Isolation of humic acid from the brown algaeAscophyllum nodosum,Fucus vesiculosus,Laminaria saccharina and the marine angiospermZostera marina

Recent discovery of humic acid (HA) in the free-living, brown algaPilayella littoralis has prompted a search for HA in other live plants. Marine algaeCodium fragile andMonostoma oxyspermum (greens),Chondrus crispus,Palmaria palmata andPolysiphonia lanosa (reds),Ascophyllum nodosum, Fucus vesiculosus andLaminaria saccharina (browns) andZostera marina (marine angiosperm) were investigated for their HA content. Only the brown algae and the marine angiosperm contained HA, which was extracted by a standard procedure augmented with necessary removal of alginic acid (where applicable). The isolated products were identified as HA by comparison of their analytical data, uv-visible, FTIR,¹H NMR spectra and morphologies with those of authentic HA isolated from municipal compost.Authors for correspondence     

Role of Auxin in Induction of Polarity in Fucus vesiculosus Zygotes

We studied the effects of auxin (indole-3-acetic acid) on formation of the primary polarity axis in zygotes of the brown algae Fucus vesiculosusL. Within the first 2.5 h after fertilization, the zygotes release this phytohormone in the ambient medium. The treatment of developing zygotes with the inhibitor of indole-3-acetic acid transport from the cell 2,3,5-triiodobenzoic acid at 5 mg/l arrests the auxin secretion and leads to its accumulation in the cells. This causes a significant delay in zygote polarization. The treatment of zygotes with the exogenous indole-3-acetic acid at 1 mg/l stimulates cell polarization and formation of a rhizoid protuberance. When auxin was added to the medium with triiodobenzoic acid, the inhibitory effect of the latter was eliminated. It has been proposed that the content of indole-3-acetic acid in the ambient medium is a key factor in the induction of polarity of the F. vesiculosus zygotes.     

Spatial variation in polyphenolic content of Ascophyllum nodosum (Fucales,Phaeophyta)

Spatial variation in polyphenolic content in annual shoots of the brown alga Ascophyllum nodosum was quantified using a hierarchical sampling design. Three sampling levels, covering distances of 10⁰–10⁶ m, were used. Comparisons were made between two areas, Tjärnö on the Swedish west-coast and the Isle of Man in the Irish Sea, with very different types of environmental conditions. No significant differences in mean polyphenolic levels were found between the two study areas (6.6% of dry mass at Tjärnö and 9.2% at the Isle of Man), whereas significant and substantial differences were found among sites within areas (range 5.7%–11.4%) and among quadrats within sites (range 3.7%–13.1 %). The extensive variation at the smaller spatial scales points out the importance of using thorough sampling procedures at all levels in large-scale studies on algal polyphenolics, e.g. biogeographical comparisons, which have been neglected in several previous studies. Moreover, the results imply that experiments on causal factors of polyphenolic variation should be designed to explain the spatial scales on which the factors are impportant. This study also investigated the relationship between polyphenolic concentration and both plant size and mean area of annual shoots. The mean area was used as an estimate of the mean growth rate of the annual shoots within an individual. No significant relationships were found between shoot growth rate, or plant size, and polyphenolic levels in annual shoots at any of the three spatial scales that were investigated.     

Antioxidant and pro-oxidant activities of the brown algae, Laminaria digitata, Himanthalia elongata, Fucus vesiculosus, Fucus serratus and Ascophyllum nodosum

The ability of Laminaria digitata, Himanthalia elongata, Fucus vesiculosus, Fucus serratus and Ascophyllum nodosum to scavenge peroxyl radicals was investigated by kinetic studies in a model system. The thermal initiated oxidation of methyl linoleate was performed at 60°C in heptanol, with or without antioxidants. When they reached 1% of the substrate, seaweed extracts exhibited antioxidant activities by extending the induction period, but they did not suppress the rate of oxygen uptake as did vitamin E. A synergistic effect occurred when both algal extracts (1.5 g L^-1) and vitamin E (0.4 mmol L^-1) were present, and the effectiveness of the combined antioxidants during the whole induction period was vitamin E effectiveness. The synergistic effect of L. digitata, however, was subject to seasonal variations: samples collected in summer were effective synergists, whereas samples collected in winter displayed a marked negative synergism. The phospholipid fractions of F. vesiculosus, F. serratus and A. nodosum, including pigments, accounted for only 6% of the total lipid fraction, and did not exhibit a large synergistic effect. The main phospholipid was not phosphatidyl ethanolamine as usually related, but phosphatidyl inositol. Fucoxanthin had some antioxidant activity per se under our experimental conditions, but did not act as a synergist of vitamin E. The most potent synergists were recognized as chlorophyll a and related compounds by the application of liquid-liquid partition and chromatography for the identification of active components. This revised version was published online in June 2006 with corrections to the Cover Date.     

