Structure-Activity Relationship of Methyl 4-Aminobenzoate Derivatives as Being Drug Candidate Targeting Glutathione Related Enzymes: in Vitro and in Silico Approaches

A thiol compound, glutathione, is essential for healthy cell defence against xenobiotics and oxidative stress. Glutathione reductase (GR) and glutathione S-transferase (GST) are two glutathione-related enzymes that function in the antioxidant and the detoxification systems. In this study, potential inhibitory effects of methyl 4-aminobenzoate derivatives on GR and GST were examined in vitro. GR and GST were isolated from human erythrocytes with 7.63 EU/mg protein and 5.66 EU/mg protein specific activity, respectively. It was found that compound 1 (methyl 4-amino-3-bromo-5-fluorobenzoate with K_i value of 0.325±0.012 μM) and compound 5 (methyl 4-amino-2-nitrobenzoate with K_i value of 92.41±22.26 μM) inhibited GR and GST stronger than other derivatives. Furthermore, a computer-aided method was used to predict the binding affinities of derivatives, ADME characteristics, and toxicities. Derivatives 4 (methyl 4-amino-2-bromobenzoate) and 6 (methyl 4-amino-2-chlorobenzoate) were estimated to have the lowest binding energies into GR and GST receptors, respectively according to results of in silico studies.     

In Silico Design of an Oseltamivir Derivative with Increased Affinity against Wild-Type and Mutant Variants of Neuraminidase and Hemagglutinin of Influenza A H1N1 Virus

Antiviral resistance has turned into a world concern nowadays. Influenza A H1N1 emerged as a problem at the world level due to the neuraminidase (NA) mutations. The NA mutants conferred resistance to oseltamivir and zanamivir. Several efforts were conducted to develop better anti-influenza A H1N1 drugs. Our research group combined in silico methods to create a compound derived from oseltamivir to be tested in vitro against influenza A H1N1. Here we show the results of a new compound derived from oseltamivir but with specific chemical modifications, with significant affinity either on NA (in silico and in vitro assays) or HA (in silico) from influenza A H1N1 strain. We include docking and molecular dynamics (MD) simulations of the oseltamivir derivative at the binding site onto NA and HA of influenza A H1N1. Additionally, the biological experimental results show that oseltamivir derivative decreases the lytic-plaque formation on viral susceptibility assays, and it does not show cytotoxicity. Finally, oseltamivir derivative assayed on viral NA showed a concentration-dependent inhibition behavior at nM, depicting a high affinity of the compound for the enzyme, corroborated with the MD simulations results, placing our designed oseltamivir derivative as a potential antiviral against influenza A H1N1.     

HPLC-DAD Analysis and Versatile Bioactivities of Turkish Sunflower Honeys Using Chemometric Approaches

Sunflower honey (SH) is bright yellow, fragrant, pollen-flavoured, slightly herbaceous and has a unique taste. The present research aims to examine the enzyme inhibitory, antioxidant, anti-inflammatory, antimicrobial and anti-quorum sensing activities and phenolic compositions of 30 sunflower honeys (SHs) produced from several regions of Turkey with chemometric study. SAH from Samsun exhibited the best antioxidant activity in β-carotene linoleic acid (IC₅₀: 7.33±0.17 mg/mL) and CUPRAC (A_0.50: 4.94±0.13 mg/mL) assays, anti-urease activity (60.63±0.87 %) and anti-inflammatory activity against COX-1 (73.94±1.08 %) and COX-2 (44.96±0.85 %). SHs exhibited mild antimicrobial activity against the test microorganisms while they showed high quorum sensing inhibition zones measured in the range of 42–52 mm against the CV026 strain. The phenolic composition was determined by high performance liquid chromatography with diode array detection (HPLC-DAD) system and levulinic, gallic, p-hydroxybenzoic, vanillic and p-coumaric acids were identified in all studied SHs. The classification of SHs was performed the using PCA and HCA. This study revealed that phenolic compounds and biological properties are effective in classification of SHs according to their geographical origin. The results suggest that studied SHs could be valued as potential agents with versatile bioactivities in oxidative stress-related disease, microbial infections, inflammation, melanoma, and peptic ulcer.     

