Biological degradation of Ascophyllum nodosum

The alginate forms the major structural component of the cell wall and the intercellular matrix of the brown alga Ascophyllum nodosum. Successful biological degradation of A. nodosum would largely depend on the dissolution of the alginate, but reactive compounds in the alga such as polyphenols may also have toxic effects on the microbial population involved. Aerobic and anaerobic batch reactors, operated at 35°C and pH 7, were fed milled A. nodosum, nutrients and inocula adapted to seaweed degradation. The dominant factor for conversion of organic matter during anaerobic digestion was the inhibitory effect of the polyphenols on alginate lyases and methane production. Probably, the relative large fraction of high molecular weight polyphenols (>10 kDa) in this alga gave efficient binding of proteins during digestion. The anaerobic degradation was greatly stimulated when the polyphenols were fixed with low amounts of formaldehyde. An accumulated content of guluronate in the remaining alginate indicated that Ca-crosslinking also limited the guluronate lyase access to the polymer. In contrast, the aerobic digestion of alga gave no increase in the guluronate content of the residual alginate. Compared to anaerobic conditions, the phenols had a much lower influence on the hydrolytic rate of organic matter during aerobic conditions. This revised version was published online in August 2006 with corrections to the Cover Date.     

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.     

Water Status Changes in Shoots of a Seaweed Ascophyllum nodosum at Subzero Temperatures

The gradient freezing and NMR spectroscopy were used to study the physical state of water in apices of the intertidal seaweed Ascophyllum nodosum at freezing temperatures. In the apices exposed to temperatures below –10°C, two fractions of bound water were revealed. The slow (T₂ 50 ms) fraction of bound water was completely frozen at –25°C, and its freezing rate was temperature-sensitive. This fraction was apparently associated with protoplasmic water and cell-wall polysaccharides. The fast fraction (T₂ < 10 ms) of bound water was presumably due to water-soluble globular proteins. The freezing rate for this fraction depended on neither the temperature nor the amount of water. The presence of unfrozen water in apical cells at –40°C was demonstrated. The role of this water fraction in maintaining the native structure of biomacromolecules and apex survival is discussed.     

Direct determination of alginate content in brown algae by near infra-red (NIR) spectroscopy

The methods used to quantify total alginate in brown algal tissue are time-consuming and may also be misleading, so faster and simpler methods for measuring alginate content would be beneficial in a variety of applications. This study reports on the use of near infra-red (NIR) analysis to monitor the alginate content of Laminaria hyperborea stipe during biodegradation. NIR reflectance spectra were recorded for 78 different freeze-dried samples of its stipe. The samples were collected during several biological degradation experiments and the total alginate content varied from 2.2 to 40.8% Na-alginate (w/w), determined by established methods based on ion exchange. Data analysis was performed using multivariate calibration methods in order to relate the spectral data to the alginate content. PLS2 analysis revealed some dependence on material type, probably reflecting differences in polyphenol content. In the end, a PLS1 model with 9 components was selected. The calculated model was validated both with internal data and with an external test set. Internal full cross validation explained 96.6% of the variance in alginate content. The external validation showed that the PLS1 model was able to predict the alginate concentration with a root mean square prediction accuracy of 2.1%. This revised version was published online in June 2006 with corrections to the Cover Date.     

Spatial distribution and quantification of Fucus species and Ascophyllum nodosum beds in intertidal zones using spot imagery

Low tide SPOT images were selected from two French coast areas characterized by important Fucaceae populations (Pleubian-Bréhat site in Northern Brittany and Ré Island on the Atlantic coast).A specific data transformation yielding the theoretical algal cover was used. This index takes the radiometric properties of the intertidal zone and of the Fucaceae into account. Satellite data cover was validated by comparing it with selected field samples.Other field data indicate that a linear relation exists between cover and biomass. This relationship is quite independent of cover patchiness. However, it can vary according to species, season and location. Hence it was possible to estimate Fucus sp and Ascophyllum nodosum harvestable biomass using appropriate segments of the intertidal zone.     

Amino acids,sugars and sterols of some Mediterranean brown algae

Free amino acids, amino sulfonic acids, sugars and sterols have been examined and quantitatively determined in 10 brown seaweeds. Acidic amino acids and their amides are the main components of the amino acid fraction. Cysteinolic acid, taurine and its N-methyl derivatives have been identified in most of the species examined. In all the algae, mannitol is present, sometimes in very large amounts. The sterol fractions of all the species contain fucosterol, cholesterol and 24-methylenecholesterol; minute amounts of 22-dehydrocholesterol and 24-methylcholesta-5, 22-dien-3β-ol have also been frequently detected.     

