Optimization of R-Phycoerythrin Extraction by Ultrasound-Assisted Enzymatic Hydrolysis: A Comprehensive Study on the Wet Seaweed Grateloupia turuturu.

Enzyme-assisted extraction (EAE) and ultrasound-assisted extraction (UAE) are both recognized as sustainable processes, but little has been done on the combined process known as ultrasound-assisted enzymatic hydrolysis (UAEH), and even less on seaweed. The present study aimed to optimize the UAEH of the red seaweed Grateloupia turuturu for the extraction of R-phycoerythrin (R-PE) directly from the wet biomass by applying a response surface methodology based on a central composite design. Three parameters were studied: the power of ultrasound, the temperature and the flow rate in the experimental system. Data analysis demonstrated that only the temperature had a significant and negative effect on the R-PE extraction yield. Under the optimized conditions, the R-PE kinetic yield reached a plateau between 90 and 210 min, with a yield of 4.28 ± 0.09 mg·g-1 dry weight (dw) at 180 min, corresponding to a yield 2.3 times higher than with the conventional phosphate buffer extraction on freeze-dried G. turuturu. Furthermore, the increased release of R-PE, carbohydrates, carbon and nitrogen can be associated with the degradation of G. turuturu constitutive polysaccharides, as their average molecular weights had been divided by 2.2 in 210 min. Our results thus demonstrated that an optimized UAEH is an efficient method to extract R-PE from wet G. turuturu without the need for expensive pre-treatment steps found in the conventional extraction. UAEH represents a promising and sustainable approach that should be investigated on biomasses where the recovery of added-value compounds needs to be improved.

Effect of the Manufacturing Process on the Microbiota, Organoleptic Properties and Volatilome of Three Salmon-Based Products.

Lightly preserved seafood products, such as cold-smoked fish and fish gravlax, are traditionally consumed in Europe and are of considerable economic importance. This work aimed to compare three products that were obtained from the same batch of fish: cold-smoked salmon (CSS) stored under vacuum packaging (VP) or a modified atmosphere packaging (MAP) and VP salmon dill gravlax (SG). Classical microbiological analyses and 16S rRNA metabarcoding, biochemical analyses (trimethylamine, total volatile basic nitrogen (TVBN), biogenic amines, pH, volatile organic compounds (VOCs)) and sensory analyses (quantitative descriptive analysis) were performed on each product throughout their storage at a chilled temperature. The three products shared the same initial microbiota, which were mainly dominated by Photobacterium, Lactococcus and Lactobacillus genera. On day 28, the VP CSS ecosystem was mainly composed of Photobacterium and, to a lesser extent, Lactococcus and Lactobacillus genera, while Lactobacillus was dominant in the MAP CSS. The diversity was higher in the SG, which was mainly dominated by Enterobacteriaceae, Photobacterium, Lactobacillus and Lactococcus. Although the sensory spoilage was generally weak, gravlax was the most perishable product (slight increase in amine and acidic off-odors and flavors, fatty appearance, slight discoloration and drop in firmness), followed by the VP CSS, while the MAP CSS did not spoil. Spoilage was associated with an increase in the TVBN, biogenic amines and spoilage associated VOCs, such as decanal, nonanal, hexadecanal, benzaldehyde, benzeneacetaldehyde, ethanol, 3-methyl-1-butanol, 2,3-butanediol, 1-octen-3-ol, 2-butanone and 1-octen-3-one. This study showed that the processing and packaging conditions both had an effect on the microbial composition and the quality of the final product.

Use of random forest methodology to link aroma profiles to volatile compounds: Application to enzymatic hydrolysis of Atlantic salmon (Salmo salar) by-products combined with Maillard reactions.

