Evaluation of antioxidative activity of extracts from a brown seaweed,Sargassum siliquastrum

Antioxidative activities of the extracts from Sargassum siliquastrum were determined using the inhibition of red blood cell (RBC) hemolysis induced by 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) radicals, suppression of lipid peroxidation using rat brain homogenate, and scavenging activity of superoxide radicals. The dichloromethane fraction isolated from the methanol crude extract by differential solvent extractions exhibited the strongest antioxidant activity in both RBC hemolysis and lipid peroxidation assays. This fraction was further fractionated into four subfractions F1-F4 by silica gel column chromatography. F1 was found to be most effective in protecting RBC against AAPH radicals and in inhibiting lipid peroxidation. On the basis of thin-layer chromatography and UV and IR spectra analyses, all subfractions contained phenolic compounds. However, there was no correlation between the above antioxidant potency and total phenolic compounds estimated by using the Folin-Ciocalteau method.     

C-phycocyanin, a very potent and novel platelet aggregation inhibitor from Spirulina platensis

The aim of this study was to systematically examine the inhibitory mechanisms of C-phycocyanin (C-PC), one of the major phycobiliproteins of Spirulina platensis (a blue-green alga), in platelet activation. In this study, C-PC concentration-dependently (0.5-10 nM) inhibited platelet aggregation stimulated by agonists. C-PC (4 and 8 nM) inhibited intracellular Ca2+ mobilization and thromboxane A2 formation but not phosphoinositide breakdown stimulated by collagen (1 microg/mL) in human platelets. In addition, C-PC (4 and 8 nM) markedly increased levels of cyclic GMP and cyclic GMP-induced vasodilator-stimulated phosphoprotein (VASP) Ser(157) phosphorylation. Rapid phosphorylation of a platelet protein of Mw 47,000 (P47), a marker of protein kinase C activation, was triggered by phorbol-12,13-dibutyrate (150 nM). This phosphorylation was markedly inhibited by C-PC (4 and 8 nM). In addition, C-PC (4 and 8 nM) markedly reduced the electron spin resonance (ESR) signal intensity of hydroxyl radicals in collagen (1 microg/mL)-activated platelets. The present study reports on a novel and very potent (in nanomolar concentrations) antiplatelet agent, C-PC, which is involved in the following inhibitory pathways: (1) C-phycocyanin increases cyclic GMP/VASP Ser157 phosphorylation and subsequently inhibits protein kinase C activity, resulting in inhibition of both P47 phosphorylation and intracellular Ca2+ mobilization, and (2) C-PC may inhibit free radicals (such as hydroxyl radicals) released from activated platelets, which ultimately inhibits platelet aggregation. These results strongly indicate that C-PC appears to represent a novel and potential antiplatelet agent for treatment of arterial thromboembolism.     

An account of the attempted control of an introduced marine alga,Sargassum muticum,in Southern England

This paper details the attempted eradication and clearance programme for the introduced brown seaweed Sargassum muticum (Yendo) Fensholt.Since the introduction of this alga to British coastal waters in the early 1970s, populations have increased rapidly, causing a number of recreational and ecological problems. Various methods including handpicking, herbicides and biological control to combat the spread of the weed have been unsuccessful. Mechanical clearance, a costly alternative, has proved to be a viable proposition; the stages in the development of a working system are presented here.     

