Improved localization of thiamine pyrophosphatase activity in mounted cryostat sections using gel fixation and gel incubation media

Summary The presence of 1% agar in the fixation and substrate solutions for the histochemical demonstration of thiamine pyrophosphatase (4.4 mM TPP; 3.6 mM Pb²⁺; 0.025 Tris-maleate buffer, pH 7.2) clearly facilitates the localization of the enzyme in Golgi apparatus in cold microtome sections prepared from unfixed specimens.     

DNA damage induction and antiproliferative activity of vanadium(V) oxido monoperoxido complex containing two bidentate heteroligands

Several peroxidovanadium(V) complexes have been shown as a potent anticancer agents. The aim of this study was to investigate the interaction of monoperoxidovanadium(V) complex Pr₄N[VO(O₂)(ox)(phen)], (Vphen), [phen = 1,10-phenantroline, ox = oxalate(2?) and Pr₄N = tetra(n-propyl)ammonium(1+)] with DNA. UV–Vis spectrophotometry and the alkaline single-cell gel electrophoresis (SCGE, the comet assay) were used to examine the possibility of the vanadium(V) complex to induce changes in DNA. The interaction of Vphen with calf thymus DNA resulted in absorption hyperchromicity in DNA spectrum and shift of the absorption band of DNA to longer wavelengths for the [complex]/[DNA] concentration ratio equals to 4 and after 60 min of incubation. The rise in DNA absorption (by 34%) and bathochromic shift (Δλ_max = 6 nm) are indicative of the interaction between DNA and the complex molecules. DNA strand breaks in cellular DNA were investigated using the comet assay. The human lymphocytes were exposed to various concentrations of Vphen for 30 min. The results revealed that Vphen contributed to the DNA damage expressed as DNA strand breaks in concentration dependent manner. The used concentrations of Vphen (ranging from 0.1 to 100 μmol/L) caused higher DNA damage in lymphocytes compared to untreated cells (from 1.2 times for 0.1 μmol/L to 1.8 times for 100 μmol/L). Vphen was screened for its potential antitumor activity towards murine leukemia cell line L1210. Vphen exhibited significant antiproliferative activity depending on its concentration and time of exposure. The IC₅₀ values were 0.247 μg/mL (0.45 μmol/L) for 24 h, 0.671 μg/mL (1.21 μmol/L) for 48 h and 0.627 μg/mL (1.13 μmol/L) for 72 h.     

Challenges facing European agriculture and possible biotechnological solutions

Agriculture faces many challenges to maximize yields while it is required to operate in an environmentally sustainable manner. In the present study, we analyze the major agricultural challenges identified by European farmers (primarily related to biotic stresses) in 13 countries, namely Belgium, Bulgaria, the Czech Republic, France, Germany, Hungary, Italy, Portugal, Romania, Spain, Sweden, UK and Turkey, for nine major crops (barley, beet, grapevine, maize, oilseed rape, olive, potato, sunflower and wheat). Most biotic stresses (BSs) are related to fungi or insects, but viral diseases, bacterial diseases and even parasitic plants have an important impact on yield and harvest quality. We examine how these challenges have been addressed by public and private research sectors, using either conventional breeding, marker-assisted selection, transgenesis, cisgenesis, RNAi technology or mutagenesis. Both national surveys and scientific literature analysis followed by text mining were employed to evaluate genetic engineering (GE) and non-GE approaches. This is the first report of text mining of the scientific literature on plant breeding and agricultural biotechnology research. For the nine major crops in Europe, 128 BS challenges were identified with 40% of these addressed neither in the scientific literature nor in recent European public research programs. We found evidence that the private sector was addressing only a few of these “neglected” challenges. Consequently, there are considerable gaps between farmer’s needs and current breeding and biotechnology research. We also provide evidence that the current political situation in certain European countries is an impediment to GE research in order to address these agricultural challenges in the future. This study should also contribute to the decision-making process on future pertinent international consortia to fill the identified research gaps.     

Interaction of cucumber mosaic virus and potato virus y with tobacco mosaic virus

A study was performed on the interaction of cucumber mosaic virus (CMV) of potato virus Y (PVY) with tobacco mosaic virus (TMV). Interference was evaluated using tobacco plantsNicotiana tabacum cv. Java responding to CMV and PVY with a systemic infection and to TMV with local necrotic lesions. The decrease in TMV — induced lesion number gave evidence of a decrease in susceptibility caused by the previous infection with CMV or PVY, the decrease of lesion enlargement demonstrated a decreased TMV reproduction in the plants previously infected with CMV or PVY. The interference concerned was incomplete, as evaluated from reproduction of the challenging TMV and from the decrease in susceptibility of the host to TMV brought about by the first infection with CMV or PVY.     