To use salmon protein hydrolysates as food ingredients and to mask the fish odor, Maillard reactions were associated with enzymatic production of hydrolysates. The study explored an original approach based on regression trees (RT) and random forest (RF) methodologies to predict hydrolysate odor profiles from volatile compounds. An experimental design with four factors: enzyme/substrate ratio, quantity of xylose, hydrolysis and cooking times was used to create a range of enzymatic hydrolysates. Twenty samples were submitted to a trained panel for sensory descriptions of odor. Hydrolysate volatile compounds were extracted by means of Headspace Solid Phase MicroExtraction (HS-SPME) and analyzed using gas chromatography/mass spectrometry (GC-MS). The results showed that RT and RF methodologies can be useful tools for predicting an entire sensory profile from volatile compounds. Four main volatile compounds made it possible to separate hydrolysates into five groups according to their specific sensory profile. 2,5-dimethylpyrazine, 1-hydroxy-2-propanone and 3-hydroxy-2-pentanone were identified as the main predictors of the roasted odor, whereas methanethiol was associated with a mud odor. These results also suggest the appropriate process conditions for obtaining a typical roasted odor.

Salmon Gravlax Biopreservation With Lactic Acid Bacteria: A Polyphasic Approach to Assessing the Impact on Organoleptic Properties, Microbial Ecosystem and Volatilome Composition.

Seafood and fishery products are very perishable commodities with short shelf-lives owing to rapid deterioration of their organoleptic and microbiological quality. Microbial growth and activity are responsible for up to 25% of food losses in the fishery industry. In this context and to meet consumer demand for minimally processed food, developing mild preservation technologies such as biopreservation represents a major challenge. In this work, we studied the use of six lactic acid bacteria (LAB), previously selected for their properties as bioprotective agents, for salmon dill gravlax biopreservation. Naturally contaminated salmon dill gravlax slices, with a commercial shelf-life of 21 days, were purchased from a French industrial company and inoculated by spraying with the protective cultures (PCs) to reach an initial concentration of 106 log CFU/g. PC impact on gravlax microbial ecosystem (cultural and acultural methods), sensory properties (sensory profiling test), biochemical parameters (pH, TMA, TVBN, biogenic amines) and volatilome was followed for 25 days of storage at 8°C in vacuum packaging. PC antimicrobial activity was also assessed in situ against Listeria monocytogenes. This polyphasic approach underlined two scenarios depending on the protective strain. Carnobacterium maltaromaticum SF1944, Lactococcus piscium EU2229 and Leuconostoc gelidum EU2249, were very competitive in the product, dominated the microbial ecosystem, and displayed antimicrobial activity against the spoilage microbiota and L. monocytogenes. The strains also expressed their own sensory and volatilome signatures. However, of these three strains, C. maltaromaticum SF1944 did not induce strong spoilage and was the most efficient for L. monocytogenes growth control. By contrast, Vagococcus fluvialis CD264, Carnobacterium inhibens MIP2551 and Aerococcus viridans SF1044 were not competitive, did not express strong antimicrobial activity and produced only few organic volatile compounds (VOCs). However, V. fluvialis CD264 was the only strain to extend the sensory quality, even beyond 25 days. This study shows that C. maltaromaticum SF1944 and V. fluvialis CD264 both have a promising potential as bioprotective cultures to ensure salmon gravlax microbial safety and sensorial quality, respectively.

Effect of vacuum and modified atmosphere packaging on the microbiological, chemical and sensory properties of tropical red drum (Sciaenops ocellatus) fillets stored at 4°C.

AIMS: The effect of vacuum (VP - 4°C) and CO2/N2-atmosphere (MAP - 4°C) packaging on the quality of red drum fillets compared with whole gutted iced fish was investigated. METHODS AND RESULTS: A metagenomic approach, bacterial enumeration and isolation, biochemical and sensory analyses were carried out. The organoleptic rejection of whole fish was observed at day 15 whereas VP and MAP fillets appeared unacceptable only after 29days. At these dates, total mesophilic counts reached 107-108CFU g-1. According to Illumina MiSeq sequencing, Arthrobacter, Chryseobacterium, Brevibacterium, Staphylococcus and Kocuria were the main genera of the fresh red drum fillets. At the sensory rejection time, lactic acid bacteria (LAB), particularly Carnobacterium sp., dominated the microbiota of both types of packaging. The pH value of fresh samples was between 5.96 and 6.37 and did not vary greatly in all trials. Total volatile basic nitrogen (TVBN) and trimethylamine (TMA) concentrations were low and not represent reliable indicators of the spoilage, contrary to some biogenic amines (cadaverine, putrescine and tyramine). CONCLUSION: Chilled packed fillets of red drum have an extended shelf-life compared to whole gutted iced fish. Overall, few differences in sensory and microbial quality were observed between the VP and MAP samples. SIGNIFICANCE AND IMPACT OF THE STUDY: Next-Generation Sequencing (NGS) provided data on the microbiota of a tropical fish.

