Antioxidant, Antibacterial Properties of Novel Peptide CP by Enzymatic Hydrolysis of Chromis notata By-Products and Its Efficacy on Atopic Dermatitis.

This study investigated the antioxidant, antimicrobial, and anti-atopic dermatitis (AD) effects of a novel peptide (CP) derived from a Chromis notata by-product hydrolysate. Alcalase, Flavourzyme, Neutrase, and Protamex enzymes were used to hydrolyze the C. notata by-product protein, and the 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical-scavenging activity was measured. Alcalase hydrolysate exhibited the highest ABTS radical-scavenging activity, leading to the selection of Alcalase for further purification. The CHAO-1-I fraction, with the highest ABTS activity, was isolated and further purified, resulting in the identification of the peptide CP with the amino acid sequence Ala-Gln-Val-Met-Lys-Leu-Pro-His-Arg-Met-Gln-His-Ser-Gln-Ser. CP demonstrated antimicrobial activity against Staphylococcus aureus, inhibiting its growth. In a 2,4-dinitrochlorobenzene (DNCB)-induced AD-like skin model in mice, CP significantly alleviated skin lesions, reduced epidermal and dermal thickness, and inhibited mast cell infiltration. Moreover, CP suppressed the elevated levels of interleukin-6 (IL-6) in the plasma of DNCB-induced mice. These findings highlight the potential of CP as a therapeutic agent for AD and suggest a novel application of this C. notata by-product in the fish processing industry.

Fermented Asterina pectinifera with Cordyceps militaris Mycelia Induced Apoptosis in B16F10 Melanoma Cells.

This prime objective of this study was to explore the anti-cancer activity of fermented Asterina pectinifera with Cordyceps militaris mycelia (FACM) in B16F10 murine melanoma cells. The effect of FACM on cell viability was assessed using MTT assay. Furthermore, the effect of FACM was compared with unfermented A. pectinifera on cell viability. The results demonstrated that the fermented FACM extract has a higher inhibitory activity on the proliferation of B16F10 murine melanoma cells than unfermented A. pectinifera. In addition, FACM also promoted the expression of pro-apoptotic protein Bax leading to stimulate apoptosis in B16F10 cells. Therefore the present study demonstrates that the FACM might be a potential effective anti-cancer agent, as a result of its stronger anti-proliferative effect and apoptosis inducing effect than A. pectinifera or C. militaris on melanoma cells.

Antioxidant Activity of Extract from the Cephalothorax of Fenneropenaeus chinensis.

We investigated the antioxidant activity of taurine rich water extract from the cephalothorax of Fenneropenaeus chinensis (FCC). The antioxidant potency of water extract from FCC was assessed using various assay methods, such as DPPH (1,1-diphenyl-2-picrylhydrazyl), alkyl radical scavenging activity, ABTS (2,2'-azinobis (3-ethylbenzothiazoline 6-sulfonic acid ammonium salt)) radical scavenging activity and Ferric reducing antioxidant power (FRAP) assay. The DPPH and alkyl radical scavenging activities of FCC were dose-dependently increased. The lipid peroxidation was estimated using ferric thiocyanate (FTC) assay and thiobarbituric acid (TBA) methods. However, a higher lipid peroxidation activity was observed in TBA method than FTC method. The results of the present study suggested that the FCC extract potentially scavenged the free radical and reduced oxidative stress. Therefore, the present study is concluded that the FCC extract could be a potential source of antioxidant activity.

Antioxidant Effect of Taurine-Rich Paroctopus dofleini Extracts Through Inhibiting ROS Production Against LPS-Induced Oxidative Stress In Vitro and In Vivo Model.

