Fluorimetric determination of febuxostat in dosage forms and in real human plasma via Förster resonance energy transfer

A rapid, simple, selective and precise fluorimetric method was developed and validated for determination of a selective xanthine oxidase inhibitor; febuxostat (FBX) in pharmaceutical formulations and in human plasma. The proposed method is based on quenching effect of FBX on the fluorescence intensity of terbium (Tb³⁺) through fluorescence resonance energy transfer (FRET) from Tb³⁺ to FBX. The formed complex was measured at λ_ex. 320 nm/λ_em. 490 nm against a reagent blank. Fluorescence intensity of Tb³⁺ was diminished when FBX was added. A linear relationship between the fluorescence quenching value of the formed complex and the concentration of FBX was investigated. The reaction conditions and the fluorescence spectral properties of the complex have been studied. The linearity range of the developed method was 1.0–16.0 μg/ml. The suggested method was applied successfully for the estimation of FBX in bulk powder, dosage forms and spiked plasma samples with excellent recoveries (96.79–98.89%). In addition, the developed method has been successfully applied for determination of FBX in real plasma samples collected from healthy volunteers with good recoveries (82.06–85.65%). All obtained results of the developed method were statistically analyzed and validated according to ICH (International Conference on Harmonization) guidelines.     

Species richness,abundance, distributional pattern and trait composition of butterfly assemblage change along an altitudinal gradient in the Gulmarg region of Jammu & Kashmir,India

Despite enormous diversity, abundance, and role in ecosystem processes, little is known about how butterflies differ across altitudinal gradients. For this, butterfly communities were investigated along an altitudinal gradient of 2700–3200 m a.s.l, along the Gulmarg region of Jammu & Kashmir, India. We aimed to determine how the altitudinal gradient and environmental factors affect the butterfly diversity and abundance. Our findings indicate that species richness and diversity are mainly affected by the synergism between climate and vegetation. Alpha diversity indices showed that butterfly communities were more diverse at lower elevations and declined significantly with increase in elevation. Overall, butterfly abundance and diversity is stronger at lower elevations and gradually keep dropping towards higher elevations because floristic diversity decreased on which butterflies rely for survival and propagation. A total of 2023 individuals of butterflies were recorded belonging to 40 species, represented by 27 genera and 05 families. Six survey sites (S I- S VI) were assessed for butterfly diversity from 2018 to 2020 in the Gulmarg region of Jammu & Kashmir. Across the survey, Nymphalidae was the most dominant family represented by 16 genera and 23 species, while Papilionidae and Hesperiidae were least dominant represented by 01 genera and 01 species each. Among the six collection sites selected, Site I was most dominant, represented by 16 genera and 21 species, while Site VI was least dominant, represented by 04 genera and 04 species.     

Unique properties of purified, Escherichia coli-expressed constitutive cytochrome P4504A5.

Cytochromes P450 of the 4A family metabolize a variety of fatty acids, prostaglandins, and eicosanoids mainly at the terminal carbon (omega-hydroxylation) and, to a lesser extent, at the penultimate carbon [(omega-1)-hydroxylation]. In the present study, cytochrome P4504A5 (4A5) has been successfully expressed in Escherichia coli, with an average yield of enzyme of approximately 80 nmol/liter of cells. Spectroscopic characterization of the purified enzyme, using electron paramagnetic resonance and absolute and substrate-perturbed optical difference spectroscopy, showed that the heme of resting 4A5 is primarily low spin, but is converted primarily to high spin by substrate binding. The kcat and Km values for laurate omega-hydroxylation were 41 min-1 and 8.5 microM, respectively, in the absence of cytochrome b5, and 138 min-1 and 38 microM, respectively, in the presence of cytochrome b5. Hydroxylation of palmitate was dependent on the presence of cytochrome b5; kcat and Km values were 48 min-1 and 122 microM, respectively. Hydroxylation of arachidonic acid was barely detectable and was unchanged by the addition of cytochrome b5.     

Zoonotic agents in small ruminants kept on city farms in southern Germany

Sheep and goats are popular examples of livestock kept on city farms. In these settings, close contacts between humans and animals frequently occur. Although it is widely accepted that small ruminants can carry numerous zoonotic agents, it is unknown which of these agents actually occur in sheep and goats on city farms in Germany. We sampled feces and nasal liquid of 48 animals (28 goats, 20 sheep) distributed in 7 city farms and on one activity playground in southern Germany. We found that 100% of the sampled sheep and 89.3% of the goats carried Shiga toxin-producing Escherichia coli (STEC). The presence of Staphylococcus spp. in 75% of both sheep and goats could be demonstrated. Campylobacter spp. were detected in 25% and 14.3% of the sheep and goats, respectively. Neither Salmonella spp. nor Coxiella burnetii was found. On the basis of these data, we propose a reasonable hygiene scheme to prevent transmission of zoonotic agents during city farm visits.     

