异枝麒麟菜人工养殖技术

为获得一种长心卡帕藻多糖的最佳提取工艺并测定抗过敏活性,该研究在单因素实验的基础上,以长心卡帕藻的多糖得率作为考察指标,选择料液比、超声波预处理时间、提取时间进行响应面优化实验,确定最佳工艺条件,并通过DEAE-52纤维素层析柱对提取的粗多糖进行纯化。随后利用RBL-2H3细胞模型,分别测定纯化后的长心卡帕藻多糖对RBL-2H3细胞活力、细胞脱颗粒抑制及细胞组胺释放水平调节的影响。结果表明,响应面优化后的长心卡帕藻多糖的最佳提取工艺如下：料液比（w/v）为1:92 g/mL,超声波预处理时间为40 min,提取时间为4 h,最佳提取工艺条件下,长心卡帕藻多糖提取率为13.92%。经DEAE-52纤维素层析纯化后的组分KSP（Kappaphycus alvarezii Sulfated Polysaccharides）在50～1200 μg/mL浓度范围内对RBL-2H3细胞活力均未表现有显著性抑制作用,无细胞毒性;浓度为25、50及100 μg/mL的KSP对经2,4-二硝基苯基白蛋白偶联物（2,4-Dinitrophenyl-Bovine Serum Albumin,DNP-BSA）刺激后的RBL-2H3细胞的β-氨基己糖苷酶酶活抑制率分别为36.3%、37.0%和51.7%,对其组胺释放抑制率分别为17.5%、18.3%和42.0%。综上,本试验得到的工艺参数具有可行性,KSP对RBL-2H3细胞无明显毒性,并表现出显著的脱颗粒和组胺的释放抑制作用,具有较好的体外抗过敏活性。该研究可为以长心卡帕藻为原料开发具有抗过敏活性的营养功能性食品提供参考。     

Anti-allergic activity of grapeseed extract (GSE) on RBL-2H3 mast cells

Grapeseed extract (GSE) is a rich source of natural phenolic compounds and possesses various pharmacological activities, including antioxidant, anticarcinogenic and anti-inflammatory properties. However, effects of GSE on immunoglobulin (Ig) E-mediated allergic responses still remain elusive. In the present study, the effects of GSE on activation and degranulation of RBL-2H3 mast cells were investigated. GSE pretreatments (20-100 μg/ml) reduced IgE-antigen mediated release of β-hexosaminidase and histamine in RBL-2H3 cells. Additionally, GSE reversibly inhibited expression of FcεRI on RBL-2H3 cells. GSE treatments caused a significant elevation of intracellular cAMP levels, whereas the Ca²⁺ influx upon antigen stimulation was inhibited. Suppression on FcεRI expression together with decreased calcium uptake and increased cAMP level might be involved in attenuated degranulation of mast cells by GSE treatment. Our results suggest a possible pharmaceutical application of GSE in treating type I allergic diseases.     

Inhibitory effect of aqueous spinach extract on degranulation of RBL-2H3 cells

The inhibitory effect of an aqueous extract from spinach on degranulation of RBL-2H3 cells is herein reported. The extract significantly suppressed antigen-induced degranulation in a dose-dependent manner without affecting cell viability. Active substances in the extract were heat-stable and trypsin-resistant with molecular weights ranging from 500 Da to 14 kDa. The extract inhibited elevation of the intracellular Ca²⁺ concentration caused by stimulation by antigen, while not suppressing degranulation induced by a calcium ionophore A23187. Immunoblot analysis revealed that the inhibitory effect results from downregulation of phosphorylation of both Syk kinase and phosphatidylinositol 3-kinase in the signalling pathways involved in degranulation caused by the antigen–antibody interaction. Taken together, these findings suggest that aqueous spinach extract has an anti-allergic activity that controls degranulation.     

Inhibition of fish oil oxidation and the radical scavenging activity of New Zealand seaweed extracts

This study describes the potential use of New Zealand seaweed extracts as antioxidants in fish oils. The extracts from two brown seaweeds (Ecklonia radiata, Macrocystis pyrifera) and two red species (Champia sp. and Porphyra sp.), were added to hoki (Macruronus novaezelandiae) oil and assessed by production of oxidation products in an elevated temperature (60 °C) storage trial. Antioxidant assays ORAC and DPPH were conducted to elucidate the radical scavenging activity of the seaweed extracts. The results showed that all the lipid-soluble seaweed extracts had some antioxidative capacity, with brown algae generally outperforming red algae species. Extracts from E. radiata performed best with significantly lower primary, secondary and total oxidation products, and higher DPPH radical scavenging ability than the commercial antioxidant, BHT. This study demonstrated the potential of brown algae seaweed extracts, in particular E. radiata, for use as antioxidants in fish oil products.     

