Rogério Mendes

Development of a sodium dodecyl sulfate-polyacrylamide gel electrophoresis reference method for the analysis and identification of fish species in raw and heat-processed samples: a collaborative study.

A collaborative study was carried out in seven European labs with the aim of achieving a sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) standard operation procedure to identify fish species in raw and cooked samples. Urea and SDS‐containing solutions were evaluated as extractants. Several preelectrophoretic operations — such as treatment with RNase/DNase, ultrafiltration and desalting — and up to ten types of gels and three SDS‐PAGE systems were considered. The SDS‐containing solution allowed a higher protein extractability than urea. Unlike urea extraction, SDS extraction seemed not to be influenced so much by the state of the sample (raw, cooked at 60oC, cooked at 85oC). Desalting, ultrafiltration or treatment with RNase/DNase did not improve the discriminatory power of the protein patterns. Commercial homogeneous 15% ExcelGels, especially when they were silver stained, yielded good results and afforded higher reproducibility, thus allowing a better matching of results among the laboratories participating in this collaborative study. Under the optimized technical conditions described above, all the fish species tested, either raw and cooked, yielded reproducible and discriminant species‐specific protein patterns.

Quality Changes During Storage of Fish Sausages Containing Dietary Fiber

ABSTRACT Two groups of fish sausages were studied regarding quality changes during an 80-day storage experiment at 2°C. Formulations were identical with the exception of pork fat, 7.8%(w/w) in the control group, replaced by 5.2% (w/w) chicory root inulin and 2.6% (w/w) extra hake mince in the other. Control sausages were more elastic, cohesive, springy, and succulent, while substituted sausages presented higher gel strength and hardness. Both sausages presented an acceptable quality not only immediately after their production but also during storage at 2 ± 1°C. However, gel strength as well as textural and sensory hardness increased, and succulence showed a clear reduction during storage time. Moreover, for substituted sausages, saltiness and astringent aftertaste increased. Formulation had no effect on the microbial growth, which was low. However, microbiological changes occurred during storage time: growth of anaerobic spore-formers since the 45th and 43rd day for control and substituted sausages, respectively.

Influence of variation in methodology on the reliability of the isoelectric focusing method of fish species identification

Abstract The reliability of isoelectric focusing (IEF) of sarcoplasmic proteins for fish species identification was evaluated by a collaborative study among eight European laboratories. Each laboratory used its own method of IEF to identify 10 unknown samples of raw muscle by means of reference material. In 93% of cases the assignment between sample and reference was correct. In a second study, the influence of extradant (water, low ionic strength buffer, or detergent) and the position of sample application on the protein pattern was examined. Working with light muscle of rainbow trout (Oncorhynchus mykiss), it was found that the type of extractant did not influence the protein pattern. Comparison of the patterns of samples, which had been applied near the anode, in the middle, or near the cathode, revealed differences in the number and position of the protein bands under the experimental conditions applied by most laboratories. This effect was not observed with the Phast System.

Species identification of cooked fish by urea isoelectric focusing and sodium dodecylsulfate polyacrylamide gel electrophoresis: a collaborative study

The suitability and reliability of urea IEF and SDS-PAGE for the identification of cooked fish flesh was tested by a collaborative study among nine laboratories. Urea IEF was performed with CleanGels as well as with ImmobilineGels, and ExcelGels were used for SDS-PAGE, enabling all three types of gels to be run in the same flat bed electrophoresis chamber. By strictly following optimised standard operation procedures (SOPs), five unknown cooked samples had to be identified with each technique using a set of 10 raw reference samples. With urea IEF, only one out of 35 identifications was incorrect, and with SDS-PAGE a similar result was obtained. It was concluded that methods, as now developed, are suitable for checking the species declaration of fishery products.

Species identification of formed fishery products and high pressure-treated fish by electrophoresis: a collaborative study

The suitability and reliability of three electrophoretic methods of fish species identification, urea isoelectric focusing (IEF), sodium dodecyl sulfate poly-acrylamide gel electrophoresis (SDS-PAGE) and native IEF, were evaluated on formed fish fillets and high pressure fish flesh by a collaborative study among four institutes. By following optimized standard operation procedures, the protein patterns of processed fish were compared to patterns of raw reference samples. The method to use depended of the effect of processing on the protein pattern. The proteins obtained from formed products were not denatured and therefore any of the three methods proved to be adequate, with a preference for native IEF which had a better discriminatory power for the species used. The high pressure process altered the proteins, and so only urea IEF and SDS-PAGE methods could be used. For these products, the chosen method should then be the one with the better discriminating power for the species being examined.

