Laurent Vandanjon

Impact of ultrafiltration and nanofiltration of an industrial fish protein hydrolysate on its bioactive properties

BACKGROUND Numerous studies have demonstrated that in vitro controlled enzymatic hydrolysis of fish and shellfish proteins leads to bioactive peptides. Ultrafiltration (UF) and/or nanofiltration (NF) can be used to refine hydrolysates and also to fractionate them in order to obtain a peptide population enriched in selected sizes. This study was designed to highlight the impact of controlled UF and NF on the stability of biological activities of an industrial fish protein hydrolysate (FPH) and to understand whether fractionation could improve its content in bioactive peptides. RESULTS The starting fish protein hydrolysate exhibited a balanced amino acid composition, a reproducible molecular weight (MW) profile, and a low sodium chloride content, allowing the study of its biological activity. Successive fractionation on UF and NF membranes allowed concentration of peptides of selected sizes, without, however, carrying out sharp separations, some MW classes being found in several fractions. Peptides containing Pro, Hyp, Asp and Glu were concentrated in the UF and NF retentates compared to the unfractionated hydrolysate and UF permeate, respectively. Gastrin/cholecystokinin-like peptides were present in the starting FPH, UF and NF fractions, but fractionation did not increase their concentration. In contrast, quantification of calcitonin gene-related peptide (CGRP)-like peptides demonstrated an increase in CGRP-like activities in the UF permeate, relative to the starting FPH. The starting hydrolysate also showed a potent antioxidant and radical scavenging activity, and a moderate angiotensin-converting enzyme (ACE)-1 inhibitory activity, which were not increased by UF and NF fractionation. CONCLUSION Fractionation of an FPH using membrane separation, with a molecular weight cut-off adapted to the peptide composition, may provide an effective means to concentrate CGRP-like peptides and peptides enriched in selected amino acids. The peptide size distribution observed after UF and NF fractionation demonstrates that it is misleading to characterize the fractions obtained by membrane filtration according to the MW cut-off of the membrane only, as is currently done in the literature.

Concentration and desalination of fish gelatin by ultrafiltration and continuous diafiltration processes

Abstract Ultrafiltration (UF) of gelatin liquor from marine source (tuna skins) has been performed on a tubular mineral membrane of cut-off 10,000 Da. An attempt was made to concentrate the fish gelatin as well as to purify it by means of the diafiltration technique (DF). Primary experiments were devoted to optimise the UF performances as a function of the main adjustable parameters. The influence of pH, transmembrane pressure, recirculating velocity, and working temperature on both volumetric permeate fluxes and protein retention rates was successively examined. Under the appropriate operating conditions, it was shown that the used membrane can achieve the initial concentration step of the gelatin solutions up to 12 wt% with a protein yield as high as 0.98. In addition, continuous DF proved useful to reduce the salt content of the solutions at an average desalting speed of 185 g/h.m2 with a few loss of protein in the permeate side.

Recovery by nanofiltration and reverse osmosis of marine flavours from seafood cooking waters

Abstract Cooking waters from buckies, shrimps and tuna have a high level of polluting load (Chemical Oxygen Demand — COD — comprised between 5 and 40 gO2/L) and have to be treated before being rejected in the environment. However, these juices seem to contain interesting flavour compounds. This recovery would allow the industrialists to diminish the waste water treatment cost and to recover high added value molecules. In this work, a membrane process system consisting in ultrafiltration (UF) followed by uanofiltration (NF) or reverse osmosis (RO) was used to reduce the pollution load and to concentrate flavour compounds of seafood cooking juices (buckies, shrimps and tuna). The first ultrafiltration step (cut-off: 0.02 μm) consisted in clarifying the cooking waters in order to increase fluxes of the following filtrations. NF (cut-off: 300 Da) did not seem to be efficient enough for flavour recovery, whereas RO was, for shrimps and buckies cooking juices. At the opposite, aromas contained in the tuna cooking juice were not perfectly retained by the RO membrane used. Nevertheless, the polluting load was considerably decreased after RO (COD reduced by 95% for shrimps and buckies and 85% for tuna).

