Final Concentration Research Articles

Field Efficacy of Antagonistic Fungi against Black Spot Disease of Chinese Kale

Background: Alternaria brassicicola (Schw.) causes black spot disease, which is one of the major diseases limiting the production of vegetable crops, especially Chinese kale in Southeast Asia and Thailand. Previous pathogen management based on synthetic fungicides is expensive, toxic for humans, and harmful to the environment. The current study investigated the efficiency of Talaromyces flavus (Klöcker) Stolk and Samson Bodhi001, Talaromyces trachyspermus (Shear) Stolk and Samson Bodhi002, Talaromyces flavus (Klöcker) Stolk and Samson Bodhi003, Neosartorya fischeri (Wehmer) Malloch and Cain Bodhi004, and Neosartorya fischeri (Wehmer) Malloch and Cain in controlling Chinese kale black spot disease caused by A. brassicicola under field conditions. Methods: A. brassicicola and four antagonistic fungus strains were cultured separately on a PDA plate and incubated at room temperature for 14 days. Ten mL of sterile water was poured into a culture plate, and the spores were gently scraped from the mycelium with a sterile loop to obtain a spore suspension and afterward adjusted to a final concentration of 106 spores mL−1. Result: The results showed that spore suspensions of 106 spores mL−1 of T. flavus Bodhi001, T. trachyspermus Bodhi002, T. flavus Bodhi003, N. fischeri Bodhi004, and N. fischeri effectively controlled black spot disease in field trials and resulted in a significant reduction in black spot incidence compared with the unprotected control. Meanwhile, the spore suspension of T. flavus Bodhi001 revealed the greatest suppression of black spot incidence, causing 10.23% and 42.93% disease reduction, compared with the negative control, indicating promising preventive activity against A. brassicicola. Based on our results, T. favus Bodhi001 is a promising biological control agent (BCA) in controlling A. brassicicola causing Chinese kale black spot disease.

Short-term preservation of canine sperm-binding ability and other metrics using the INRA-96 in comparison to Tris-egg yolk extender.

The use of assisted reproductive techniques, such as chilled semen, contributes to the maintenance and genetic improvement of canine breeding. The INRA-96 extender is a commercially available, chemically defined medium that was initially developed for the preservation of equine semen and exhibits preservation potential in the canine species. This research aims to evaluate the INRA-96 extender as an alternative for the short-term preservation of canine semen in terms of sperm quality parameters such as motility and kinetic parameters, integrity and functionality of the plasma membrane in fresh and chilled-rewarmed samples, as well as the sperm-binding ability using the perivitelline membrane of the chicken egg as an indicator of the fertilizing capacity of the preserved semen. A total of 18 ejaculates from 9 French bulldogs (two ejaculates per dog) were collected and divided into two aliquots that were diluted in Tris-egg yolk 20% (control) or INRA-96 to a final concentration of 100 × 106 sperm/mL. Samples were refrigerated in a biological incubator at 5°C and evaluated at 0, 24 and 48 h time points. Comparing the two treatments after 48 h of refrigeration, both extenders showed similar values (p < .5) for the majority of kinetic parameters, with the INRA-96 group promoting a total motility of 88.1 ± 2.9%. In addition, the morphology, integrity and functionality of the plasma membrane were preserved above 70% in this group. Dilution with INRA-96 also provided a significantly higher amount of sperm bound (256.2 ± 21.1) to the perivitelline membrane of the egg yolk compared to the sperm-binding rates (p < .05) achieved at the use of Tris-egg yolk (215.2 ± 21 bound spermatozoa) at 48 h. Our study proved similar functional properties of dog sperm cells treated with INRA-96 in comparison to commonly used home-made Tris-based extender during short-time storage.

Peptide Fractions from Pepsin-digested Moringa oleifera Seed Proteins Inhibit Hemoglobin Glycation and Carbohydrate-hydrolyzing Enzymes

The multidirectional abilities of peptide digests and fractions obtained from the hydrolysis of food-based proteins have been investigated in recent times. This study aims to evaluate the effects of pepsin-derived Moringa oleifera seed protein hydrolysates and fractions on hemoglobin glycation and the carbohydrases - ?-amylase and ?-glucosidase. Proteins were extracted from M. oleifera seeds and consequently digested using pepsin. The hydrolysates obtained were separated into fractions of &lt;1 kD, 1-3 kD, and 3-5 kD ranges using size-exclusion chromatography and comparison with elution volumes of known standards. The activities of the hydrolysates and peptide fractions against both the non-enzymatic glycation of hemoglobin and the carbohydrases were determined in vitro. Results revealed that the hydrolysate and its peptide fractions demonstrated varying abilities against the glycation of hemoglobin, with the unfractionated hydrolysate showing better activities (78.230 ± 0.774 % at a maximum concentration of 1.0 mg/ml) than its peptide fractions. Also, the hydrolysates and fractions demonstrated higher inhibitory effects on a-amylase (with all fractions displaying above 50% inhibition at a final concentration of 1.0 mg/mL) than against a-glucosidase. Kinetic analysis of a selected fraction showed that it inhibited ?-amylase via a mixed mechanism (Ki = 0.029 mg/mL) but displayed an uncompetitive mode for ?-glucosidase inhibition (Ki = 0.333 mg/mL). Therefore, it is inferred that M. oleifera seed proteins encode potentially therapeutic peptide sequences that could be further processed to formulate potential antidiabetic agents.