Taurine is an essential amino acid to improve the function of cardiovascular, skeletal muscle, retina, and central nervous system. It also plays a role as an antioxidant agent against reactive oxygen species (ROS) generated by various substances. The aim of the current study was to examine the antioxidant capacity of water extracts of Paroctopus dofleini. Radical scavenging activity of P. dofleini extracts was performed using an ESR spectrophotometer. Protective effects of P. dofleini extracts against lipopolysaccharide (LPS)-induced oxidative stress in RAW264.7 cells were evaluated using flow cytometry. The P. dofleini extracts showed a potent antioxidant activity against LPS-induced oxidative stress on RAW264.7 cells. Furthermore, the in vivo antioxidant activity of P. dofleini extract on LPS-induced oxidative stress was assessed using zebrafish embryos. P. dofleini successfully scavenged the LPS-induced intracellular ROS and prevented lipid peroxidation in zebrafish embryos. The results obtained in this study clearly demonstrate that the P. dofleini significantly scavenge the ROS and prevent lipid peroxidation in both in vitro and in vivo models.

Taurine Attenuates Doxorubicin-Induced Toxicity on B16F10 Cells.

This study aimed to investigate the effect of doxorubicin co-treatment with taurine on B16F10 melanoma cells. Frequently, Doxorubicin is used in the treatments of many different kinds of cancers, some of which are soft tissue sarcomas, hematological malignancies and carcinomas. However, the clinical application of doxorubicin is compromised by its severe adverse effects, including cardiotoxicity. In the present study, the efficacy of doxorubicin co-treatment with taurine was investigated. B16F10 cell viability was evaluated using MTT assays, trypan blue dye exclusion assays, and fluorescent staining technique. Apoptotic cells were detected by flow cytometry and the proteins associated with apoptosis and cellular differentiations were assessed by immunoblotting. Doxorubicin inhibited cell growth and induced cell death in B16F10 cells. Interestingly, doxorubicin co-treatment with taurine inhibited apoptosis in B16F10 cells. These results indicate that doxorubicin co-treatment with taurine attenuates doxorubicin-induced cytotoxicity and reduces ROS production in B16F10 cells.

Neuroprotective Effect of Taurine-Rich Cuttlefish (Sepia officinalis) Extract Against Hydrogen Peroxide-Induced Oxidative Stress in SH-SY5Y Cells.

Oxidative stress mediates the cell damage in several neurodegenerative diseases, some of which are Alzheimer's disease (AD), multiple sclerosis and Parkinson's disease (PD). In this study, we investigated whether the taurine-rich cuttlefish extract could exert a protective effect on damaged human neuroblastoma SH-SY5Y cells induced by hydrogen peroxide (H2O2). Our results revealed that pre-treatment with cuttlefish extract effectively increased the cell viability by protecting the cells from intracellular reactive oxygen species (ROS) induced by H2O2 exposure. Furthermore, apoptosis related proteins Bcl-2 and Bax were investigated by western-blot analysis and results indicated that cuttlefish extract promoted the expression of anti-apoptotic Bcl-2 protein while inhibiting the expression of pro-apoptotic Bax protein. Therefore, cuttlefish extract containing the ability of scavenging excessive ROS, the capacity of anti-oxidative stress, could be employed in neurodegenerative disease prevention. In conclusion, the results suggest that cuttlefish extract could be used as a potential candidate for preventing several human neurodegenerative and other disorders caused by oxidative stress.

Biosynthesis of Oligomeric Anthocyanins from Grape Skin Extracts.

We synthesized oligomeric anthocyanins from grape skin-derived monomeric anthocyanins such as anthocyanidin and proanthocyanidin by a fermentation technique using Aspergillus niger, crude enzymes and glucosidase. The biosyntheses of the oligomeric anthocyanins carried out by the conventional method using Aspergillus niger and crude enzymes were confirmed by ESI-MS. The molecular weight of the synthesized anthocyanin oligomers was determined using MALDI-MS. The yield of anthocyanin oligomers using crude enzymes was higher than that of the synthesis using Aspergillus fermentation. Several studies have been demonstrated that oligomeric anthocyanins have higher antioxidant activity than monomeric anthocyanins. Fermentation-based synthesis of oligomeric anthocyanins is an alternative way of producing useful anthocyanins that could support the food industry.

Trapa japonica Pericarp Extract Reduces LPS-Induced Inflammation in Macrophages and Acute Lung Injury in Mice.

