The present study aims to screen some pharmacological activities of three promising ethacrynic acid (EA) derivatives (P3, P4, and P5) containing a piperazine moiety. The EA derivatives were tested against MDA-MB-468 cancer cell line as a model for the triple-negative breast cancer and MCF7 cancer cell line as a model subtype of luminal breast cancer using the methyl tetrazolium test (MTT). Additionally, the modulation of the glutathione and thioredoxin enzymatic systems, as well as the antioxidant and cytoprotective potential of each compound, were investigated. Moreover, initial in vitro safety screening was conducted using human blood cells. As a result, EA derivatives showed clear dose-dependent antiproliferative activity in the micromolar range (between 1.13 and 2.51 mu M), with high selectivity to cancer cells by orchestrating oxidative stress. Derivative P3 has the most promising potential for further preclinical investigation, owing to its safety profile and significant cytoprotective and antitumor properties.

Class II water-soluble chlorophyll proteins (WSCPs) from Brassicaceae are non-photosynthetic proteins that bind with chlorophyll (Chl) and its derivatives. The physiological function of WSCPs is still unclear, but it is assumed to be involved in stress responses, which is likely related to their Chl-binding and protease inhibition (PI) activities. Yet, the dual function and simultaneous functionality of WSCPs must still be better understood. Here, the biochemical functions of Brassica napus drought-induced 22-kDa protein (BnD22), a major WSCP expressed in B. napus leaves, were investigated using recombinant hexahistidine-tagged protein. We showed that BnD22 inhibited cysteine proteases, such as papain, but not serine proteases. BnD22 was able to bind with Chla or Chlb to form tetrameric complexes. Unexpectedly, BnD22-Chl tetramer displays higher inhibition toward cysteine proteases, indicating (i) simultaneous Chl-binding and PI activities and (ii) Chl-dependent activation of PI activity of BnD22. Moreover, the photostability of BnD22-Chl tetramer was reduced upon binding with the protease. Using three-dimensional structural modeling and molecular docking, we revealed that Chl binding favors interaction between BnD22 and proteases. Despite its Chl-binding ability, the BnD22 was not detected in chloroplasts but rather in the endoplasmic reticulum and vacuole. In addition, the C-terminal extension peptide of BnD22, which cleaved off post-translationally in vivo, was not implicated in subcellular localization. Instead, it drastically promoted the expression, solubility and stability of the recombinant protein.

Women's breast cancer is one of the most significant healthcare issues for the human race that demands a proactive strategy for a cure. In this study, the cytotoxic activity (MTT assay) of two natural steroidal compounds, protodioscin and dioscin, against two major subtypes of human breast cancer estrogen receptor-positive (ER-positive)/MCF-7 and triple-negative breast cancer (TNBC)/MDA-MB-468), was assessed. The clonogenic capacity was evaluated using the clonogenic assay. Oxidative stress was determined by measuring the formation of malondialdehyde and H2O2 and the assessment of total antioxidant enzyme activities (SOD, GPx, GR, and TrxR). Protodioscin and dioscin were highly cytotoxic against the tested cell lines (1.53 μM <IC50< 6 μM) with low cytotoxicity on normal cells (PBMC; IC50 ≥ 50 μM). Interestingly, these compounds were responsible for a substantial decrease in the clonogenic capacity of both cell lines. Moreover, dioscin was able to reduce the cell motility of the invasive breast cancer cells (MDA-MB-468). At the molecular level, the two treatments resulted in an increase of reactive oxygen species. Notably, both compounds were responsible for decreasing the enzymatic activities of glutathione reductase and thioredoxin reductase. On the basis of such considerations, protodioscin and dioscin may serve as promising natural compounds to treat TNBC and ER-positive breast cancer through the induction of oxidative stress.

Sorghum, the fifth most important cereal crop, is a well-adapted cereal to arid/semi-arid regions. Sorghum is known for multiple end-uses as food, feed, fuel, forage, and as source of bioactive compounds that could be used for medical applications. Although the great improvement in the process of sorghum breeding, the average yield of this crop is still very low. Therefore, exploring the genetic diversity in sorghum accessions is a critical step for improving this crop. The main objective of the current work was to study the genetic variation existing in a Moroccan sorghum collection. Indeed, 10 sorghum ecotypes were characterized based on agromorphological descriptors. Both quantitative (25) and qualitative (7) traits revealed variability (p < 0.05) among the studied ecotypes. At the seedling stage, most of the ecotypes showed good to high vigor (70%). However, as the sorghum plants grow, the difference between genotypes become more apparent, especially at the generative phase. For instance, three different panicle shapes have been observed, erect (50%), semi-bent (30%), and bent (20%) with different degree of compactness (20% for loose, semi-compact, and compact panicles, and 30% for semi-loose panicles). In another part of this study, the phytochemical composition and antioxidant activities of the sorghum ecotypes have been determined. The results showed variable total phenolic contents, and total flavonoid contents ranging from 125.86 ± 1.36 to 314.91 ± 3.60 mg GAE/g dw and 114.0 ± 13.2 to 138.5 ± 10.8 (mg catechin equivalent/100 g, dw) respectively, with a differential antioxidant activities as well. These results indicate that for any crop breeding program, it is preferable to take into consideration both morphological and biochemical traits for a better selection of high yielding varieties with high added value compounds. Therefore, the implication of these results in the context of sorghum breeding activities could be a resourceful option for farmers.