The synergism of andisol soil and foundry sand mixture as Cu(II) ion adsorbent

The aims of this study were to determine Cu(II) metal ion adsorption ability by using andisol soil and foundry sand as an adsorbent. Andisol soil and foundry sand were activated with 3 M NaOH. Adsorption of copper (II) metal ion performed on the composition of andisol soil and foundry sand 100:0, 75:25, 50:50, 25:75, and 0:100 (wt%), pH solution variation pH 3-7, and contact time 15-135 minutes with batch system. The final concentration at the time of equilibrium was measured using Atomic Absorption Spectrometry (AAS). Characterizations of the adsorbent was conducted by Fourier Transform-Infrared Analyzer (FT-IR), Scanning Electron Microscope-Energy Dispersive X-Ray (SEM-EDX), and Surface Area Analyzer (SAA). The optimum condition for Cu(II) ion adsorption was obtained at the composition of the mixture andisol soil and foundry sand 75:25 (wt%) at pH 6 and contact time of 90 minutes with an adsorption capacity of 3.839 mg/g and adsorption percentage 94.55%. The adsorption isotherm followed the Langmuir isotherm model and the adsorption kinetics are pseudo-second-order.

Process for nutrient recycling from intensive aquaculture through microalgae-bacteria consortium

This work studies a biological process based on a microalgae-bacteria consortium for recycling nutrients in a recirculating aquaculture system (RAS) implanted in an intensive marine aquaculture farm. Additionally, some techniques were used for microalgae biomass harvesting and tested the effectiveness of filtration by a column with multi-layer sand to reduce the solids concentrations in the effluent.The consortium was grown in photobioreactors in batch and semi-continuous operation modes using the solids concentrated stream generated in the RAS system. The semi-continuous operation showed a high percentage of TDN and TDP removal, achieving final concentrations of 1.09 ± 0.02 mg·L−1 and 0.01 ± 0.01 mg·L−1, respectively, while DOC was reduced to 3.87 ± 0.06 mg·L−1. The values of productivity 44 ± 9 mg TSS·L−1 indicated that the studied stream is a suitable culture medium for the growth of the microalgae-bacteria consortium.A combination of harvesting techniques was studied, coagulation-flocculation-settling and coagulation-flocculation-flotation. The first step was to optimise the dose of FeCl3 through the coagulation-flocculation test to pre-concentrate the biomass generated, achieving an optimal dose of 0.106 mg Fe·mg TSS−1. Then, two separation processes were applied to the stream and compared: settling and flotation. The maximum removal efficiency (90.2 ± 0.3 %) was obtained in the settling process, so the coagulation-flocculation-settling was select as the best combination of harvesting techniques.Finally, sand filtration was studied as an effluent refining process to improve solids reduction of the water obtained in the harvesting step resulting in an effluent with 17.18 ± 1.49 mg TSS·L−1.The proposed sequence process is capable of recycling nutrients from an intensive marine aquaculture farm by using these resources via transformation into microalgae biomass and generating quality effluent.

Discovery and characterization of a novel PKD-Fn3 domains containing GH44 endoglucanase from a Tibetan metagenomic library.