Influenza virus neuraminidase inhibitory activity of phlorotannins from the edible brown alga Ecklonia cava

Influenza A virus infections continue to pose a major threat to humans and several animal species. Neuraminidase (NA) is one of the most promising targets for the development of drugs against influenza viruses because of its critical role in the viral life cycle. During the course of a search for NA inhibitors from edible natural sources, we found that the ethyl acetate layer of ethanol extracts of Ecklonia cava showed extremely high NA-inhibitory activity (72.1% inhibition at 30 μg/mL). Bioactivity-guided fractionation of the ethyl acetate layer yielded five phlorotannins, identified as phloroglucinol (1), eckol (2), 7-phloroeckol (3), phlorofucofuroeckol (4), and dieckol (5). The inhibitory activities of these compounds (1-5) against NAs from group-1 (A/Bervig_Mission/1/18 [H1N1], A/PR/8/34 [H1N1]) and group-2 (A/Hong Kong/8/68 [H3N2], A/Chicken/Korea/MS96/96 [H9N2]) influenza A were evaluated to determine potencies and kinetic behavior. Analyses using various in vitro influenza A virus NA assays showed that all five phlorotannin derivatives were selective NA inhibitors. Of the phlorotannins, phlorofucofuroeckol (4) exhibited the most potent inhibitory activities toward group-1 NAs (IC?? values, 4.5 and 14.7 μM), whereas dieckol (5) potently inhibited group-2 NAs. Kinetic analyses indicated that compounds 1-5 were all noncompetitive. Notably, these noncompetitive inhibitors synergized with oseltamivir to enhance the NA-inhibitory effects of oseltamivir.     

Isolation and identification of 2-methoxy-3,5-dimethylpyrazine, a potent musty compound from wine corks

The compound responsible for a "fungal must" taint evident in industry assessments of wine corks was identified as 2-methoxy-3,5-dimethylpyrazine. The identification was made on the basis of gas chromatography/odor analyses, collection of material using micropreparative techniques, determination of chemical properties of collected material, and comparison by gas chromatography/mass spectrometry with an authentic sample, synthesized from 2-hydroxy-3,5-dimethylpyrazine. 2-Methoxy-3,5-dimethylpyrazine is an extremely potent compound with an unpleasant, musty, moldy aroma and an aroma threshold in a white wine of 2.1 ng/L. While its contribution to the frequency and intensity of cork taint in bottled wine is yet to be established, it has been assessed by some wine industry personnel as second only to 2,4,6-trichloroanisole as a cause of cork taint in Australian wine.     

Phenylboronic acid--a potent inhibitor of lipase from Oryza sativa

The kinetics of inhibition of rice bran lipase (RBL) by phenylboronic acid (PBA) was studied to elucidate the nature of inhibition and the effect of the inhibitor on the structure-function of RBL. The effectiveness of an inhibitor is normally expressed by the constant K(i), which is calculated from the Lineweaver-Burk plot and found to be 1.7 mM at pH 7.4. The kinetics of inhibition by PBA was competitive, indicating the presence of serine in the active site of the enzyme. The loss of activity of RBL was concentration dependent on the inhibitor (PBA), and the inactivation followed a pseudo-first-order kinetics. Fluorescence emission measurements indicated a decrease in the fluorescence emission intensity and a red shift in the emission maximum as the inhibitor concentration was increased. The inhibition of the enzyme by PBA was also confirmed by thermal denaturation measurements, which indicated a shift in the thermal denaturation temperature of the enzyme toward lower temperatures. The far-UV-CD data suggest that there were no significant changes in the conformation of the enzyme as a result of binding of PBA. These results indicate that PBA is a potential inhibitor of RBL and binds to the enzyme in bringing about inhibition without any structural alterations.     

Resveratrol is a potent inhibitor of the dioxygenase activity of lipoxygenase

Resveratrol is a naturally occurring phytoalexin, present in grapes and other food products, with important antioxidant properties. Although still under debate, it is generally assumed that resveratrol has protective effects against heart diseases and probably tumor development. Lipoxygenase is a dioxygenase with peroxidase activity involved in the synthesis of mediators in inflammatory, atherosclerotic, and carcinogenic processes. Lipoxygenase activity is also involved in the generation of flavors and aromas in foods from animal or vegetal sources. The results presented here show that resveratrol was a potent inhibitor of the dioxygenase activity of lipoxygenase, with an IC(50) = 13 microM. Simultaneously, resveratrol was oxidized by the peroxidase activity of lipoxygenase with a V(max) = 0.28 microM min(-1) and a k(M) = 16.6 microM. Furthermore, oxidized resveratrol was as efficient a lipoxygenase inhibitor as in its reduced form. From the data obtained it can be concluded that both resveratrol and its oxidized form can act as inhibitors of the dioxygenase activity of lipoxygenase. In contrast, the hydroperoxidase activity of lipoxygenase was not inhibited by resveratrol. These results suggest that resveratrol may be used as an antioxidant food additive and as a pharmacological agent to prevent the generation of eicosanoids involved in pathological processes.     