Seasonal changes in situ grazing of the mesoherbivores Idotea baltica and Gammarus oceanicus on the brown algae Fucus vesiculosus and Pylaiella littoralis in the central Gulf of Finland, Baltic Sea

The in situ grazing experiments were performed in the shallow water rocky habitat of the northern Baltic Sea during ice-free season 2002. In the experiments the effects of algal species and choice on the grazing of the mesoherbivores Idotea baltica (Pallas) and Gammarus oceanicus Segerstråle were tested. Salinity, temperature, concentration of nutrients in water and macroalgae and net production of macroalgae were considered as random effects in the analysis. The invertebrate feeding rate was mainly a function of the net photosynthetic activity of Pylaiella littoralis (L.) Kjellman and Fucus vesiculosus L. Feeding rate increased significantly with decreasing algal photosynthetic activity. When the two algal species were incubated together invertebrates fed primarily on P. littoralis. Low selectivity towards P. littoralis coincided with its high photosynthetic activity. The presence of F. vesiculosus did not modify the invertebrate feeding on P. littoralis. The results indicated that (1) the grazing on F. vesiculosus depended on the availability of P. littoralis, (2) the photosynthetic activity of algae explained the best the variation in grazing rate and (3) the grazers are not likely to control the early outbreak of filamentous algae in the northern Baltic Sea by avoiding young and photosynthetically active algae. The likely mechanism behind the relationship is that the increased photosynthetic activity of macroalgae coincides with their higher resistance to herbivory.     

Effects of filamentous algae and deposited matter on the survival of Fucus vesiculosus L. germlings in the Baltic Sea

As a result of increased nutrient levels in the Baltic Sea during thepast 50 years, mass developments of filamentous algae have become a commonfeature along the Swedish east coast and deposition of organic matter has alsoincreased. To test whether these two factors have any effects on the early lifestages of Fucus vesiculosus a number of laboratory andfield studies were conducted. The amount of epilithic and epiphytic filamentousalgae on F. vesiculosus and the amount of deposited matterin the littoral zone were quantified during the two reproductive periods ofF. vesiculosus, early summer (May–June) and lateautumn (September–October). Both filamentous algae (Cladophoraglomerata) and deposited matter (introduced either before or aftersettlement of fertilized eggs) were shown to significantly decrease the numberof surviving germlings. The survival of germlings seeded on stones withfilamentous algae, or seeded on culture dishes concurrently with the lowestconcentration of deposited matter (0.1 g dm^–2),was 5% or less. In the field, the amount of filamentous algae was significantlyhigher during F. vesiculosus summer reproduction, whereasthe amount of deposited matter collected in traps was significantly higherduring the period of autumn reproduction. The greatest biomass of filamentousalgae was observed at sheltered sites. Based on the negative effects offilamentous algae and deposited matter on Fucusrecruitmentand the observation of local and seasonal differences in abundance offilamentous algae and deposition, we suggest that the prerequisites for thesurvival of either summer or autumn-reproducing populations of F.vesiculosus in the Baltic Sea may differ locally.     