To explore novel microbial endoglucanases with unique properties derived from extreme environments by using metagenomics approach. A Tibetan soil metagenomic library was applied for screening cellulase-active clones by function-based metagenomics. The candidate genes in the active clones were identified through bioinformatic analyses and heterologously expressed using an Escherichia coli system. The recombinant endoglucanases were purified and characterized using enzyme assays to determine their bioactivities, stabilities, substrate specificities, and other enzymatic properties. A novel endoglucanase gene Zfeg1907 was identified, which consisted of a glycoside hydrolase family 44 (GH44) catalytic domain along with a polycystic kidney disease (PKD) domain and a fibronectin type Ⅲ (Fn3) domain at the C terminal. Recombinant enzyme ZFEG1907 and its truncated mutant ZFEG1907t (ΔPKDΔFn3) were successfully expressed and purified. The two recombinants exhibited catalytic activities towards carboxymethyl cellulose, konjac glucomannan and lichenan. Both enzymes had an optimal temperature of 50°C and an optimal pH value of 5.0. The catalytic activities of both recombinant enzymes were promoted by adding Zn2+ and Ca2+ at the final concentration of 10mM. The Km value of ZFEG1907 were lower while the kcat/Km value of ZFEG1907 was higher than those of of ZFEG1907t when using carboxymethyl cellulose, konjac glucomannan and lichenan as substrates. Structure prediction of two recombinants revealed that PKD-Fn3 domains consisted of a flexible linker and formed a β-sandwich structure. A novel endoglucanase ZFEG1907 contained a GH44 catalytic domain and a PKD-Fn3 domain was characterized. The PKD-Fn3 domains were not indispensable for the activity but contributed to the enzyme binding of the polysaccharide substrates as a carbohydrate-binding module (CBM).

Development and Characterization of Probiotic Beers with Saccharomyces boulardii as an Alternative to Conventional Brewer's Yeast.

The development of new non-dairy probiotic foods is interesting, given lactose intolerance, milk allergies, and the growing trend of vegetarianism. In this paper, beer has been used as a probiotic delivery matrix, using Saccharomyces boulardii as an alternative to conventional brewer's yeast. The strain was able to grow in worts prepared with hops containing different alpha-acid concentrations, attaining in all cases a final cell concentration above 1·108 cells mL-1. Some differences were found in the physicochemical parameters of beers brewed with S. boulardii compared to those brewed with a standard brewer's yeast. Probiotic beers turned out to be less cloudy, which could help with a possible filtering step; less alcoholic in some cases; a healthier alternative; and with a slightly lower pH, interesting for the reduction of spoilage risk. Thirty volatile compounds were determined in the samples, and, in general, the beers brewed with the probiotic yeast presented significantly higher concentrations for the majority of the studied volatile compounds. In addition, multivariate statistical analysis was successfully performed to differentiate the beers obtained in terms of their volatile composition. Probiotic and standard beers were also subjected to sensory analysis, and they presented similar results in their overall impression.

UPLC-ESI-QTOF-MS assisted targeted metabolomics to study the enrichment of vinca alkaloids and related metabolites in Catharanthus roseus plants grown under controlled LED environment

Enrichment of pharmaceutically important vinca alkaloids, vinblastine and vincristine, in the leaves of Madagascar periwinkle (Catharanthus roseus) plants through different pre- or postharvest treatments or cultivation conditions, e.g., exposing the plants to UV-irradiation, has been in focus for decades. Controlled LED environment in the visible light range offers the possibility of monitoring the changes in the concentration of metabolites in the vinca alkaloid-related pathway without involving UV-related abiotic stress. In the frame of our targeted metabolomics approach, 64 vinca alkaloids and metabolites were screened with the help of a UPLC-ESI-QTOF-MS instrumental setup from the leaf extracts of C. roseus plants grown in chambers under control (medium light), low light, and high blue / high red/ high far-red conditions. Out of the 14 metabolites that could be assigned either unambiguously with authentic standards or tentatively with high resolution mass spectrometry-based methods, all three dimer vinca alkaloids, that is, 3’,4’-anhydrovinblastine, vinblastine and vincristine showed an at least nine-fold enrichment under high blue irradiation when compared with the control conditions: final concentrations of 961 mg kg-1 dry weight, 33.8 mg kg-1 dry weight, and 11.7 mg kg-1 dry weight could be achieved, respectively. As supported by multivariate statistical analysis, the key metabolites of the vinca alkaloid pathway were highly represented among the metabolites that were specifically stimulated by high blue light application.

Exposure pathways matter: Aquatic phototrophic communities respond differently to agricultural run‐off exposed via sediment or water