In this study, we found that chloroform fraction (CF) from TJP ethanolic extract inhibited lipopolysaccharide (LPS)-induced production of nitric oxide (NO) and intracellular ROS in RAW264.7 cells. In addition, expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) genes was reduced, as evidenced by western blot. Our results indicate that CF exerts anti-inflammatory effects by down-regulating expression of iNOS and COX-2 genes through inhibition of MAPK (ERK, JNK and p38) and NF-κB signaling. Similarly we also evaluated the effects of CF on LPS-induced acute lung injury. Male Balb/c mice were pretreated with dexamethasone or CF 1 h before intranasal instillation of LPS. Eight hours after LPS administration, the inflammatory cells in the bronchoalveolar lavage fluid (BALF) were determined. The results indicated that CF inhibited LPS-induced TNF-α and IL-6 production in a dose dependent manner. It was also observed that CF attenuated LPS-induced lung histopathologic changes. In conclusion, these data demonstrate that the protective effect of CF on LPS-induced acute lung injury (ALI) in mice might relate to the suppression of excessive inflammatory responses in lung tissue. Thus, it can be suggested that CF might be a potential therapeutic agent for ALI.

Antioxidant and Anti-Inflammatory Effects of Chaenomeles sinensis Leaf Extracts on LPS-Stimulated RAW 264.7 Cells.

The fruit of Chaenomeles sinensis has been traditionally used in ethnomedicine for the treatment of various human ailments, including pneumonia, bronchitis, and so on, but the pharmacological applications of the leaf part of the plant have not been studied. In this study, we evaluated the various radical scavenging activities and anti-inflammatory effects of different Chaenomeles sinensis leaf (CSL) extracts. The water extract showed a higher antioxidant and radical scavenging activities. However the ethanolic extracts showed higher NO scavenging activity than water extract, therefore the ethanolic extract of CSL was examined for anti-inflammatory effects on lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. The 70% ethanol extract of CSL (CSLE) has higher anti-inflammatory activity and significantly inhibited the production of nitric oxide (NO), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). In addition, CSLE suppressed LPS-stimulated inducible nitric oxide synthase (iNOS) and NO production, IL-1ß and phospho-STAT1 expression. In this study, we investigated the effect of CSLE on the production of inflammatory mediators through the inhibition of the TRIF-dependent pathways. Furthermore, we evaluated the role of CSLE on LPS-induced expression of pro-inflammatory cytokines, such as TNF-α, IL-1ß and IL-6. Our results suggest that CSLE attenuates the LPS-stimulated inflammatory responses in macrophages through regulating the key inflammatory mechanisms, providing scientific support for its traditional uses in treating various inflammatory diseases.

First evidence that Ecklonia cava-derived dieckol attenuates MCF-7 human breast carcinoma cell migration.

We investigated the effect of Ecklonia cava (E. cava)-derived dieckol on movement behavior and the expression of migration-related genes in MCF-7 human breast cancer cell. Phlorotannins (e.g., dieckol, 6,6'-biecko, and 2,7″-phloroglucinol-6,6'-bieckol) were purified from E. cava by using centrifugal partition chromatography. Among the phlorotannins, we found that dieckol inhibited breast cancer cell the most and was selected for further study. Radius™-well was used to assess cell migration, and dieckol (1-100 µM) was found to suppress breast cancer cell movement. Metastasis-related gene expressions were evaluated by RT-PCR and Western blot analysis. In addition, dieckol inhibited the expression of migration-related genes such as matrix metalloproteinase (MMP)-9 and vascular endothelial growth factor (VEGF). On the other hand, it stimulated the expression of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2. These results suggest that dieckol exerts anti-breast cancer activity via the regulation of the expressions of metastasis-related genes, and this is the first report on the anti-breast cancer effect of dieckol.

Anti-inflammatory effects of extract from Haliotis discus hannai fermented with Cordyceps militaris mycelia in RAW264.7 macrophages through TRIF-dependent signaling pathway.

In this study, Haliotis discus hannai (H. discus hannai) fermentation was attempted with Cordyceps militaris (C. militaris) mycelia using a solid culture. We tried to ferment H. discus hannai to determine the optimal conditions fermentation with regards to its anti-inflammatory effects. The extracts of H. discus hannai fermented with C. militaris mycelia (HFCM-5) showed higher nitric oxide inhibitory effects than H. discus hannai and C. militaris alone. HFCM-5 also decreased pro-inflammatory cytokines, TNF-α and IL-6 in a dose-dependent manner. HFCM-5 did not affect the MyD88-dependent pathway, but decreased phosphorylation of IRF3 and STAT1 which are involved in TRIF-dependent pathway. Taken together, our results suggest that HFCM-5 exerts its anti-inflammatory effects via TRIF signaling pathway and could potentially be used as a functional food in the regulation of inflammation.