Characterization of Aloeride, a new high-molecular-weight polysaccharide from Aloe vera with potent immunostimulatory activity

We have characterized a new immunostimulatory polysaccharide called Aloeride from commercial aloe vera (Aloe barbadensis) juice. Aloeride is between 4 and 7 million Da, and its glycosyl components include glucose (37.2%), galactose (23.9%), mannose (19.5%), and arabinose (10.3%). At 0.5 microg/mL Aloeride increased NF-kappa B directed luciferase expression in THP-1 human monocytic cells to levels 50% of those achieved by maximal concentrations (10 microg/mL) of LPS. Aloeride induced the expression of the mRNAs encoding IL-1beta and TNF-alpha to levels equal to those observed in cells maximally activated by LPS. Acemannan, the major carbohydrate component from aloe, used at 200 microg/mL in the macrophage assay resulted in negligible NF-kappa B activation. Analysis of acemannan and Aloeride using size-exclusion chromatography suggests that the low activity of acemannan is due to trace amounts of Aloeride. Although Aloeride comprises only 0.015% of the aloe juice dry weight, its potency for macrophage activation accounts fully for the activity of the crude juice.     

Depolymerized products of lambda-carrageenan as a potent angiogenesis inhibitor

Since angiogenesis is involved in initiating and promoting several diseases such as cancer and cardiovascular events, this study was designed to evaluate the anti-angiogenesis of low-molecular-weight (LMW), highly sulfated lambda-carrageenan oligosaccharides (lambda-CO) obtained by carrageenan depolymerization, by CAM (chick chorioallantoic membrane) model and human umbilical vein endothelial cells (HUVECs). Significant inhibition of vessel growth was observed at 200 microg/pellet. A histochemistry assay also revealed a decrease of capillary plexus and connective tissue in lambda-CO treated samples. lambda-CO inhibited the viability of cells at the high concentration of 1 mg/mL, whereas it affected the cell survival slightly (>95%) at a low concentration (<250 microg/mL), and HUVEC is the most sensitive to lambda-CO among three kinds of cells. Furthermore, the inhibitory action of lambda-CO was also observed in the endothelial cell invasion and migration at relatively low concentration (150-300 microg/mL), through down-regulation of intracellular matrix metalloproteinases (MMP-2) expression on endothelial cells. Taken together, these findings demonstrate that lambda-CO is a potential angiogenesis inhibitor with combined effects of inhibiting invasion, migration, and proliferation.     

Isolation, characterization, and antioxidant activity of bromophenols of the marine red alga Rhodomela confervoides

A total of 19 naturally occurring bromophenols, with six new and 13 known structures, were isolated and identified from the methanolic extract of the marine red alga Rhodomela confervoides. The new compounds were identified by spectroscopic methods as 3,4-dibromo-5-((methylsulfonyl)methyl)benzene-1,2-diol (1), 3,4-dibromo-5-((2,3-dihydroxypropoxy)methyl)benzene-1,2-diol (2), 5-(aminomethyl)-3,4-dibromobenzene-1,2-diol (3), 2-(2,3-dibromo-4,5-dihydroxyphenyl)acetic acid (4), 2-methoxy-3-bromo-5-hydroxymethylphenol (5), and (E)-4-(2-bromo-4,5-dihydroxyphenyl)but-3-en-2-one (6). Each compound was evaluated for free radical scavenging activity against DPPH (α,α-diphenyl-β-dipicrylhydrazyl) and ABTS [2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)diammonium salt] radicals. Most of them exhibited potent activities stronger than or comparable to the positive controls butylated hydroxytoluene (BHT) and ascorbic acid. The results from this study suggest that R. confervoides is an excellent source of natural antioxidants, and inclusion of these antioxidant-rich algal components would likely help prevent the oxidative deterioration of food.     