Abstract Small shallow ponds are widespread but understudied water bodies in agricultural landscapes. Agricultural run‐off (ARO) transports pesticides and nutrients into adjacent aquatic ecosystems where they occur dissolved in the water column or are bound to sediments. Consequently, aquatic communities are affected by ARO via different exposure pathways. We hypothesize that sediment‐bound ARO mainly affects submerged rooted macrophytes, while phytoplankton and periphyton are more prone to ARO in water. These primary producers compete for resources resulting in a regime shift between alternative stable states of macrophyte or phytoplankton dominance. We hypothesize that warming increases nutrient release from sediments and thereby facilitates the occurrence of phytoplankton dominance. Using a full‐factorial microcosm design, we exposed aquatic primary producers to either sediment or water application of a mixture of common pesticides (terbuthylazine, pirimicarb, tebuconazole and copper) and nitrate at two concentrations and two temperatures (22°C and 26°C) for 4 weeks. Initial and final concentrations of pesticides and nitrate, final biomass of macrophytes, periphyton and phytoplankton, pesticide accumulation in macrophytes and changes in carbon, nitrogen and phosphorus content and selected exoenzyme activities in the sediment were measured. We found lower final macrophyte biomass for both ARO treatments compared to controls, indicating a prevalence of negative effects by herbicides and competition for light with other phototrophs. In contrast, phytoplankton and periphyton biomass increased, but only when exposed to ARO via the water column, indicating a prevalence of positive effects by nutrient supply. Microbial carbon and nutrient cycling in sediments was not affected by ARO. Higher temperature mitigated ARO‐related effects on macrophytes under sediment exposure. Synthesis and application. ARO poses a strong risk of submerged macrophyte loss and establishment of turbid conditions with phytoplankton dominance in aquatic ecosystems. In conclusion, exposure pathways as well as indirect and interacting effects of multiple stressors need to be considered when designing appropriate mitigation measures. Under climate change, we suggest to prioritize local measures as buffer strips a reduced use of pesticides and fertilizers, and sediment removal as appropriate measures to protect these vulnerable but widespread aquatic systems, which are highly relevant for biodiversity in agricultural landscapes.

Cryopreservation of black seabream (Acanthopagrus schlegelii) sperm

In recent years, biotechnology has had a significant impact on the aquaculture industry, particularly in the field of breeding. Molecular selection breeding has emerged as a novel approach to breeding. Reducing the cost of genetic information for individuals with desirable traits after breeding has become an important research direction. Cryopreservation technology allows bypassing time and space constraints in genetic breeding, simplifying broodstock management. This study presents a detailed cryopreservation method for black seabream sperm, evaluating extender type, glucose concentration, cryoprotectant type and concentration, sperm-dilution ratio, and cooling protocols. Sperm motility parameters were analyzed using computer-assisted sperm analysis (CASA) before and after two days of freezing. This involved using an RS solution with a glucose concentration of 15 g/L and adding a 5% final concentration of EG as the sperm cryoprotectant. After mixing the sperm and solution at a ratio of 1:2, we subjected it to 5 min fumigation at 5 cm above the liquid nitrogen surface before plunging it into the nitrogen. Sperm motility reached 85.46 ± 7.32% after two days. Various enzymatic activities showed changes over 20 days post-cryopreservation. This improved cryopreservation protocol for black seabream sperm is beneficial for genetic breeding and reproduction and provides reference for studying the cryodamage mechanisms of black seabream sperm.

Self- immobilized bio-nanomaterials based on hybridized green reduced graphene and Burkholderia vietnamiensis C09V for enhanced removal of Sb species from mine wastewater

Elevated levels of antimony (Sb) in the environs of smelters or mines can pose significant human and environmental health risks, where efficient removal of Sb species from mine wastewater is currently a challenge. In this paper, a functional reduced graphene hybrid Burkholderia vietnamiensis C09V biomaterial (RGO-B.V. C09V) was initially synthesized by self-immobilization and subsequently used to remove 100% of Sb(III) within 24 h and 90.0% of Sb(V) within 120 h. This removal efficiency was also much higher than that of free B.V. C09V alone, which removed 100% of Sb(III); but only 26.7% of Sb(V); within 120 h. Scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) confirmed that Burkholderia vietnamiensis C09V (B.V. C09V) was firmly immobilized on the RGO surface with an abundant distribution of extracellular polymeric substances (EPS). Three-dimensional excitation-emission matrix (3D-EEM) analysis and electrochemical techniques were used to confirm the function of RGO as a potential electron shuttle and the composition of EPS to identify its key role in any redox reactions. Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) analysis confirmed that polysaccharides and protein-like substances on the surface of RGO-B.V. C09V participated in the biosorption of Sb via complexation. When RGO-B.V. C09V was practically used to treat a mine wastewater the final Sb concentration decreased by 96% from 0.12 to 0.005 mg·L−1. RGO-B.V. C09V also exhibited high stability, with removal efficiencies of up to 100% for Sb(III), and 72% for for Sb(V) after 4 reuse cycles, with no regeneration. The results showed that RGO-B.V. C09V was a stable, environmentally friendly bionanomaterial suitable for the practical removal of Sb species from mine wastewater over a sustained period.

A Novel and Low-cost Approach for Intravitreal Injection in an Experimental Model of Endophthalmitis.