Assessment of the spatial variability of phenolic contents and associated bioactivities in the invasive alga Sargassum muticum sampled along its European range from Norway to Portugal

Sargassum muticum, an invasive brown macroalga presently distributed along European Atlantic coasts from southern Portugal to the south coast of Norway, was studied on a large geographical scale for its production of phenolic compounds with potential industrial applications and their chemical and biological activities. S. muticum can produce high biomass in Europe, which could be exploited to supply such compounds. S. muticum was collected in Portugal, Spain, France, Ireland and Norway (three sites/country) to examine the effect of the latitudinal cline and related environmental factors. Assays focused particularly on polyphenols and their activities. Crude acetone–water extracts were purified using solid phase extraction (SPE) and antioxidant and antimicrobial activities of crude extracts and semi-purified fractions measured. Total phenolic content was assessed by colorimetric Folin–Ciocalteu assay and reactive oxygen species activities by 2,2-diphenyl-1-picrylhydrazyl, reducing power, β-carotene bleaching method and xanthine oxidase assay. Antibacterial activities were tested on terrestrial and marine strains to evaluate potential use in biomedical and aquaculture fields. Purified active phlorotannins, isolated by SPE, were identified using NMR. Phenolic contents differ clearly among countries and among sites within countries. Quality did not change between countries, however, although there were some slight differences in phlorethol type. Additionally, some fractions, especially from the extreme north and south, were very active. We discuss this in relation to environmental conditions and the interest of these compounds. S. muticum represents a potential natural source of bioactive compounds and its collection could offer an interesting opportunity for the future management of this species in Europe.

Application of response surface methodology to optimise the antioxidant activity of a saithe (Pollachius virens) hydrolysate.

The objective of this study was to produce, by an enzymatic hydrolysis process at a pilot scale, a saithe (Pollachius virens) hydrolysate with a high antioxidant activity. Design of experiment methodology, based on laboratory-scale experiments, was used to obtain a behavioral reduced model that allows one to determine the optimal operating conditions maximizing the antioxidant activity. Two specifications were studied: the degree of hydrolysis and the antioxidant activity. The effects of the following hydrolysis parameters (temperature, pH, enzyme concentration, and operating time) were studied and presented as response surfaces. From these results, a multifactorial optimization was performed and the Pareto optimal set of efficient solutions was evaluated. The optimal conditions were tested at laboratory scale and then validated by comparison with tests carried out on a pilot plant.

Rapid geographical differentiation of the European spread brown macroalga Sargassum muticum using HRMAS NMR and Fourier-Transform Infrared spectroscopy

Two recent techniques based on chemical footprinting analysis, HRMAS NMR and FTIR spectroscopy, were tested on a brown macroalgal model. These powerful and easily-to-use techniques allowed us to discriminate Sargassum muticum specimens collected in five different countries along Atlantic coasts, from Portugal to Norway. HRMAS NMR and FTIR permitted the obtaining of an overview of metabolites produced by the alga. Based on spectra analysis, results allowed us to successfully group the samples according to their geographical origin. HRMAS NMR and FTIR spectroscopy respectively point out the relation between the geographical localization and the chemical composition and demonstrated macromolecules variations regarding to environmental stress. Then, our results are discussed in regard of the powerful of these techniques together with the variability of the main molecules produced by Sargassum muticum along the Atlantic coasts.