Isolation and properties of a Kunitz-type protein inhibitor obtained from Pithecellobium dulce seeds

We report for the first time the isolation and characterization of a protease inhibitor from the seeds of Pithecellobium dulce, which is a Leguminosae tree native to Mexico. The purification of the P. dulce trypsin inhibitor (PDTI) was a direct process. After its extraction (pH 8.0) and precipitation (80% (NH(4))(2)SO(4)), the pH was adjusted to 4.0, the supernatant was loaded onto a CM-Sepharose column, and a single peak of trypsin inhibitory activity was eluted (CM-TIA). The main component of CM-TIA was PDTI, a protein composed of two polypeptide chains joined by disulfide bridge(s), with a pI of 4.95 and a molecular weight determined by electrospray mass spectrometry of 19 614 Da. The N-terminal sequence of PDTI has the highest similarity with the seed inhibitor of Acacia confusa. PDTI lacks chymotrypsin inhibitory activity. A low rate of cytotoxicity of CM-TIA toward RINm5F cells contrasted with a high rate of the active fraction G75-TIA (gel filtration chromatography; LC(50) of 0.04 mg/mL).     

Metal bioavailability and risk assessment from edible brown alga Laminaria japonica, using biomimetic digestion and absorption system and determination by ICP-MS

A new biomimetic digestion and absorption system, including in vitro bionic digestion and biomimetic membrane extraction, was used for the first time for the pretreatment of edible Laminaria japonica . After bionic digestion, 11 species of trace metals (V, Cr, Mn, Fe, Ni, Cu, Zn, Se, As, Cd, and Pb) in the resulting chyme were transformed into their final coordinated complexes and then absorbed by the biomembrane. Similar to the biomembrane between gastrointestinal tract and blood vessels, monolayer liposome was used for the first time as a biomembrane model. Affinity-monolayer liposome metals (AMLMs) were separated, determined by ICP-MS, and then used for the metal bioavailability assessment as the bioassimilated part. The action of gastrointestinal acidity and components (including digestive enzymes) was assessed on the basis of the concentration of AMLMs; the safe dosage and tolerable upper intake level of L. japonica for adults were proposed as 33.3 and 230.8 g/day, respectively.     

A specific and potent inhibitor of brassinosteroid biosynthesis possessing a dioxolane ring

Screening for brassinosteroid biosynthesis inhibitors was performed to find azole derivatives that induced dwarfism, to resemble brassinosteroid-deficient mutants in Arabidopsis, and which could be rescued by brassinosteroid. Through this screening experiment, propiconazole fungicide was selected as a likely inhibitor of brassinosteroid biosynthesis and, thus, propiconazole derivatives with optimized activity and selectivity were synthesized. The biological activity of these compounds was evaluated by examining cress stem elongation. Among the compounds tested, 2RS,4RS-1-[2-(4-trifluoromethylphenyl)-4-n-propyl-1,3-dioxolan-2-ylmethyl]-1H-1,2,4-triazole (12) showed the most potent capability to retard cress stem elongation in the light. The compound-induced hypocotyl dwarfism was restored by the coapplication of 10 nM brassinolide but not by 1 microM gibberellin. These results suggest that 12 should affect brassinosteroid biosynthesis. The potency and specificity of 12 were greater than those of brassinazole, a previously reported brassinosteroid biosynthesis inhibitor.     

