Novel Antioxidant Peptides Identified from Arthrospira platensis Hydrolysates Prepared by a Marine Bacterium Pseudoalteromonas sp. JS4-1 Extracellular Protease.

Crude enzymes produced by a marine bacterium Pseudoalteromonas sp. JS4-1 were used to hydrolyze phycobiliprotein. Enzymatic productions showed good performance on DPPH radical and hydroxyl radical scavenging activities (45.14 ± 0.43% and 65.11 ± 2.64%, respectively), especially small peptides with MWCO <3 kDa. Small peptides were fractioned to four fractions using size-exclusion chromatography and the second fraction (F2) had the highest activity in hydroxyl radical scavenging ability (62.61 ± 5.80%). The fraction F1 and F2 both exhibited good antioxidant activities in oxidative stress models in HUVECs and HaCaT cells. Among them, F2 could upregulate the activities of SOD and GSH-Px and reduce the lipid peroxidation degree to scavenge the ROS to protect Caenorhabditis elegans under adversity. Then, 25 peptides total were identified from F2 by LC-MS/MS, and the peptide with the new sequence of INSSDVQGKY as the most significant component was synthetized and the ORAC assay and cellular ROS scavenging assay both illustrated its excellent antioxidant property.

Purification, Characterization, and Application for Preparation of Antioxidant Peptides of Extracellular Protease from Pseudoalteromonas sp. H2.

The study reported on the isolation of a metalloprotease named EH2 from Pseudoalteromonas sp. H2. EH2 maintained more than 80% activity over a wide pH range of 5-10, and the stability was also nearly independent of pH. Over 65% activity was detected at a wide temperature range of 20-70 °C. The high stability of the protease in the presence of different surfactants and oxidizing agents was also observed. Moreover, we also investigated the antioxidant activities of the hydrolysates generated from porcine and salmon skin collagen by EH2. The results showed that salmon skin collagen hydrolysates demonstrated higher DPPH (1,1-diphenyl-2-picrylhydrazyl) (42.88% ± 1.85) and hydroxyl radical (61.83% ± 3.05) scavenging activity than porcine skin collagen. For oxygen radical absorbance capacity, the hydrolysates from porcine skin collagen had higher efficiency (7.72 ± 0.13 µmol·TE/µmol). Even 1 nM mixed peptides could effectively reduce the levels of intracellular reactive oxygen species. The two types of substrates exerted the best antioxidant activity when hydrolyzed for 3 h. The hydrolysis time and type of substrate exerted important effects on the antioxidant properties of hydrolysates. The hydrolyzed peptides from meat collagens by proteases have good antioxidant activity, which may have implications for the potential application of marine proteases in the biocatalysis industry.

A Method for Detecting Antioxidant Activity of Antioxidants by Utilizing Oxidative Damage of Pigment Protein.

A simple and effective method for detecting the antioxidant activity by utilizing oxidative damage of pigment proteins was developed. In this method, phycocyanin and bovine hemoglobin pigment proteins were used as substrates attacked by free radicals; AAPH was used as a free radical initiator; and Trolox as a positive control; and the fermentation products of Lactobacillus plantarum 793, phycocyanin hydrolysates, salmon skin collagen hydrolysates, and synthetic peptides PMRGGYHY and FCVLRP are antioxidants inspected in this study. Because of being attacked by free radicals, the absorbance of the pigment proteins at their characteristic absorption peak changes with time. By recording the time-varying curve at the characteristic absorption peak of the pigment protein in the blank/negative control sample, the Trolox positive control sample, and the samples of inspected antioxidants, the antioxidant activity could be calculated by using the specific equation. The linear detection ranges of Trolox in the phycocyanin assay and the bovine hemoglobin assay were 1-4 µM and 4-24 µM, respectively. Compared with the ORAC assay, the antioxidant activities of the samples measured by this method were slightly lower. The method proposed in this study reflects the protective effects of inspected antioxidants on pigment proteins, which could potentially serve as new biomarkers of oxidative damage processes.

New insights into the structure-activity relationships of antioxidative peptide PMRGGGGYHY.

The sequence and structure of antioxidant peptides play fundamental roles in their antioxidant functions. However, the structural mechanism of antioxidant peptides is still unclear. In this study, we used quantum calculations to reveal the antioxidant mechanism of the peptide PMRGGGGYHY. PMRGGGGYHY has multiple antioxidant active sites, and two tyrosine residues were determined to be the major active sites. Based on the structure-activity relationships of PMRGGGGYHY, the antioxidant activity of the modified peptide significantly improved by 4.8-fold to 9.73 ± 0.61 µmol TE/µmol. In addition, the removal of glycine residues from PMRGGGGYHY would increase the energy of the HOMOs and simplify the hydrogen bonding network, causing a significant increase in antioxidant activity. The intracellular ROS scavenging ability gradually decreased with decreasing glycine content. This same peptide has very different effects in vitro versus as a cellular antioxidant. This paper provides new insights into the structural mechanism and rational design/modification of novel antioxidant peptides.

Antidiabetic Function of Lactobacillus fermentum MF423-Fermented Rice Bran and Its Effect on Gut Microbiota Structure in Type 2 Diabetic Mice.