The water-soluble chlorophyll-proteins (WSCP) of class II from Brassicaceae are non-photosynthetic proteins that bind chlorophylls (Chls) and chlorophyll derivatives. Their physiological roles, biochemical functions and mode of action are still unclear. It is assumed that the WSCPs have a protection function against Chl photodamage during stressful conditions. WSCPs are subdivided into class IIA and class IIB according to their apparent Chla/b binding ratio. Although their Chla/Chlb binding selectivity has been partly characterized, their Chl affinities are not yet precisely defined. For instance, WSCPs IIA do not show any Chl binding preference while WSCPs IIB have greater affinity to Chlb. In this study, we present a novel method for assessment of Chl binding to WSCPs based on the differences of Chl photobleaching rates in a large range of Chl/protein ratios. The protein we have chosen to study WSCP is BnD22, a WSCP IIA induced in the leaves of Brassica napus under water deficit. BnD22 formed oligomeric complexes upon binding to Chla and/or Chlb allowing a protective effect against photodamage. The binding constants indicate that BnD22 binds with high affinity the Chls and with a strong selectivity to Chla. Moreover, dependending of Chl/protein ratio upon reconstitution, two distinct binding events were detected resulting from difference of Chl stoichiometry inside oligomeric complexes.

Improved crop yield is proportional to nitrogen (N) fertilizer applied to agricultural land. However, N fertilizers are also major source of environmental pollution mainly in soils, water and the atmosphere. Determining the optimal amount ofNfertilizer needed for crop growth is therefore essential. Arbuscular mycorrhizal fungi (AMF) characterized by their capacity to provide mineral nutrients are also exploited by the soil beyond the zone ofinfluence of the plant root and by an important role in reducing the amount of N fertilizer. Thus, three Moroccan sorghum ecotypes (5p3, 3p9 and 4p11), cereal with agronomic and economic interest, were cultivated with or without AMF. Growth parameters were measured and key enzymes responsible for carbon/nitrogen interaction such as glutamine synthetase (GS), glutamate dehydrogenase (GDH), NADPtIsocitrate dehydrogenase (ICDH), and NADtmalate dehydrogenase (MDH) have been determined.In the three ecotypes, mycorrhizal plants showed a longer plant length compared to control plants. The biochemical parameters showed a significant increase in GS and ICDH activity in the leaves and roots of mycorrhizal plants. However, mycorrhizal fungi appear to affect the activity of GDH and MDH only in the root of sorghum plants. AMF can be an effective way to optimize nitrogen uptake by the plant and thus improve crop yields with lower amounts of nitrogen fertilizers.

The first part of this work consists in the identification of a prokaryotic-type bifunctional enzyme (aspartate aminotransferase, PT-AAT) in photosynthetic cells of Sorghum–sudangrass hybrid seeds Sorghum × drummondii (Nees ex Steud.) Millsp. and Chase. The results obtained show that the enzyme is constituted by a subunit of 45 kDa immunologically similar to the enzymes of C3 plants and gymnosperms. This result suggests a high degree of conservation of the PT-AAT in higher plants. To investigate whether the expression of AAT is associated with the differentiation of leaves, we analyzed the chlorophyll levels and AAT, glutamine synthetase (GS), glutamate synthase (GOGAT), isocitrate dehydrogenase (IDH), Rubisco and nitrate reductase (NR) contents in serial sections of the leaves. The chlorophyll content was high in the top sections; and then decreased toward basal sections in a continuous gradient. The Rubisco and NR contents matched the pattern of chlorophyll in the leaves with high levels in the aged section and very low levels in the basal sections. In contrast to chlorophyll, Rubisco and NR contents, GS2 and Fd-GOGAT polypeptides markedly increased from the top to the basal sections of leaves. However the patterns of AAT and IDH polypeptides did not change in different sections. In the second part of this work, to determine the relative localization of AAT, mesophyll cells (MCs) and bundle-sheath cells (BSCs) of sorghum leaves were separated by cellulase and macerase digestion. PT-AAT protein was most abundant in MC fraction than BSC fraction.

