Impact of temperature stress on secondary metabolite profile and phytotoxicity of Amaranthus cruentus L. leaf extracts

In this study Amaranthus cruentus plants were grown under controlled optimal conditions (28/21 °C) for three months and then subjected to cold (14/7 °C) and hot (33/40 °C) temperatures. We investigated the influence of these temperature regimes on the metabolite profile of the leaves through analyses of data by TLC, HPLC and GC-MS spectrometry. The phytotoxic potential of a methanol-water (MW) and dichloromethane (DCM) extract from the aerial parts were examined through in vitro screening of germination and growth of lettuce and pepper. The optimal extracts displayed the highest diversity of secondary metabolites, and the highest total phenolics and flavonoids content. Through TLC and HPLC analysis the significantly lower phenolic content in the hot temperature treated samples was confirmed. A wide range of metabolites were detected in the DCM extracts through GC-MS analyses. The phytotoxicity of both the MW and DCM extracts were demonstrated, as germination and growth of pepper and lettuce were significantly inhibited, indicating the presence of more than one allelochemical compound which may affect the allelopathic activity of A. cruentus during changes in environmental temperatures

Influence of altered temperatures on allelopatic properties of Amaranthus cruentus L.

The relationships between allelochemicals and environmental factors are a key factor for the growth of plants under rotation. We investigated the allelopathic potential of Amaranthus cruentus L, grown under different temperature conditions in in vitro bioassays. An inhibitory effect on germination and growth of lettuce (Lactuca sativa L.), tomato (Solanum lycopersicum L.), pepper (Capsicum annuum L.) and cucumber (Cucumis sativus L.) was observed when seeds were subjected to the leaf litter of Amaranthus cruentus. Analysis from our study indicated that germination percentage was significantly affected by growth temperatures (T) of the amaranth (P ˂ 0.0001), litter concentration (C) (P ˂ 0.0001), vegetable type (V) (P ˂ 0.0001), the T × V interaction (P = 0.0041) and V × C interaction (P ˂ 0.0001). Pepper was the most sensitive with a decline in germination percentage at increasing concentrations (0, 1 and 5 mg ml-1) of litter. Hypocotyl and seminal root lengths were adversely influenced by the plant litter for all the temperature treatments, although effects were most severe when exposed to the leaf litter of the hot temperature treatment. The inhibition caused by the litter was dependent on growth temperature and concentration, while each vegetable species showed different levels of sensitivity.</p

Conformationally restricted calpain inhibitors

The cysteine protease calpain-I is linked to several diseases and is therefore a valuable target for inhibition. Selective inhibition of calpain-I has proved difficult as most compounds target the active site and inhibit a broad spectrum of cysteine proteases as well as other calpain isoforms. Selective inhibitors might not only be potential drugs but should act as tools to explore the physiological and pathophysiological roles of calpain-I. α-Mercaptoacrylic acid based calpain inhibitors are potent, cell permeable and selective inhibitors of calpain-I and calpain-II. These inhibitors target the calcium binding domain PEF(S) of calpain-I and -II. Here X-ray diffraction analysis of co-crystals of PEF(S) revealed that the disulfide form of an α-mercaptoacrylic acid bound within a hydrophobic groove that is also targeted by a calpastatin inhibitory region and made a greater number of favourable interactions with the protein than the reduced sulfhydryl form. Measurement of the inhibitory potency of the α-mercaptoacrylic acids and X-ray crystallography revealed that the IC50 values decreased significantly on oxidation as a consequence of the stereo-electronic properties of disulfide bonds that restrict rotation around the S–S bond. Consequently, thioether analogues inhibited calpain-I with potencies similar to those of the free sulfhydryl forms of α-mercaptoacrylic acids

Chemoenzymatic synthesis of the alarm pheromone (+)-verbenone from geranyl diphosphate

The enzyme-guided asymmetric synthesis of (+)-verbenone from geranyl diphosphate in a simple two-step, one pot transformation highlights the potential of chemoenzymatic procedures for the generation of high-value terpenoids

Bisphosphonate inhibitors of squalene synthase protect cells against cholesterol‐dependent cytolysins

Certain species of pathogenic bacteria damage tissues by secreting cholesterol‐dependent cytolysins, which form pores in the plasma membranes of animal cells. However, reducing cholesterol protects cells against these cytolysins. As the first committed step of cholesterol biosynthesis is catalyzed by squalene synthase, we explored whether inhibiting this enzyme protected cells against cholesterol‐dependent cytolysins. We first synthesized 22 different nitrogen‐containing bisphosphonate molecules that were designed to inhibit squalene synthase. Squalene synthase inhibition was quantified using a cell‐free enzyme assay, and validated by computer modeling of bisphosphonate molecules binding to squalene synthase. The bisphosphonates were then screened for their ability to protect HeLa cells against the damage caused by the cholesterol‐dependent cytolysin, pyolysin. The most effective bisphosphonate reduced pyolysin‐induced leakage of lactate dehydrogenase into cell supernatants by >80%, and reduced pyolysin‐induced cytolysis from >75% to <25%. In addition, this bisphosphonate reduced pyolysin‐induced leakage of potassium from cells, limited changes in the cytoskeleton, prevented mitogen‐activated protein kinases cell stress responses, and reduced cellular cholesterol. The bisphosphonate also protected cells against another cholesterol‐dependent cytolysin, streptolysin O, and protected lung epithelial cells and primary dermal fibroblasts against cytolysis. Our findings imply that treatment with bisphosphonates that inhibit squalene synthase might help protect tissues against pathogenic bacteria that secrete cholesterol‐dependent cytolysins

The structural basis of differential inhibition of human calpain by indole and phenyl α--mercaptoacrylic acids

Excessive activity of neutrophils has been linked to many pathological conditions, including rheumatoid arthritis, cancer and Alzheimer’s disease. Calpain-I is a Ca2+-dependent protease that plays a key role in the extravasation of neutrophils from the blood stream prior to causing damage within affected tissues. Inhibition of calpain-I with small molecule mercaptoacrylic acid derivatives slows the cell spreading process of live neutrophils and so these compounds represent promising drug leads. Here we present the 2.05 and 2.03 Å co-crystal X-ray structures of the pentaEF hand region, PEF(S), from human calpain with (Z)-3-(4-chlorophenyl)-2-mercaptoacrylic acid and (Z)-3-(5-bromoindol-3-yl)-2-mercaptoacrylic acid. In both structures, the α-mercaptoacrylic acid derivatives bind between two α-helices in a hydrophobic pocket that is also exploited by a leucine residue of the endogenous regulatory calpain inhibitor calpastatin. Hydrophobic interactions between the aromatic rings of both inhibitors and the aliphatic residues of the pocket are integral for tight binding. In the case of (Z)-3-(5-bromoindol-3-yl)-2-mercaptoacrylic acid, hydrogen bonds form between the mercaptoacrylic acid substituent lying outside the pocket and the protein and the carboxylate group is coplanar with the aromatic ring system. Multiple conformations of (Z)-3-(5-bromoindol-3-yl)-2-mercaptoacrylic acid were found within the pocket. The increased potency of (Z)-3-(5-bromoindol-3-yl)-2-mercaptoacrylic acid relative to (Z)-3-(4-chlorophenyl)-2-mercaptoacrylic acid may be a consequence of the indole group binding more deeply in the hydrophobic pocket of PEF(S) than the phenyl ring

Evaluation of Plectranthus esculentus N.E.Br. as a potential vegetable crop

Please read the abstract in the section 00front of this documentThesis (PhD (Agronomy))--University of Pretoria, 2007.Plant Production and Soil Scienceunrestricte