Compared to other analytical methods, matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) presents several unique advantages for the structural characterization of degradation products of carbohydrates. Our final goal is to implement this technique as a high-throughput platform, with the aim of exploring natural bio-diversity to discover new carbohydrate depolymerizing enzymes. In this approach, a variety of carbohydrates will be used as enzymes substrates and MALDI-MS will be employed to monitor the oligosaccharides produced. One drawback of MALDI, however, is that the choice of the matrix is largely dependent on the chemical properties of the analyte. In this context, our objective in the present work was to find the smallest set of MALDI matrices able to detect chemically heterogeneous oligosaccharides. This was done through the performance evaluation of more than 40 MALDI matrices preparations. Homogeneity of analyte-matrix deposits was considered as a critical feature, especially since the final objective is to fully automate the analyses. Evaluation of the matrices was done by means of a rigorous statistical approach. Amongst all tested compounds, our work proposes the use of the DHB/DMA ionic matrix as the most generic matrix, for rapid detection of a variety of polysaccharides including neutral, anionic, methylated, sulfated, and acetylated compounds. The selected matrices were then used to screen crude bacterial incubation media for the detection of enzymatic degradation products.
Tisochrysis lutea (T. lutea, ex T. Isochrysis galbana or T-Iso) is a marine haptophyte that was first isolated from Tahiti seawater. Because of its high content in lipids, this tropical species is commonly used in aquaculture to feed fishes, crustaceans and molluscs larvae. It is also a rich source of fucoxanthin with a high potential for nutraceutical, cosmetic and pharmaceutical applications. The purpose of the present study was to detail the pigment composition of T. lutea and to develop an efficient process to recover highly purified fucoxanthin. Using ultra performant liquid chromatography coupled to diode arrays and high-resolution mass spectrometry detectors (UPLC-DAD-MS/MS), we demonstrated for the first time the presence of echinenone, 3-hydroxy-echinenone and chlorophyll c2-monogalactosyldiacylglycerol [18:4/14:0] in unstressed cultures of T. lutea. The chemotaxonomic relevance of this updated pigment composition was discussed in relation to the Haptophyta phylum. A two-step purification of fucoxanthin was then optimized using centrifugal partition chromatography coupled to flash chromatography. This process allowed the efficient isolation of fucoxanthin (purity > 99%), that was further assessed as a lowtoxicity antineoplastic and chemosensitizing natural product in human chemoresistant melanoma cells. This carotenoid exerted an antiproliferative activity in A2058 melanoma cells and reversed in vitro their chemoresistance to dacarbazine, a DNA-alkylating agent clinically used for the treatment of metastatic melanoma.Please note that this is an author-produced PDF of an article accepted for publication following peer review. The definitive publisher-authenticated version is available on the publisher Web site.
To promote efficient separation and structural analysis of glycosaminoglycan oligosaccharides, we developed a straightforward method that combined gel electrophoresis and mass spectrometry (MS). Potential limitations of this approach (e.g., low extraction yields and weak compatibility with MS) were resolved by developing an active extraction procedure that yielded a quantitative amount of sulfated oligosaccharides from excised gel bands. The compatibility of obtained oligosaccharides for subsequent MS analysis was ensured using a single, simple clean-up step on a mixed C18/graphite carbon solid-phase column that was fully effective for polymerization degrees ranging from di- to dodecasaccharides. The reported combination of carbohydrates-polyacrylamide gel electrophoresis with MS was successfully applied to glucosamino- (heparin) and galactosamino- (dermantan sulfate) glycans, demonstrating the potential of our method for structural analysis of bioactive sulfated carbohydrates extracted from biological matrices. Graphical Abstract ᅟ.
With the increase in life expectancy, reducing the visible signs of skin aging has become a major issue. A reduction in collagen and hyaluronic acid synthesis by fibroblasts is a feature of skin aging. The green seaweed, Ulva intestinalis, is an abundant and rich source of nutrients, especially proteins and peptides. The aim of this study was to assess the potential cosmetic properties of a protein fraction from Ulva intestinalis (PROT-1) containing 51% of proteins and 22% of polysaccharides, and its enzymatic peptide hydrolysates on human dermal fibroblasts. PROT-1 was extracted using a patented acid- and solvent-free process (FR2998894 (B1)). The biochemical characterization and chromatographic analysis showed a main set of proteins (25 kDa). To demonstrate the anti-aging potential of PROT-1, fibroblast proliferation and collagen and hyaluronic acid production were assessed on fibroblast cell lines from donors aged 20 years (CCD-1059Sk) and 46 years (CCD-1090Sk). PROT-1 induced a significant increase in collagen and hyaluronic acid production per cell, and a reduction in cell proliferation without increasing cell mortality. These effects were reversed after protein hydrolysis of PROT-1, showing the central role of proteins in this promising anti-aging property.
The variegated scallop (Mimachlamys varia) is a filter feeder bivalve encountered in marine regions of the Atlantic coast. In particular, it is present in the La Rochelle marina (France), where it is used for the biomonitoring of marine pollution, due to its ability to strongly bioaccumulate pollutants. In this semi-closed environment, contamination generated by port activities leads to an accumulation of both organic and metal pollutants. Zinc is one of these pollutants, present at a dose of up to 150 to µg.L -1 . This study investigated the effects of 48 h zinc exposure upon the metabolic profiles of Mimachlamys varia using UHPLC/QToF (ultrahigh performance liquid chromatography-quadrupole time-of-flight) tandem mass spectrometry metabolomics. After acclimation in mesocosms recreating in situ conditions, both controls and exposed with Zn 2+ (150 μg.L -1) bivalves were dissected to recover the gills after 48 h and stored at -80°C before metabolites extraction. UHPLC/QToF tandem mass spectrometry was performed to study metabolite composition of samples. Statistical analysis of results using multivariate techniques showed a good classification between control and exposed groups. Eleven identified metabolites were found to be downmodulated in exposed scallops. These variations could reflect potential zinc effects on several of the biological processes, such as energy metabolism, osmoregulation and defense against oxidative stress. Among the eleven metabolites highlighted, four were reported for the first time in an aquatic organism exposed to Zn. This study demonstrates once again the diversity of interactions between bivalves and metals and the complexity of the physiological response of marine bivalves to pollutants.
Ports are a good example of how coastal environments, gathering a set of diverse ecosystems, are subjected to pollution factors coming from human activities both on land and at sea. Among them, trace element as copper represents a major factor. Abundant in port ecosystem, copper is transported by runoff water and results from diverse port features (corrosion of structures, fuel, anti-fouling products, etc.). The variegated scallop Mimachlamys varia is common in the Atlantic port areas and is likely to be directly influenced by copper pollution, due to its sessile and filtering lifestyle. Thus, the aim of the present study is to investigate the disruption of the variegated scallop metabolism, under a short exposure (48 h) to a copper concentration frequently encountered in the waters of the largest marina in Europe (82 μg/L). For this, we chose a non-targeted metabolomic approach using ultra-high performance liquid chromatography coupled to high resolution mass spectrometry (UHPLC-HRMS), offering a high level of sensitivity and allowing the study without a priori of the entire metabolome. We described 28 metabolites clearly modulated by copper. They reflected the action of copper on several biological functions such as osmoregulation, oxidative stress, reproduction and energy metabolism.
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