Structural and functional properties of soluble Antarctic krill proteins covalently modified by rutin.

New polyphenol-protein conjugates were successfully prepared by covalently crosslinking soluble Antarctic krill proteins with rutin (SAKPs-rutin). The physico-chemical and functional properties of SAKPs-rutin conjugates were systematically evaluated by measuring the changes in interfacial tension, structural conformation, and emulsifying ability, etc. The results showed that SAKPs-rutin conjugates possessed higher surface hydrophobicity, surface charge, and thermal denaturation temperature, and lower ß-sheet conformation compared to native SAKPs. On this basis, the interfacial tension of SAKPs-rutin conjugates was reduced, which greatly contributed to the formation of denser and more ordered networks at the oil-water interface. Meanwhile, the emulsifier endowed the fabricated high internal phase emulsions (HIPEs) with excellent physical performance and oxidative stability, evidenced by low peroxide values (POV) and malondialdehyde (MDA) after the treatment of long-term storage (15d), heating (65 °C) and UV light treatment. These findings suggest that SAKPs-rutin conjugates are a novel and promising food resource for preparing food-grade emulsions.

Fenton reaction-assisted photodynamic inactivation of calcined melamine sponge against Salmonella and its application.

Photodynamic inactivation (PDI) is an effective alternative to traditional antibiotics to broadly kill bacteria. This study aimed to develop a potent PDI system by coupling calcinated melamine sponges (CMSs) with the Fenton reaction. The results showed that CMS calcined at 350 ℃ was successfully carbonized with intact and porous structures, and it possessed excellent hydrophilicity and photothermal conversion performance. When Fe2+ was added and internalized, the Fenton reaction in which Fe2+ reacted with H2O2 in cells occurred to produce reactive oxygen species (ROS) (OH, OOH, etc.) and O2, and notably, the O2 molecules could serve as a raw material to absorb the photothermal energy of CMS to generate highly reactive 1O2. Under synergistic effects, CMS-350 coupled with Fe2+ potently inactivated > 6 Log CFU/mL (>99.9999%) of Salmonella under 201.6 J/cm2 blue LED illumination by destroying Na+/K+-ATPase and Ca2+/Mg2+-ATPase, DNA synthesis-related enzymes, cell membranes, etc. Meanwhile, the composite photocatalyst was proven to be nontoxic and could inactivate Salmonella in various foods, including vegetables (Brassica chinensis L), eggs and fresh cucumber juice. As a result, CMS coupled with the Fenton reaction greatly improves the inactivation potency of PDI against harmful bacteria.

Anti-tyrosinase, antioxidant and antibacterial activities of gallic acid-benzylidenehydrazine hybrids and their application in preservation of fresh-cut apples and shrimps.

A series of gallic acid-benzylidenehydrazine hybrids were synthesized and evaluated for their tyrosinase inhibitory activity. Thereinto, compounds 5d and 5f potently inhibited tyrosinase with IC50 of 15.3 and 3.3 µM, compared to kojic acid (44.4 µM). The inhibition mechanism suggested that 5d and 5f not only chelated with Cu2+, but also reduced Cu2+ to Cu1+ in the tyrosinase active site. Additionally, 5d and 5f exhibited strong DPPH scavenging and antibacterial activities against Vibrio parahaemolyticu and Staphylococcus aureus, which can be attributed to the function of gallic acid and hydrazone moiety. These compounds also exhibited capacity to preserve fresh-cut apples and shrimps. Finally, 5d and 5f exhibited low cytotoxic activity in a human cell line (HEK293). Therefore, these compounds possess anti-tyrosinase, antioxidant, and antibacterial activities, and can be used in the development of novel food preservatives.

A systematic review of synthetic tyrosinase inhibitors and their structure-activity relationship.