The effect of algae diets (Skeletonema costatum and Rhodomonas baltica) on the biochemical composition and sensory characteristics of Pacific cupped oysters (Crassostrea gigas) during land-based refinement.

Oyster refinement, a common practice in France, is aimed at increasing the weight of oyster tissue and influencing the taste properties of the refined oysters. Refinement usually takes place in land-based systems where the oysters are fed with relatively high concentrations of microalgae. In this study the impact of feeding Skeletonema costatum and Rhodomonas baltica on the biochemical composition and sensory characteristics of Pacific cupped oysters (Crassostrea gigas) from the Eastern Scheldt during land-based refinement was studied. After a feeding period of four and seven weeks market-sized oysters were sampled for the analysis of fatty acids, free amino acids and volatile organic compounds and for a sensory evaluation by consumers and an expert panel. The algae Skeletonema costatum showed a lower ∑PUFA, ∑n-3, ∑n-6, C18:2n6, C18:3n3, C18:4n3, C22:6n3 content as compared with Rhodomonas baltica. These differences were also reflected in the fatty acid profile of the oysters fed with the corresponding algae diets. Furthermore, general linear model and principal component analysis showed marked differences in free amino acids and volatile organic compound content between Skeletonema, Rhodomonas fed oysters and reference oysters. For example, threonine, glutamine, leucine, histidine, (E)-2-hexenal, (E)-2octenal, (E)-2-octen-1-ol, (E,E)-2,4-octadien-1-ol, (E,Z)-3,6-nonadien-1-ol and (Z,E)-2,6-nonadienal contents were higher in Skeletonema fed oysters compared to Rhodomonas fed oysters. Sensory differences between the experimental oyster groups were shown. Skeletonema fed Pacific cupped oysters were characterized by a stronger seaweed flavor, higher perceived sweetness and a firmer texture in comparison with Rhodomonas fed oysters. Naïve consumers were only able to differentiate between Rhodomonas fed oysters and reference oysters.

Improving simultaneously the quality and safety of cooked and peeled shrimp using a cocktail of bioprotective lactic acid bacteria.

Tropical shrimp is of considerable economic importance in the world but is highly perishable due to microbial and chemical degradation. Biopreservation is a food preservation technology based on the addition of "positive" bacteria able to kill or prevent the growth of undesirable microorganisms. Two strains of lactic acid bacteria (LAB) have previously been selected for a biopreservation strategy: Lactococcus piscium CNCM I-4031, for its ability to prevent the sensory deterioration of seafood and Carnobacterium divergens V41, which inhibits growth of Listeria monocytogenes. The objective was to test the association of the two strains to improve both the quality and safety of shrimp. In a first trial, the two LAB were inoculated alone or in a cocktail in cooked and peeled shrimp (CPS) Penaeus vannamei at 5×105CFU/g. Chemical, sensory and microbiological analyses by culture-dependent and -independent methods were performed during storage under modified atmosphere packaging (MAP) at 8°C. The results were compared to a non-inoculated batch. In a second trial, the same experiments were repeated in the presence of 102CFU/g of L. monocytogenes RF191. The microbiota of CPS was composed of LAB, Shewanella spp. and Enterobacteriaceae. Brochothrix thermosphacta was not detected. L. piscium and C. divergens reached 108 and 109CFU/g, respectively, in 7days and did not inhibit each other when co-inoculated. L. piscium reduced L. monocytogenes by 1Log (CFU/g) for 28days. C. divergens had an immediate listericidal effect lasting 7days. A regrowth of L. monocytogenes was then observed but the count was always 2 to 5Log (CFU/g) lower than in the control. No additional or synergic effect between protective strains was observed and the cocktail had the same inhibitory effect as C. divergens alone. C. divergens was very effective at preventing the sensory deterioration of CPS. This may be related to the inhibition of Shewanella and Enterobacteriaceae. However, the panelists could detect the presence of C. divergens during the first 10days of storage, with slight unpleasant odors and flavors. L. piscium improved the sensory quality of CPS for 14days only. In co-culture, L. piscium eliminated the off-odors and flavors released by C. divergens in the early stage of storage and the co-culture allowed maintaining a good quality of CPS throughout the storage. Therefore, the use of a cocktail of L. piscium CNCM I-4031 and C. divergens V41 is recommended in a strategy of biopreservation of shrimp.

