Glyoxalase I Assay as a Possible Tool for Evaluation of Biological Activity of Antioxidant-Rich Plant Extracts.

The health-promoting properties of natural plant bioactive compounds are mainly attributable to their ability to counteract oxidative stress. This is considered a major causative factor in aging and aging-related human diseases, in which a causal role is also ascribed to dicarbonyl stress. This is due to accumulation of methylglyoxal (MG) and other reactive dicarbonyl species, leading to macromolecule glycation and cell/tissue dysfunction. The glyoxalase (GLYI) enzyme, catalyzing the rate-limiting step of the GSH-dependent MG detoxification pathway, plays a key role in cell defense against dicarbonyl stress. Therefore, the study of GLYI regulation is of relevant interest. In particular, GLYI inducers are important for pharmacological interventions to sustain healthy aging and to improve dicarbonyl-related diseases; GLYI inhibitors, allowing increased MG levels to act as proapoptotic agents in tumor cells, are of special interest in cancer treatment. In this study, we performed a new in vitro exploration of biological activity of plant bioactive compounds by associating the measurement of their antioxidant capacity (AC) with the evaluation of their potential impact on dicarbonyl stress measured as capability to modulate GLYI activity. AC was evaluated using TEAC, ORAC, and LOX-FL methods. The GLYI assay was performed using a human recombinant isoform, in comparison with the recently characterized GLYI activity of durum wheat mitochondria. Different plant extracts were tested, obtained from plant sources with very high phytochemical content ('Sun Black' and wildtype tomatoes, black and 'Polignano' carrots, and durum wheat grain). Results showed high antioxidant properties of the tested extracts, associated with different modes (no effect, activation, and inhibition) and effectiveness in modulating both GLYI activity sources. Overall, results indicate the GLYI assay as an advisable and promising tool for researching plant foods as a source of natural antioxidant compounds acting as GLYI enzymatic regulators to be used for dietary management associated the treatment of oxidative/dicarbonyl-promoted diseases.

Influence of Drought and Salt Stress on Durum Wheat Grain Quality and Composition: A Review.

Durum wheat is a staple crop for the Mediterranean diet because of its adaptability to environmental pressure and for its large use in cereal-based food products, such as pasta and bread, as a source of calories and proteins. Durum wheat whole grains are also highly valued for their peculiar amount of dietary fiber and minerals, as well as bioactive compounds of particular interest for their putative health-beneficial properties, including polyphenols, carotenoids, tocopherols, tocotrienols, and phytosterols. In Mediterranean environments, durum wheat is mostly grown under rainfed conditions, where the crop often experiences environmental stresses, especially water deficit and soil salinity that may induce a hyperosmotic stress. In particular, changes in C and N accumulation due to these abiotic conditions, during grain filling, can influence starch and storage protein amount and composition in durum wheat caryopsis, thus influencing yield and quality traits. Recent advancements regarding the influence of water deficit and salinity stress on durum wheat are critically discussed. In particular, a focus on stress-induced changes in (a) grain protein content and composition in relation to technological and health quality; (b) starch and dietary fiber accumulation and composition; (c) phytochemical composition; (d) health-related grain micronutrient accumulation, such as Fe and Zn.

Changes in Antioxidant Defence System in Durum Wheat under Hyperosmotic Stress: A Concise Overview.

Durum wheat is one of the most commonly cultivated species in the world and represents a key commodity for many areas worldwide, as its grain is used for production of many foods, such as pasta, bread, couscous, and bourghul. Durum wheat grain has a relevant role in the human diet, providing carbohydrates, proteins, lipids, fibres, vitamins, and minerals, as well as highly valued bioactive compounds contributing to a healthy diet. Durum wheat is largely cultivated in the Mediterranean basin, where it is mainly grown under rain-fed conditions, thus currently undergoing drought stress, as well as soil salinity, which can hamper yield potential and influence the qualitative characteristics of grain. When plants suffer drought and/or salinity stress, a condition known as hyperosmotic stress is established at cellular level. This leads to the accumulation of ROS thus generating in turn an oxidative stress condition, which can ultimately result in the impairment of cellular integrity and functionality. To counteract oxidative damage due to excessive ROS production under stress, plants have evolved a complex array of both enzymatic and non-enzymatic antioxidant mechanisms, working jointly and synergically for maintenance of ROS homeostasis. Enhancement of antioxidant defence system has been demonstrated as an adaptive mechanism associated to an increased tolerance to hyperosmotic stress. In the light of these considerations, this review provides a concise overview on recent advancements regarding the role of the ascorbate-glutathione cycle and the main antioxidant enzymes (superoxide dismutase, catalase, and peroxidases) in durum wheat response to drought and salt stresses that are expected to become more and more frequent due to the ongoing climate changes.