Tyrosinase is a copper-containing oxidation enzyme, which is responsible for the production of melanin. This enzyme is widely distributed in microorganisms, animals and plants, and plays an essential role in undesirable browning of fruits and vegetables, antibiotic resistance, skin pigment formation, sclerotization of cuticle, neurodegeneration, etc. Hence, it has been recognized as a therapeutic target for the development of antibrowning agents, antibacterial agents, skin-whitening agents, insecticides, and other therapeutic agents. With great potential application in food, agricultural, cosmetic and pharmaceutical industries, a large number of synthetic tyrosinase inhibitors have been widely reported in recent years. In this review, we systematically summarized the advances of synthetic tyrosinase inhibitors in the literatures, including their inhibitory activity, cytotoxicity, structure-activity relationship (SAR), inhibition kinetics, and interaction mechanisms with the enzyme. The collected information is expected to provide a rational guidance and effective strategy to develop novel, potent and safe tyrosinase inhibitors for better practical applications in the future.

Application of the curcumin-mediated photodynamic inactivation for preserving the storage quality of salmon contaminated with L. monocytogenes.

The effect of curcumin-mediated blue light-emitting diode (LED) photodynamic inactivation (PDI) for preserving the quality of salmon contaminated with Listeria monocytogenes was investigated by microbiological, physical, chemical and histological methods during sample storage at 4 â„ƒ and 25 â„ƒ. The results showed that PDI decelerated the proliferation of L. monocytogenes on salmon during storage at 25 â„ƒ, with the maximum inhibition reaching 4.0 log10 CFU/g (99.99%), compared to the negative control. Moreover, PDI greatly retarded the increase in pH (P < 0.05) and the production of TVB-N, retarded the accumulation of free fatty acids, and decelerated the degradation of proteins, ultimately preserving the high nutritional value of the salmon. In addition, PDI effectively prevented a change in colour and retarded the loss of water from the salmon, thereby conserving its texture and sensory properties. Therefore, PDI is a promising and valid non-thermal technology to use for fish preservation.

Effects of ultrasound-assisted basic electrolyzed water (BEW) extraction on structural and functional properties of Antarctic krill (Euphausia superba) proteins.

A novel protein extraction method of ultrasound-assisted basic electrolyzed water (BEW) was proposed, and its effects on the structural and functional properties of Antarctic krill proteins were investigated. Results showed that BEW reduced 30.9% (w/w) NaOH consumption for the extraction of krill proteins, and its negative redox potential (-800 ~ -900 mV) protected the active groups (carbonyl, free sulfhydryl, etc.) of the proteins from oxidation compared to deionized water (DW). Moreover, the ultrasound-assisted BEW increased the extraction yield (9.4%), improved the solubility (8.5%), reduced the particle size (57 nm), favored the transition of α-helix and ß-turn to ß-sheet, promoted the surface hydrophobicity and disulfide bonds formation of krill proteins when compared to BEW without ultrasound. These changes contributed to the enhanced foam capacity, foam stability and emulsifying capacity of the krill proteins. Notably, all the physicochemical, structural and functional properties of the krill proteins were comparable to those extracted by the traditional ultrasound-assisted DW. This study suggests that the ultrasound-assisted BEW can be a potential candidate to extract proteins, especially offering an alternative way to produce marine proteins with high nutritional quality.

Synthesis, antioxidant and anti-tyrosinase activity of 1,2,4-triazole hydrazones as antibrowning agents.

A series of 1,2,4-triazole hydrazones (1-16) were synthesized, and their inhibitory activities and mechanisms on tyrosinase were investigated by ultraviolet spectrophotometry, fluorescence quenching, molecular docking study, etc. Most of compounds possessed potent tyrosinase inhibitory activity. Thereinto, compound 9 presented the superior activity with IC50 of 0.9 µM, which was markedly lower than the standard kojic acid (IC50 = 64.1 µM). Compound 9 not only interacted with copper ions in the active center of the enzyme but also bound to the enzyme-substrate complex, indicating that it was a competitive-noncompetitive mixed inhibitor. Additionally, it also displayed potent DPPH scavenging activity. Antibrowning test showed that compound 9 effectively reduced the enzymatic browning of fresh-cut potatoes. Furthermore, compound 9 exhibited low cytotoxic activity against human normal cell line with IC50 of 49.9 µM. Overall, the present study suggests that these compounds may serve as lead molecules for developing novel antibrowning agents in food industry.

Characterization of recombinant rice quiescin sulfhydryl oxidase and its improvement effect on wheat flour-processing quality.