Selection of bioprotective cultures for preventing cold-smoked salmon spoilage.

Biopreservation is a natural technology of food preservation, which consists of inoculating food with microorganisms selected for their antibacterial properties. The objective of this study was to select lactic acid bacteria (LAB) to improve the quality of cold-smoked salmon (CSS). In this work, different strains representative of the 4 dominant species, identified in a previous study by pyrosequencing the 16S rRNA gene, were isolated and their spoiling potential in CSS blocks, sterilized by ionization, was assessed by twelve trained panelists along the vacuum storage at 8°C. Photobacterium phosphoreum, Brochothrix thermosphacta and Serratia proteamaculans released strong off-odors whereas the spoiling potential of Carnobacterium divergens was weaker. The spoiling capacity of Lactococcus piscium EU2241, Leuconostoc gelidum EU2247, Lactobacillus sakei EU2885, Staphylococcus equorum S030674 and 4 commercial starters was tested by the same method and 2 strains were eliminated due to off-odor production. The effect of the 6 selected LAB against the 4 specific spoiling organisms (SSOs) selected was tested by challenge tests in sterile CSS blocks. The protective effect of the LAB differed from one SSO to another and no correlation could be established between the sensory improvement, SSO inhibition, and the implantation or acidification of protective cultures (PCs). All the PCs except L. piscium reduced the off-odors released by P. phosphoreum although some of them had no effect on its growth. S. equorum, which did not grow in CSS, favored the implantation of P. phosphoreum but prevented its off-odor formation. L. piscium was the only strain that prevented the spoilage of B. thermosphacta and S. proteamaculans although it did not grow very well and did not acidify the product. L. gelidum EU2247 inhibited the growth of these 2 SSOs and lowered the pH but had no effect on the sensory quality. Finally, L. piscium was tested in 2 naturally contaminated products, with a positive effect on 1 batch. This effect was not correlated with the microbial ecosystem as determined by acultural and cultural techniques. Based on these results, the selection strategy is discussed.

Evaluation of the spoilage potential of bacteria isolated from spoiled cooked whole tropical shrimp (Penaeus vannamei) stored under modified atmosphere packaging.

The spoilage potential of isolates belonging to five bacterial groups/species (Shewanella baltica, Carnobacterium maltaromaticum, Aeromonas salmonicida, Vibrio sp., "other Gamma-Proteobacteria" [containing one strain of Pseudoalteromonas sp. and one strain of Psychrobacter sp.]) isolated from spoiled cooked and whole tropical shrimp stored under modified atmosphere packaging (MAP) was evaluated by inoculation into ionized cooked and peeled tropical shrimp followed by storage for 32 days at 8 °C. Microbial growth and sensory changes were monitored during the storage period. The major spoilage bacterial isolate groups were C. maltaromaticum and S. baltica. In order to characterize their spoilage potential further and to study the effect of their interactions, each of these two specific spoilage organisms (SSO) and one mixed-culture, C. maltaromaticum/S. baltica, were tested using a combination of complementary methods: molecular (PCR-TTGE), sensory, chemical, and conventional microbiological analyses. It was concluded that, in the mixed-culture-inoculated samples, both species groups imposed their spoilage characteristics.