Rice bran is an industrial byproduct that exerts several bioactivities despite its limited bioavailability. In this study, rice bran fermented with Lactobacillus fermentum MF423 (FLRB) had enhanced antidiabetic effects both in vitro and in vivo. FLRB could increase glucose consumption and decrease lipid accumulation in insulin resistant HepG2 cells. Eight weeks of FLRB treatment significantly reduced the levels of blood glucose and lipids and elevated antioxidant activity in type 2 diabetic mellitus (T2DM) mice. H&E staining revealed alleviation of overt lesions in the livers of FLRB-treated mice. Moreover, high-throughput sequencing showed notable variation in the composition of gut microbiota in FLRB-treated mice, especially for short-chain fatty acids (SCFAs)-producing bacteria such as Dubosiella and Lactobacillus. In conclusion, our results suggested that rice bran fermentation products can modulate the intestinal microbiota and improve T2DM-related biochemical abnormalities, so they can be applied as potential probiotics or dietary supplements.

Impact of Lactobacillus plantarum 423 fermentation on the antioxidant activity and flavor properties of rice bran and wheat bran.

Rice bran (RB) and wheat bran (WB) fermented with L. plantarum 423 had enhanced odor intensity, especially for sulfides and aromatics. The hydroxyl radical-scavenging activity (73.28 ± 3.18%) and oxygen radical-scavenging activity (2.12 ± 0.08 mmol·TE/g) of RB fermentation broth were better than those of WB fermentation broth. Even at 2 µg/ml, the purified antioxidant fractions from the WB fermentation broth showed strong intracellular ROS-scavenging activity in human umbilical vein endothelial cells (HUVECs), and the purified antioxidant fractions (200 µg/ml) from the RB fermentation broth had a good antiaging effect. The dominant antioxidant components in the RB and WB fermentation broths were acids (70.21%) and ketones (10.64%), these components jointly give the RB and WB fermentation broths a variety of antioxidant properties. These results are beneficial for developing RB and WB deep-processing technology and laid the foundation for the preparation of antioxidant fractions with L. plantarum 423.

Comparison of the diversity of cultured and total bacterial communities in marine sediment using culture-dependent and sequencing methods.

Despite recent great advances in microbial culture, most microbes have not yet been cultured, and the impact of medium composition on the isolation of microbes from natural systems has not been elucidated. To optimize media for culturing marine microbes, microbial communities in three sediment samples were described using high-throughput sequencing (HTS) and culture-dependent techniques. HTS revealed communities dominated by Gammaproteobacteria, and culture-based methods revealed communities dominated by Actinobacteria. Among the total operational taxonomic units (OTUs) from the HTS dataset, 6% were recovered in the culture collection. Four potentially novel bacterial strains belonging to Oceaniovalibus, Psychrobacter and Salegentibacter were isolated. The combination of media cultured more taxa than any single medium. Nutrient-rich and single-carbon/nitrogen-source media supported the growth of relatively few taxa, and the quality of nitrogen strongly influenced the types of bacteria isolated.

Genome Sequencing of Mesonia algae K4-1 Reveals Its Adaptation to the Arctic Ocean.

The special ecological environment of the Arctic has brought about a large number of salt-tolerant and psychrotolerant microorganisms. We isolated two culturable bacterial strains of the genus Mesonia; one from the Arctic ocean, Mesonia algae K4-1, and one from the tropical sea, Mesonia sp. HuA40. Our genome analysis and phenotypic experiments indicated that Mesonia algae K4-1 is a moderately halophilic and psychrophilic bacterium. Mesonia algae K4-1 can tolerate 3-14% NaCl and grow at a wide range of temperatures from 4 to 50°C. Mesonia sp. HuA40 is a mesophilic bacterium that can only grow with 3-9% NaCl. In addition, the salt adaptation strategy of Mesonia algae K4-1 accumulates organic osmolytes in the cell. RNA helicases, glutathione and organic compatible solutes may play important roles in maintaining the metabolism and physiological function of Mesonia algae K4-1 under cold stress. Moreover, the ability of Mesonia algae K4-1 to adapt to an oligotrophic marine environment is likely due to the synthesis of a large number of extracellular polysaccharides and the secretion of various families of extracellular proteases. This study systematically analyzed the relationship between genomic differentiation and environmental factors of the Mesonia genus and revealed the possible adaptation mechanism of Mesonia algae K4-1 in the extreme Arctic marine environment at the genomic level.

Overexpression of SmLEA enhances salt and drought tolerance in Escherichia coli and Salvia miltiorrhiza.

Salinity and drought are important abiotic stresses limiting plant growth and development. Late embryogenesis abundant (LEA) proteins are a group of proteins associated with tolerance to water-related stress. We previously cloned an LEA gene, SmLEA, from Salvia miltiorrhiza Bunge. Phylogenetic analysis indicated that SmLEA belongs to Group LEA14, which is involved in the dehydration response. To determine its function in detail, we have now overexpressed SmLEA in Escherichia coli and S. miltiorrhiza. The logarithmic increase in accumulations of SmLEA proteins in E. coli occurred earlier under salinity than under standard conditions. SmLEA-transformed S. miltiorrhiza plants also showed faster root elongation and a lower malondialdehyde concentration than the empty vector control plants did when cultured on MS media supplemented with 60 mM NaCl or 150 mM mannitol. Moreover, SmLEA-overexpressing transgenics experienced a less rapid rate of water loss. Under either salinity or drought, overexpressing plants had greater superoxide dismutase activity and a higher glutathione concentration. These results suggest that SmLEA may be useful in efforts to improve drought and salinity tolerance in S. miltiorrhiza. Our data also provide a good foundation for further studies into the stress resistance mechanism and molecular breeding of this valuable medicinal plant.