Magma intrusion in the upper crust of the Abu Dabbab area, South East of Egypt from Vp and Vp/Vs tomography

3-Dimages of P-wave velocity and Vp/Vs ratio have been produced for the upper crust of the Abu Dabbab area, North Mars Alam city. The inversion of local travel times of high quality data recorded at eleven mobile seimic stations around the study area is carried out. The best, in the least-squares sense, 1-D Vp model and the average value of Vp/Vs (1.72) were computed as prerequisites of the 3-D inversion that reaches a depth of 14 km. From the 3-D model it is evident that the distributions of Vp and Vp/Vs are characterized by marked lateral and vertical variations delineating structural heterogeneities. Due to the presence of a thin layer of sedimentary rocks saturated with surface water, low P-wave velocity and high Vp/Vs values are noticed near the surface. At greater depths, high Vp and low Vp/Vs zones may indicate crustal rocks with relatively higher rigidity and brittle behavior, while high Vp/Vs and low Vp may identify zones of relatively softer rocks, with ductile behavior. Low P-wave velocity values are observed at the intersections among the faults. A possibility of some magma intrusion could be associated with the high Vp/Vs values which form an elongated anomaly, in the central part of the study area, which extends from a depth of 12 km to about 1–2 km of depth. If the obtained 3-D model is used in the relocation of selected events, they turn out to be strongly clustered in correspondence with the high velocity anomalies detected in the central part of the study area. Most of the seismicity tends to occur at the boundaries between the high and low velocity anomalies and at pre-existing weakness zones, i.e. the areas of intersection among different faults. The occurrence of the seismic activity in the vicinity of low velocity anomalies and at the boundary between velocity contrast could also be explained by the occurrence of serpentinization processes in the crust of the study area.      

miR-615-5p is restrictedly expressed in cirrhotic and cancerous liver tissues and its overexpression alleviates the tumorigenic effects in hepatocellular carcinoma

microRNAs aberrant behavior in heptocellular carcinoma (HCC) plays a major role in HCC pathogenesis. miR-615-5p expression has never been evaluated in HCC. We showed that miR-615-5p was preferentially expressed in HCC, cirrhotic liver tissues and HCC cell lines, but undetected in normal livers. Forced miR-615-5p expression in HCC cell lines led to significant decrease in cell growth and migration. In-silico predication revealed insulin-like growth factor-II (IGF-II) as a potential downstream target for miR-615-5p. Forcing the expression of miR-615-5p showed downregulation of IGF-II mRNA, as well as inhibition of the luciferase activity in a luciferase reporter vector harboring the IGF-II-3′UTR target sequence. miR-615-5p acts as tumor-suppressor in HCC through targeting IGF-II.     

Dendrocyin: an isocucurbitacin with novel cyclic side chain from Dendrosicyos socotrana

Dendrosicyos socotrana Balf.f. is a unique species (Cucurbitaceae) native to Socotra island in the horn of Africa. From the chloroform extract of the stems, A new isocucurbitacin (Dendrocyin) with unusual cyclization in the side chain; 24beta-ethoxy-20-25-epoxy-3alpha,16alpha-dihydroxy-9-methyl-19-norlanost-5(6) ene-2,11,22-trione has been isolated alongside isocucurbitacin R. Their structural configuration were established by usual spectroscopic (1H NMR, 13C NMR and DEPT) and two-dimensional NMR techniques (1H-1H Cosy, HMBC and HMQC).     

Regulators of G-protein signaling form a quaternary complex with the agonist, receptor, and G-protein. A novel explanation for the acceleration of signaling activation kinetics

Regulators of G-protein signaling (RGS) proteins modulate signaling through heterotrimeric G-proteins. They act to enhance the intrinsic GTPase activity of the Galpha subunit but paradoxically have also been shown to enhance receptor-stimulated activation. To study this paradox, we used a G-protein gated K+ channel to report the dynamics of the G-protein cycle and fluorescence resonance energy transfer techniques with cyan and yellow fluorescent protein-tagged proteins to report physical interaction. Our data show that the acceleration of the activation kinetics is dissociated from deactivation kinetics and dependent on receptor and RGS type, G-protein isoform, and RGS expression levels. By using fluorescently tagged proteins, fluorescence resonance energy transfer microscopy showed a stable physical interaction between the G-protein alpha subunit and RGS (RGS8 and RGS7) that is independent of the functional state of the G-protein. RGS8 does not directly interact with G-protein-coupled receptors. Our data show participation of the RGS in the ternary complex between agonist-receptor and G-protein to form a "quaternary complex." Thus we propose a novel model for the action of RGS proteins in the G-protein cycle in which the RGS protein appears to enhance the "kinetic efficacy" of the ternary complex, by direct association with the G-protein alpha subunit.     

Balanitoside, a furostanol glycoside, and 6-methyldiosgenin from Balanites aegyptiaca.

In addition to a known spirostanol glycoside, balanitin-3, and a new sapogenol, 6-methyldiosgenin, a new furostanol saponin, balanitoside has been isolated from the fruits (mesocarp) of Balanites aegyptiaca. The structure of the glycoside has been determined as 26-O-beta-D-glucopyranosyl-3 beta, 22,26-trihydroxy-furost-5-ene 3-O-alpha-L-rhamnopyranosyl-(1-->2)-beta-D- glucopyranosyl-(1-->4)-beta-D-glucopyranoside, on the basis of spectroscopic and chemical evidence.