Multi-spectroscopies and molecular docking insights into the interaction mechanism and antioxidant activity of isoquercetin and zein nanoparticles.

The purpose of this study was to compare and characterize the theoretical properties and interaction mechanisms of zein and isoquercetin (ISO) from experimental and theoretical perspectives. Zein nanoparticles with different ISO concentrations (ZINPs) were prepared by the antisolvent precipitation method. The experimental results indicated all particles appeared spherical. When the mass ratio of zein to ISO was 10:1, the encapsulation efficiency of ZINPs reached 88.19 % with an average diameter of 126.67 nm. The multispectral method and molecular docking results confirmed that hydrogen bonding and van der Waals force played a dominant role for the binding of ISO to zein, and the primary fluorescence quenching mechanism for zein by ISO was static quenching. Furthermore, ZINPs had greater solubility and antioxidant activity, as well as inhibited the release of ISO during simulated gastrointestinal digestion processes. This research contributes to the understanding of the non-covalent binding mechanism between zein and ISO, providing a theoretical basis for the construction of ISO active carriers.

Hemp seed globulin-alginate nanoparticles for encapsulation of Cannabisin A with enhanced colloidal stability and antioxidant activity.

This study develops hemp seed globulin (GLB)-alginate (ALG) nanoparticles (GANPs) for Cannabisin A (CA) stabilization under environmental stress and during pepsin digestion. The optimal GLB: ALG mass ratio of 1: 1.5 was determined for GANPs formation at pH 3.5, resulting in a high yield of 95.13 ± 0.91 %, a ζ-potential of -35.73 ± 1.04 mV, a hydrodynamic diameter of 470.67 ± 11.36 nm, and a PDI of 0.298 ± 0.016. GANPs were employed to encapsulate CA, achieving a high loading capacity of 13.48 ± 0.04 µg mg-1. FTIR analysis demonstrated that the formation of CA-GLB-ALG nanoparticles (CGANPs) involves electrostatic interactions, hydrogen bonding, and hydrophobic interactions. XRD and DSC analyses revealed that CA is amorphous within the CGANPs. CGANPs demonstrated remarkable dispersion stability as well as resistance to high ionic strength and high-temperature treatments, indicating their potential as efficient hydrophobic drug-delivery vehicles. When compared to free CA, CA coated within CGANPs displayed greater DPPH/ABTS scavenging activity. Furthermore, the ALG-shelled nanoparticles protected GLB from pepsin digestion and slowed the release of CA throughout the release process, extending their stay on the intestinal wall mucosa. These findings imply that CGANPs is an ideal delivery vehicle for CA as they may expand the application of CA in food items.

Non-covalent interaction between hemp seed globulin and two hemp seed phenolic compounds: Mechanism and effects on protein structure, bioactivity, and in vitro simulated digestion.

This study focused on elucidating the non-covalent interactions between hemp seed globulin (GLB) and two hemp seed phenolic compounds, Cannabisin A (CA) and Cannabisin B (CB), and to explore these interactions on the protein's structure, conformation, and functionality. Fluorescence quenching and thermodynamic analysis revealed that static quenching governed non-covalent interaction processes, with hydrogen bonds and van der Waals forces functioning as major forces. This was further substantiated by molecular docking studies. The binding affinity order was CA > CB, indicating that the specific phenolic compound had a notable impact on the binding affinity. Furthermore, when complexed with CA, Tyr and Trp residues were exposed to a more hydrophilic environment than when complexed with CB. It was noted that the complexation with either CA or CB consistently affects GLB's secondary structure, particle size, and ζ-potential. GLB treated with the phenolic compounds exhibited enhanced ABTS and DPPH scavenging activities and improved digestibility compared to untreated GLB. Furthermore, the non-covalent interactions significantly increased CA's water solubility, highlighting GLB as a promising natural carrier for hydrophobic bioactive components. These findings hold potential implications for enhancing hemp seed protein applications within the food industry by positively influencing its functional properties and bioactivity.

Effect of hydrochloric acid and citric acid with ultrasound processing on characteristics of superfine-ground pectic polysaccharides from okra (Abelmoschus esculentus L.) peel.