In order to better understand the biochemical mechanisms of proline metabolism under different nitrogen (N) level and source treatments in sorghum plants, different biochemical parameters on shoots and roots of two Moroccan sorghum Ecotype (3P4 and 4P11) such as photosynthetic pigments content, protein content, amino acid content, proline content and two proline metabolizing enzymes were investigated. Proline content reached its highest value under N deficiency and ammonium excess. Furthermore, the proline degradation enzyme (ProDH) activity was lower under N deficiency and ammonium excess. The proline dehydrogenase activity was inversely proportional to the proline content and to the proline biosynthesis enzyme (P5CS) activity which explains the proline accumulation under N stress. We showed also that 4P11 has higher photosynthetic pigment, amino acid and protein contents with less proline accumulation which may indicate a higher tolerance to N stress for this ecotype compared to the 3P4. Therefore, the biochemical analysis reported here may be used for the selection of genotypes with higher tolerance to N stress.

In order to evaluate the antioxidant properties of aqueous and methanol extracts of needles and berries of Juniperus oxycedrus subsp. oxycedrus (Joo) species, various antioxidant capacity assessment tests (free radical scavenging assays (DPPH• and ABTS•+ tests), ferrous ions (Fe2+) chelating activity and reducing power assay (FRAP) were conducted. In all of the tests, the extracts exhibited strong antioxidant activity. Furthermore, in-vitro cytotoxic activity assays of the methanolic extracts showed potent cytotoxic effects against two breast cancer cell lines (MDA-MB-468 and MCF-7), with no cytotoxicity towards normal cells (PBMCs). Reactive oxygen species generation was presumed to be a potential reason for the observed cytotoxic effects. According to all the above, and considering its appropriate composition of mineral elements and phenolic compounds, Joo could offer a beneficial and natural source of bioactive compounds that can be either used on the preventive side as it could potentially be used in the clinic without toxicity.

This paper reviewed the beneficial effects of the major phenolic acid compounds of Sorghum bicolor seeds. Different studies were reviewed to determine the major phenolic acid components of sorghum seeds. Several kinds of literature were then analyzed to discuss the different beneficial effects of these molecules. S. bicolor is an important source for food and feed. It is among the top five crops regarding its production and consumption throughout the world. Till date, many studies highlighted different aspects of the biochemical and physiological properties of sorghum grain. However, studies concerning the pharmacological properties of sorghum grain are scarce. The predominant phenolic acids of sorghum seeds are ferulic, p-coumaric, and protocatechuic acids. The bioactive effects of these phenolic acids are mainly related to their antioxidant, antitumor, antidiabetic, antimicrobial, cardiovascular, and gastrointestinal activities. The data collected from recent studies indicate that these molecules have a promising future as natural agents for the treatment of various diseases, and this is particularly due to their strong antioxidant properties. This review provides evidence for the importance of sorghum seeds and their phenolic compounds in the prevention and treatment of several diseases. This work showed that sorghum grains are a good source of beneficial and therapeutic molecules. It also recommended the addition of sorghum grains to human diet as other cereals because of its high nutritional value.

BACKGROUND:In cancer cells, the intracellular antioxidant capacity and the redox homeostasis are mainly maintained by the glutathione- and thioredoxin-dependent systems which are considered as promising targets for anticancer drugs. Pyridazinones constitute an interesting source of heterocyclic compounds for drug discovery. The present investigation focused on studying the in-vitro antitumor activity of newly synthesized Pyridazin-3(2h)-ones derivatives against P815 (Murin mastocytoma) cell line.METHODS:The in-vitro cytotoxic activities were investigated toward the P815 cell line using tetrazolium-based MTT assay. Lipid peroxidation and the specific activities of antioxidant enzymes were also determined.RESULTS:The newly compounds had a selective dose-dependent cytotoxic effect without affecting normal cells (PBMCs). Apoptosis was further confirmed through the characteristic apoptotic morphological changes and DNA fragmentation. Two compounds (6F: and 7H: ) were highly cytotoxic and were submitted to extend biological testing to determine the likely mechanisms of their cytotoxicity. Results showed that these molecules may induce cytotoxicity via disturbing the redox homeostasis. Importantly, the anticancer activity of 6F: and 7H: could be due to the intracellular reactive oxygen species hypergeneration through significant loss of glutathione reductase and thioredoxin reductase activities. This eventually leads to oxidative stress-mediated P815 cell apoptosis. Furthermore, the co-administration of 6F: or 7H: with Methotrexate exhibited a synergistic cytotoxic effect.CONCLUSIONS:considering their significant anticancer activity and chemosensitivity, 6F: and 7H: may improve the therapeutic efficacy of the current treatment for cancer.