In this study, recombinant rice quiescin sulfhydryl oxidase (rQSOX) was expressed and characterized, and its performance in flour-processing quality was further evaluated. The purified rQSOX exhibited the highest sulfhydryl oxidation activity (1.96 IU/mg) using dithiothreitol as a substrate, accompanying the production of H2O2. The optimal temperature and pH were 60 °C and pH 8.0 for rQSOX catalyzing oxidation of dithiothreitol. And rQSOX retained 50% of its maximum activity after incubation at 80 °C for 1 h. Moreover, rQSOX supplementation improved the farinograph properties of dough, indicated by the increased dough stability time and decreased degree of softening, and enhanced viscoelastic properties of the dough. Addition of rQSOX (10 IU/g flour) provided remarkable improvement in specific volume (37%) and springiness (17%) of the steamed bread, and significantly reduced the hardness by half, which was attributed to the strengthened gluten network. The results provide an understanding for rQSOX using in flour-processing industry.

Recovery and identification bioactive peptides from protein isolate of Spirulina platensis and their in vitro effectiveness against oxidative stress-induced erythrocyte hemolysis.

BACKGROUND: Spirulina platensis is recognized as one of the most nutritious foods, containing a high protein content of up to 70%. Meanwhile, he interest in using natural protein resources to develop bioactive peptides is steadily increasing. Therefore, this study released the bioactive peptides from S. platensis by enzymatic hydrolysis using pepsin (1:3000 U g-1 ), and their amino acid sequences were determined by de novo sequencing. On this basis, the antioxidant activities of synthesized bioactive peptides were comprehensively evaluated by 2,2'-azinobis-3-ethylbenzothiazolin-6-sulfonic acid assay, 1,1-diphenyl-2-picryhydrazyl assay, and cell hemolysis assay induced by 2,2'-azobis-(2-amidino-propane) dihydrochloride (AAPH). RESULTS: The degree of hydrolysis and recovery percentage of pepsin hydrolysis were 172 and 825 g kg-1 respectively, and FFEFF (P1: m/z 736.4, 8%), EYFDALA (P2: m/z 828.4, relative intensity 18.5%), and VTAPAASVAL (P3: m/z 899.5, relative intensity 17.3%) were purified and identified. P2 possessed an excellent radical scavenging activity compared with P1, P3, and vitamin C, which was contributed to by its high ß-sheet conformation and specific amino acid compositions. Moreover, P2 significantly attenuated AAPH-induced oxidative hemolysis of erythrocytes and protected the erythrocytes, because it reduced the formation of malondialdehyde and increased the enzyme activities of superoxide dismutase, catalase, and glutathione peroxidase in erythrocytes. CONCLUSION: This study provided insights into the potential antioxidant function of the synthesized peptides originated from the bioactive peptides of S. platensis proteins, which would contribute to the development of natural antioxidant from new protein resources. © 2020 Society of Chemical Industry.

Dissecting the Disulfide Linkage of the N-Terminal Domain of HMW 1Dx5 and Its Contributions to Dough Functionality.

The N-terminal domain of HMW-GS 1Dx5 (1Dx5-N) contains three cysteine residues (Cys10, Cys25, Cys40), which are the basis of gluten network formation through disulfide bonds. Disulfide linkage in 1Dx5-N was dissected by site-directed mutagenesis and LC-MS/MS, and its contributions to structural and conformational stability of 1Dx5-N and dough functionality were investigated by circular dichroism, intrinsic fluorescence, surface hydrophobicity determination, size exclusion chromatography, nonreducing/reducing SDS-PAGE, atomic force microscopy, and farinographic analysis. Results showed that Cys10 and Cys40 of 1Dx5-N were the active sites for intermolecular linkage. Meanwhile, Cys40 also exhibited the ability to form intrachain disulfide linkage with Cys25. Moreover, Cys10 and Cys40 played a functionally important role in maintaining the structural and conformational stability and high surface hydrophobicity of the N-terminal domain of HMW-GS, which in turn facilitated the formation of HMW polymers and massive disulfide linkage of HMW-GS through hydrophobic interaction. Additionally, the 1Dx5-N mutants in which Cys were replaced by serine (Ser) presented different effects on dough functionality, while only the C25S mutant produced positive effects compared with wild type 1Dx5-N. Na2CO3-induced ß-elimination of cystine might occur in glutenin without heating, which would make it much easier to reduce the nutritional quality of flour products by the cost of lysine. Therefore, these results give a deep understanding of the disulfide linkage of the N-terminal domain of HMW-GS and its functional importance, which will provide a practical guide to effectively generate a superior HMW-GS allele by artificial mutagenesis.