Alginate degradation during anaerobic digestion of Laminaria hyperborea stipes

Polyphenols and divalent metal ions present in the tissue may seriously affect the degradation of alginate during anaerobic digestion of brown seaweeds. Laminaria hyperborea stipes, harvested at 59 °N off the Norwegian coast in the autumn, were degraded at different concentrations of polyphenols in anaerobic batch reactors at 35 °C and pH 7. This was done by removing or adding the mechanically peeled outer phenolic layer of the algae, and using methanogenic and alginate degrading inocula already adapted to L. hyperborea degradation. Initial alginate released from the algal particles was affected by NaOH titrations because the Ca/Na-ratio was reduced. After a rapid consumption of the mannitol, alginate lyases were induced, and guluronate lyases showed the highest extracellular activity. Then the microbes digested 0.12–0.23 g Na-alginate L⁻¹ h⁻¹. Later the degradation rate of alginates declined almost to zero, and 13–50% of the alginate remained insoluble. The total solubilisation of alginates was apparently limited by both Ca-crosslinked guluronate residues and complexation with compounds such as polyphenols. The methane production had a lag phase that increased at higher amounts of soluble polyphenols, and the total fermentation probably also became product inhibited if soluble compounds such as acetate, ethanol and butyrate were accumulated. This revised version was published online in September 2006 with corrections to the Cover Date.     

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.     

Phenolic lipids from related marine algae of the order dictyotales

2-(1′-Oxo-dodeca-5′, 8′, 11′, 14′, 17′(all Z)-pentaenyl)-5-methoxy-1, 3-dihydroxybenzene, 2- (1′-oxo-dodeca-5′, 8′, 11′, 14′, 17′(all Z)-pentaenyl)-1, 3, 5-trihydroxybenzene, 2-(17′-hydroxy-1′-oxo-dodeca-5′, 8′, 11′, 14′(all Z)-tetraenyl)-1, 3, 5-trihydroxybenzene and 2-(1′oxo-hexadecyl)-1, 3, 5-trihydroxybenzene have been isolated from the related brown algae Zonaria farlowii, Z diesingiana and Lobophora papenfussii. The structures of these new metabolites are based on extensive spectral analyses and comparisons with model compounds. The isolation of (+)-7, 8-dimethyltocol, from L. papenfussii, is also reported.     

围体外循环期血小板保护的研究进展

在体外循环过程中,血小板可经各种途径被激活,导致α-颗粒释放,发生粘附、聚集、收缩、释放等反应,导致术后血小板数量和质量的下降。通过在围体外循环期使用某些药物可对血小板进行功能性保护,而血小板分离技术可使血小板避免体外循环的打击,得到数量和功能的双重保护。本文将就体外循环期间血小板保护的研究进展作一综述。     

Microbial degradation of pollutants at high salt concentrations

Though our knowledge on microbial degradation of organic pollutants at high salt concentrations is still limited, the list of toxic compounds shown to be degraded or transformed in media of high salinity is growing. Compounds transformed aerobically include saturated and aromatic hydrocarbons (by certain archaeobacteria), certain aromatic compounds, organophosphorus compounds, and formaldehyde (by halotolerant eubacteria). Anaerobic microbial transformations of toxic compounds occurring at high salt concentrations include reduction of nitroaromatic compounds, and possibly transformation of chlorinated aromatic compounds.     

Maintenance of biodegradation capacities of aerobic bacteria during long-term preservation

Six strains of aerobic Gram negative bacteria degrading toluene, 2,4-dichlorophenoxyacetate, 2,2-dichloropropionate or 3-chlorobenzoate were freeze-dried and liquid-dried in the presence or absence of a protective agent. Survival and maintenance of the biodegradation capability was checked before and after drying, and after storage of the ampoules for one year at 4° or 25°C. In many cases, stability of the degradation potential was low although viability was high. Survival and stability of all strains was always highest after preservation by liquid drying in the presence of myo-inositol and activated charcoal as protective agents. Losses of biodegradation abilities were highest after freeze-drying using no protective agents. Cells grown on complex medium were less sensitive to drying than cells grown under selective pressure (on mineral medium with a special compound as the sole carbon source). A choice of the most appropriate preservation method and the use of an effective protectant is recommended to avoid genetic alterations, and to maintain biodegradation capacities during long-term preservation.     

长双歧杆菌DD98冻干保护剂优化及菌粉保存稳定性研究

以长双歧杆菌DD98为研究对象,通过对冻干保护剂配方的优化,冻干菌粉的存活率提高到90%以上。通过进一步稳定性研究,采用保护剂优化配方制备的冻干菌粉在4℃保存24个月后,活菌数仍在1.0×10^10 CFU/g以上,在25℃条件下可以保存12个月,双歧杆菌的存活率在1.0×10^6CFU/g以上,符合FAO/WHO建议食品益生菌活菌数应在1.0×10^6 CFU/g^1.0×10^7CFU/g的标准。     

Identification of associating carbohydrate sequences with labelled oligosaccharides