The structural properties and biological activities of okra pectic polysaccharides (OPs) were impacted by various extraction methods. Based on commonly grinding (40, 100 meshes) and superfine grinding okra powders, two extraction solvents (hydrochloric acid, HA; citric acid, CA) were used firstly. Next, the extraction yield, physical and chemical properties, molecular structure and functional properties of OPs were analyzed by non-ultrasonic treatment and ultrasound-assisted superfine grinding method. The outcomes demonstrated that the extraction yield of OPs rose as the particle size of the powder decreased. HA-OPs had higher molecular weight (Mw), apparent viscosity and emulsification ability than CA-OPs. CA-OPs had higher esterification degree (DE), solubility and total sugar content, and higher amounts of rhamnogalacturonan-I (RG-I) segments. Compared with OPs without ultrasound-assisted extraction, ultrasound-assisted superfine grinding extraction exhibited higher sugar content, antioxidant capacity, emulsification ability, lower Mw, DE and apparent viscosity. Finally, the correlation between structure and function of OPs was further quantified. The antioxidant capacity was positively correlated with RG-I content, and negatively correlated with DE and Mw. The emulsification ability was mainly positively correlated with the GlcA of OPs. This study provides a theoretical basis for the development of OPs foods with clear structure-function relationship, which would be instructive for the application of OPs in food and cosmetics.

Bioactive polyphenols separated from hemp seed shells ameliorate H2 O2 -induced oxidative stress injury in human umbilical vein endothelial cells.

In the present study, we investigated the protection of hemp seed polyphenols on human umbilical vein endothelial cells (HUVEC) from H2 O2 -mediated oxidative stress injury. Fractions with different polarities were obtained by separating the hemp seed extract using HPD300 macroporous resin-packed column. The fraction, desorbed by 50% ethanol, is rich in polyphenol (789.51 ± 21.92 mg GAE/g) and has the highest antioxidant activity in vitro. HPLC-QTOF-MS/MS identified the main polyphenol components in hemp seed shells: 4 hydroxycinnamic acid amides and 15 lignanamides. The protective effects of hemp seed polyphenol against oxidative-stress injury in HUVEC cells were evaluated by cell viability, intracellular antioxidant parameters, and cell apoptosis assay. After HUVEC cells were precultured with 50 µg/ml hemp seed polyphenols, the cell viability increased significantly from 53.07 ± 2.46% (model group) to 80.65 ± 1.32% (p < 0.01). In addition, the pretreatment of HUVEC cells with polyphenol could substantially increase their intracellular superoxide dismutase activity and reduce their intracellular reactive oxygen species level, malondialdehyde content, and lactate dehydrogenase leakage index. These findings demonstrate the defensive potential of hemp seed polyphenol in reducing the incidence of cardiovascular disease. PRACTICAL APPLICATION: Hemp seed shell waste is produced while producing hemp seed kernel and has abundant phenolic compounds. This research showed that hemp seed polyphenol has potent antioxidant activity in vitro and protects HUVEC cells against H2 O2 -induced oxidative stress injury, suggesting that hemp seed polyphenol has the defensive potential to reduce the incidence of cardiovascular disease. These results indicated that polyphenol separated from hemp seed shells is valuable for further research and development, which will improve the utilization rate of hemp seed.

Enantioselective Synthesis of Cα-Tetrasubstituted N-Hydroxyl-α-amino Nitriles via Cyanation of Ketonitrones Using Me2(CH2Cl)SiCN.

Here, we report an unprecedented catalytic enantioselective cyanation of ketonitrones enabled by the bifunctional cyanating reagent Me2(CH2Cl)SiCN. This approach allows facile access to optically active N-hydroxyl-α-amino nitriles that are of high synthetic value but difficult to acquire by other methods. The use of bifunctional cyanating reagent Me2(CH2Cl)SiCN not only achieves an enantioselectivity higher than that with TMSCN but also enables various diversification reactions of the resulting silylated adducts. This represents the first enantioselective catalytic nucleophilic addition reaction of unactivated ketone-derived nitrones, exhibiting the potential of such tetrasubstituted C═N bonds for asymmetric synthesis of N-hydroxy α-amino acids and other N-hydroxy tertiary amines.

Structural Changes and Antibacterial Activity of Epsilon-poly-l-lysine in Response to pH and Phase Transition and Their Mechanisms.

ε-Poly-l-lysine (ε-PL) consists of 25-35 lysine residues which are linked by an isopeptide bond formed by dehydration condensation of α-carboxyl and ε-amino groups and has good antibacterial activity and broad-spectrum inhibition range. However, there is no clear conclusion about the structure and antibacterial mechanism of ε-PL in aqueous solution. Herein, a high purity of ε-PL was prepared using Amberlite IRC-50 ion-exchange resin. Membrane filtration and dynamic light scattering were used to study the variations of ε-PL aggregation in aqueous solution with pH value. The conformational changes and antibacterial activities of ε-PL and carbamoylated ε-PL in different water environments were studied with circular dichroism (CD) and inhibition zone. The structural changes during the spray-drying process were determined by Fourier transform infrared spectroscopy. The results indicated that the side chain amino charge played a decisive role in the ε-PL conformation and aggregation. ε-PL exhibited the properties of a ß-sheet during spray drying from acidic liquids to solids. The cation enhanced the antibacterial activity of ε-PL but did not play a key role. Instead, the backbone of ε-PL might determine the mechanism of ε-PL antibacterial.