Energy Metabolism During Late Gestation and Lactation in Muciparous Sows in Relation to Backfat Thickness and the Interval from Weaning to First Oestrus

Ten crossbred, fourth or fifth parity sows were divided into 2 groups - high (H) and low (L) - according to their backfat thickness 9 days before parturition. Body weight, backfat thickness and litter weight were recorded repeatedly during a 5 week lactation period. The length of the interval from weaning to first oestrus was also noted. All sows were fed a commercial diet (11.9 MJ/kg, 14.5% crude protein). During gestation, daily food intake was 2.2 kg/sow, while during lactation it was 3.0 kg/sow plus 0.4 kg/piglet. Blood samples were drawn on day 9 before parturition and on days 2,7,14 and 21 of lactation. The samples were analysed to determine concentrations of glucose, urea nitrogen, creatinine, triglycerides, free fatty acids and beta-hydroxybutyric acid. In both groups, concentrations of free fatty acids and urea nitrogen were low on day 9 before parturition while those of triglycerides were high, indicating anabolism regardless of backfat thickness. During the first week of lactation, concentrations of free fatty acids increased in the H-group but not in the L-group, and concentrations of urea nitrogen were higher in the H-group. These differences, together with the greater loss of weight observed in the H-group, indicate that catabolism of maternal fat and protein depots was more pronounced in the Η-group than in the L-group during this time. On day 14 of lactation, both groups showed equally low concentrations of free fatty acids, decreasing creatinine concentrations and stable triglyceride and urea nitrogen concentrations. Furthermore, weight loss during the second and third weeks of lactation was low in both groups. These facts, taken together, indicate that the catabolic rate was decreasing in both groups during this period. No differences in return to oestrus interval were noted between the groups. The present study indicates that under a restricted feeding regime the catabolic rate during the first week of lactation is higher in sows with higher backfat thickness in late gestation. As lactation progresses, a more balanced metabolism is achieved regardless of backfat thickness, which may tend to reduce differences in return to oestrous interval.     

Nutraceutical Activity of Anthocyanins from the Edible Berries of Rhamnus pompana

We present the inhibitory properties of the R. pompana anthocyanin fraction (RPAF) and its major constituents on alpha-glucosidase (AG), pancreatic lipase (PL), HMG-CoA reductase, and ornithine decarboxylase (ODC). The effect of RPAF was also evaluated in ICR male mice subjected to oral glucose tolerance test (OGTT) and hypercaloric/atherogenic diet for 30 days. RP-HPLC/MS profiling revealed that RPAF contained five major anthocyanins and induced slight inhibition on PL and HMG-CoA reductase (IC₅₀, 245–338 μg mL⁻¹) whereas strong activity on AG and ODC (IC₅₀, 130–133 μg mL⁻¹) was observed. Kinetic studies and molecular docking with pelargonidin-3-O-rutinoside (P3R) on ODC, revealed changes in K_m (0.9514–0.9746 mM) and V_max (1.96–2.32 μmol mg⁻¹ min⁻¹) suggesting mixed inhibition and molecular interaction with two active sites of ODC. P3R showed antiproliferative activity (IC_50, 46.5 μM) and decreased polyamine accumulation in DLD-1 cells. The results of OGTT confirmed that RPAF regulates postprandial glucose levels in diabetic animals which experienced a significant glucose depletion (30 %; p<0.001) from 30 to 120 min post-treatment. Prolonged supplementation of RPAF caused significant decrease (p<0.001) in plasma glucose, total cholesterol, LDL-c and triglycerides as well as significant increase (p<0.001) of HDL-c compared with normoglycemic untreated animals.     

A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex-determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.     

Genome-wide RAD sequencing data suggest predominant role of vicariance in Sino-Japanese disjunction of the monotypic genus Conandron (Gesneriaceae)

Disjunct distribution is a key issue in biogeography and ecology, but it is often difficult to determine the relative roles of dispersal vs. vicariance in disjunctions. We studied the phylogeographic pattern of the monotypic Conandron ramondioides (Gesneriaceae), which shows Sino-Japanese disjunctions, with ddRAD sequencing based on a comprehensive sampling of 11 populations from mainland China, Taiwan Island, and Japan. We found a very high degree of genetic differentiation among these three regions, with very limited gene flow and a clear Isolation by Distance pattern. Mainland China and Japan clades diverged first from a widespread ancestral population in the middle Miocene, followed by a later divergence between mainland China and Taiwan Island clades in the early Pliocene. Three current groups have survived in various glacial refugia during the Last Glacial Maximum, and experienced contraction and/or bottlenecks since their divergence during Quaternary glacial cycles, with strong niche divergence between mainland China + Japan and Taiwan Island ranges. Thus, we verified a predominant role of vicariance in the current disjunction of the monotypic genus Conandron. The sharp phylogenetic separation, ecological niche divergence among these three groups, and the great number of private alleles in all populations sampled indicated a considerable time of independent evolution, and suggests the need for a taxonomic survey to detect potentially overlooked taxa.     

