Anticancer Properties of Different Varieties of Date Palm (Phoenix dactylifera L.) Leaf Extracts in Human Tumor Cells: a Comparative Study.

Plant polyphenols are nutraceutical components with relevant biological effects on human health. They act against development of several diseases including cancer. In this study, the methanolic extracts of four date palm Phoenix dactylifera leaves (Deglet Noor (DN), Barhee (B), Khalas (KS) and Khunezi (KZ)) collected from south Tunisia were preliminary analyzed for their effects against U87 (human glioblastoma) and MDA-MB-231 (human breast cancer) cell line development. Results showed that Barhee extract (30 µg/mL) was the most efficient to reduce the growth of both tumor cells to about 40% (p < 0.05) without inducing cytotoxicity. Significantly, KS, KZ, DN and B extracts (30 µg/mL) decreased MDA-MB-231 and U87 cell adhesion towards fibrinogen and fibronectin. Using integrin blocking antibodies, leaf extracts competitively decreased human glioblastoma cell attachment to immobilized antibodies by interfering to αvß3 and α5ß1 integrin receptors. At the same concentration, extracts decreased MDA-MB-23 and U87 cell migration performed with wound healing assay. Particularly, Barhee and Deglet Noor leaf extracts (30 µg/mL) significantly reduced U87 cell invasion by 52.92% (p < 0.01) and 74.56% (p < 0.01), respectively. Collegially, our findings revealed beneficial proprieties of four varieties of date palm leaf especially those displayed by DN and B extracts that may serve as active candidates against human glioblastoma and breast cancer progression.

Insights into the mechanisms governing P01 scorpion toxin effect against U87 glioblastoma cells oncogenesis.

The emerging concept of small conductance Ca2+-activated potassium channels (SKCa) as pharmacological target for cancer treatment has significantly increased in recent years. In this study, we isolated the P01 toxin from Androctonus australis (Aa) scorpion venom and investigated its effect on biological properties of glioblastoma U87, breast MDA-MB231 and colon adenocarcinoma LS174 cancer cell lines. Our results showed that P01 was active only on U87 glioblastoma cells. It inhibited their proliferation, adhesion and migration with IC50 values in the micromolar range. We have also shown that P01 reduced the amplitude of the currents recorded in HEK293 cells expressing SK2 channels with an IC50 value of 3 pM, while it had no effect on those expressing SK3 channels. The investigation of the SKCa channels expression pattern showed that SK2 transcripts were expressed differently in the three cancer cell lines. Particularly, we highlighted the presence of SK2 isoforms in U87 cells, which could explain and rely on the specific activity of P01 on this cell line. These experimental data highlighted the usefulness of scorpion peptides to decipher the role of SKCa channels in the tumorigenesis process, and develop potential therapeutic molecules targeting glioblastoma with high selectivity.

The Impact of Exercise Training Intensity on Physiological Adaptations and Insulin Resistance in Women with Abdominal Obesity.

Abdominal obesity has emerged globally as a major public health issue due to its high prevalence and morbidity. The benefits of physical exercise among the obese population are well documented. However, the optimal exercise intensity for reducing body fat and preventing insulin resistance and metabolic disorders is still under debate. This study aimed to examine the effects of three different intensities of combined endurance and strength training programs on anthropometric variables, physiological and muscular adaptations, and insulin sensitivity. Forty-three obese young women (age 26.4 ± 4.7 years, BMI 33.1 ± 2.5 kg/m2) were randomly assigned to one of four groups: a control group (G0), a moderate-intensity training group (G50, exercising brisk walking at 50% heart rate reserve HRR), a high-intensity training group (G75, exercise jogging at 75% HRR), and an alternated-intensity training group (G50/75, exercise brisk-walking/jogging at 50−75% HRR) with additional strength training once a week for each group. Body composition, waist circumference (WC), fasting blood glucose, insulin sensitivity and resistance (Homa-IR), resting heart rate (RHR), 6-min walk distance (6MWD), 1-repetition maximum (1-RM), and time to exhaustion (TTE) at 45% and 75% maximal voluntary contraction (MVC) for both the flexor and extensor muscle groups of the knees, were recorded before and after three months of exercise training. All training groups showed significant decreases in body mass, BMI, total body fat, body fat percentage, WC, abdominal and visceral mass (p < 0.001), with a greater reduction of body mass and BMI in G75 (p < 0.05). Lean mass increased significantly only in G50/75 (p < 0.05). The insulin sensitivity and Homa-IR decreased in the three training groups (p < 0.01), with greater enhanced resistance in G50 compared to G75 and G50/75 (p < 0.05). In contrast, there were no pre-post changes in all groups for fasting blood glucose (p > 0.05). 1-RM and TTE of the knee flexor and extensor muscles were improved in the three groups (p < 0.01), with greater improvement in G50/75 for 1RM and G75 in most of the TTE parameters (p < 0.05). RHR decreased and 6MWD increased significantly in the three training groups (p < 0.01), with greater 6MWD improvement in G75 (p < 0.05). In conclusion, the three training intensities seem to generate benefits in terms of body composition, physiological and muscular adaptations, and insulin resistance. High training intensity resulted in greater improvements in body mass, BMI, and endurance and strength, whereas moderate training intensity resulted in greater improvements of insulin resistance and homo-IR. Following alternate-intensity training, greater improvements were observed in lean mass and maximal strength performance.