Inhibitory effects of water extracts of fermented rice bran on allergic response

In this study, the effects of water extracts of fermented rice bran (FRBE) on immediate-type allergic reactions in vitro (RBL-2H3 cell line) and in vivo [ovalbumin (OVA) sensitized mice model] were investigated. FRBE inhibited β-hexosaminidase release from RBL-2H3 cells activated by anti-dinitrophenyl (DNP) IgE while FRBE pretreatment profoundly inhibited the systemic allergic reaction induced by compound 48/80. Moreover, FRBE had a significant inhibitory effect on the mRNA expression of anti-DNP IgE-induced tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-4, IL-6, and IL-10, all of which mediate inflammatory reactions triggered by allergic reaction. To determine the mRNA levels of cytokines in an OVA-sensitized mice model after FRBE treatment, spleen cells were harvested with several concentrations of FRBE. The FRBE-treated group showed decreased levels of TNF-α, IFN-γ, IL-6, and IL-10. These results suggest that FRBE suppressed allergic and inflammatory reactions through inhibition of degranulation, histamine release, and pro-inflammatory cytokine production from mast cells.     

Inhibitory effect of zinc on beta-hexosaminidase release from RBL-2H3 cells by synthetic chemicals

The effects of foods and chemicals related to food hygiene on degranulation were evaluated using a method for assaying the enzyme activity of beta-hexosaminidase as an index of chemical mediator release from RBL-2H3 cells in vitro. Using a previously developed assay system, we had found a large number of inhibitors and promoters of degranulation of RBL-2H3 cells. In the present study, we examined the inhibitory effect of zinc chloride on the degranulation (beta-hexosaminidase release) from RBL-2H3 cells with or without antigen in the presence of the degranulation-promotive chemicals, namely, 4 food additives, 7 pesticides and 2 veterinary drugs. These promotive chemicals were classified into two types on the basis of inhibitory profile by zinc chloride: 1) those which showed marked degranulation-inhibitory action when the cells were stimulated with antigen, such as butylhydroxyanisole, dibutylhydroxytoluene, EPN, cis- and trans-permethrin, prothiofos, pyridaben, terbufos, 2) those which showed marked degranulation-inhibitory action whether the cells were stimulated with antigen or not, such as butyl p-hydroxybenzoate, o-phenylphenol, bitertanol, salinomycin. In conclusion, zinc had a dramatic inhibitory effect on enhanced degranulation induced by synthetic chemicals in vitro.     

The fucoidans from brown algae of Far-Eastern seas: Anti-tumor activity and structure–function relationship

The sulfated polysaccharides from brown algae – the fucoidans – are known to be a topic of numerous studies, due to their beneficial biological activities including anti-tumour activity. In this study the effect of fucoidans isolated from brown algae Saccharina cichorioides, Fucus evanescens, and Undaria pinnatifida on the proliferation, neoplastic transformation, and colony formation of mouse epidermal cells JB6 Cl41, human colon cancer DLD-1, breast cancer T-47D, and melanoma RPMI-7951 cell lines was investigated. The algal fucoidans specifically and markedly suppressed the proliferation of human cancer cells with less cytotoxic effects against normal mouse epidermal cells. The highly sulfated (1 → 3)-α-l-fucan from S. cichorioides was found to be vitally important in the inhibition of EGF-induced neoplastic transformation of JB6 Cl41 cells. In colony formation assay the fucoidans from different species of brown algae showed selective anti-tumour activity against different types of cancer, which depended on unique structures of the investigated polysaccharides. These results provide evidence for further exploring the use of the fucoidans from S. cichorioides, F. evanescens, and U. pinnatifida as novel chemotherapeutics against different types of cancer.     