Role of N-terminal domain of HMW 1Dx5 in the functional and structural properties of wheat dough.

Effects of N-terminal domain of high molecular weight glutenin subunit (HMW-GS) 1Dx5 (1Dx5-N) on functional and structural properties of wheat dough were determined by farinographic and rheological analysis, size exclusion chromatography, non-reducing/reducing SDS-PAGE, total free sulfhydryl determination, scanning electron microscopy and Fourier transform infrared spectroscopy. Results showed that 1Dx5-N improved the quality of dough with the increased water absorption, dough stability time, elastic and viscous modulus, and the decreased degree of softening, loss tangent. These improvements could be attributed to the formation of the macro-molecular weight aggregates and massive protein networks, which were favored by 1Dx5-N through disulfide bonds and hydrophobic interactions. Additionally, 1Dx5-N drove the transition of α-helix and random coil conformations to ß-sheet and ß-turn conformations, further demonstrating the formation of HMW-GS polymers and the enhancement of dough strength. Moreover, all the positive effects of 1Dx5-N were reinforced by edible salt NaCl.

Anti-inflammatory, antioxidant and antitumor activities of ingredients of Curcuma phaeocaulis Val.

Curcuma phaeocaulis Val. is used in Chinese Pharmacopoeia as health food and folk medicine for removing blood stasis, alleviating pain and tumor therapy. This research was aimed to explore and compare three main bioactivities including anti-oxidant, antitumor and anti-inflammatory activities between the ethanol extract of C. Phaeocaulis and its fractions using different in vitro models. Firstly, 70 % ethanol was used to extract C. Phaeocaulis, and then the crude extract was re-extracted, resulting in petroleum ether (EZ-PE), ethyl acetate (EZ-EA), and water fractions (EZ-W), respectively, and then a series of index was detected. Results showed that all the extracts had medium DPPH radical scavenging activity when the concentration was 200 µg/ml and their DPPH radical scavenging activity was in a concentration-dependent manner. The extracts except ethanol extract of C. Phaeocaulis had almost no cytotoxicity to the survival of RAW264.7 cell when the concentration reached 80 µg/ml, and all of them had medium inhibitory effect on nitrite release. Extracts of C. Phaeocaulis had medium intensity antitumor activity, EZ-PE and EZ-EA fractions significantly inhibited the proliferation of four tumor cells (SMMC-7721 cell lines, HepG-2 cell lines, A549 cell lines and Hela cell lines). C. Phaeocaulis had antioxidant and anti-inflammatory activities, which did not carry out centralized phenomenon when re-extracted. EZ-PE and EZ-EA were active antitumor sites of C. Phaeocaulis.

The soluble recombinant N-terminal domain of HMW 1Dx5 and its aggregation behavior.

This study seeks to clarify and determine the fundamental properties of N-terminal domain of high molecular weight glutenin subunits (HMW-GS) 1Dx5 (1Dx5-N). 1Dx5-N was expressed in E. coli and its solubility was measured by spectrophotometry. Effects of edible salts (NaCl, Na2CO3), disulfide bond reductant dithiothreitol (DTT) and hydrophobic interactions of denaturant sodium dodecyl sulfonate (SDS) on 1Dx5-N polymer were investigated by native polyacrylamide gelelectrophoresis (PAGE), nonreducing/reducing SDS-PAGE, intrinsic fluorescence, size exclusion chromatography (SEC), dynamic light scattering (DLS) and circular dichroism (CD). Results showed that 1Dx5-N formed a soluble aggregate in aqueous solutions by native-PAGE, clarifying the role of N-terminal of HMW-GS in the insolubility of the whole subunits. Meanwhile, the hydrophobic interaction was more potent in promoting the aggregation of 1Dx5-N in aqueous solutions from the results of SEC, DLS and CD. Edible salts, NaCl and Na2CO3, could improve the polymer formation of 1Dx5-N through disulfide bonds. Moreover, Na2CO3 at high concentrations (>200mM) greatly favored polymer formation by disulfide bonds, and it induced other types of cross-links between amino acids in 1Dx5-N according to nonreducing/reducing SDS-PAGE and fluorescence spectrum. Moreover, the formation of covalent bonds was reinforced by hydrophobic interactions between 1Dx5-N. Therefore, these results provide much novel information on the N-terminal domain of HMW-GS to facilitate the understanding of its functional properties in wheat flour.