The cell cycle inhibitor p21 promote mouse lung fibrosis by activating alveolar macrophages / 药学学报

This study was to determine the expression of the cell cycle inhibitor p21 in alveolar macrophages (AMs) and the role of p21 in activation of AMs in bleomycin (BLM) injury-induced lung fibrosis. The expression of CD206 in AMs was measured by immunofluorescence staining. Reverse transcription-polymerase chain reaction (RT-PCR) assay was used to detect the expression of macrophage activation markers. The coculture assay for macrophage and fibroblast was employed to explore the effect of macrophage on fibroblast activation. Immunofluorescence staining and western blotting assay were adopted to detect the expression of p21 in fibrotic tissues. AMs were treated with p21 knockdown or overexpression virus, RT-PCR and the co-culture system were used to explore the effect of p21 expression on macrophage activation. The Experimental Animal Welfare Ethics Committee of the Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College approved all of the protocols for this research. Our results showed that the expression of CD206 and macrophage activation markers was increased in AMs from fibrotic mice, indicating that AMs from fibrotic mice were associated with a profibrotic phenotype. Moreover, the expression of p21 was upregulated in AMs after BLM treatment. Depletion of p21 suppressed macrophage activation, while overexpression of p21 promoted the profibrotic phenotype of AMs from healthy mice. In summary, BLM injury causes the progressive accumulation of p21 in AMs, which induces the production of a number of profibrotic factors promoting the development of pulmonary fibrosis.

Author Correction: Draft genomic and transcriptome resources for marine chelicerate Tachypleus tridentatus.

In the original version of this Data Descriptor the word "Gulf" was incorrectly spelled in the affiliation "Ocean College, Beibu Gulf University, Qinzhou, 535011, Guangxi, China". This has now been corrected in both the HTML and PDF versions.

Draft genomic and transcriptome resources for marine chelicerate Tachypleus tridentatus.

Chinese horseshoe crabs (Tachypleus tridentatus), ancient marine arthropods dating back to the mid-Palaeozoic Era, have provided valuable resources for the detection of bacterial or fungal contamination. However, excessive exploitation for the amoebocyte lysate of Tachypleus has dramatically decreased the population of the Chinese horseshoe crabs. Thus, we present sequencing, assembly and annotation of T. tridentatus, with the hope of understanding the genomic feature of the living fossil and assisting scientists with the protection of this endangered species. The final genome contained a total size of 1.943 Gb, covering 90.23% of the estimated genome size. The transcriptome of three larval stages was constructed to investigate the candidate gene involved in the larval development and validate annotation. The completeness of the genome and gene models was estimated by BUSCO, reaching 96.2% and 95.4%, respectively. The synonymous substitution distribution of paralogues revealed that T. tridentatus had undergone two rounds of whole-genome duplication. All genomic and transcriptome data have been deposited in public databases, ready to be used by researchers working on horseshoe crabs.

Grape seed proanthocyanidin extract alleviates high-fat diet induced testicular toxicity in rats.

The present study aimed to investigate the protective effects of grape seed proanthocyanidin extract (GSPE) on high-fat diet (HFD) induced testicular damage, oxidative stress, and apoptotic germ cell death. Male rats (n = 40) were randomly divided into four groups: the control group (treated with physiological saline), HFD group, HFD + GSPE (100 mg kg-1) group and HFD + GSPE (300 mg kg-1) group. Compared with the HFD group the rats of the GSPE-treated group showed improved serum testosterone levels, sperm quality and histological appearance of the testis tissue. Significant elevation of antioxidant enzyme (SOD, GSH, and GSH-Px) activities and remarkable reduction in MDA were also observed by GSPE administration, indicating that GSPE can decrease testicular oxidative stress. Finally, a significant reduction in spermatogenic cell apoptosis was detected by TUNEL assay. In summary, these results indicated that GSPE can suppress testicular dysfunction and this effect may be attributed to its antioxidant and anti-apoptotic properties. The current study indicates that GSPE can be considered a promising candidate for use as a drug or a food supplement to alleviate HFD-induced testicular dysfunction.

One-Pot Sequential [3 + 3] Dipolar Cycloaddition of Aldehyde or Ketone and Hydroxylamine with Spirocyclopropyl Oxindole.