Inhibitory effects of flavonoid glycosides isolated from the peel of Japanese persimmon (Diospyros kaki Fuyu) on antigen-stimulated degranulation in rat basophilic leukaemia RBL-2H3 cells

We found that two distinct flavonoid glycosides isolated from the peel of Japanese persimmon (Diospyros kaki Fuyu), isoquercitrin (Isq) and hyperin (Hyp), are capable of inhibiting antigen-stimulated degranulation in rat basophilic leukaemia RBL-2H3 cells. In order to elucidate the underlying mechanisms, we examined effects of Isq and Hyp on cellular responses induced by antigen stimulation. Treatment with both Isq and Hyp markedly inhibited antigen-stimulated elevation of intracellular free Ca²⁺ concentration and reactive oxygen species (ROS). Isq and Hyp did not affect NADPH oxidase (NOX) activity, but they possessed DPPH radical-scavenging activity similar to that of epigallocatechin gallate, a potent anti-oxidant, Finally, Isq and Hyp showed little or no effects on Ag-stimulated Syk activation or phosphorylation of signalling molecules. These results indicate that inhibition of antigen-stimulated degranulation by Isq and Hyp is mainly due to suppression of intracellular Ca²⁺ elevation, which is caused by direct scavenging of ROS that are generated by NOX. Our findings suggest that Isq and Hyp, isolated from the peel of persimmon, would be beneficial for alleviating symptoms of type I allergy.     

Varieties of antioxidant and antibacterial properties of Ecklonia stolonifera and Ecklonia kurome products harvested and processed in the Noto peninsula,Japan

Ecklonia stolonifera and Ecklonia kurome are traditional edible brown algae in the fisheries towns in Far East Asia. In the Noto Peninsula area, Ishikawa, Japan, both the Ecklonia are called ‘kajime’ and people believe that the algae improve the property of blood. To determine the varieties of antioxidant and antibacterial properties E. stolonifera and E. kurome products, assays for total phenolic content and antioxidant activities, including DPPH radical-scavenging activity, superoxide anion radical-generated by non-enzymatic system, ferrous-reducing power and WST-8, a tetrazolium salt, redox activity of four dried and two boiled E. stolonifera, and four dried and two raw E. kurome preparations were tested in this study. Furthermore, antibacterial activity of the products was tested. Though the total phenolic content, the antioxidant activity and the antibacterial activities of E. stolorifera and E. kurome products were high, these properties were varied by manufacturers or each product. Especially, two dried and two boiled products of E. stolonifera showed low activities. The results of this study indicate that the contents of functional compounds and their activities were affected or decreased greatly by the processing method.     

Minerals,polysaccharides and antioxidant properties of aqueous solutions obtained from macroalgal beach-casts in the Noto Peninsula,Ishikawa, Japan

To clarify the useful properties of the macroalgal beach-casts for food, we determined mineral, water-soluble polysaccharide and total phenolic compound contents and antioxidant properties of aqueous solutions obtained from frond of eight brown and four green algae. These algae were washed ashore from the East Sea (the Sea of Japan) to the northeast beaches of the Noto Peninsula in autumn. There were fresh and large amounts of Ecklonia stolonifera (Es), Ecklonia kurome (Ek), Sargassum ringgoldianum subsp. coreanum (Sr) and Sargassum macrocarpum on the beaches. Potassium, magnesium and calcium ions were high in Sr-solution. High molecular weight water-soluble polysaccharide, mainly alginates, and relative viscosity were also high in Sr-solution. Both Es and Ek solutions showed high contents of total phenolic compounds and high antioxidant activities, including DPPH radical- and hydroxyl radical-scavenging and ferrous reducing power. On the other hand, the highest superoxide anion radical-scavenging activity was in the Sr solution. In the case of Sr, the same antioxidant properties were also found in the aqueous solution obtained from stem. These results suggest that some macroalgal beach-casts, especially Sr, can be utilised as new natural resources for functional foods, cosmetics, medical applications and fertilizer instead of being processed to landfill or incineration.     

Effective extraction of carotenoids from brown seaweeds and vegetable leaves with edible oils

Factors affecting carotenoid extraction with edible oils were examined using edible brown seaweed, Sargassum horneri, as main sample. The results indicate that drying was essential to extract fucoxanthin (Fx) from S. horneri and physical (boiling) and chemical (acid/alkali) pretreatment of the wet sample increased the extraction rate of Fx. Additionally, more Fx was found from the dried S. horneri powder with a smaller particle size. The extraction rate of Fx is affected by the extraction temperature and time, showing that the effective extraction would be obtained at 50 °C within 12 hr extraction. Among the oils used, short-chain (C4 and C6) triacylglycerol (TAG) (SCT) and medium-chain (C8) TAG (MCT) could extract more Fx from S. horneri and more β-carotene and lutein from spinach and olive leaves. The relatively lower viscosity of SCT and MCT would be the most likely reason for the higher extraction rates of both TAGs.     