Isolation of two anti-inflammatory and one pro-inflammatory polyunsaturated fatty acids from the brown seaweed Undaria pinnatifida

Two anti-inflammatory omega-3 polyunsaturated fatty acids (PUFAs) of stearidonic acid (SA) and eicosapentaenoic acid (EPA) and one pro-inflammatory omega-6 PUFA of arachidonic acid (AA) were isolated from the edible brown seaweed Undaria pinnatifida. SA was active against mouse ear inflammation induced by phorbol myristate acetate, with IC50 values of 160, 314, and 235 microg per ear for edema, erythema, and blood flow, respectively. EPA was also active against edema, erythema, and blood flow, with IC50 values of 230, 462, and 236 microg per ear, respectively. Although AA at low concentrations showed anti-inflammatory activities when measured 10 h later, AA doses of more than 243 microg per ear induced inflammatory symptoms 1 h later. Mature thalli generally had larger amounts of PUFAs than young thalli. The algal blade contained more omega-3 PUFAs than were found in other parts, while the holdfast contained extremely high amounts of AA. Late-season thalli showed increased amounts of PUFAs, especially AA.     

Emulsifying ability of porphyran prepared from dried nori, Porphyra yezoensis, a red alga

A suspension of low-quality dried nori processed from Porphyra yezoensis, a red alga, was autoclaved at 120 degrees C for 30 min, and from the supernatant, five preparations of porphyran of differing molecular masses and chemical compositions were obtained by preprecipitation with ethanol at stepwise-increasing concentrations of 50 and 67% followed by size-exclusion chromatography. The porphyran preparations exhibited a high emulsifying activity index and high emulsion stability over a wide range of pH and temperature and also in the presence of sodium chloride. An adequately high coefficient of correlation between the median diameter of oil droplets and their 3,6-anhydrogalactose content suggests that 3,6-anhydrogalactose could take part in emulsification with porphyran.     

Isolation and characterization of enzymes involved in lysine catabolism from sorghum seeds

Lysine is an essential amino acid synthesized in plants via the aspartic acid pathway. The catabolism of lysine is performed by the action of two consecutive enzymes, lysine 2-oxoglutarate reductase (LOR, EC 1.5.1.8) and saccharopine dehydrogenase (SDH, EC 1.5.1.9). The final soluble lysine concentration in cereal seeds is controlled by both synthesis and catabolism rates. The production and characterization of high-lysine plants species depends on knowledge of the regulatory aspects of lysine metabolism and manipulation of the key enzymes. We have for the first time isolated, partially purified, and characterized LOR and SDH from developing sorghum seeds, which exhibited low levels of activity. LOR and SDH were only located in the endosperm and were very unstable during the isolation and purification procedures. LOR and SDH exhibited some distinct properties when compared to the enzymes isolated from other plant species, including a low salt concentration required to elute the enzymes during anion-exchange chromatography and the presence of multimeric forms with distinct molecular masses.     

Seasonal variations of bromophenols in brown algae (Padina arborescens,Sargassum siliquastrum,and Lobophora variegata) collected in Hong Kong

Distributions and seasonal variations of the key seafood flavor compounds including 2-bromophenol, 4-bromophenol, 2,4-dibromophenol, 2,6-dibromophenol, and 2,4,6-tribromophenol in three species of brown algae (Padina arborescens, Sargassum siliquastrum, and Lobophora variegata) found in Hong Kong waters were investigated. Bromophenols were extracted by simultaneous steam distillation and solvent extraction apparatus and analyzed by gas chromatography-mass spectrometry. On a dried weight basis, the total bromophenol content (TBC) determined varied widely with seasons (from 40.9 to 7030 ng/g). The TBCs detected were higher in winter and lower in summer. Except for 2-bromophenol, the rest of the bromophenols were detected in all of the algal samples. The TBC of L. variegata was generally the highest among all of the algae collected. Relatively high concentrations of bromophenols in algae supported the fact that marine algae were major producers of bromophenols in the marine environment.