Investigations into the Toxicology of Spirolides, a Group of Marine Phycotoxins

Spirolides are marine phycotoxins produced by the dinoflagellates Alexandrium ostenfeldii and A. peruvianum. Here we report that 13-desmethyl spirolide C shows little cytotoxicity when incubated with various cultured mammalian cell lines. When administered to mice by intraperitoneal (ip) injection, however, this substance was highly toxic, with an LD50 value of 6.9 µg/kg body weight (BW), showing that such in vitro cytotoxicity tests are not appropriate for predicting the in vivo toxicity of this toxin. Four other spirolides, A, B, C, and 20-methyl spirolide G, were also toxic to mice by ip injection, with LD50 values of 37, 99, 8.0 and 8.0 µg/kg BW respectively. However, the acute toxicities of these compounds were lower by at least an order of magnitude when administration by gavage and their toxic effects were further diminished when administered with food. These results have implications for future studies of the toxicology of these marine toxins and the risk assessment of human exposure

A comparative study on innate immune parameters in the epidermal mucus of various fish species.

Peer reviewed: YesNRC publication: Ye

Myxinidin, a Novel Antimicrobial Peptide from the Epidermal Mucus of Hagfish, Myxine glutinosa L.

Fish epidermal mucus contains innate immune components that provide a first line of defense against various infectious pathogens. This study reports the bioassay-guided fractionation and characterization of a novel antimicrobial peptide, myxinidin, from the acidic epidermal mucus extract of hagfish (Myxine glutinosa L.). Edman sequencing and mass spectrometry revealed that myxinidin consists of 12 amino acids and has a molecular mass of 1,327.68 Da. Myxinidin showed activity against a broad range of bacteria and yeast pathogens at minimum bactericidal concentration (MBC) ranging from 1.0 to 10.0 \ub5g/mL. Screened pathogens, Salmonella enterica serovar Typhimurium C610, Escherichia coli D31, Aeromonas salmonicida A449, Yersinia ruckeri 96-4, and Listonella anguillarum 02-11 were found to be highly sensitive to myxinidin at the MBC of 1.0\u20132.5 \ub5g/mL; Staphylococcus epidermis C621 and yeast (Candida albicans C627) had an MBC of 10.0 \ub5g/mL. The antimicrobial activity of myxinidin was found to be two to 16 times more active than a potent fish-derived antimicrobial peptide, pleurocidin (NRC-17), against most of the screened pathogens. The microbicidal activity of myxinidin was retained in the presence of sodium chloride (NaCl) at concentrations up to 0.3 M and had no hemolytic activity against mammalian red blood cells. These results suggest that myxinidin may have potential applications in fish and human therapeutics.Peer reviewed: YesNRC publication: Ye

Comparison of antimicrobial activity in the epidermal mucus extracts of fish

The mucus layer on the surface of fish consists of several antimicrobial agents that provide a first line of defense against invading pathogens. To date, little is known about the antimicrobial properties of the mucus of Arctic char (Salvelinus alpinus), brook trout (S. fontinalis), koi carp (Cyprinus carpio sub sp. koi), striped bass (Morone saxatilis), haddock (Melanogrammus aeglefinus) and hagfish (Myxine glutinosa). The epidermal mucus samples from these fish were extracted with acidic, organic and aqueous solvents to identify potential antimicrobial agents including basic peptides, secondary metabolites, aqueous and acid soluble compounds. Initial screening of the mucus extracts against a susceptible strain of Salmonella enterica C610, showed a significant variation in antimicrobial activity among the fish species examined. The acidic mucus extracts of brook trout, haddock and hagfish exhibited bactericidal activity. The organic mucus extracts of brook trout, striped bass and koi carp showed bacteriostatic activity. There was no detectable activity in the aqueous mucus extracts. Further investigations of the activity of the acidic mucus extracts of brook trout, haddock and hagfish showed that these fish species had specific activity for fish and human pathogens, demonstrating the role of fish mucus in antimicrobial protection. In comparison to brook trout and haddock, the minimum bactericidal concentrations of hagfish acidic mucus extracts were found to be 3c 1.5 to 3.0 times lower against fish pathogens and 3c 1.6 to 6.6 folds lower for human pathogens. This preliminary information suggests that the mucus from these fish species may be a source of novel antimicrobial agents for fish and human health related applications.Peer reviewed: YesNRC publication: Ye

Selected bioactivities of vaccinium berries and other fruit crops in relation to their phenolic contents

Antioxidant activity, urinary tract protective activity, and cardioprotective anti-platelet effects are among the bioactivities associated with dietary phenolics. These bioactivities were measured in vitro in fruit extracts from seven Vaccinium species and five non-Vaccinium species to determine their relationship to total phenolic content and to anthocyanin and proanthocyanidin content. Berries belonging to the genus Vaccinium were particularly high in antioxidant activity and urinary tract protective anti-adhesion activity, while anti-platelet activity varied among species. There was a positive relationship between antioxidant activity (using the oxygen radical absorbing capacity (ORAC) assay) and both the total phenolic (R2 = 0.76) and anthocyanin content (R2 = 0.43) of the fruit, although there was no relationship between ORAC and proanthocyanidin content. There were no relationships between anti-adhesion activity and total phenolic content, anthocyanin content, or proanthocyanidin content. Likewise, no relationships were observed between anti-platelet activity and total phenolic content, anthocyanin content, or proanthocyanidin content. These results suggest that while antioxidant properties are characteristic of all fruit phenolics, in vitro anti-adhesion and anti-platelet bioactivities may be due to less abundant phenolic subgroups.Peer reviewed: YesNRC publication: Ye