Anti-allergy effect of marc from silver vine Actinidia polygama prepared by supercritical fluid extraction

Anti-allergic effect of marc extract after supercritical fluid extraction SFE comparing the crude extract from silver vine was investigated to increase its potential for use as a new source of functional food that reduced non-favorable flavor. The results tested by thiazolyl blue tetrazolium bromide MTT assay indicated that the ethanol extract of silver vine ETOH and ethanol extract of SFE marc SFEM showed no cytotoxic effects on IgE-sensitized rat basophilic leukemia RBL-2H3 cells under the conditions. ETOH, as well as SFEM inhibited antigen-induced degranulation in RBL-2H3 mast cells dose dependently. The morphological change of the cells treated with compound 48/80 was prevented by the extracts. Production of tumor necrosis factor TNF-α on the activated RBL-2H3 cells was also reduced by ETOH and SFEM, similarly. Moreover, the total phenolic content of the extracts seems to correlate with the anti-allergic activity. In conclusion, the results suggest that the flavor-improved SFE marc extract suppressed the antigen-induced allergic reactions in RBL-2H3 mast cells.     

Bioactivities of low-grade green coffee and spent coffee in different in vitro model systems

Methanolic extracts of low-grade green coffee beans (LCB) and spent coffee were analysed for radical-scavenging activity (α,α-diphenyl-β-picrylhydrazyl radical) and oxygen radical absorbance capacity (ORAC). The extracts were also evaluated for anti-tumour (P388 cell assay), anti-inflammatory (J774A.1 cell assay) and anti-allergenic (RBL-2H3 cell line) activities in vitro. LCB extract was found to exhibit a radical-scavenging activity of 92.0% followed by spent Arabica (86.9%) and spent Robusta (82.0%) at a concentration of 50 ppm. The antioxidant activity of LCB extract, measured as Trolox equivalents (4416 μM/g) was significantly (p < 0.05) higher than that of the spent coffee extracts. However, extracts of spent coffee exhibited significantly (p < 0.05) more anti-tumour activity than the LCB extract in terms of cell viability. This could be due to the possible role of brown pigments (melanoidins and phenolic polymers), formed during roasting, which may protect cells from oxidative damage in the biological system. However, both the extracts of LCB and spent coffee showed limited anti-inflammatory and anti-allergic activities. The presence of phenolics and chlorogenic acids in appreciable quantities along with brown pigments makes these coffee by-products a source for natural antioxidants.     

Antimetastatic activity of polyphenol-rich extract of Ecklonia cava through the inhibition of the Akt pathway in A549 human lung cancer cells

An ethyl acetate extract (ECE) of a brown alga, Ecklonia cava, was examined for its anti-metastatic effect, using A549 human lung carcinoma cells. ECE treatment significantly suppressed the migration and invasion of A549 cells in a concentration-dependent manner. It also strongly down-regulated the matrix metalloproteinase (MMP)-2 activity of the cancer cells by gelatin zymography assay. For elucidating its mechanism of action in cancer cell metastasis, ECE was further investigated for various cell signalling pathways, including JNK, ERK, p38, and Akt. In this, ECE showed an anti-metastatic effect in a concentration- and time-dependent manner by the mechanism of suppression of Akt and p38, but not JNK and ERK. These results, for the first time, suggest that ECE (a polyphenol-enriched, highly anti-oxidative fraction of brown alga, E. cava) may have therapeutic potential in metastatic lung cancer, based on its strong inhibitory effects on the migration and invasiveness of A549 human lung adenocarcinoma cells.     

Purification of novel anti-inflammatory peptides from enzymatic hydrolysate of the edible microalgal Spirulina maxima

Natural bioactive peptides have been known as ingredients in nutraceuticals and pharmaceuticals. In this study, the potential anti-inflammatory peptides were purified from enzymatic hydrolysate of the edible microalgae Spirulina maxima using gastrointestinal endopeptidases (trypsin, α-chymotrypsin, and pepsin). Purification of the hydrolysate resulted in two peptides with amino acid sequences of LDAVNR (P1, 686 Da) and MMLDF (P2, 655 Da). It was found that peptides P1 and P2 exhibited significant inhibition on the release and production of histamine from antigen-stimulated RBL-2H3 mast cells. Moreover, the suppressive effect of P1 and P2 on production as well as expression of interleukin-8 in histamine-stimulated EA.hy926 endothelial cells was determined. Notably, the production of intracellular reactive oxygen species from mast cells and endothelial cells was decreased in the presence of P1 or P2. Collectively, the peptides P1 and P2 from S. maxima could be used as functional ingredients with potent anti-inflammatory benefits.     