A Sc(OTf)3-catalyzed highly diastereoselective one-pot sequential [3 + 3] dipolar cycloaddition reaction of aldehyde or ketone, N-alkyl hydroxylamine, and spirocyclopropyl oxindole is developed, allowing facile construction of spirocyclic oxindole-tetrahydro-1,2-oxazines with sufficient structural diversity. The corresponding catalytic enantioselective one-pot protocol of aldehydes is also reported, affording the desired adducts in up to 97% ee. The biological evaluation of selected oxindole-based spirocyclic tetrahydro-1,2-oxazines revealed that they exerted cytotoxic effects on human prostate cancer cells with the capacity to inhibit NFκB signaling in prostate cancer cells.

Diastereo- and enantioselective [3 + 3] cycloaddition of spirocyclopropyl oxindoles using both aldonitrones and ketonitrones.

Optically active spirocyclic compounds play an important role in drug discovery, and new synthetic strategies for the efficient generation of spiro stereocenters are in much demand. Here we report a catalytic enantioselective cycloaddition using spirocyclic donor-acceptor cyclopropanes as a promising approach for the generation of spiro stereocenters. A diastereo- and enantioselective [3 + 3] cycloaddition of spirocyclopropyl oxindoles with both aldonitrones and ketonitrones is developed. The key to reaction development is the activation of spirocyclopropyl oxindoles by a suitable electron-withdrawing N-protecting group. This activation approach offers the promise of a general solution to enable spirocyclopropyl oxindoles as synthons for catalytic enantioselective synthesis of spirocyclic oxindoles featuring a C3 spiro stereocenter, a prominent structural motif in drugs and pharmaceutically active compounds. This protocol also constitutes the catalytic enantioselective reaction using unactivated achiral ketonitrones to construct tetrasubstituted carbon stereocenters.

Characterization of a new Lactobacillus salivarius strain engineered to express IBV multi-epitope antigens by chromosomal integration.

To obtain adhesive and safe lactic acid bacteria (LAB) strains for expressing heterologous antigens, we screened LAB inhabitants in intestine of Tibetan chickens by analyzing their adhesion and safety properties and the selected LAB was engineered to express heterologous antigen (UTEpi C-A) based on chromosomal integration strategy. We demonstrated that a new Lactobacillu salivarius TCMM17 strain is strongly adhesive to chicken intestinal epithelial cells, contains no endogenous plasmids, is susceptible to tested antimicrobials, and shows no toxicities. In order to examine the potential of TCMM17 strain as heterogenous antigen delivering vehicle, we introduced a UTEpi C-A expression cassette in its chromosome by constructing a non-replicative plasmid (pORI280-UUTEpi C-AD). The recombinant TCMM17 strain (∆TCMM17) stably was found to keep the gene cassette through 50 generations, and successfully displayed EpiC encoded by the cassette on its surface. This work provides a universal platform for development of novel oral vaccines and expression of further antigens of avian pathogens.

Assembly and immunogenicity of baculovirus-derived infectious bronchitis virus-like particles carrying membrane, envelope and the recombinant spike proteins.

OBJECTIVE: To assemble infectious bronchitis virus (IBV)-like particles bearing the recombinant spike protein and investigate the humoral immune responses in chickens. RESULTS: IBV virus-like particles (VLPs) were generated through the co-infection with three recombinant baculoviruses separately encoding M, E or the recombinant S genes. The recombinant S protein was sufficiently flexible to retain the ability to self-assemble into VLPs. The size and morphology of the VLPs were similar to authentic IBV particles. In addition, the immunogenicity of IBV VLPs had been investigated. The results demonstrated that the efficiency of the newly generated VLPs was comparable to that of the inactivated M41 viruses in eliciting IBV-specific antibodies and neutralizing antibodies in chickens via subcutaneous inoculation. CONCLUSIONS: This work provides basic information for the mechanism of IBV VLP formation and develops a platform for further designing IBV VLP-based vaccines against IBV or other viruses.

Development of a multi-epitope antigen of S protein-based ELISA for antibodies detection against infectious bronchitis virus.

An indirect enzyme-linked immunosorbent assay (ELISA) method based on a novel multi-epitope antigen of S protein (SE) was developed for antibodies detection against infectious bronchitis virus (IBV). The multi-epitope antigen SE protein was designed by arranging three S gene fragments (166-247 aa, S1 gene; 501-515 aa, S1 gene; 8-30 aa, S2 gene) in tandem. It was identified to be approximately 32 kDa as a His-tagged fusion protein and can bind IBV positive serum by western blot analysis. The conditions of the SE-ELISA method were optimized. The optimal concentration of the coating antigen SE was 3.689 µg/mL and the dilution of the primary antibodies was identified as 1:1000 using a checkerboard titration. The cut-off OD450 value was established at 0.332. The relative sensitivity and specificity between the SE-ELISA and IDEXX ELISA kit were 92.38 and 89.83%, respectively, with an accuracy of 91.46%. This assay is sensitive and specific for detection of antibodies against IBV.

