Plant P5C reductase as a new target for aminomethylenebisphosphonates

A series of N-substituted derivatives of aminomethylenebisphosphonic acid were evaluated as potential inhibitors of delta1-pyrroline-5-carboxylate reductase (EC 1.5.1.2), the enzyme that catalyzes the last step in proline biosynthesis, partially purified from Arabidopsis thaliana suspension cultured cells. At millimolar concentrations, three compounds out of 26 were found to interfere with the catalytic mechanism. One of them, namely, 3,5-dichloropyridyl-aminomethylenebisphosphonic acid, retained such inhibitory activity in the micromolar range. Kinetic analyses ruled out the possibility that the inhibition could simply rely upon the chelating properties of bisphosphonates and showed mechanisms of a noncompetitive type against NADH and an uncompetitive type against delta1-pyrroline-5-carboxylic acid, with KI values of 199 +/- 6 and 10.3 +/- 1.5 microM, respectively. A computer-aided docking analysis, performed on the basis of the crystal structure of the enzyme from Streptococcus pyogenes, suggested that this phosphonate may interact with amino acid residues near the binding site of delta1-pyrroline-5-carboxylic acid, thus blocking the substrate in a pocket and preventing its interaction with NADH. Because in higher plants the step catalyzed by delta1-pyrroline-5-carboxylate reductase is shared by all pathways leading to proline synthesis, such a compound may represent a lead structure to be exploited for the design of new substances endowed with herbicidal activity.     

STABILITY,NITROGEN MINERALIZATION CAPACITY AND AGRONOMIC VALUE OF COMPOST-BASED GROWING MEDIA FOR LETTUCE CULTIVATION

Seven composts were used as growing media for lettuce mixed with peat at 25 and 50% (v/v). On the unblended composts and the resulting 14 mixes were determined the main physical-chemical characteristics and the stability by means of a respirometric test in a liquid environment. The potential nitrogen (N)-mineralization capacity of the mixes was measured during the respirometric test solely by the ammonium (NH⁺ ₄-N) determination. The lettuce pot-growing test was performed with and without fertilization; plant biomass and total Kjeldahl nitrogen (TKN) tissue content were measured after cultivation for two months. Compost origin, rate in substrates and fertilization affected plant growth and nitrogen uptake. Multiple regression analysis showed that the stability, proved to be a good predictor for plant growth inhibition. Moreover N-mineralization capacity showed a good fitting with plant uptake. The coupled stability and nitrogen mineralization test gives reliable information about the potential constraints in compost-based growing media.     

Circulating Salicylic Acid and Metabolic and Inflammatory Responses after Fruit Ingestion

We hypothesized that fruit ingestion provides measurable amounts of salicylic acid (SA) and produces different metabolic and inflammatory responses compared to mere fruit sugars. In a randomized-crossover study, 26 healthy subjects received a peach shake meal (PSM) (SA: 0,06 ± 0,001 mg/100 g) and a mixed sugar meal (MSM), consisting in an aqueous solution with the same sugars found in the peach shake. In order to control for the SA contribution from meals in the previous day, 16 subjects (Group 1) abstained from fruits and vegetables consumption the evening before trials, and 10 subjects (Group 2) maintained their usual diet. Circulating SA, glucose, insulin, free fatty acids, and interleukin-6 were determined. Basal SA was lower in Group 1 than in Group 2 (0.09 ± 0.02 vs. 0.30 ± 0.03 μmol/l, p < 0.001), peaked at 90 min in both groups (0.18 ± 0.01 vs. 0.38 ± 0.02 μmol/l, p < 0.01) and remained above baseline (p < 0.05) up to 3 h. Glycemia increased less after PSM at 15 min (p < 0.01) with a lower average glucose excursion (p < 0.05). Insulin peaked at 45 min with both meals but decreased less rapidly with PSM. Free fatty acids decreased more (p < 0.01), and interleukin-6 increased less (p < 0.05) with PSM. Dietary fruit intake increases the concentration of SA in vivo, and provides non-nutrients capable to modulate the inflammatory and metabolic responses to carbohydrates.     

Characterization of Fungal Biodiversity In Compost and Vermicompost

The ability of fungi to break down complex carbon sources makes them of vital importance in both the generation and application of compost. This paper illustrates the diversity and functions of the mycoflora of both a green compost (made solely thermophilically from plant debris) and a vermicompost (made mesophilically by the action of earthworms on plant and animal wastes). The soil dilution plate technique was applied on 3 media (PDA, CMC, PDA plus cycloheximide) and at 3 incubation temperatures (24°C, 37°C and 45°C) to isolate and identify fungal entities. Enzymatic activities (amylase, cellulase, chitinase, esterase, ligninase, pectinase, phosphatase, plastic degradation and xylanase) of most species from both composts were evaluated with a semiquantitative method on 15 substrates. There were substantial qualiquantitative differences in the species composition of the two composts. A total of 193 entities were isolated: 54 from green compost only, 77 from vermicompost only, 62 from both. This taxonomic diversity was reflected in the metabolic potential. Amilase, cellulase, protease and esterase activities were significantly higher in the green compost. Lignin and plastic polymer degradation were significantly higher in the vermicompost. Structural and functional characterization of this kind is of assistance in determining both the most appropriate application of composts and their hygienic quality.     