Variation in natural selection for growth and phlorotannins in the brown alga Fucus vesiculosus

Directional selection for plant traits associated with resistance to herbivory tends to eliminate genetic variation in such traits. On the other hand, balancing selection arising from trade-offs between resistance and growth or spatially variable selection acts against the elimination of genetic variation. We explore both the amount of genetic variation and variability of natural selection for growth and concentration of phenolic secondary compounds, phlorotannins, in the brown alga Fucus vesiculosus. We measured variation in selection at two growing depths and two levels of nutrient availability in algae that had faced two kinds of past growing environments. Genetic variation was low for growth but high for phlorotannins. The form and strength of selection for both focal traits depended on the past growing environment of the algae. We found strong directional selection for growth rate in algae previously subjected to higher ultraviolet radiation, but not in algae previously subjected to higher nutrient availability. Stabilizing selection for growth occurred especially in the deep growing environment. Selection for phlorotannins was generally weak, but in some past-environment-current-environment combinations we detected either directional selection against phlorotannins or stabilizing selection. Thus, phlorotannins are not selectively neutral but affect the fitness of F. vesiculosus. In particular, there may be a fitness cost of producing phlorotannins, but the realization of such a cost varies from one environment to another. Genetic correlations between selective environments were high for growth but nonexistent for phlorotannins, emphasizing the high phenotypic plasticity of phlorotannin production. The highly heterogeneous selection, including directional, stabilizing, and spatially variable selection as well as temporal change in selection due to responses to past environmental conditions, probably maintains a high amount of genetic variation in phlorotannins. Such variation provides the potential for rapid evolutionary response of phlorotannins under directional selection.     

Competitive interactions between Fucus spiralis L. and F. vesiculosus L. (Fucales,Phaeophyta)

Fucus spiralis forms a conspicuous belt in the upper intertidal on rocky shores of the NW Atlantic. The objective of this study was to determine whether competition among congeners plays a role in restricting the distribution of F. spiralis to the upper shore. A replacement series design was used to test the growth performance of F. spiralis and F. vesiculosus in monocultures and in mixtures set up in the mid shore level on an exposed rocky coast. For F. spiralis and F. vesiculosus, all individual and relative crowding coefficients were < 1 and > 1, respectively. Hence, F. vesiculosus succeeded relative to F. spiralis in all mixtures tested. This finding was confirmed by graphical analysis. Ratio diagrams showed that the outcome of long term interaction among the species is extinction of F. spiralis. The results lead to the conclusion that interaction among congeners is a major determinant of the restricted distribution of F. spiralis.     

The antibacterial effect of a polyhydroxylated fucophlorethol from the marine brown alga,Fucus vesiculosus

An antibacterial compound was isolated from the brown alga Fucus vesiculosus. Purification consisted of extraction of plant material with 0.05% trifluoroacetic acid, concentration on a C₁₈ cartridge, and reverse-phase high performance liquid chromatography on a C₁₈ semi-preparative column. The isolated compound exhibited antibacterial activity against both the Gram-positive and the Gram-negative bacteria tested. Killing studies conducted indicated that the activity was bactericidal. The compound showed no haemolytic effect against human red blood cells. Results obtained by electrospray ionization mass spectrometry indicated that the antibacterial activity was caused by a polyhydroxylated fucophlorethol.     

Isolation and cross‐species amplification of microsatellite loci from the fucoid seaweeds Fucus vesiculosus,F. serratus and Ascophyllum nodosum (Heterokontophyta,Fucaceae)

The Fucaceae is a family of brown seaweeds that dominate and frequently co‐occur on North Atlantic rocky shores. We developed nine polymorphic microsatellite markers for the fucoid seaweeds Fucus vesiculosus, F. serratus and Ascophyllum nodosum using a combined, enriched library. Six of these loci were polymorphic in at least two species, showing from two to eight alleles with heterozygosities ranging from 0.41 to 0.85. Loci were also tested on F. spiralis, revealing five polymorphic microsatellite loci in this species.     