Attenuation of inflammatory-mediated neurotoxicity by Saururus chinensis extract in LPS-induced BV-2 microglia cells via regulation of NF-κB signaling and anti-oxidant properties.

BACKGROUND: A Saururus chinensis Baill (SC) has been used by Native Americans, early colonists and practitioners of Korean traditional medicine for treating several diseases including cancer, rheumatoid arthritis and edema. The objective of this study was to evaluate the effects of SC extract in lipopolysaccharide (LPS)-stimulated neuroinflammatory responses in BV-2 microglial cells. METHODS: The effects of SC on the LPS-induced neuroinflammatory responses in BV-2 microglial cells were assessed by Western blotting, RT-PCR and immunofluorescence labeling techniques. DPPH and alkyl radical scavenging assay was performed to evaluate the anti-oxidant effects. Comparisons between groups were analyzed using one-way analysis of variance followed by Dunnett's multiple comparisons test using GraphPad Prism V5.01 software. RESULTS: Pre-treatment with SC extract (1, 5 and 10 µg/mL) significantly (p < 0.001 at 10 µg/mL) and concentration dependently inhibited LPS-induced production of nitric oxide (NO), inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2) and suppressed the inflammatory cytokine levels such as tumor necrosis factor-alpha and interleukin (IL)-6 in BV-2 microglial cells (p < 0.001 at 10 µg/mL). Further, SC suppressed the nuclear factor-kappa B (NF-κB) activation by blocking the degradation of IκB-α. SC also exhibited profound anti-oxidant effects by scavenging 1, 1-diphenyl-2-picrylhydrazyl (DPPH) (IC50: 0.055 mg/mL) and alkyl radicals (IC50: 0.349 mg/mL). High performance liquid chromatography finger printing analysis of SC revealed quercetin (QCT) as one of the major constituents compared with reference standard. QCT also inhibited the excessive release of NO, and inhibited the increased expressional levels of IL-6, iNOS and COX-2 in LPS-stimulated BV-2 cells. CONCLUSIONS: Our results indicated that SC inhibited the LPS-stimulated neuroinflammatory responses in BV-2 microglia via regulation of NF-κB signaling. The antioxidant active constituents of SC might be partly involved in delivering such effects. Based on the traditional claims and our present results SC can be potentially used in treating inflammatory-mediated neurodegenerative diseases.

Sulfated chitosan oligosaccharides suppress LPS-induced NO production via JNK and NF-κB inactivation.

Various biological effects have been reported for sulfated chitosan oligosaccharides, but the molecular mechanisms of action of their anti-inflammatory effects are still unknown. This study aimed to evaluate the anti-inflammatory effects of sulfated chitosan oligosaccharides and to elucidate the possible mechanisms of action. The results showed that pretreated low molecular weight sulfated chitosan oligosaccharides inhibited the production of nitric oxide (NO) and inflammatory cytokines such as IL-6 and TNF-α in lipopolysaccharide (LPS)-activated RAW264.7 cells. The sulfated chitosan oligosaccharides also suppressed inducible nitric oxide synthase (iNOS), phosphorylation of JNK and translocation of p65, a subunit of NF-κB, into the nucleus by inhibiting degradation of IκB-α. Our investigation suggests sulfated chitosan oligosaccharides inhibit IL-6/TNF-α in LPS-induced macrophages, regulated by mitogen-activated protein kinases (MAPKs) pathways dependent on NF-κB activation.

Purification of a novel peptide derived from Mytilus coruscus and in vitro/in vivo evaluation of its bioactive properties.