Chemical characterization and photoprotective activity measurement of extracts from the red macroalga Solieria chordalis

Abstract The photoadaptive responses of macroalgal communities to abiotic stresses have been studied, and a number of UV-absorbing molecules have been identified. Among these compounds, photoprotective compounds such as mycosporine-like amino acids and carotenoids have been isolated from various red macroalgal species. However, several substances still need to be characterized. We describe the preparation of photoprotective extracts obtained from Solieria chordalis. Two solvents, 2-octyl dodecanol and octyldodecyl ester of l-pyrrolidone carboxylic acid, were selected based on their cosmetic functions for performing an ultrasound-assisted extraction. The efficiency of extraction was monitored by spectrophotometry and in vitro photoprotective activity measurements. 2-Octyl dodecanol and octyldodecyl ester of l-pyrrolidone carboxylic acid extracts showed maximum absorption wavelengths ranging from 280 to 340 nm and 270 to 350 nm, respectively. The anti UV-B capacity for protecting a synthetic chlorophyll solution was assessed by measuring its pseudo first-order degradation kinetics at room temperature. Under irradiation at 312 nm, chlorophyll introduced in the 2-octyl dodecanol S. chordalis extract showed the slowest degradation kinetics with a half-life t1/2 of 121.0 min. Several compounds were detected in the seaweed extract by high-performance liquid chromatography. Among them, the mycosporine-like amino acid, palythenic acid, was detected in the algal extract.

Seasonal Variation of Sargassum Muticum Biochemical Composition Determined by Fourier Transform Infra-Red Spectroscopy

Proliferation of the introduced brown macro alga Sargassum muticum is known as a natural and hard to control phenomenon occurring along the Atlantic coasts. The phenomenon causes serious troubles for local ecosystems including the alteration of ecosystem structure, the reduction in indigenous biodiversity and economic losses (tourism, aquaculture). However, despite the serious troubles caused by S. muticum, this species contains highly remarkable bioactive metabolites. This macro alga is at present under-exploited and the valorization of its metabolites to give a positive value to this seaweed could be a solution of ecosystemic service. Biorefinery process could be one solution to valorize S. muticum. Comprehensive knowledge concerning the biochemical composition of S. muticum and the impact of environmental factors, particularly seasons, on its composition is a prerequisite before its valorization. In this study, the biochemical composition of S. muticum was evaluated by using classic colorimetric methods based on chemical analysis and Fourier Transform Infra-Red spectrometry (FTIR), and was used as a rapid and safe method that could bring advantages in screening studies and a more comprehensive management and use of seaweed products. Our results are globally in accordance, notably for phenolic compounds, showing the relevance of the use of infrared spectrometry. Moreover, based on the absorption bands of some specific and valuable compounds shown by FTIR, there was a seasonal variation in the polysaccharides, i.e., uronic acids and sulphated compounds, together with phenolic contents of S. muticum.

Valorization of the Macroalgae Sargassum Muticum by Enzymatic Hydrolysis, Interest of Surfactants to Improve the Extraction of Phlorotannins and Polysaccharides

The use of surfactants to improve enzymatic hydrolysis of the macroalgae Sargassum muticum has been investigated. Visible absorption spectroscopy has been used to quantify the solubilization of both polysaccharides and phlorotannins in the hydrolysates.   After total extraction, results showed that Sargassum muticum contained 2.74% (expressed in percent of the dry weight of the algae) of phlorotannins whose 32 % were in the cell wall. This result shows that it is important to access to the parietal phlorotannins. To reach this objective, we chose the enzymatic approach for destructurating the cell wall of the algae. The use of 5% dry weight (DW - 5% by weight of hydrolyzed algae) of an enzymatic mix containing a commercial beta-glucanase, a commercial protease and an alginate lyase extracted from Pseudomonas alginovora led after 3 hours of hydrolysis to the solubilization of 2.43% DW polysaccharides and 0.52% DW phlorotannins. The use of 0.5% volume of the surfactant Triton® X-100 with 10% DW of the enzymatic mix has allowed to reaching the value of 2.63% DW of solubilized phlorotannins, that is 96% of the total phenolic content.   The use of non-ionic surfactant, combined to enzymatic hydrolysis, showed an increased efficiency in disrupting cell wall and solubilizing phlorotannins in Sargassum muticum.