Isolation and identification of ingredients inducing cancer cell death from the seeds of Alpinia galanga, a Chinese spice.

This study was carried out to isolate ingredients from the seeds of a Chinese spice (Alpinia galangal) and to evaluate their cytotoxic activity on cancer cell lines. Isolation and purification of the phytochemical constituents were conducted using silica gel, Sephadex LH-20 and ODS columns. After extraction using 95% ethanol, the total extracts were re-extracted, resulting in petroleum ether (PE), ethyl acetate (EA) and water fractions, respectively. Activity tests showed that the EA fraction exhibited obvious (p < 0.05) protective effects on H2O2 damaged PC-12 cells at 20 µg mL(-1), and showed much higher (p < 0.05) cytotoxic activity on cancer cell lines than other fractions. Five compounds, 1'-S-1'-acetoxyeugenol acetate (I), 1'-S-1'-acetoxychavicol acetate (II), 2-propenal, 3-[4-(acetyloxy)-3-methoxyphenyl] (III), isocoronarin D (IV) and caryolane-1, 9ß-diol (V), were obtained from the EA fraction and identified by HPLC, UV, MS, and NMR spectroscopic analyses. Compounds III and V were isolated from A. galangal for the first time. Moreover, compounds I, II, IV and V were the main active ingredients for inducing death of the tested cancer cells, and their IC50 values ranged from 60 to 90 µg mL(-1), indicating that these compounds possessed a wide anti-cancer capability. Therefore, A. galangal seeds could be a potential source of healthy food for tumor prevention.

HIRA is essential for the development of gibel carp.

HIRA is one of the chaperones of histone H3.3. Mutation of Hira results in embryonic lethality in mice, suggesting a critical role in embryogenesis. However, Hira-mutated Drosophila may survive to adults, indicating that it is dispensable in Drosophila development. The role of Hira in fish development is unknown. In this study we first investigated the expression of Hira during embryogenesis of gibel carp (Carassius auratus gibelio) by whole-mount in situ hybridization. We found that Hira signal appeared ubiquitously in the early embryos. After gastrulation, it appeared mainly along the anterior-posterior axis, including the tail bud. In hatching period, the signal was detected in head, heart, and the endoderm region on the back of yolk. Then by microinjection with morpholino-HIRA at the beginning of development, we observed delayed gastrulation and abnormal somitogenesis in gibel carp embryos. The HIRA morphants exhibited short trunk, limited yolk extension, and twisted tail. Most of the mutants died during embryogenesis or shortly after hatching. The rest of the HIRA morphants could survive to larvae but with severe defects in organogenesis. These data suggest that HIRA may be essential for the development of gibel carp, and this function is conserved in vertebrates.

Application of fluorescently labeled tracer technique for detection of natural active macromolecules in Chinese medicine.

Active substances in traditional Chinese Medicine (TCM) contain not only a variety of small molecules, but also many other macromolecules (TCMMs), such as proteins, peptides and polysaccharides. Active TCMM can achieve good therapeutic effects by regulating the body's overall function with lower side effects. This review summarized the literatures published in recent years on the application of fluorescently labeled tracer technique for detection of natural active macromolecules in TCM. Classified by fluorescent markers, applications of fluorescein, rhodamine, and quantum dots (QDs) in TCMM active tracer are reviewed, and the methods and principles of TCMM fluorescent marker are illustrated. Studies on active TCMMs and their action mechanism are quite difficult due to a multitarget, multicomponent, and multipath system of TCM. However, the development of fluorescently labeled active tracer technique (FLATT) provides this research with new tools. Traditional fluorescent markers have many deficiencies, such as easily quenched, short luminous cycle, and intrinsic toxicity. Relatively, FLATT has many obvious advantages, and its application in TCMM is still at the early stage. In order to improve the overall level of fluorescence labeling in TCMM active tracer, the improvement on FLATT's detection sensitivity and biological affinity is urgent and critical to allow study of these interesting molecules.