Evaluation of the spoilage potential of bacteria isolated from spoiled raw salmon (Salmo salar) fillets stored under modified atmosphere packaging.

The spoilage potential of eight bacterial groups/species (Serratia spp., Hafnia alvei, Brochothrix thermosphacta, Carnobacterium maltaromaticum, Shewanella baltica, Lactococcus piscium, Photobacterium phosphoreum, "other Enterobacteriaceae" [containing one strain of Moellerella sp., Morganella sp. and Pectobacterium sp.]) isolated from spoiled raw salmon fillets stored under modified atmosphere packaging (MAP) was evaluated by inoculation into sterile raw salmon cubes followed by storage for 12days at 8°C. Microbial growth and sensory changes were monitored during the storage period. The dominant spoilage bacteria were C. maltaromaticum, H. alvei and P. phosphoreum. In order to further characterize their spoilage potential and to study the effect of their interactions, each of these 3 specific spoilage organisms (SSO) and two mixed-cultures, C. maltaromaticum/H. alvei and C. maltaromaticum/P. phosphoreum were tested in the sterile salmon model system using a combination of complementary methods: molecular (PCR-TTGE), sensory, chemical and conventional microbiological analyses. It was concluded that, in the mixed-culture inoculated samples, the dominant species determined the spoilage characteristics. The volatile fraction of P. phosphoreum inoculated samples was analyzed by solid-phase microextraction (SPME) followed by gas chromatography coupled to mass spectrometry (GC-MS). Among the specific volatile compounds present on P. phosphoreum spoiled inoculated samples, acetic acid was correlated with sensory analysis and can be proposed as a raw salmon spoilage marker.

Perception of oyster-based products by French consumers. The effect of processing and role of social representations.

The search for new markets in the seafood sector, associated with the question of the continuity of raw oyster consumption over generations can be an opportunity for processors to extend their ranges with oyster-based products. The twofold aim of this study was to evaluate the impact of processing and social representation on perception of oyster-based products by French consumers and to identify the best means of development in order to avoid possible failure in the market. Five products with different degrees of processing (cooked oysters in a half-shell, hot preparation for toast, potted oyster, oyster butter and oyster-based soup) were presented within focus groups and consumer tests, at home and in canteens with the staff of several companies in order to reach consumers with different ages and professional activities. The results showed that social representation had a strong impact and that behaviours were contrasted according to the initial profile of the consumer (traditional raw oyster consumers or non-consumers) and their age distribution (younger and older people). The degree of processing has to be adapted to each segment. It is suggested to develop early exposure to influence the food choices and preferences of the youngest consumers on a long-term basis.

Characterisation of the spoilage microbiota in raw salmon (Salmo salar) steaks stored under vacuum or modified atmosphere packaging combining conventional methods and PCR-TTGE.

In order to characterise the spoilage related to microbiota of raw salmon, a combination of culture-dependent and -independent methods, including PCR-TTGE, was used to analyse 3 raw salmon batches stored for 3 days at chilled temperature in modified atmosphere packaging (MAP) (50% CO2/50% N2) or under vacuum. Sensory evaluation, microbiological enumeration and chemical analysis were performed after 3, 7 and 10 days of storage. At the onset of spoilage, 65 bacterial isolates were picked from the plates. Thus, 13 different genera or species were identified by phenotypic and molecular tests: Serratia spp., Photobacterium phosphoreum, Yersinia intermedia, Hafnia alvei, Buttiauxella gaviniae, Pseudomonas sp., Carnobacterium maltaromaticum, Carnobacterium divergens, Lactococcus piscium, Lactobacillus fuchuensis, Vagococcus carniphilus, Leuconostoc gasicomitatum and Brochothrix thermosphacta. The PCR-TTGE profiles and band identification enabled a shift of the dominant populations during the storage to be visualised for all the batches, probably due to the temperature change and the packaging. At the beginning of storage, Pseudomonas sp. dominated the raw salmon microbiota while in the following days (7 and 10), P. phosphoreum and L. piscium were identified as the main bacterial groups. This study enhances the knowledge of MAP and vacuum-packed raw salmon spoilage microbiota.