In-site and Ex-site Date Palm Exposure to Heavy Metals Involved Infra-Individual Biomarkers Upregulation.

As a tree of considerable importance in arid regions-date palm, Phoenix dactylifera L. survival in contaminated areas of Sfax city has drawn our attention. Leaf samples of the plants grown in the study area showed high levels of cadmium (Cd), copper (Cu), and chromium (Cr). On the basis of this finding, the cellular mechanisms that explain these metal accumulations were investigated in controlled conditions. After four months of exposure to Cd, Cr, or Cu, high bioconcentration and translocation factor (TF>1) have been shown for date palm plantlets exposed to Cd and low TF values were obtained for plantlets treated with Cr and Cu. Moreover, accumulation of oxidants and antioxidant enzyme activities occurred in exposed roots to Cu and Cd. Secondary metabolites, such as polyphenols and flavonoids, were enhanced in plants exposed at low metal concentrations and declined thereafter. Accumulation of flavonoids in cells may be correlated with the expression of the gene encoding Pdmate5, responsible for the transport of secondary metabolites, especially flavonoids. Other transporter genes responded positively to metal incorporation, especially Pdhma2, but also Pdabcc and Pdnramp6. The latter would be a new candidate gene sensitive to metallic stress in plants. Expressions of gene coding metal chelators were also investigated. Pdpcs1 and Pdmt3 exhibited a strong induction in plants exposed to Cr. These modifications of the expression of some biochemical and molecular based-markers in date palm helped to better understand the ability of the plant to tolerate metals. They could be useful in assessing heavy metal contaminations in polluted soils and may improve accumulation capacity of other plants.

The extremely halophilic archaeon Halobacterium salinarum ETD5 from the solar saltern of Sfax (Tunisia) produces multiple halocins.

The extremely halophilic archaeon Halobacterium salinarum strain ETD5 was previously isolated from the solar saltern of Sfax (Tunisia) and shown to encode and express halocin S8. The Hbt. salinarum ETD5 culture supernatant was shown here to exhibit high antimicrobial activity against several halophilic archaea and bacteria of different genera, showing a cross-domain inhibition. The antimicrobial activity was destroyed by proteases, thus pointing to halocins. A bioguided purification procedure was applied using two chromatography steps and antimicrobial assays directed against Halorubrum chaoviator ETR14. In-gel screening assay showed the presence of two antimicrobial bands of approximately 8 and 14 kDa, for which characterization was investigated by N-terminal sequencing and mass spectrometry. The full-length form of halocin S8 that contains 81 amino acids and differs from the 36 amino acid short-length halocin S8 previously described from an uncharacterized haloarchaeon S8a, was identified in the 8 kDa halocin band. A novel halocin that we termed halocin S14 was found in the 14 kDa band. It exhibits amino acid sequence identities with the N-terminally truncated region of the archaeal Mn-superoxide dismutase. These results show that Hbt. salinarum ETD5 produces multiple halocins, a feature that had not been described until now in the domain Archaea.