Animal models are necessary in understanding the pathogenesis of endophthalmitis and are also necessary to assist the development of new therapeutics for this sight-threatening ocular inflammation. Hamilton syringes are usually preferred to inject pathogens when performing experiments on test subjects, however, this method has technical and financial disadvantages. In this study, we report the findings and assess the related benefits of applying a novel low-cost intravitreal injection technique to initiate endophthalmitis in a mouse model while using the Eppendorf tip and a 26G needle. The 18-hr culture of clinical isolates of bacteria (Staphylococcus aureus and Pseudomonas aeruginosa) and fungus (Aspergillus flavus and Candida albicans) were resuspended to a final concentration of 10,000 colony forming units (CFU)/1 µL which were separately injected intravitreally into C57BL/6 mice (6-8 weeks) using a 0.1-2.5µL pipette attached to the modified Eppendorf tip with a 26G needle. The contralateral eye served as vehicle/uninjected control. Disease progression was determined by assessing the corneal haze, opacity, bacterial burden, and retinal histology of the eyes used in the model. Following euthanization, bacteria-infected mice were enucleated after 24 hr of the initial injection, and fungus-infected mice after 72 hr. Of the 50 mice injected, the modified technique was successful in 48 mice. Two mice were excluded due to cataract formed by accidental injury to the lens. The experimental endophthalmitis mice model successfully mimicked the natural clinical course. Clinical assessment and histopathology confirmed the influx of inflammatory cells into the posterior segment of the eye along with dissolution of retinal architecture. Our novel method of injection using a modified Eppendorf tip and 26G needle yielded a cost-effective mouse model of clinical endophthalmitis, resulting in reproducible infection for understanding various aspects of its pathobiology.

The Effect of Xanthohumol Derivatives on Apoptosis Induction in Canine Lymphoma and Leukemia Cell Lines.

Xanthohumol is a cancer chemopreventive agent that can interfere with the initiation, promotion, and progression phase of carcinogenesis via a variety of inhibitory mechanisms. Xanthohumol was reported as an effective agent against leukemia/lymphoma cells. In the present study, we investigated the effect of xanthohumol and its natural and semisynthetic derivatives against various canine leukemia/lymphoma cell lines. Xanthohumol, three hops minor prenylflavonoids (xanthohumol C, xanthohumol D, α,β-dihydroxanthohumol) and four derivatives obtained by biotransformation (xanthohumol 4'-O-β-D-(4‴-O-methyl)-glucopyranoside) as well as by chemical modification (1″,2″-dihydroxanthohumol K, 2,3-dehydroisoxanthohumol, (Z)-6,4'-dihydroxy-4-methoxy-7-prenylaurone) were tested for their antiproliferative and pro-apoptotic activities against the following canine leukemia/lymphoma cell lines: CLBL-1 (B-cell lymphoma), CLB70 (B-cell leukemia), and GL-1 (B-cell leukemia). The compounds were tested at a final concentration range of 0.1-30 µM for 48 h. All eight of the tested flavonoids exerted concentration-dependent cytotoxicity in the selected canine lymphoma/leukemia cell lines. Three compounds markedly decreased the viability of all cell lines with IC50 in the range of 0.5 to 8 μM. Double-staining of the treated cells with AnnexinV and propidium iodide revealed that the dying cells were mostly in the late apoptosis stage. ROS production and changes in mitochondrial potential were detected. Western blot analysis showed a decreased expression of Bcl-2. Canine lymphoma and leukemia cell lines are sensitive to xanthohumol derivatives, and the compounds acted through an apoptotic cell-death mechanism. These compounds, either used alone or in combination with other therapies, may be useful for the treatment of canine leukemia/lymphoma.

Adsorptive desulphurization of fuels by hypercrosslinked nanoporous polymers derived from polycyclic aromatic hydrocarbons

A sustainable method is employed to remove polycyclic aromatic sulphur heterocycles (PASHs) from fuels by using a series of high surface area nanoporous adsorbents synthesized from polycyclic aromatic hydrocarbons (PAHs). The hypercrosslinking of PAHs has been used to synthesize high surface area (SABET of 620–1565 m2 g−1) nanoporous polymeric materials via a microwave assisted method. The adsorbent poly-naph (highest SABET, i.e., 1565 m2 g−1) display best performance for desulphurization of fuels from batch to fixed bed column mode. In batch mode, the adsorption capacity of 12.1 (up to 9 ppm sulphur) and 8.9 mgS g−1 (up to 43 ppm sulphur), has been estimated for model (100 ppm sulphur) and real fuels (110 ppm sulphur), respectively. Furthermore, from ultra-low sulphur model (10 ppm sulphur) and real fuel (9 ppm sulphur), the adsorption capacity of 16.1 (up to 0.3 ppm sulphur) and 10 mgS g−1 (up to 3 ppm sulphur) has been estimated. In fixed-bed column mode, the high concentrated model and real fuel have been desulphurized to final concentration of 1 and 12 ppm, respectively. While, at low concentrations, the final concentration of 100 ppb and 1 ppm is achieved for model and real fuels, respectively. The adsorption mechanism revealed that π-electron density in the polymeric framework plays prominent role in the adsorption process apart from high surface area. This approach to control carcinogenic aromatic pollutants like PAHs and PASHs can be sustained over time through technological advancement, financial assistance and governmental regulation.