Low mislabeling rates indicate marked improvements in European seafood market operations

Over the span of a decade, genetic identification methods have progressively exposed the inadequacies of the seafood supply chain, revealing previously unrecognized levels of seafood fraud, raising awareness among the public, and serving as a warning to industry that malpractice will be detected. Here we present the outcome of the latest and largest multi-species, transnational survey of fish labeling accuracy to date, which demonstrates an apparent sudden reduction of seafood mislabeling in Europe. We argue that recent efforts in legislation, governance, and outreach have had a positive impact on industry regulation. Coordinated, technology-based, policy-oriented actions can play a pivotal role in shaping a transparent, sustainable global seafood market and in bolstering healthier oceans.

Changes of Enzymes Activity and Protein Profiles Caused by High-Pressure Processing in Sea Bass (Dicentrarchus labrax) Fillets

High-pressure processing (HPP) is a technology of growing interest for food preservation, due to its ability to control the activity of degradative enzymes. The effect of three variables (pressure levels of 100, 250, and 400 MPa; pressure holding times of 0, 5, 15, and 30 min; and pressurization rates of 8 and 14 MPa/s) on the activity of the enzymes acid phosphatase, cathepsins (B and D), lipase, and calpains was studied using sea bass fillets as a case study model. Additionally, the effect of HPP on sarcoplasmic proteins was studied by SDS-PAGE and isoelectric focusing electrophoreses. The increase in pressure level and holding time decreased the protein concentration in sarcoplasmic extracts, and also the activity of calpains. As compared to nontreated samples, acid phosphatase activity was lower at 400 MPa, and for cathepsin D lower activities were observed at 100 and 400 MPa. The increase in pressurization rate increased the activity of cathepsin D, lipase, and calpains, although it was not always significant. In contrast, cathepsin B and lipase activities were less affected by HPP treatments. Electrophoresis separation of sarcoplasmic proteins showed that the intensity of many protein bands changed mainly due to pressure level and holding time. The results of this study suggest that HPP causes lysosomes disruption and also denaturation, aggregation, and fragmentation of sarcoplasmic proteins, and this evidence might be related to the decrease in enzymes activity especially at 400 MPa. In conclusion, HPP can be a potential tool to control the activity of degradative enzymes, which might prevent the softening of sea bass muscle due to autolytic reactions.

Effect of Oxygen Absorber on the Shelf Life of Gilthead Seabream (Sparus aurata)

ABSTRACT The effectiveness of oxygen absorber on the shelf life of scaled and gutted seabream packed in air during 10 days chilled storage (∼ 5°C) was studied. The quality changes were evaluated by sensory, microbial and chemical methods. Absorbers reduced oxygen concentration by 54% within the first two days and levels attained 0.1% after 6 days of storage. Significant differences in K values were not observed between packed batches with and without O2 absorber. Positive effects of O2 absorber on the reduction of lipid oxidation and sensory attributes, mainly on the appearance, were observed.

Quality characteristics of high pressure-induced hake (Merluccius capensis) protein gels with and without MTGase.

The effects of microbial transglutaminase (MTGase; 0.5%, w/w) and high hydrostatic pressure (HHP) on the quality of protein gels from unwashed mince of hake (Merluccius capensis) trimmings were studied. MTGase incorporation improved texture. Protein solubility was lower for those gels containing MTGase, as a result of myosin heavy chain cross-linking. HHP improved texture. Pressure level was the most important HHP parameter, since higher levels (300 vs. 100 MPa) augmented gel strength (GS). A positive synergistic effect of MTGase and HHP was found for some properties, such as GS, yielding improved gels from a raw material that otherwise shows poor gelation.

Quality Changes During Storage of Minced Fish Products Containing Dietary Fiber and Fortified with ω3 Fatty Acids

Two ready-to-eat minced fish products from hake were developed, their proximate composition and fatty acid profiles determined and their quality changes followed during 3.5 months under refrigeration at 2 ± 1 °C and 10 ± 1 °C. These products contain dietary fiber and are innovative and healthy. The formulation was identical, except vegetable oil (VO), 5.6% (w/w) in one group and 2.7% (w/w) plus 2.9% (w/w) cod liver oil (CLO) in the other. CLO products had a higher ω3/ω6 ratio (0.54 ± 0.02 versus 0.08 ± 0.02) and ensured, per 100 g serving, the 500 mg recommended daily intake of eicosapentaenoic and docosahexaenoic acids. CLO products showed lower gel strength (p ≤ 0.05), however, other textural properties were similar to those of the VO group. Thiobarbituric acid reactive substances values were higher in CLO products. All groups presented acceptable sensory scores and no microbiological growth. During storage products became redder and less yellow, while seafood aroma and flavor declined and saltiness perception augmented. Temperature had a negative effect on sensory elasticity and instrumental texture.