A
A
A
5
B
A
A
8
B
B
A
12
B
B
B
Eyes
C-0
C-400
C-800
A
A
A
A
A
A
A
A
A
B
B
B
C-0
C-400
C-800
A
A
A
A
A
A
B
B
A
C
C
B
B
B
A
C
C
B
External
odour
477
478
Icing
condition
Muscle
C-0
A
B
odour and
C-400
A
B
taste
C-800
A
A
* Starting fish were category A for all attributes considered.
** Icing conditions are expressed as in Table 1.
23
479
Figure Legends
480
481
Fig. 1. Evolution of mesophilic aerobes in hake muscle (Merluccius merluccius) for
482
chilled fish specimens stored under C-0, C-400 and C-800 conditions.
483
Fig. 2. Evolution of mesophilic aerobes in the muscle of chilled megrim
484
(Lepidorhombus whiffiagonis) specimens stored under C-0, C-400 and C-800
485
conditions.
486
Fig. 3. Evolution of mesophilic aerobes (A), psychrotrophs (B) and proteolytic bacteria
487
(C) in the muscle of chilled angler (Lophius piscatorius) specimens stored under C-0,
488
C-400 and C-800 conditions.
489
Fig. 4. Changes in the muscle pH of chilled megrim (A) and angler (B) specimens
490
stored under C-0, C-400 and C-800 conditions.
24
FIGURE 1
Mesophilic aerobic log CFU/g
9,00
8,00
7,00
6,00
5,00
4,00
treatment C
3,00
treatment 8
2,00
treatment 4
1,00
0,00
0
1
5
8
12
15
Storage time (days)
25
FIGURE 2
Mesophilic aerobic log CFU/g
9,00
8,00
7,00
6,00
5,00
4,00
treatment C
3,00
treatment 8
2,00
treatment 4
1,00
0,00
0
1
5
8
12
15
Storage time (days)
26
FIGURE 3
A
B
Mesophilic aerobic log CFU/g
9,00
8,00
7,00
C
6,00
5,00
4,00
treatment C
3,00
treatment 8
2,00
treatment 4
1,00
0,00
0
1
5
8
Storage time (days)
12
15
27
FIGURE 4
A
Mesophilic aerobic log CFU/g
9,00
8,00
7,00
B
6,00
5,00
4,00
treatment C
3,00
treatment 8
2,00
treatment 4
1,00
0,00
0
1
5
8
12
15
Storage time (days)
28