Monoclonal antibodies as molecular probes for cell wall antigens of the brown alga,Fucus

Monoclonal antibodies to cell wall carbohydrates were produced against carbohydrates extracted from the brown alga, Fucus distichus ssp. edentatus (de la Pyl.) Powell. Mouse spleen cells were immunized in vitro with alginate and fucans, and hybridoma cultures were screened by enzyme immunoassay. Most antibodies were immunoglobulin (Ig)M and one was IgA. Antigens were localized on methacrylate sections of Fucus tissues by indirect immunofluorescence. Each antibody labelled tissues with a distinctive distribution pattern in cell walls and extracellular matrix regions, demonstrating that each antibody was specific for a different extracellular epitope (i.e., antigenic determinant). Most antibodies also labelled intracellularly on at least one cell type. Punctate, fibrous or clumped labelling was characteristic of individual antibodies and provided information related to carbohydrate structure and solubility. These antibodies are molecular probes for small regions on cell wall polymers and should be valuable in studies of cell wall synthesis, secretion, assembly and modification as well as carbohydrate fine structure and function.Abbreviations EDTA ethylenediaminetetraacetic acid - EIA enzyme immunoassay - Ig immunoglobulin (IgG, IgM and IgA are immunoglobulin types)     

Bromoperoxidase activity and vanadium level of the brown alga Ascophyllum nodosum

Vanadium-dependent peroxidase activity in extracts of Ascophyllum nodosum growing in the intertidal region close to Roscoff/France, and algal vanadium levels, followed approximately similar seasonal variation, as deduced from a study lasting from April 2005 to March 2006. High peroxidase (PO) activity was found in extracts obtained from algae collected in between midwinter to spring [∼100-190 U per g dry mass (dm), triiodide assay] with a maximum in April. Periods of reduced PO activity lasted from summer to early winter (∼50-90 U per g dm). High vanadium levels (1.5-2.2 mg kg⁻¹ dm) were found in algae collected from midwinter to spring, whereas reduced levels (0.6-1.4 mg kg⁻¹ dm) were found in summer to early winter.     

Asymmetrical jelly secretion of zygotes of Pelvetia and Fucus: An early polarization event

Five hours before germination the zygotes of Pelvetia fastigiata adhere to their substrate. A jelly layer covers the entire cell but most of the transparent jelly, artificially outlined by a layer of resin beads, is secreted at the prospective rhizoid pole. If the direction of the growth-orienting light is shifted after the asymmetrical secretion has already started, the direction of the secretion is also shifted. The polarization axis can be predicted by the site of the intensive jelly secretion. The germination of Fucus vesiculosus and F. spiralis is also preceded by an intensive asymmetrical jelly secretion. However, at the rhizoid pole of F. serratus the jelly secretion does not increase until the germinating zygote becomes pear-shaped. Fucoid zygotes do not adhere, neither do they have a jelly cover as long as they develop in sulfate-free sea water.Abbreviations a.f. after fertilization     

Calcium and potassium currents in the Fucus egg

The electrical properties of unfertilized eggs of Fucus serratus L. were characterized using voltage clamp and current clamp with single electrodes. The plasma membrane of the unfertilized egg is excitable. Depolarizing the egg in current clamp induced a transient depolarizing voltage response, the amplitude of which was dependent on the presence of external Ca²⁺ or Ba²⁺ and was blocked by La³⁺. The repolarizing phase was blocked by tetraethylammonium ions. Repeated stimulation at frequencies greater than 0.5 Hz caused a transient loss of excitability. Voltage-clamp experiments revealed that an inward current with an activation threshold of -35 mV underlies the depolarizing phase of the voltage response. This current showed rapid activation and slow inactivation. The current was blocked by La³⁺ and could be carried by Ca²⁺ and Ba²⁺ but not by Sr²⁺ or Na⁺. Further depolarization to values more positive than-5 mV induced a slowly activating outward K⁺ current in addition to the inward current, which corresponded to the repolarizing phase of the voltage response. This K⁺ current showed little or no inactivation during stimulation and slow deactivation on return to the resting potential. Hyperpolarizing the egg elicited an inward current. On fertilization, the Fucus egg generates a depolarizing fertilization potential. Voltage-clamp experiments revealed an inward fertilization current underlying the fertilization potential. Within 15 min of fertilization a dramatic, irreversible increase in resting K⁺ permeability developed. The roles of the plasma-membrane channels in generation of the fertilization potential and egg activation are discussed.Abbreviations and Symbols ASW artificial seawater - SECC single-electrode current clamp - SEVC single-electrode voltage clamp - TEA tetraethylammonium - Vm membrane potential This work was supported by The Marine Biological Association U.K., Science and Engineering Research Council U.K. and The Royal Society of London.