Antioxidant, Anti-Inflammatory, and Antitumoral Effects of Aqueous Ethanolic Extract from Phoenix dactylifera L. Parthenocarpic Dates.

The aim of this study was to evaluate the antioxidant, the anti-inflammatory, and the antitumoral activities of the aqueous ethanolic extract from Phoenix dactylifera L. parthenocarpic dates. The antioxidant activity was carried using DPPH radical scavenging activity. The result showed that parthenocarpic dates had strongly scavenging activity on DPPH reaching 94% with an IC50 value of 0.15 ± 0.011 mg/mL (p < 0.05). The anti-inflammatory potential was determined by the inhibitory effect of the aqueous ethanolic extract on phospholipase A2 activity as well as on carrageenan-induced paw oedema in mice. The in vitro study showed that the extract inhibited the phospholipase A2 activity with an IC50 value of 130 µg/mL and the in vivo study showed a significantly decrease in the paw oedema after 1 h compared to the control group. Finally, the antiproliferative activity of the aqueous ethanolic extract was assessed by MTT test against MCF-7 and MDA-MB-231 cancer cell lines. This extract was effective in inhibiting MDA-MB-231 and MCF-7 cancer cells growth with IC50 values of 8 and 18 mg/mL, respectively, after 72 h treatment. These results confirm the ethnopharmacological significance of Phoenix dactylifera L. parthenocarpic dates, which could add support for its pharmaceutical use.

Insight into the expression variation of metal-responsive genes in the seedling of date palm (Phoenix dactylifera).

Phytochelatin synthase and metallothionein gene expressions were monitored via qPCR in order to investigate the molecular mechanisms involved in Cd and Cr detoxification in date palm (Phoenix dactylifera). A specific reference gene validation procedure using BestKeeper, NormFinder and geNorm programs allowed selection of the three most stable reference genes in a context of Cd or Cr contamination among six reference gene candidates, namely elongation factor α1, actin, aldehyde dehydrogenase, SAND family, tubulin 6 and TaTa box binding protein. Phytochelatin synthase (pcs) and metallothionein (mt) encoding gene expression were induced from the first days of exposure. At low Cd stress (0.02 mM), genes were still up-regulated until 60th day of exposure. At the highest metal concentrations, however, pcs and mt gene expressions decreased. pcs encoding gene was significantly up-regulated under Cr exposure, and was more responsive to increasing Cr concentration than mt encoding gene. Moreover, exposure to Cd or Cr influenced clearly seed germination and hypocotyls elongation. Thus, the results have proved that both analyzed genes participate in metal detoxification and their expression is regulated at transcriptional level in date palm subjected to Cr and Cd stress. Consequently, variations of expression of mt and pcs genes may serve as early-warning biomarkers of metal stress in this species.

Copper toxicity and date palm (Phoenix dactylifera) seedling tolerance: Monitoring of related biomarkers.

Date palm (Phoenix dactylifera) seeds were exposed to different copper (Cu) solutions to examine plant stress responses. Low Cu concentrations (0.02 and 0.2 mM) caused an increase of seed germination, whereas higher Cu amounts (2 mM) significantly inhibited seed germination, delayed hypocotyl elongation, increased seedling mortality, and reduced the germination index by more than 90%. Metal-related toxicity symptoms appeared after 15 d of 2 mM of Cu exposure. Biochemical activities such as amylase activity and redox balance elements were examined to study the relationship between external Cu amount and internal plant response. The present study showed that amylolytic activity was dose- and time-dependent. Likewise, H2 O2 production increased after exposure to Cu, which was correlated with thiobarbituric acid reactive substance (TBARS) accumulation. Furthermore at low Cu concentrations, superoxide dismutase (SOD) and catalase (CAT) activities increased, suggesting that date palm seed stimulated its metal homeostasis networks. However, the highest cupric ion amounts increased cell oxidant accumulation and reduced enzyme production. Gene expression level measures of P. dactylifera phytochelatin synthase (Pdpcs) and P. dactylifera metallothionein (Pdmt) encoding genes have been carried out to investigate the implication of PdPCS and PdMT proteins in Cu homeostasis and/or its sequestration. Phoenix dactylifera metallothionein induction reached a peak after 30 d of exposure to 0.2 mM of Cu. However, it was down-regulated in plants exposed to higher Cu concentrations. In the same conditions, Pdpcs was overexpressed during 1 mo of exposure before it decreased thereafter. These observations provide a new insight into date palm cell response to Cu, a metal that can be toxic but that is also an essential element. Environ Toxicol Chem 2018;37:797-806. © 2017 SETAC.