Sensory characteristics of spoilage and volatile compounds associated with bacteria isolated from cooked and peeled tropical shrimps using SPME-GC-MS analysis.

The spoilage potential of six bacterial species isolated from cooked and peeled tropical shrimps (Brochothrix thermosphacta, Serratia liquefaciens-like, Carnobacterium maltaromaticum, Carnobacterium divergens, Carnobacterium alterfunditum-like and Vagococcus penaei sp. nov.) was evaluated. The bacteria were inoculated into shrimps, packaged in a modified atmosphere and stored for 27 days at 8 °C. Twice a week, microbial growth, as well as chemical and sensory changes, were monitored during the storage period. The bacteria mainly involved in shrimp spoilage were B. thermosphacta, S. liquefaciens-like and C. maltaromaticum whose main characteristic odours were cheese-sour, cabbage-amine and cheese-sour-butter, respectively. The volatile fraction of the inoculated shrimp samples was analysed by solid-phase microextraction (SPME) and gas chromatography coupled to mass spectrometry (GC-MS). This method showed that the characteristic odours were most likely induced by the production of volatile compounds such as 3-methyl-1-butanal, 2,3-butanedione, 2-methyl-1-butanal, 2,3-heptanedione and trimethylamine.

Effect of bacterial interactions on the spoilage of cold-smoked salmon.

Cold-smoked salmon is a lightly preserved fish product in which a mixed microbial flora develops during storage and where the interactive behaviour of micro-organisms may contribute to their growth and spoilage activity. The aim of this study was to assess the effect of the bacterial interactions between the main species contaminating the cold-smoked salmon on bacterial growth, chemical and sensory changes, and spoilage. First, Carnobacterium piscicola, Photobacterium phosphoreum, Lactobacillus sakei, Vibrio sp., Brochothrix thermosphacta and Serratia liquefaciens-like were inoculated as pure cultures on sterile cold-smoked salmon. All bacterial species grew well; Vibrio sp. was the fastest and L. sakei strains developed very rapidly as well with a high maximum cell density on cold-smoked salmon blocks (up to 10(9) cfu g(-1) after 10 days at 8 degrees C). Based on sensory analysis, Vibrio sp. was identified as non-spoilage bacteria, C. piscicola as very lightly and B. thermosphacta as lightly spoiling. L. sakei and S. liquefaciens-like were found to be the most spoiling bacteria. Secondly, C. piscicola and L. sakei, two species frequently occurring in the lactic flora of the product, were inoculated together and each of them in mixed cultures with respectively P. phosphoreum, Vibrio sp., B. thermosphacta, and S. liquefaciens-like. The growth of L. sakei was shown to strongly inhibit most of the co-inoculated strains i.e. P. phosphoreum, B. thermosphacta, S. liquefaciens-like and, to a lesser extent, Vibrio sp. The growth of C. piscicola seemed to be enhanced with B. thermosphacta and to develop earlier with P. phosphoreum and Vibrio sp. Conversely, S. liquefaciens-like and P. phosphoreum were weakly inhibited by C. piscicola. The main observation resulting from the sensory evaluation was the delay in the appearance of the spoilage characteristics in the mixed cultures with L. sakei, in particular L. sakei/ S. liquefaciens-like. On the other hand, the spoilage activity of the non-spoiler strains Vibrio sp. or the moderate spoilage strains B. thermosphacta and C. piscicola was increased when they were associated together. It is concluded that the spoilage behaviour of micro-organisms in mixed culture is significantly different from pure culture and explain the difficulty to find robust quality indices for this product.