Nitrogen stress as well as other stresses can negatively impact the plant development and metabolism. Generally, stress factors increase the reactive oxygen species (ROS) and methylglyoxal (MG) production, which may, in the absence of effective protective mechanisms, induce irreparable metabolic dysfunction and death. The effect of different amounts (from deficiency to excess) of nitrate, ammonium or nitrate combined to ammonium, on enzyme activities of antioxidant and methylglyoxal detoxification systems of two sorghum ecotypes (3P4 and 4P11) was studied. The N supply was performed per pot during the sowing step using potassium nitrate and/or ammonium sulfate. Six N treatments were applied using 120, 240 and 480 Kg ha-1 of ammonium or nitrate and three other treatments were applied using 120 kg ha-1 nitrate combined to 120, 240 and 480 kg ha-1of ammonium. The specific activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), glutathione reductase (GR), glyoxalase I (Gly I) and glyoxalase II (Gly II) were investigated. Results showed that, ammonium excess and N-deficient conditions increased the contents of malondialdehyde (MDA), and induced the enzyme activities of ROS and MG detoxification systems, supporting the sorghum’s ability to counteract the negative effect of N stress (deficit and excess). We have also shown that the SOD, CAT, GR and Gly I enzyme activities were higher in the 4P11 ecotype compared to the 3P4 ecotype. These results indicate that sorghum ecotypes exhibit differential tolerance to N stress and suggest that the 4P11 ecotype has higher capacity to cope with N stress.

Sorghum (Sorghum bicolor (L.) Moench) is a species of great socio-economic and ecological importance for countries in arid and semi-arid climate. In C4 plants like sorghum, phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) plays a key role in seed development and germination. In this work, the PEPC activity shows an increase followed by a decrease at the early and later stages of maturation, respectively. In germinating seeds, the PEPC activity quickly increases after soaking. The L-malate test and the ratio of PEPC activity determined at pH 8.0 and 7.1, indicates, that PEPC is phosphorylated at the early stages of maturation then becomes dephosphorylated at the later stages and during seed germination, PEPC takes back its phosphorylated form. The determination of the affinity constant showed different KM depending on the seed developmental stage. As there is no PEPC-C4 isoform in developing sorghum seeds, this result indicates that the different KM observed during seed maturation could be a result of a post-translational regulation such as phosphorylation or ubiquitination of a pre-existing isoform. This regulation enhances the PEPC activity at early stages of seed development.

Carbon and nitrogen metabolism pathways are regulated by complex mechanisms in order to optimize growth and development of plants and play a major role in determining the amount of protein stored in cereal grains. This study was conducted to contribute to determine the roles of certain key enzymes of carbon and nitrogen metabolism in developing sorghum seeds. Sorghum bicolor (L.) Moench was grown in a growth chamber. Seeds were harvested at various stages of development and used to the analysis of certain key enzymes of carbon and nitrogen metabolism. The results were subjected to stastistical analysis using one-way analysis of variance (ANOVA) followed by the Tukey test. The results showed that during the seed development the highest activities of glutamine synthetase, glutamate dehydrogenase, phosphoenolpyruvate carboxylase, malate dehydrogenase, isocitrate dehydrogenase, and aspartate aminotransferase were detected at the time of maximum seed filling stages (15-20 DAP). Hence, higher activities of these enzymes at the time of maximum seed-filling stages could be effectively linked to amino acids synthesis and therefore the deposition of protein reserves.

Carbon and nitrogen metabolism are regulated by complex mechanisms in order to optimize growth and development of plants and play a major role during germination and post-germinative growth. The objective of the current work was to establish the degree of changes in activities of key enzymes implicated in these two metabolism pathways during germination and post-germinative growth such as; glutamine synthetase, glutamate dehydrogenase, phosphoenolpyruvate carboxylase, malate dehydrogenase, isocitrate dehydrogenase and aspartate aminotransferase. We have observed that during germination and seedlings development the most activities of nitrogen and carbon metabolism enzymes were correlated (nearly show similar kinetic changes) and influenced by their cellular environment and developmental stage. Therefore, higher activities of these enzymes were observed in the seed, root and shoot at early stages of the post-germinative phase could be effectively linked to the demand of de novo proteins to ensure a good development of seedlings.