The gelling subunit of alginate, the major cell-wall polysaccharide of brown algae, was used as a molecular marker for identification of this cell-wall carbohydrate subunit at the cellular level. Short polyguluronate chains were conjugated to fluorescein and used as a probe to identify the gelling regions of alginate in tissue sections from a brown alga. The specificity of the probe for gelling subunits was demonstrated by lack of cell-wall labelling in the absence of calcium, correlation between divalent-cation binding affinities of polyguluronate with labelling intensity, and lack of labelling by fluorescein-conjugated nongelling subunits. The probe labelling-pattern also differed from sulfated fucan distribution. Extracellular matrix and cell walls were labelled on sections of vegetative blade, stipe and reproductive frond of Fucus gardneri Silva. Probe labelling was rapid, being virtually complete within 5 min. Probe labelling in seawater differed markedly from labelling at lower ionic strength and is interpreted as reflecting alginategelling properties in natural conditions. High-and low-affinity binding sites are discussed in terms of gelling-subunit length and steric availability. Fluorescein-conjugated polygalacturonate, which also forms calcium dimers, labelled extracellular alginate by formation of mixed polygalacturonate-polyguluronate dimers. Binding by the alginate hybridization probe differs from nucleic-acid hybridization in divalent-cation bridging and the lack of both a conformational transition and polymer polarity.Abbreviations EDTA ethylenedinitrilo tetraacetic acid (ethylenediaminetraacetic acid) - NMR nuclear magnetic resonance spectroscopy     

The effect of elevated temperature and reactor shutdown on the benthic marine flora of the millstone thermal quarry,Connecticut

Forty-nine species and one variety of benthic blue-green, red, brown and green algae were found over a 1.5 year period in a thermal sea water dump where temperatures average 10°C above ambient Long Island Sound waters. Of these, 58% can survive temperatures exceeding 30°C, but only six show survival after prolonged excessive temperature. At temperatures less than 27°C, the number of taxa is independent of temperature, but at greater temperatures there is a significant negative correlation of temperature to taxa count, reaching a minimum of 3 species. Rapid temperature drops cause concomitant drops in taxa counts, 14% of this variation being attributed to drastic temperature change which affects the algae.     

Contribution of surface salt bridges to protein stability: guidelines for protein engineering

The small globular protein, ubiquitin, contains a pair of oppositely charged residues, K11 and E34, that according to the three-dimensional structure are located on the surface of this protein with a spatial orientation characteristic of a salt bridge. We investigated the strength of this salt bridge and its contribution to the global stability of the ubiquitin molecule. Using the "double mutant cycle" analysis, the strength of the pairwise interactions between K11 and E34 was estimated to be favorable by 3.6kJ/mol. Further, the salt bridge of the reverse orientation, i.e. E11/K34, can be formed and is found to have a strength (3.8kJ/mol) similar to that of the K11/E34 pair. However, the global stability of the K11/E34 variant of ubiquitin is 2.2kJ/mol higher than that of the E11/K34 variant. The difference in the contribution of the opposing salt bridge orientations to the overall stability of the ubiquitin molecule is attributed to the difference in the charge-charge interactions between residues forming the salt bridge and the rest of the ionizable groups in this protein. On the basis of these results, we concluded that surface salt bridges are stabilizing, but their contribution to the overall protein stability is strongly context-dependent, with charge-charge interactions being the largest determinant. Analysis of 16 salt bridges from six different proteins, for which detailed experimental data on energetics have been reported, support the conclusions made from the analysis of the salt bridge in ubiquitin. Implications of these findings for engineering proteins with enhanced thermostability are discussed.     

Isolation of water-soluble alginate from brown algae

Summary Water-soluble alginate was obtained from an aqueous extract of Kjellmaniella crassifolia by precipitation with HCl, calcium acetate or 20% ethanol in the presence of 0.05 M MgCl₂ Of these precipitation procedures, MgCl₂-ethanol gave the purest alginate preparation as judged by electrophoresis. The thin-layer and gas-liquid chromatography of its acid hydrolysate, and the IR spectra analysis of the whole alginate, suggested that the water-soluble alginate is similar to ordinary water-insoluble and alkali-soluble alginate such as Kelco alginate.However, the alginate obtained in the present work contained a great excess of mannuronic acid residues, giving an M:G ratio of about 13. Its molecular weight distribution was rather broad as with Kelco alginate, but the molecular weight of its major component was estimated to be 500 000 amu, whereas that Kelco alginate measured on the same column under the same condition was 1 700 000 amu. This suggests that water-soluble alginate was far smaller in average molecular size than Kelco alginate.     

Modulating biochemical perturbations during 72-hour machine perfusion of kidneys: role of preservation solution

This study documents renal biochemistry during hypothermic machine perfusion of kidneys. It is intended to demonstrate that a comprehensive evaluation of organ viability during ex-vivo preservation is needed to increase the number of organs available for transplantation and to reduce the current renal discard rate. Porcine kidneys were hypothermically machine perfused for 72 h with either Unisol-UHK or Belzer-Machine Perfusion Solution, (Belzer-MPS). Renal perfusate samples were periodically collected and biochemically analyzed. Significant differences were measured in the renal metabolic activity between the two experimental groups while similar values for traditional parameters such as renal flow rate and vascular resistance values were recorded. The effluent of UHK perfused kidneys showed strong metabolites and NH(4)(+) dynamics (P<0.05 vs. baseline), while the Belzer-MPS kidneys metabolic activity led to little or no change of the effluent biochemistry relative to baseline.