Regulatory effects of the Nocardia rubra cell wall skeleton on the biological function of human neutrophils

Objective: To investigate the regulatory effects and mechanism of Nocardia rubra cell wall skeleton (Nr-CWS) on the biological function of human neutrophils. Methods: The experimental research method was used. Fifteen healthy adult volunteers (7 males and 8 females, aged 24 to 45 years) were recruited from Suzhou Physical Examination Center for physical examination from May to October 2022, the peripheral venous blood was collected, and neutrophils were extracted by immunomagnetic bead sorting. The cells were divided into normal control group without any treatment, Nr-CWS alone group treated with Nr-CWS of final mass concentration 60 ng/mL alone, endotoxin/lipopolysaccharide (LPS) alone group stimulated with LPS of final mass concentration 1 μg/mL alone, and LPS+Nr-CWS group stimulated with LPS first and then treated with Nr-CWS as before. After 1 h of culture, the chemotaxis distance, chemotactic cell percentage, chemotactic index, maximum chemotactic speed, and chemotactic function score of neutrophils were detected using the modified agarose chemotactic model; the proportion and fluorescence intensity of phagocytosis cells, the level of reactive oxygen species (ROS), the protein expression levels of granular protein CD35, CD66b, and CD63, and the concentrations of inflammatory cytokines of interleukin 2 (IL-2), IL-4, IL-6, IL-10, IL-17A, tumor necrosis factor alpha (TNF-α), and interferon-γ in cell culture supernatant were detected by flow cytometry. The number of samples in each group in the above experiments was 15. Data were statistically analyzed with analysis of variance for factorial design and independent sample t test. Results: After 1 h of culture, the chemotactic function score of cells in normal control group, Nr-CWS alone group, LPS alone group, and LPS+Nr-CWS group were 15.0, 14.5±0.5, 1.5±0.5, 12.0±1.5, respectively. Compared with those in normal control group, the chemotaxis distance, chemotactic cell percentage, chemotactic index, maximum chemotactic speed, and chemotactic function score of cells were significantly decreased in LPS alone group and LPS+Nr-CWS group (with t values of 18.36, 18.88, 54.28, 18.36, 46.77, 10.58, 14.74, 6.84, 10.58, and 4.24, respectively, P<0.05); compared with those in LPS alone group, the five chemotactic function indexes as above in LPS+Nr-CWS group were significantly increased (with t values of 11.47, 14.65, 11.62, 11.47, and 13.75, respectively, P<0.05). After 1 h of culture, compared with those in normal control group, the proportion and fluorescence intensity of phagocytosis cells were significantly increased in Nr-CWS alone group (with t values of 6.86 and 6.73, respectively, P<0.05), and the above two indexes were significantly decreased in LPS alone group (with t values of 7.35 and 22.72, respectively, P<0.05) and LPS+Nr-CWS group (with t values of 21.37 and 13.10, respectively, P<0.05). After 1 h of culture, compared with that in normal control group, the level of ROS of cells in LPS alone group was significantly increased (t=6.64, P<0.05); compared with that in LPS alone group, the level of ROS of cells in LPS+Nr-CWS group was significantly decreased (t=5.46, P<0.05). After 1 h of culture, compared with those in normal control group, the protein expressions of CD35, CD66b, and CD63 of cells were significantly increased in LPS alone group and LPS+Nr-CWS group (with t values of 16.75, 17.45, 10.82, 5.70, 19.35, and 15.37, respectively, P<0.05); compared with those in LPS alone group, the protein expressions of CD35, CD66b, and CD63 of cells were significantly decreased in LPS+Nr-CWS group (with t values of 4.92, 5.72, and 3.18, respectively, P<0.05). After 1 h of culture, compared with those in normal control group, the concentrations of IL-2, IL-4, IL-6, IL-10, IL-17A, TNF-α, and interferon-γ in cell culture supernatant were significantly increased in LPS alone group (with t values of 22.10, 9.50, 7.21, 10.22, 24.88, 8.43, and 47.48, respectively, P<0.05), and the concentrations of IL-6, IL-10, IL-17A, TNF-α, and interferon-γ in cell culture supernatant were significantly increased in LPS+Nr-CWS group (with t values of 4.68, 5.12, 8.02, 5.58, and 7.13, respectively, P<0.05); compared with those in LPS alone group, the concentrations of IL-2, IL-4, IL-6, IL-10, IL-17A, TNF-α, and interferon-γ in cell culture supernatant were significantly decreased in LPS+Nr-CWS group (with t values of 5.39, 2.83, 5.79, 2.90, 5.87, 4.88, and 39.64, respectively, P<0.05). Conclusions: Nr-CWS can enhance the phagocytosis ability of neutrophils in normal condition and improve the chemotactic function, ROS level, degranulation protein level, and inflammatory factor level of human neutrophils in infectious condition. Nr-CWS can enhance the anti-infection ability of human neutrophils by regulating its biological behavior in innate immunity.