Effect of exogenous gibberellin A4/7 on tracheid production, longitudinal growth and the levels of indole‐3‐acetic acid and gibberellins A4, A7 and A9 in the terminal shoot of Pinus sylvestris seedlings

Synchronously dividing cultures of the unicellular green alga Scenedesmus obtusiusculus were cultivated for 24 or 70 h in medium high (1000 μM) or low (60 μM) in phosphorus. Aliquots of AlCl₃ (0, 37, 74, 111, 148, 185, or 222 μmol) were added daily to 1 l cell suspension at the end of the cell division phase. Algae were also grown in media with different pH, adjusted with HCl, in the absence of AlCl₃.
Effects of Al on cell metabolism vary with the intracellular Al concentration and with the concentration of Al available per cell. When the concentration of phosphorus is low, internal concentrations of Al are high and the chlorophyll content and the net dry matter production per cell increase, whereas the photosynthesis and the cell division are increased. Presence of Al in a low P medium decreases the pH of the medium down to 4.5. There are only small effects of Al in the presence of P, due to precipitation of most of the Al with P in the medium.
Despite the Al-induced decrease of the pH of the culture medium, effects caused by Al cannot be explained as a pH effect. Instead, the Al effect may, at least to some extent, be related to a decrease in availability of P in the metabolism, due to formation of aluminium phosphate inside the cell.     

Lincomycin-induced alteration in the contents of chlorophyll-protein complexes of dimorphic maize chloroplasts and its effect on the temperature-induced spectral changes

The difference spectroscopy technique has been utilized to investigate the temperature-induced spectral changes in mesophyll and bundle sheath chloroplasts of maize ( Zea mays L. cv. Ganga-5) in order to assess the role of different pigment-protein complexes in the manifestation of temperature effect on the chloroplast membranes. Cooling and heating of both mesophyll and bundle sheath chloroplasts resulted in absorbance difference (AA) bands at similar wavelengths but the degree of absorb-ance changes were significantly higher in bundle sheath chloroplasts. For example, upon cooling to 7-8°C, positive AA bands were observed at 440, 490 and 680 nm in mesophyll chloroplasts and at 440, 495–500 and 680 nm in bundle sheath chloroplasts but the absorbance change at 680 nm was ca 2% in mesophyll chloroplasts, whereas it was ca 5% in bundle sheath chloroplasts, which have a lower content of light-harvesting pigment-protein complex. The role of chlorophyll-protein complexes was further investigated by monitoring the temperature-induced spectral changes of mesophyll and bundle sheath chloroplasts isolated from lincomycin-treated maize plants where lincomycin selectively inhibits the biosynthesis of specific chlorophyll-protein complexes. Results indicated that depletion of certain pigment-protein complexes in mesophyll chloroplasts made them more susceptible (a ca 4% vs ca 2% absorbance change upon cooling and a ca 6% vs ca 4% absorbance change upon heating) and less tolerant to temperature variation (a 76% vs 39% reversibility during ambient→Cooling→ambient temperature cycle). The data indicate that pigment-protein complexes play a significant role in protecting the chloroplast membranes against temperature variation.     

Fine Structure of the Tentacles of Phoronis australis Haswell (Phoronida,Lophophorata)

The epidermis of the tentacles of Phoronis australis consists of six cell types: supporting cells, choanocyte-like sensory cells, both types monociliated, secretory A-cells with a mucous secretion, and three kinds of B-cells with mucoprotein secretions. On cross-sections of the tentacle, one can distinguish four faces: the frontal one, heavily ciliated and located between the two frontolateral rows of sensory cells, the lateral and the abfrontal ones. The orientation of the basal structures of the cilia is related to the direction of their beat. The basiepidermal nervous system is grouped mainly at the frontal and abfrontal faces. The basement membrane is thickest on the frontal face and consists of circular collagen fibrils near the epidermis and longitudinal ones near the peritoneum. All peritoneal cells surrounding the mesocoel are provided with smooth longitudinal myofibrils, and isolated axons are situated between these cells and the basement membrane. The wall of the single blood capillary in each tentacle consists of epitheliomuscular cells with circular myofilaments, lying on a thin internal basal lamina; there is no endothelium.