Antagonistic Properties of Some Halophilic Thermoactinomycetes Isolated from Superficial Sediment of a Solar Saltern and Production of Cyclic Antimicrobial Peptides by the Novel Isolate Paludifilum halophilum.

This study has focused on the isolation of twenty-three halophilic actinomycetes from two ponds of different salinity and the evaluation of their ability to exert an antimicrobial activity against both their competitors and several other pathogens. From the 23 isolates, 18 strains showed antagonistic activity, while 19 showed activities against one or more of the seven pathogen strains tested. Six strains exhibited consistent antibacterial activity against Gram-negative and Gram-positive pathogens characterized at the physiological and molecular levels. These strains shared only 94-95% 16S rRNA sequence identity with the closely related species of the Thermoactinomycetaceae family. Among them, the potent strain SMBg3 was further characterized and assigned to a new genus in the family for which the name Paludifilum halophilum (DSM 102817T) is proposed. Sequential extraction of the antimicrobial compounds with ethyl acetate revealed that the crude extract from SMBg3 strain had inhibitory effect on the growth of the plant pathogen Agrobacterium tumefaciens and the human pathogens Staphylococcus aureus, Salmonella enterica, Escherichia coli, and Pseudomonas aeruginosa. Based on the HRESI-MS spectral data, the cyclic lipopeptide Gramicidin S and four cyclic dipeptides (CDPs) named cyclo(L-4-OH-Pro-L-Leu), cyclo(L-Tyr-L-Pro), cyclo(L-Phe-L-Pro), and cyclo(L-Leu-L-Pro) were detected in the fermentation broth of Paludifilum halophilum. To our knowledge, this is the first report on the isolation of these compounds from members of the Thermoactinomycetaceae family.

Extraction optimization and in vitro and in vivo anti-postprandial hyperglycemia effects of inhibitor from Phoenix dactylifera L. parthenocarpic fruit.

Phoenix dactylifera L. plays an important role in social, economic, and ecological Tunisian sectors. Some date palms produce parthenocarpic fruit named Sish. The aqueous ethanolic extract from P. dactylifera parthenocarpic dates demonstrated a potent inhibition of the enzymes related to type II diabetes. In this work, extraction optimization of amylase inhibitors was carried out using Box-Behnken Design. Bioactivity-guided fractionation of the 70% aqueous ethanol extract was performed to identify the active compounds. The physicochemical results by liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis showed the presence of 13 phenolic compounds. The in vitro study showed that the extract exhibited a more specific inhibitor of α-glucosidase than α-amylase with an IC50 value of 0.6 and 2.5mg/mL, respectively. The in vivo study of this extract effect on the postprandial hyperglycemia activity showed a decrease in plasma glucose levels after 30min stronger than the Acarbose effect. These results confirmed the anti-postprandial hyperglycemia activity of the aqueous ethanolic extract from P. dactylifera parthenocarpic dates, which could lend support for its pharmaceutical use.

Extraction, partial purification and characterization of amylase from parthenocarpic date (Phoenix dactylifera): effect on cake quality.

BACKGROUND: Phoenix dactylifera L. plays an important role in social, economic and ecological Tunisian sectors. Some date palms produce parthenocarpic fruit named Sish. The objective of the present study was to extract biomolecules from parthenocarpic fruit by producing value-added products from the fruits. RESULTS: The extraction of amylolytic activity from parthenocarpic fruit (AmyPF) was optimized using Box-Behnken design (BBD). Partial purification of about 250-fold with an activity yield of 47% was achieved. The amylase exhibited a specific activity of 80 U mg-1 protein. The optimum pH and temperature for enzyme activity were 5 and 55 °C respectively. The enzyme was highly active over a wide range of pH (5-10), and significant stabilization was observed at 60 °C. The purified enzyme belongs to the exo type of amylases. Given the economic and industrial relevance of amylases used in the food industry, three different concentrations of AmyPF (0.007, 0.014 and 0.018 U g-1 ) were incorporated into a cake formulation, resulting in a decrease in density, moisture retention and water activity and an increase in hardness. CONCLUSION: The beneficial effect of AmyPF on the technological characteristics of cakes was confirmed by sensory evaluation. © 2017 Society of Chemical Industry.