Improvement situation on indexes of the zebrafish disease model of non-alcoholic fatty liver disease with FGF21 analogues

Objective: To detect the therapeutic efficacy of FGF21 analogues on the zebrafish model of non-alcoholic fatty liver disease. Methods: A zebrafish model of non-alcoholic fatty liver disease was established by providing the normal diet fed to wild-type zebrafish three times daily. PF-05231023 was administered exogenously at a final concentration of 0.5 μmol/L. Body length, body weight, triglycerides, and other indexes were measured after 20 days. Pathological changes were evaluated in liver tissue sections by HE staining. Quantitative PCR was used to identify expressional changes in genes related to lipid metabolism, endoplasmic reticulum stress, and inflammation. Results: QPCR and immunofluorescence staining results showed that FGF21 was highly expressed in the zebrafish model group. The addition of the FGF21 analogue PF-05231023 significantly reduced the body length and body weight (P < 0.01), and the triglyceride content (P < 0.05) in the zebrafish model group. The liver HE staining results showed that PF-05231023 had alleviated the large and tiny bullae fat, lesions, and others in the zebrafish model group. The quantitative PCR results demonstrated that PF-05231023 reduced the expression of lipogenic factors (P < 0.01), inflammatory-related factors (P < 0.001), and genes related to endoplasmic reticulum stress (P < 0.05), but raised lipid-oxidation-related factors (P < 0.05) in the zebrafish model group. The addition of PF-05231023 reduced oleic acid-induced lipid and triglyceride levels in HepG2 cells. Conclusion: FGF21 analogue addition can improve indexes in the zebrafish disease model of non-alcoholic fatty liver disease.

Effect of a dark-colored substrate on the production of phycocyanin by the cyanobacterium Phormidium sp

As the biotechnological production of valuable compounds has recently gathered research interest, more and more photosynthetic microorganisms are being examined for the synthesis of proteins, fatty acids, pigments, and other biopolymers. In this work, a local isolate (lagoons of Western Greece), of the blue-green cyanobacterium Phormidium sp., was evaluated. Its cultures were considered to be promising for the recovery of the phycobiliprotein, called phycocyanin, even during mixotrophic growth. Interestingly, phycocyanin accumulation was favored by substrates with a low organic load. In order to minimize substrate cost, anaerobic digestion effluent was used as a nutrient and organic carbon source, replacing a synthetic medium containing glucose, to meet the cyanobacterium’s nutrient needs. The results from the cultures containing liquid digestate were surprising, as its addition at low concentrations not only allowed Phormidium sp. to grow but also enhanced the phycocyanin synthesis. The experiments that exhibited a notable phycocyanin production showed a final biomass concentration that barely exceeded 0.7 g L−1 regardless of the nature of the substrate. When Phormidium sp. was inoculated in a pure synthetic medium with 1 g L−1 of glucose, phycocyanin concentration approached 10 mg L−1, while it was measured being 1.75 times higher in the culture containing 10% v/v anaerobic effluent, due to greater cellular accumulation. The present work comes as evidence of the successful bioremediation of digestate accompanied by the production of a high added-value pigment, leading to the removal of organic carbon, total nitrogen, and phosphorous in percentages of 60%, 90%, and 40% respectively.

Understanding Drug Skin Permeation Enhancers Using Molecular Dynamics Simulations.

Our skin constitutes an effective permeability barrier that protects the body from exogenous substances but concomitantly severely limits the number of pharmaceutical drugs that can be delivered transdermally. In topical formulation design, chemical permeation enhancers (PEs) are used to increase drug skin permeability. In vitro skin permeability experiments can measure net effects of PEs on transdermal drug transport, but they cannot explain the molecular mechanisms of interactions between drugs, permeation enhancers, and skin structure, which limits the possibility to rationally design better new drug formulations. Here we investigate the effect of the PEs water, lauric acid, geraniol, stearic acid, thymol, ethanol, oleic acid, and eucalyptol on the transdermal transport of metronidazole, caffeine, and naproxen. We use atomistic molecular dynamics (MD) simulations in combination with developed molecular models to calculate the free energy difference between 11 PE-containing formulations and the skin's barrier structure. We then utilize the results to calculate the final concentration of PEs in skin. We obtain an RMSE of 0.58 log units for calculated partition coefficients from water into the barrier structure. We then use the modified PE-containing barrier structure to calculate the PEs' permeability enhancement ratios (ERs) on transdermal metronidazole, caffeine, and naproxen transport and compare with the results obtained from in vitro experiments. We show that MD simulations are able to reproduce rankings based on ERs. However, strict quantitative correlation with experimental data needs further refinement, which is complicated by significant deviations between different measurements. Finally, we propose a model for how to use calculations of the potential of mean force of drugs across the skin's barrier structure in a topical formulation design.