Antioxidative activity of selected fruits and vegetables

Recent studies have demonstrated that dietary plants are rich source of antioxidants and can contribute to the protection from age-related diseases. The aim of our study was to determine the total antioxidant capacity of extracts from different kinds of fruits and vegetables, and to examine their inhibitory effect on the oxidative damage to proteins in vitro. For determination of antioxidant capacity we used two direct methods. Among the food materials chosen for the present study, blueberries and red beets gave the maximum antioxidant activity. The lowest activity was determined in pears and green beans. Some extracts were more active in one method, while their activity was lower using the other method. To investigate inhibitory effects of fruits and vegetables extracts on the oxidative damage to proteins in vitro, we induced the oxidative damage to plasma proteins by sodium hypochlorite leading to formation of carbonyl compounds detected by spectrophotometric method. All extracts of fruits and vegetables showed inhibitory activity on the oxidative damage to proteins with raspberries and leek as most effective. Results of this study will be useful as an aid for dietary choices to increase antioxidant intake and will allow the investigation of the relation between dietary antioxidants and oxidative stress-induced diseases.     

Rocks and Walls: Natural Versus Secondary Habitats

Walls are often considered secondary habitats for vegetation of natural rock surfaces. Compared with rocks, walls differ in many features, for example the presence of a binding material (mainly calcareous mortar), location in settlements and exposure to human impact. A data set of 1,205 phytosociological relevés recorded on horizontal wall tops, wall verticals and rock verticals in the Czech Republic was used to compare their vegetation with regard to i) species composition (frequent species, species diversity, endangered and alien species) and ii) the ecological requirements of the respective species. Gamma diversity of vascular plant species was comparable in all habitat types (242 species on wall tops, 212 species on wall verticals and 197 species on rock verticals). Wall verticals had higher beta diversity, but lower alpha diversity than rocks. Remarkable differences were found comparing the diversity of alien species. Whereas alpha and gamma diversities of aliens were higher on both wall habitats, beta diversity of aliens was the highest on rocks. The high floristic heterogeneity of walls is mainly attributable to the large pool of species from the surrounding urbanized landscape (e.g., cultivated ornamental species and synanthropic weeds) that are favoured by high nutrient inputs. In contrast, species characteristic of rocks are mainly substrate specialists. Walls and rocks share the frequent occurrence of ferns, grasses and herbs typical of forest understorey and clearings. Compared with rocks, walls are generally colonized by species requiring higher nutrient content, soil reaction, temperature and moisture. Secondary wall habitats might be suitable for some rare and endangered species, but contrary to rocks their occurrences are only accidental and temporary. The representation of aliens was considerably higher on walls (approximately 35%) than on rocks (9%).     

Single and multiple dose pharmacokinetics of maritime pine bark extract (Pycnogenol) after oral administration to healthy volunteers

Background

Since plant extracts are increasingly used as phytotherapeutics or dietary supplements information on bioavailability, bioefficacy and safety are warranted. We elucidated the plasma kinetics of genuine extract components and metabolites after single and multiple ingestion of the standardized maritime pine bark extract Pycnogenol (USP quality) by human volunteers.

Methods

Eleven volunteers received a single dose of 300 mg pine bark extract, five volunteers ingested 200 mg daily for five days to reach steady state concentrations. Plasma samples were obtained before and at defined time points after intake of the extract. Samples were analyzed by HPLC with ion-pair reagents and simultaneous UV and electrochemical detection.

Results

We quantified total plasma concentrations of catechin, caffeic acid, ferulic acid, taxifolin and the metabolite M1 (δ-(3,4-dihydroxy-phenyl)-γ-valerolactone). Additionally, we describe plasma time courses and steady state appearance of ten so far unknown compounds, U1 to U10. After single ingestion, compounds derived from the extract were rapidly absorbed and the majority of them were detectable over whole experimental period of 14 h. The analysis of steady state plasma samples revealed significant phase II metabolism.

Conclusion

We present the first systematic pharmacokinetic analysis of compounds derived from maritime pine bark extract. Beyond the known constituents and metabolites we uncovered the plasma time courses of ten unknown compounds. In concert with our previous detection of anti-inflammatory bioefficacy of these plasma samples ex vivo we suggest that constituents and metabolites of Pycnogenol bear potential for disclosure of novel active principles.     

Survival of Bacteria in Lyophilized Plasma

Although lyophilization reduces the bacterial population in plasma, some species survive for several months. Sensitivity or resistance to lyophilization in plasma is a characteristic of each morphological type, with gram-negative species being more sensitive to destruction than cocci or sporulating gram-positive species. Reconstituted preparations of lyophilized plasma must be incubated for 24 hr to prevent false-negative sterility tests. The killing process in lyophilized plasma continues with time if the preparations are stored at room temperature. Strict asepsis must be realized during processing of plasma because lyophilization alone does not destroy the microorganisms present.