Selected bioactivities of vaccinium berries and other fruit crops in relation to their phenolic contents

Antioxidant activity, urinary tract protective activity, and cardioprotective anti-platelet effects are among the bioactivities associated with dietary phenolics. These bioactivities were measured in vitro in fruit extracts from seven Vaccinium species and five non-Vaccinium species to determine their relationship to total phenolic content and to anthocyanin and proanthocyanidin content. Berries belonging to the genus Vaccinium were particularly high in antioxidant activity and urinary tract protective anti-adhesion activity, while anti-platelet activity varied among species. There was a positive relationship between antioxidant activity (using the oxygen radical absorbing capacity (ORAC) assay) and both the total phenolic (R2 = 0.76) and anthocyanin content (R2 = 0.43) of the fruit, although there was no relationship between ORAC and proanthocyanidin content. There were no relationships between anti-adhesion activity and total phenolic content, anthocyanin content, or proanthocyanidin content. Likewise, no relationships were observed between anti-platelet activity and total phenolic content, anthocyanin content, or proanthocyanidin content. These results suggest that while antioxidant properties are characteristic of all fruit phenolics, in vitro anti-adhesion and anti-platelet bioactivities may be due to less abundant phenolic subgroups.Peer reviewed: YesNRC publication: Ye

Improved methods of analysis for betaines in Ascophyllum nodosum and its commercial seaweed extracts

Beneficial effects of seaweeds and their extracts on crop performance have been attributed to a variety of compounds, including the betaines which are quaternary ammonium betaines. Methods of analysis of betaines published thus far suffer from low sensitivity, lack of baseline separation of individual betaines and from interference from other sample constituents. A rapid cleanup protocol and a sensitive LC- MS/MS method of analysis were developed to afford baseline separation of four betaines in the brown alga Ascophyllum nodosum and its commercial seaweed extract. Using this method, the presence of glycine betaine, \u3b4-aminovaleric acid betaine, \u3b3-aminobutyric acid betaine and laminine in A. nodosum, and commercial extracts derived from A. nodosum, were confirmed and quantified. The major betaine present was \u3b3-aminobutyric acid betaine accounting for 0.008-0.014% of the dry weight of the seaweed and 0.014-0.027% of the dry weight of the commercial extracts. Seasonal variation in betaine content was observed. Differences in the total betaine content were observed between A. nodosum of the yellow (0.011-0.017% dry weight) and the olive green (0.017-0.021% dry weight) coloured morphologies.Peer reviewed: YesNRC publication: Ye

Metabolic changes in Atlantic salmon exposed to Aeromonas salmonicida detected by 1H-nuclear magnetic resonance spectroscopy of plasma

1H-NMR (nuclear magnetic resonance)-based chemometric methods have been applied for the first time to investigate changes in the plasma metabolite profiles of Atlantic salmon Salmo salar as a result of exposure to Aeromonas salmonicida subsp. salmonicida, a Gram-negative bacterium that is the etiological agent of furunculosis. Plasma samples were obtained from salmon that survived 21 d post exposure to A. salmonicida, and from a control group maintained under similar conditions. 1D 1H-NMR spectra were acquired and principal components analysis (PCA) was used to assess differences between the spectral profiles of plasma from salmon that survived an A. salmonicida challenge, and non-infected controls. PCA enables simultaneous comparison of spectra, presenting a simplified overview of the relationship between spectral data, where spectra cluster based on metabolite profile similarities and differences; information regarding the metabolite variations can therefore be readily deciphered. The major metabolite changes responsible for the spectral differences were related to modification in the lipoprotein profile and choline-based residues, with minor changes in carbohydrates, glycerol, trimethylamine-N-oxide and betaine. These changes indicated that exposure to A. salmonicida induced a characteristic biochemical response which could be used to determine the health status of salmon. This study suggests that with further development this metabolite profiling technique may be a useful tool for diagnosis of disease states in salmon and could provide a better understanding of the host-pathogen relationship which at present is poorly understood for A. salmonicida and Atlantic salmon.Peer reviewed: YesNRC publication: Ye

Biosynthesis of 13-Desmethyl Spirolide C by the dinoflagellate Alexandrium ostenfeldii

Biosynthetic origins of the cyclic imine toxin 13-desmethyl spirolide C were determined by supplementing cultures of the toxigenic dinoflagellate Alexandrium ostenfeldii with stable isotope-labeled precursors [1,2-13C2]acetate, [1-13C]acetate, [2-13CD3]acetate, and [1,2-13C2,15N]glycine and measuring the incorporation patterns by 13C NMR spectroscopy. Despite partial scrambling of the acetate labels, the results show that most carbons of the macrocycle are polyketide-derived and that glycine is incorporated as an intact unit into the cyclic imine moiety. This work represents the first conclusive evidence that such cyclic imine toxins are polyketides and provides support for biosynthetic pathways previously defined for other polyether dinoflagellate toxins.Peer reviewed: YesNRC publication: Ye