Melatonin supplementation to the freezing medium enhances post-thaw sperm quality and fertility of giant grouper (Epinephelus lanceolatus)

Giant grouper (Epinephelus lanceolatus) is not only favored in the international live reef-fish trade but also a promising species for mariculture in coastal communities throughout Asia. Sperm cryopreservation contributes to the extensive utilization of artificial insemination in grouper aquaculture industry. Inevitable damage during the process of freezing can negatively affect post-thaw sperm quality and fertility, thus hindering its routine application in practice. Melatonin is an antioxidant that can protect sperm from the adverse effects of cryopreservation in a variety of species. This study was aimed to assess the effect of melatonin addition on post-thaw sperm function of giant grouper. Semen samples were cryopreserved with freezing medium containing different final concentrations of melatonin (1, 2 and 4 μM) and the control group (without melatonin). After thawing, multiple sperm functional parameters were evaluated. Generally, a positive effect on cryopreservation was observed for all doses of melatonin. Melatonin treatment at 1 and 2 μM seemed to be efficient in preventing sperm cryodamage, which yielded significantly higher total motility, progressive motility, mitochondrial membrane potential (P < 0.05) and improved velocity (VCL, VAP, VSL) compared to controls. Lower apoptotic-like changes were detected at 2 μM treatment. Samples frozen with melatonin also contributed to greater sperm viability and lesser DNA damage. Although not statistically different from control, a reduction in intracellular ROS was found under the melatonin treatments. Moreover, melatonin supplementation improved the fertility of frozen-thawed sperm, which increased significantly the hatching rate under a limited sperm/egg ratio (10,000:1). Hence, our study suggests inclusion of melatonin to semen extender can improve the post-thaw quality of cryopreserved giant grouper spermatozoa, the recommended concentration of melatonin is 1–2 μM in the cryomedium.

Experimental upscaling analyses for a surfactant-enhanced in-situ chemical oxidation (S-ISCO) remediation design

Surfactant-enhanced in-situ chemical oxidation (S-ISCO) is an emerging innovative remediation technology for the treatment of dense non-aqueous phase liquids (DNAPLs). S-ISCO combines the solubilization of contaminants by means of surfactants with the chemical oxidation by an oxidizing agent, thus, potentially increasing the efficiency of the state-of-the-art ISCO technique. Scientific investigations are needed to enable the technology transfer for potential field applications based on the development of a remediation design under well-defined boundary conditions.For this purpose, experimental upscaling analyses were performed using the special infrastructure of the research facility for subsurface remediation (VEGAS). Batch tests showed that oxidation of the selected surfactant E-Mulse 3® (EM3) by activated persulfate (Na-PS) reduced the solubilization of the model contaminants 1,4-DCB, naphthalene, and PCE. As a consequence, the processes of contaminant solubilization and degradation were temporally and spatially separated in the developed remediation design.A proof of concept was provided by performing an S-ISCO medium-scale experiment (100 cm length, 70 cm height, 12.5 cm width), with 1,2-DCB as model DNAPL contaminant to be treated. A groundwater circulation well (GCW) was used to inject a 60 g/L Na-PS solution and to effectively mix the reagents. Sampling of the experiment's outflow and the soil material after treatment showed that neither rebound effects nor residual mass loadings on the soil material could be detected after termination of the S-ISCO treatment.To further evaluate the S-ISCO remediation design under field-like conditions, a large-scale S-ISCO experiment was conducted (6 m length, 3 m height, 1 m width), allowing for an extensive sampling campaign to monitor relevant processes. An efficient contaminant removal from the former source zone could be reached by surfactant solubilization, decreasing contaminant levels from initially over 2000 mg/L 1,2-DCB to final concentrations below 5 mg/L 1,2-DCB. The heterogeneously distributed contaminant degradation, implemented by a three-filter GCW, was attributed to density-induced migration processes that impeded an optimal reaction zone. A density-dependent numerical transport could qualitatively match the observations. By comparing different simulation scenarios, an adapted operation of the GCW was established that provides for a more efficient distribution of the density-influenced oxidant injection.