The uniqueness of the plant mitochondrial potassium channel.

The ATP-inhibited Plant Mitochondrial K(+) Channel (PmitoKATP) was discovered about fifteen years ago in Durum Wheat Mitochondria (DWM). PmitoKATP catalyses the electrophoretic K(+) uniport through the inner mitochondrial membrane; moreover, the co-operation between PmitoKATP and K(+)/H(+) antiporter allows such a great operation of a K(+) cycle to collapse mitochondrial membrane potential (ΔΨ) and ΔpH, thus impairing protonmotive force (Δp). A possible physiological role of such ΔΨ control is the restriction of harmful reactive oxygen species (ROS) production under environmental/oxidative stress conditions. Interestingly, DWM lacking Δp were found to be nevertheless fully coupled and able to regularly accomplish ATP synthesis; this unexpected behaviour makes necessary to recast in some way the classical chemiosmotic model. In the whole, PmitoKATP may oppose to large scale ROS production by lowering ΔΨ under environmental/oxidative stress, but, when stress is moderate, this occurs without impairing ATP synthesis in a crucial moment for cell and mitochondrial bioenergetics.

Plant inner membrane anion channel (PIMAC) function in plant mitochondria.

To date, the existence of the plant inner membrane anion channel (PIMAC) has been shown only in potato mitochondria, but its physiological role remains unclear. In this study, by means of swelling experiments in K(+) and ammonium salts, we characterize a PIMAC-like anion-conducting pathway in mitochondria from durum wheat (DWM), a monocotyledonous species phylogenetically far from potato. DWM were investigated since they possess a very active potassium channel (PmitoK(ATP)), so implying a very active matching anion uniport pathway and, possibly, a coordinated function. As in potato mitochondria, the electrophoretic uptake of chloride and succinate was inhibited by matrix [H(+)], propranolol, and tributyltin, and was insensitive to Mg(2+), N,N'-dicyclohexylcarbodiimide (DCCD) and mercurials, thus showing PIMAC's existence in DWM. PIMAC actively transports dicarboxylates, oxodicarboxylates, tricarboxylates and Pi. Interestingly, a novel mechanism of swelling in ammonium salts of isolated plant mitochondria is reported, based on electrophoretic anion uptake via PIMAC and ammonium uniport via PmitoK(ATP). PIMAC is inhibited by physiological compounds, such as ATP and free fatty acids, by high electrical membrane potential (Delta Psi), but not by acyl-CoAs or reactive oxygen species. PIMAC was found to cooperate with dicarboxylate carrier by allowing succinate uptake that triggers succinate/malate exchange in isolated DWM. Similar results were obtained using mitochondria from the dicotyledonous species topinambur, so suggesting generalization of results. We propose that PIMAC is normally inactive in vivo due to ATP and Delta Psi inhibition, but activation may occur in mitochondria de-energized by PmitoK(ATP) (or other dissipative systems) to replace or integrate the operation of classical anion carriers.

Reactive oxygen species inhibit the succinate oxidation-supported generation of membrane potential in wheat mitochondria.

In order to gain a first insight into the effects of reactive oxygen species (ROS) on plant mitochondria, we studied the effect of the ROS producing system consisting of xanthine plus xanthine oxidase on the rate of membrane potential (DeltaPsi) generation due to either succinate or NADH addition to durum wheat mitochondria as monitored by safranin fluorescence. We show that the early ROS production inhibits the succinate-dependent, but not the NADH-dependent, DeltaPsi generation and oxygen uptake. This inhibition appears to depend on the impairment of mitochondrial permeability to succinate. It does not involve mitochondrial thiol groups sensitive to either mersalyl or N-ethylmaleimide and might involve both protein residues and/or membrane lipids, as suggested by the mixed nature. We propose that, during oxidative stress, early generation of ROS can affect plant mitochondria by impairing metabolite transport, thus preventing further substrate oxidation, DeltaPsi generation and consequent large-scale ROS production.