Excess oxidant can promote inflammatory responses. Moreover, chronic inflammation accompanied by oxidative stress is connected various steps involved in many diseases. From the aspect, we investigated an antioxidant peptide to prevent inflammatory response against oxidant overexpression. To prepare the peptide, eight proteases were employed for enzymatic hydrolysis, and the antioxidant properties of the hydrolysates were investigated using free radical scavenging activity by electron spin resonance (ESR) spectrometry. Papain hydrolysates, which showed clearly superior free radical scavenging activity, were further purified using consecutive chromatographic methods. Finally, a novel antioxidant peptide was obtained, and the sequence was identified as Ser-Leu-Pro-Ile-Gly-Leu-Met-Ile-Ala-Met at N-terminal. Oral administration of the peptide to mice effectively inhibited malondialdehyde (MDA) levels in a thiobarbituric acid reactive substances (TBARS) assay, and we also confirmed the antioxidative enzyme activities in superoxide dismutase (SOD) and glutathione-s-transferase (GST) assays. This is the first report of an antioxidant peptide derived from the hydrolysate of Mytilus coruscus, and also these results suggest that the peptide possesses potent antioxidant activity, and potential to enhance anti-inflammatory response.

Purification of a novel nitric oxide inhibitory peptide derived from enzymatic hydrolysates of Mytilus coruscus.

Shellfish contain significant levels of high quality protein and are therefore a potential source for biofunctional high-value peptides. To purify a novel anti-inflammatory peptide from Mytilus coruscus (M. coruscus), we applied enzymatic hydrolysis and tangential flow filtration (TFF) and investigated its nitric oxide inhibitory property. To prepare the peptide, eight proteases were employed for enzymatic hydrolysis. Flavouzyme hydrolysates, which showed clearly superior nitric oxide inhibitory activity on lipopolysaccharide (LPS)-stimulated RAW264.7, were further purified using a TFF system and consecutive chromatographic methods. Finally, a novel anti-inflammatory peptide composed of 10 amino acid residues was obtained, and the sequence was identified as Gly-Val-Ser-Leu-Leu-Gln-Gln-Phe-Phe-Leu at N-terminal position. The peptide from M. coruscus effectively inhibited nitric oxide production on macrophage cells. This is the first report of an anti-inflammatory peptide derived from the hydrolysates of M. coruscus.

Chitooligosaccharides induce apoptosis in human myeloid leukemia HL-60 cells.

In this study we propose a novel anticancer agent using hetero-chitooligosaccharide (hetero-COS). To examine the possibility of the hetero-COS as a anticancer agent, we prepared nine kinds of hetero-COS with relatively higher molecular weights (90, 75 and 50-COS I, 5-10kDa), medium molecular weights (90, 75 and 50-COS II, 1-5kDa), and lower molecular weights (90, 75 and 50-III, below 1kDa), and their anticancer properties were investigated on HL-60 cells using flow cytometry and morphological analysis. The results obtained indicate that 90-COS III, which is relatively higher degree of deacetylation and lower molecular weights, showed the highest anticancer activity, and the data showed the anticancer property of the hetero-COSs depended on their degree of deacetylation values and molecular weight.

Purification and characterization of a novel peptide with inhibitory effects on colitis induced mice by dextran sulfate sodium from enzymatic hydrolysates of Crassostrea gigas.

The anti-inflammatory activity of purified peptides from Crassostrea gigas (C. gigas) hydrolysates was studied. To prepare hydrolysates from C. gigas, we used eight different proteinases and the anti-inflammatory activities were determined using a nitric oxide (NO) assay in RAW264.7 cells. Among the hydrolysates, Protamex hydrolysates showed the highest anti-inflammatory activity. We separated and purified the total hydrolysate using an ultrafiltration membrane system and consecutive chromatographic methods. Finally, we obtained a peptide with the following sequence: Gln-Cys-Gln-Cys-Ala-Val-Glu-Gly-Gly-Leu at N-terminal position. The anti-inflammatory peptide purified from C. gigas inhibited NO production by 72.2% compared to the lipopolysaccharide (LPS) treated group. In addition, the Protamex hydrolysates from C. gigas showed decreased serum IgE levels and increased spleen CD4(+)/CD8(+) levels on dextran sulfate sodium (DSS) induced colitis in mice. These results suggest the peptide and hydrolysate from C. gigas possess potent anti-inflammatory effect.