Recycle waste salt as reagent: a one-pot substitution/Krapcho reaction sequence to α-fluorinated esters and sulfones.

A "one-pot" tandem substitution/Krapcho reaction is reported for the facile synthesis of α-fluorinated esters and sulfones, which utilizes the byproduct salt formed in the substitution step as an indispensible reagent to facilitate the Krapcho reaction step. This represents the first sustainable tandem reaction that internally recycles the waste salt formed in the upstream step as the reagent for the downstream step.

[Expression of neuraminidase gene of influenza virus H1N1 in baculovirus-expression system].

OBJECTIVE: To construct the recombinant baculovirus with NA gene of Influenza H1N1 virus. METHODS: Full-length NA gene of Influenza virus H1N1 (A/PR/8/34) was amplified by PCR and inserted into pFastBacdual vector to construct the recombinant baculovirus transfer vector pFBD-NA. Recombinant shuttle vectors rBacmid-NA was then obtained after transforming DH10B competent cells containing bacmid plasmids. After transfecting into sf9 cells, recombinant baculovirus rBac-NA was obtained. The rBac-NA genome was extracted and identified by PCR. NA protein expressed by recombinant baculovirus-infected sf9 cells was determined by IFA, Western Bolt and ELISA. RESULTS: PCR results proved that recombinant shuttle vectors rBacmid-NA was successfully constructed. NA protein was detected by IFA and showed strong specific green fluorescence on the surface of infected cells. NA protein was recognized by two polyclonal antibodies specific for NA in Western Blot. ELISA showed specific reaction of recombinant NA protein with mouse polyclonal antibody against influenza virus (PR8), indicating high antigenicity. CONCLUSION: Recombinant baculovirus rBac-NA that expresse NA protein of influenza virus was successfully constructed. This work provides a basis for further study on NA protein function and novel influenza vaccine development.

Genome analysis and signature discovery for diving and sensory properties of the endangered Chinese alligator.

Crocodilians are diving reptiles that can hold their breath under water for long periods of time and are crepuscular animals with excellent sensory abilities. They comprise a sister lineage of birds and have no sex chromosome. Here we report the genome sequence of the endangered Chinese alligator (Alligator sinensis) and describe its unique features. The next-generation sequencing generated 314 Gb of raw sequence, yielding a genome size of 2.3 Gb. A total of 22 200 genes were predicted in Alligator sinensis using a de novo, homology- and RNA-based combined model. The genetic basis of long-diving behavior includes duplication of the bicarbonate-binding hemoglobin gene, co-functioning of routine phosphate-binding and special bicarbonate-binding oxygen transport, and positively selected energy metabolism, ammonium bicarbonate excretion and cardiac muscle contraction. Further, we elucidated the robust Alligator sinensis sensory system, including a significantly expanded olfactory receptor repertoire, rapidly evolving nerve-related cellular components and visual perception, and positive selection of the night vision-related opsin and sound detection-associated otopetrin. We also discovered a well-developed immune system with a considerable number of lineage-specific antigen-presentation genes for adaptive immunity as well as expansion of the tripartite motif-containing C-type lectin and butyrophilin genes for innate immunity and expression of antibacterial peptides. Multifluorescence in situ hybridization showed that alligator chromosome 3, which encodes DMRT1, exhibits significant synteny with chicken chromosome Z. Finally, population history analysis indicated population admixture 0.60-1.05 million years ago, when the Qinghai-Tibetan Plateau was uplifted.

[Construction of recombinant baculovirus co-expressing M1 and HA of influenza A virus].

The M1 and HA genes of H1N1 influenza virus were amplified and then cloned into the pFastBac dual donor plasmid. The recombinant pFastBac Dual-M1-HA was identified by restriction enzyme digestion. After the pFastBacdual-M1-HA was transformed into the baculovirus shuttle plasmid (bacmid) in DH10Bac competent cells, the colonies were identified by antibiotics and blue-white selection. The rBac-mid-M1-HA was verified by PCR and transfected into S f9 cells to produce recombinant baculovirus (rBac-M1-HA). Gene insertion of rBac-M1-HA was verified and the expression of M1 and HA genes was analyzed by IFA and Western-blot, demonstrating M1 and HA were co-expressed successfully. This study provides the foundation for researching the formation mechanism of influenza VLP and developing new influenza vaccines.