Evaluation of anti-diabetic and anti-tumoral activities of bioactive compounds from Phoenix dactylifera L's leaf: In vitro and in vivo approach.

Among various chronic disorders, cancer and diabetes mellitus are the most common disorders. This study was designed to evaluate the effectiveness of hydroalcoholic extract of Phoenix dactylifera L. leaves (HEPdL) in animal models of type II diabetes in vitro/in vivo and in a human melanoma-derived cell line (IGR-39). A liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis was also performed to determine the amount of phenolic and flavonoid compounds in this plant. The physicochemical results by LC-MS/MS analysis of HEPdL showed the presence of 10 phenolic compounds. The in vitro study showed that the extract exhibited a more specific and potent inhibitor of α-glucosidase than α-amylase with an IC50 value of 20±1µg/mL and 30±0.8µg/mL, respectively. More importantly, the in vivo study of the postprandial hyperglycemia activity with (20mg/kg) of HEPdL showed a decrease in plasma glucose levels after 60min in resemblance to the glucor (acarbose) (50mg/kg) effect. The oral administration of HEPdL (20mg/kg) in alloxan-induced diabetic mices for 28days showed a more significant anti-diabetic activity than that of the drug (50mg/kg). Moreover, cytotoxicity effects of HEPdL in IGR-39 cancer cell lines were tested by MTT assay. This extract was effective in inhibiting cancer cells growth (IGR-39) at dose 35 and 75µg/mL. These results confirm ethnopharmacological significance of the plant and could be taken further for the development of an effective pharmaceutical drug against diabetes and cancer.

Combat sports practice favors bone mineral density among adolescent male athletes.

The aim of this study was to determine the impact of combat sports practice on bone mineral density (BMD) and to analyze the relationship between bone parameters and anthropometric measurements, bone markers, and activity index (AI). In other words, to detect the most important determinant of BMD in the adolescent period among combat sports athletes. Fifty athletes engaged in combat sports, mean age 17.1±0.2 yr, were compared with 30 sedentary subjects who were matched for age, height, and pubertal stage. For all subjects, the whole-body BMD, lumbar spine BMD (L2-L4), and BMD in the pelvis, arms, and legs was measured by dual-energy X-ray absorptiometry, and anthropometric measurements were evaluated. Daily calcium intake, bone resorption, and formation markers were measured. BMD measurements were greater in the combat sports athletes than in the sedentary group (p<0.01). Weight, body mass index, and lean body mass were significantly correlated with BMD in different sites. Daily calcium consumption lower than daily calcium intake recommended in both athletes and sedentary group. AI was strongly correlated with all BMD measurements particularly with the whole body, legs, and arms. Negative correlations were observed between bone markers and BMD in different sites. The common major predictor of BMD measurements was AI (p<0.0001). AI associated to lean body mass determined whole-body BMD until 74%. AI explained both BMD in arms and L2-L4 at 25%. AI associated to height can account for 63% of the variance in BMD legs. These observations suggested that the best model predicting BMD in different sites among adolescent combat sports athletes was the AI. Children and adolescents should be encouraged to participate in combat sports to maximize their bone accrual.

AaTX1, from Androctonus australis scorpion venom: purification, synthesis and characterization in dopaminergic neurons.