[miR-125b promotes proliferation of human acute myeloid leukemia cells by targeting Bak1].

OBJECTIVE: To investigate miR- 125b regulation mechanism by identifying miR-125b target genes and its function in acute myeloid leukemia (AML). METHODS: The bioinformatics software and database were applied to predict and analyze target genes of miR-125b. The vector contained the target gene 3'-UTR portion cloned into a luciferase reporter construct. A luciferase reporter assay was performed following co-transfection of small molecular miR-125b mimics and target gene wild-type or mutant plasmid into HEK-293T cells. Further in leukemia cell lines NB4 and HL-60, the protein level of target gene was measured by Western blot after overexpression miR-125b. Finally, the viabilities of NB4 and HL-60 cells were measured by CCK-8 assay at 24 h, 48 h, 72 h, 96 h after electroporation. RESULTS: Bcl-2-antagonist/killer 1 (Bak1), a pro-apoptotic gene, was a target gene of miR-125b by software predicts. Reporter vector containing the 3'-UTR Bak1 wild and mutation sites were co-transfected with small molecule analogues of miR-125b in HEK-293T cells. Dual luciferase reporter gene assay system showed that miR-125b significantly suppresses the reporter gene activity containing Bak1 3'-UTR by about 53.8% (P<0.05), but it didn't suppresses the reporter gene activity containing 3'-UTR Bak1 mutation. Western blot showed that miR-125b mimics significantly down-regulated the expression of Bak1 in human leukemia cell lines NB4 and HL-60. Meanwhile, the growth rate of cells treated with miR-125b obviously increased compared with that in control by CCK-8 test (P<0.05). CONCLUSION: Our findings strongly indicated that BAK1 was a downstream target gene of miR-125b, and miR-125b promoted proliferation in human AML cells at least partially by targeting Bak1, so we speculated that miR-125b as an oncogene could be a potential therapeutic target for treating AML.

Antioxidant and anticomplement functions of flavonoids extracted from Penthorum chinense Pursh.

Penthorum chinense Pursh is rich in flavonoids, which have strong antioxidant and anticomplement activities. In order to optimize their extraction conditions, various extraction parameters were chosen to identify their effects on flavonoids extraction. Single factor and Box-Behnken experimental designs consisting of 24 experimental runs and five replicates at zero point were applied to obtain the optimal extraction yield. The optimization conditions for flavonoids extraction were determined as follows: ethanol concentration 60.89%, extraction time 68.15 min, temperature 52.89 °C and liquid/solid ratio 19.70 : 1. The corresponding flavonoids content was 7.19%. The regression equation was found to fit well with the actual situation. Furthermore, the antioxidant activity (the free radical scavenging ability and ferric reducing/antioxidant power) and anticomplement ability of the flavonoids from P. chinense were determined. Results showed that the flavonoids of P. chinense displayed significant antioxidant and anticomplement activities. Its antioxidant activity can compete with ascorbic acid (Vc), whereas its anticomplement activity (IC50 = 111.6 µg ml(-1)) surpassed the effect of heparin (IC50 = 399.7 µg ml(-1)) which was used as the positive control, suggesting that P. chinense flavonoids and their related products could potentially be used as a promising natural agent in the treatment of humoral effector inflammation.

[McCune-Albright syndrome: a difficult and complicated case study].

McCune-Albright syndrome is a rare G proteins alpha disorder. The disorder is characterized by polyostotic fibrous dysplasia, sexual precocity and hyperpigmented macules. It is caused due to mutations in the gene Gsalpha that incodes the alpha subunit of the trimeric guanosine triphate-binding protein. There is no specific treatment for this syndrome. Treatment is generally symptomatic. This paper reported three cases of McCune-Albright syndrome and reviewed the relevant literatures regarding to the pathogenesis, pathological features, diagnosis and treatment. All three cases presented with a characteristic triad: polyostotic fibrous dysplasia, sexual precocity and hyperpigmented macules and were thus definitely diagnosed with McCune-Albright syndrome.