Protective effects of extra virgin olive oil phenolics on oxidative stability in the presence or absence of copper ions

The antioxidant activity of the phenolic fraction of extra virgin olive oil was assessed in samples that had a decreasing content of antioxidants in the presence and absence of copper ions as a catalyst of autoxidation. The oxidation process was evaluated by measuring primary and secondary oxidation products. Changes in phenols and tocopherols were investigated by high-performance liquid chromatography. Both the total phenol content and their antioxidant activity were monitored by spectrophotometric assays (with Folin-Ciocalteu and ABTS*+ reagents). The important role of phenolic compounds (particularly the o-diphenols) in protection from autoxidation was confirmed. However, the tocopherols were more quickly consumed in oils that had the lowest content of o-diphenols, which also showed evidence of an ability to chelate copper. In particular, a dramatic decrease was observed in the isomeric form of decarboxymethyl-oleuropein aglycone after addition of the metal, despite its significant increase in samples stored in the absence of copper.     

Minor components of olive oil modulate proatherogenic adhesion molecules involved in endothelial activation

The Mediterranean diet reduces the risk of coronary artery disease as a consequence of its high content of antioxidants, namely, hydroxytyrosol (HT) and oleuropein aglycone (OleA), typical of virgin olive oil. Because intercellular and vascular cell adhesion molecules (ICAM-1 and VCAM-1) and E-selectin are crucial for endothelial activation, the role of the phenolic extract from extra virgin olive oil (OPE), OleA, HT, and homovanillyl alcohol (HVA) on cell surface and mRNA expression in human umbilical vascular endothelial cells (HUVEC) was evaluated. OPE strongly reduced cell surface expression of ICAM-1 and VCAM-1 at concentrations physiologically relevant (IC50 < 1 microM), linked to a reduction in mRNA levels. OleA and HT were the main components responsible for these effects. HVA inhibited cell surface expression of all the adhesion molecules, whereas the effect on mRNA expression was weaker. These results supply new insights on the protective role of olive oil against vascular risk through the down-regulation of adhesion molecules involved in early atherogenesis.     

Effectiveness and mode of action of phosphonate inhibitors of plant glutamine synthetase

BACKGROUND: Aiming at the rational design of new herbicides, the availability of the three‐dimensional structure of the target enzyme greatly enhances the optimisation of lead compounds and the design of derivatives with increased activity. Among the most widely exploited herbicide targets is glutamine synthetase. Recently, the structure of a cytosolic form of the maize enzyme has been described, making it possible to verify whether steric, electronic and hydrophobic features of a compound are in agreement with inhibitor–protein interaction geometry. RESULTS: Three series of compounds (aminophosphonates, hydroxyphosphonates and aminomethylenebisphosphonates) were evaluated as possible inhibitors of maize glutamine synthetase. Aminomethylenebisphosphonate derivatives substituted in the phenyl ring retained the inhibitory potential, whereas variations in the scaffold, i.e. the replacement of the second phosphonate moiety with a hydroxyl or an amino residue, resulted in a significant loss of activity. A kinetic characterisation showed a non‐competitive mechanism against glutamate and an uncompetitive mechanism against ATP. A docking analysis suggested the mode of bisphosphonate binding to the active site. CONCLUSION: Results made it possible to define the features required to maintain or enhance the biological activity of these compounds, which represent lead structures to be further exploited for the design of new substances endowed with herbicidal activity. Copyright © 2009 Society of Chemical Industry     

The disappearance kinetics of fenhexamid in sterile and non-sterile conditions,as revealed by a mixed-effects model analysis

A kinetic model was used to evaluate rate and capacity of both sorption and degradation of fenhexamid in four soils under laboratory conditions. This molecule was chosen for its stability to chemical hydrolysis at environmental pH values and its very fast disappearance in soil. The biotic contribution to disappearance was determined in experiments conducted on four soils that were either unsterilised or sterilised by: chloroform fumigation (CHCl₃), autoclaving or phenylmercuric acetate (PMA) addition. Sterilisation with CHCl₃ was the least efficient method as proven by microbial counts. The clayey soils sterilised by autoclave showed a sudden down-step around 50–75 h in disappearance kinetics. Biotic contribution should be minimal because soils were sterilised as shown by plate counts and the disappearance might be due to new sorption surfaces, available after 50–75 h as a consequence of the disruption of some aggregates. Sterilisation of the soil with the highest organic matter content by PMA increased the instantaneous sorption, and in some cases, the fast sorption of fenhexamid. The treatment of soil A with PMA did not completely inhibit microbial activity. A non-linear regression analysis with mixed-effects models was a valuable tool in describing the disappearance kinetics in soils because it showed small differences among treatments. The analysis showed that three processes were important in the disappearance of fenhexamid: instantaneous sorption, fast sorption and slow sorption/degradation. The biotic contribution probably depended on soil type, being predominant only in one soil, whereas sorption (instantaneous and fast) was prevailing in the other soils.