Antibacterial,anti-inflammatory and anti-allergic activities of standardised pomegranate rind extract

Antibacterial, anti-inflammatory and anti-allergic activities of standardised pomegranate rind extract (SPRE) containing 13% w/w ellagic acid were studied in vitro. The antibacterial activity of SPRE was determined using the disc diffusion and broth microdilution methods. SPRE exhibited a potent bacteriostatic effect against Propionibacterium acnes, a Gram-positive anaerobe, with a MIC of 15.6 μg/ml, and Gram-positive facultative anaerobic bacteria, Staphylococcus aureus and Staphylococcus epidermidis, with MICs of 7.8–15.6 μg/ml. Anti-inflammatory activity of SPRE was evaluated by measuring the inhibition of nitric oxide (NO) production by murine macrophage-like RAW264.7 cells. SPRE exhibited a potent NO inhibitory effect, with an IC₅₀ of 10.7 μg/ml. Evaluation of the anti-allergic activity showed that SPRE inhibited the release of β-hexosaminidase from antigen-stimulated rat basophilic leukemia (RBL-2H3) cells with an IC₅₀ of 20.9 μg/ml. In addition, SPRE exhibited only moderate cytotoxicity on human keratinocyte cells, with CC₅₀ of 33.6 μg/ml. These findings support the potential use of SPRE as a nutraceutical for antibacterial, anti-inflammatory and anti-allergic proposes.     

Novel anticoagulant compound from fermented red alga <Emphasis Type="Italic">Pachymeniopsis elliptica</Emphasis>

Natural fermentation was tested as a method of releasing active compounds during screening for potential anticoagulant activity in three types of algae (Pachymeniopsis elliptica, Sargassum horneri, and Ulva pertusa). Freeze dried algae samples (2.5 g) were fermented by adding 75 g of sugar and 500 mL of water and thereafter kept at room temperature (25 °C) for 3 months. Activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT) were measured every 2 weeks for 3 months to determine the optimum time for the highest activity. Fermented P. elliptica, (which had the highest activity) was subjected to anion exchange chromatography (DEAE-cellulose) and sepharose 4B gel permeation chromatography. The purified sample was analyzed by agarose-gel electrophoresis and polyacrylamide gel electrophoresis (PAGE) to confirm the purification and to determine the molecular mass, respectively. The 360 μg/mL of purified compound (Mwt > 500,000 Da) had both APTT and PT activities (>1,000 s). However, at the concentrations of 180 μg/mL, purified compound and heparin showed 540 and >1,000 s APTT activity, respectively. Though, the purified compound of P. elliptica considered as a weaker anticoagulant than heparin, this purified anticoagulant polysaccharide could be considered as a good alternative source as an anticoagulant. Moreover, the technique of fermentation is an inexpensive and feasible, this purified anticoagulant polysaccharide compound could be used in pharmaceutical and biomedical industry. Further investigations need to be performed to determine the mechanism of this novel anticoagulant compound. The authors Prashani Mudika Ekanayake and Chamilani Nikapitiya contributed equally to this work.     

基于RBL-2H3模型分析过敏反应效应细胞激活的差异基因

构建IgE介导的大鼠嗜碱性粒细胞（rat basophilic leukemia，RBL）-2H3作为过敏反应细胞模型，并利用转录组测序分析比较RBL-2H3细胞激活前后的转录组基因差异，富集相关信号通路。IgE介导的RBL-2H3细胞激活后，过敏介质和细胞因子分泌显著提高；转录组测序得到RBL-2H3细胞激活前后的232 个差异表达基因，其中有127 个（54.74%）可进行基因功能注释；RBL-2H3细胞激活主要影响肿瘤坏死因子、丝裂原活化蛋白激酶、Janus激酶-信号转导与转录激活子、Toll样受体等信号通路，涉及的转录因子有MAP3K8、Nfkbia、Junb、Jun、Fos等。本研究建立了IgE介导的过敏反应体外细胞模型，并且通过对效应细胞转录组信息的全面分析和潜在靶点及信号通路的富集为过敏性疾病的防控治疗提供参考。