We have purified the AaTX1 peptide from the Androctonus australis (Aa) scorpion venom, previously cloned and sequenced by Legros and collaborators in a venom gland cDNA library from Aa scorpion. AaTX1 belongs to the α-Ktx15 scorpion toxins family (αKTx15-4). Characterized members of this family share high sequence similarity and were found to block preferentially IA-type voltage-dependent K(+) currents in rat cerebellum granular cells in an irreversible way. In the current work, we studied the effects of native AaTX1 (nAaTX1) using whole-cell patch-clamp recordings of IA current in substantia nigra pars compacta dopaminergic neurons. At 250 nM, AaTX1 induces 90% decrease in IA current amplitude. Its activity was found to be comparable to that of rAmmTX3 (αKTx15-3), which differs by only one conserved (R/K) amino acid in the 19th position suggesting that the difference between R19 and K19 in AaTX1 and AmmTX3, respectively, may not be critical for the toxins' effects. Molecular docking of both toxins with Kv4.3 channel is in agreement with experimental data and suggests the implication of the functional dyade K27-Y36 in toxin-channel interactions. Since AaTX1 is not highly abundant in Aa venom, it was synthesized as well as AmmTX3. Synthetic peptides, native AaTX1 and rAmmTX3 peptides showed qualitatively the same pharmacological activity. Overall, these data identify a new biologically active toxin that belongs to a family of peptides active on Kv4.3 channel.

Characterization of the bacteriocin BacJ1 and its effectiveness for the inactivation of Salmonella typhimurium during turkey escalope storage.

During a screening programme for bacteriocin producers, a new lactic acid bacterium called J1, isolated from chicken gizzard, was noted to produce a bacteriocin (BacJ1) that inhibited Gram-positive and Gram-negative food-borne pathogens, especially Salmonella typhimurium, and it was, therefore, selected for identification and further study. The results, from 16S rRNA gene nucleotide sequencing (1583 pb accession no HE861352) and phylogenetic analysis, suggested that the new isolate be assigned as Weissella paramesenteroides J1. BacJ1 was purified from the culture supernatant of the J1 strain using heat treatment (15 min at 90 °C), 80% ammonium sulfate precipitation, and gel filtration (Sephadex G25). MALDI-TOF analyses revealed that BacJ1 had an exact molecular mass of 1881.036 Da. The analysis of the N-terminal sequence (GPAGPFGKLY) of this active peptide revealed no significant similarity to currently available antimicrobial peptides. The addition of pre-purified BacJ1, at a final concentration of 400 AU per gramme, may open new promising opportunities for the prevention of contamination from and growth of pathogenic bacteria, particularly S. typhimurium, during turkey escalope storage at 4 °C.

Enzymatic properties of stingray Dasyatis pastinaca group V, IIA and IB phospholipases A(2): a comparative study.

In the present study, we have purified the group V phospholipase from the heart of cartilaginous fish stingray Dasyatis pastinaca and compared its biochemical properties with group IIA (sPLA2-IIA) and IB (sPLA2-IB) phospholipases previously purified from pancreas and intestine, respectively. Group V phospholipase (sPLA2-V) was purified to homogeneity by heat treatment, ammonium sulphate precipitation and RP-HPLC. The N-terminal sequence of the purified sPLA2-V exhibits a high degree of homology with those of mammal. The enzyme was found to be monomeric with a molecular mass estimation of 14 kDa. The specific activity of the purified enzyme, measured at pH 8 and 37 °C was 52 U/mg. Like sPLA2-IB and sPLA2-IIA, the sPLA2-V is found to be stable between pH 3 and 11 after 30 min of incubation. The purified sPLA2-V retained 65% of its activity after 10 min of incubation at 70 °C and it absolutely requires Ca(2+) for enzymatic activity. In addition it displayed high tolerance to organic solvents. Kinetic parameters Kmapp, kcat and the deduced catalytic efficiency (kcat/Kmapp) of the purified group-V, -IB and -IIA PLA2s were determined using phosphatidylethanolamine (PE), phosphatidylcholine (PC) or phosphatidylserine (PS) as substrate. The three enzymes hydrolyze the zwiterionic PE and PC substrates more efficiently than anionic PS substrate.

Morphological and biochemical behavior of fenugreek (Trigonella foenum-graecum) under copper stress.

The effects of copper on germination and growth of fenugreek (Trigonella foenum-graecum) was investigated separately using different concentrations of CuSO4. The germination percentage and radical length had different responses to cupric ions: the root growth increased with increasing copper concentration up to 1 mM Cu²âº and was inhibited thereafter. In contrast, the germination percentage was largely unaffected by concentrations of copper below 10 mM. The reduction in root growth may have been due to inhibition of hydrolytic enzymes such as amylase. Indeed, the average total amylolytic activity decreased from the first day of treatment with [Cu²âº] greater than 1 mM. Furthermore, copper affected various plant growth parameters. Copper accumulation was markedly higher in roots as compared to shoots. While both showed a gradual decrease in growth, this was more pronounced in roots than in leaves and in stems. Excess copper induced an increase in the rate of hydrogen peroxide (H2O2) production and lipid peroxidation in all plant parts, indicating oxidative stress. This redox stress affected leaf chlorophyll and carotenoid content which decreased in response to augmented Cu levels. Additionally, the activities of proteins involved in reactive oxygen species (ROS) detoxification were affected. Cu stress elevated the ascorbate peroxidase (APX) activity more than two times at 10 mM CuSO4. In contrast, superoxide dismutase (SOD) and catalase (CAT) levels showed only minor variations, only at 1 mM Cu²âº. Likewise, total phenol and flavonoid contents were strongly induced by low concentrations of copper, consistent with the role of these potent antioxidants in scavenging ROS such as H2O2, but returned to control levels or below at high [Cu²âº]. Taken together, these results indicate a fundamental shift in the plant response to copper toxicity at low versus high concentrations.

Purification and characterization of a phospholipase A2-IIA from common stingray (Dasyatis pastinaca) intestine.

A phospholipase A2 belonging to IIA group secretory PLA2 was isolated and purified to homogeneity from the intestine of common stingray (Dasyatis pastinaca) using acidic treatment (pH 1.5) and ammonium sulphate precipitation methods combined with single-column ion-exchange chromatography. The purified enzyme was found to be a glycosylated monomeric protein with a molecular mass of about 14 kDa. The stingray sPLA2-IIA had optimum activity at 45 degrees C, unlike known mammalian PLA2-IIAs, which show optimum activity at 37 degrees C. The purified enzyme exhibited a specific activity of 290 U/mg at optimal conditions (pH 9.5 and 45 degrees C) in the presence of 6 mM NaDC and 8 mM CaCl2 with egg yolk as substrate. The NH2-terminal sequence of the enzyme and some protein fragments obtained from its tryptic digestion were also determined. All sequences obtained were similar to those of sPLA2-IIA. The enzyme also showed good stability in the presence of organic solvents, acidic and alkaline pH media and high temperature conditions. Thus, the purified enzyme exhibited a number of unique and promising properties, making it a potential possible candidate for future applications in the treatment of phospholipid-rich industrial effluents and synthesis of useful preparations for the food production and processing industry.

Microbial production of levanase for specific hydrolysis of levan.

A newly isolated bacterial strain from Tunisian thermal source was selected for its ability to produce extracellular levanase when grown on levan substrate. The optimization of carbon source, nitrogen source, temperature and initial pH of the growth medium in submerged liquid cultures were investigated. In fact, levan was found to be a good inducer of levanase enzymes. The optimal temperature and pH of the levanase activity were 40 °C and 6.4, respectively. This enzyme exhibited a remarkable stability and retained 75% of its original activity at 55 °C for more than 1 h at pH 6.4. Crude enzyme of the strain rich in levanase was established for the hydrolysis of levan in order to produce fructooligosaccharides with variable degrees of polymerization which could be used in important fields such medicine, food-processing industry and cosmetic. The extracellular levanase of the strain was then, partially purified as determined by SDS-PAGE. The purification was achieved by ammonium sulfate precipitation, gel filtration and DEAE cellulose chromatographies.

Structural determination and NMR characterization of a bacterial exopolysaccharide.

A strain of Bacillus licheniformis with high exopolysaccharide (EPS) production ability was isolated and identified. A new type of EPS was isolated from the strain fermentation and its structural characteristics were investigated and elucidated by partial and total acid hydrolysis, Fourier transform infrared, and (1)H and (13)C NMR spectroscopy including 2D (1)H, COSY, NOESY, XHCOR and HMBC experiments. Based on obtained data, the EPS was found to be a levan composed of linear chains of (2→6)-linked ß-d-fructofuranosyl residues with connections ß (2→6).