Phytochemical profiling, in vitro biological activities, and in silico (molecular docking and absorption, distribution, metabolism, excretion, toxicity) studies of Polygonum cognatum Meissn.

Polygonum cognatum Meissn, a perennial herbaceous belonging to the Polygonaceae family, is an aromatic plant. High-performance liquid chromatography/diode array detector method was developed and validated for the phytochemical analysis of the plant. Also, various methods were used to investigate the antioxidant, antimicrobial, and cytotoxic activities of the methanolic extracts. Antioxidant activities were researched by 2,2'-diphenyl-1-picrylhydrazyl and cupric reducing antioxidant capacity methods. Among the tested standard microbial strains, Candida albicans was found to be more sensitive with a 24.60 ± 0.55 mm inhibition zone according to the diffusion tests. In the microdilution tests, the minimum inhibitory concentration and minimum bactericidal/fungicidal concentration values were 4.75 and ≥ 4.75 mg/mL, respectively, for all tested pathogens. Human colon carcinoma cells were used to investigate cytotoxicity by using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide analysis (IC50  = 2891 µg/mL for Plant A, IC50  = 3291 µg/mL for Plant B). Molecular docking and absorption, distribution, metabolism, excretion, and toxicity analysis were used to explain inhibition mechanisms of major phenolic compounds of plants against Tankyrase 1, Tankyrase 2 enzymes, and deoxyribonucleic acid gyrase subunit B and found compatible with experimental results.

The combined effects of polyethylene microplastics and benzoanthracene on Manila clam Ruditapes philippinarum.

Microplastic (MP) toxicity has recently been explored in various marine species. Along with the toxicity of plastics polymer itself, additional substances or pollutants that are absorbed onto it may also be harmful. In the present study, we investigated the combined impacts of polyethylene microplastics (PE MPs) and an organic pollutant (Benzo(a)anthracene, BaA) on Manila clam Ruditapes philippinarum during a one-week exposure. Two PE MPs concentrations (26 µg L-1 and 260 µg L-1) and one BaA concentration (3 µg L-1) were tested. The clams were exposed to BaA and PE MPs either alone or in combination. BaA and PE MPs were incubated before the combined exposure. The biological effects of PE MPs and BaA on the clams were evaluated by considering several assays such as feeding rate, anti-oxidant enzyme activities, and the expression levels of stress-related genes. The feeding rate significantly decreased in individual PE MPs and individual BaA groups while it remained unchanged in combined groups. Superoxide dismutase (SOD) was the most affected among the biochemical parameters. Malondialdehyde (MDA), and glutathione peroxidase (GPx) activities were slightly affected, whereas no changes were observed in glutathione s-transferase (GST) activities. CYP1A1, CYP3A4, and HSP70 gene expressions displayed slightly significant changes. Considering all stressor groups, high PE MPs exposure (260 µg L-1 PE MPs) more effectively altered the biological parameters in the clams compared to individual low PE MPs and BaA exposure, and their combination. The results also indicated the negligible vector role of PE MPs to transport BaA into the clam tissues.

Hyperglycemia with or without insulin resistance triggers different structural changes in brain microcirculation and perivascular matrix.

Both type-1 and type-2 DM are related to an increased risk of cognitive impairment, neurovascular complications, and dementia. The primary triggers for complications are hyperglycemia and concomitant insulin resistance in type-2 DM. However, the diverse mechanisms in the pathogenesis of diabetes-related neurovascular complications and extracellular matrix (ECM) remodeling in type-1 and 2 have not been elucidated yet. Here, we investigated the high fat-high sucrose (HFHS) feeding model and streptozotocin-induced type-1 DM model to study the early effects of hyperglycemia with or without insulin resistance to demonstrate the brain microcirculatory changes, perivascular ECM alterations in histological sections and 3D-reconstructed cleared brain tissues. One of the main findings of this study was robust rarefaction in brain microvessels in both models. Interestingly, the HFHS model leads to widespread non-functional angiogenesis, but the type-1 DM model predominantly in the rostral brain. Rarefaction was accompanied by basement membrane thickening and perivascular collagen accumulation in type-1 DM; more severe blood-brain barrier leakage, and disruption of perivascular ECM organization, mainly of elastin and collagen fibers' structural integrity in the HFHS model. Our results point out that the downstream mechanisms of the long-term vascular complications of hyperglycemia models are structurally distinctive and may have implications for appropriate treatment options.

Identification of natural compounds of Jurinea species by LC-HRMS and GC-FID and their bioactivities.

The Jurinea Cass. is one of the most important genera within Asteraceae and it comprises about 250 species in total. This genus is known for its numerous biological activities such as antioxidant, antimicrobial, antilipid peroxidation, anticholinesterase, antileishmanial activities. The aim of this study was to determine chemical composition and biological activities of ethanol and n-hexane extracts of three different Jurinea species. For this purpose, different parts of J. mollis, J. cadmea and J. pontica were extracted and totally six n-hexane and six ethanol extracts were obtained. Fatty acid content of n-hexane extracts was determined by GC-FID whereas phenolic and flavonoid content of ethanol extracts by LC-HRMS. Palmitic acid (16:0) was detected as the most abundant fatty acid in all n-hexane extracts with the rates ranging from 42.16%-55.08%, except flowers of J. mollis (JMF) and J. cadmea (JCF). LC-HRMS analysis showed the rutin content of all extracts was higher than other flavonoids, except of J. cadmea flowers, whereas apigenin-7-glucoside was found the most abundant in JCF. Cytotoxic effects of the extracts on HeLa and HEK-293 cells were determined by MTT method, and antioxidant activities were evaluated by DPPH and CUPRAC assays. Ethanol extract of J. mollis flowers significantly inhibited cancerous HeLa cells, with the IC50 value of 9.683 µg/mL while it was more less toxic on healthy HEK-293 cells. Ethanol extracts of J. mollis flowers and J. mollis steams-leaves (JMSL) showed the highest antioxidant activity by a DPPH inhibition % of 45.516 ± 2.497 and 56.671 ± 1.496, respectively. JMF and JMSL have also the highest CUPRAC values (0.880 ± 0.067 and 1.085 ± 0.152 mmol TR/g DWE, respectively). Total flavonoid content was determined using aluminum chloride colorimetric assay while total tannin and phenolic content by Folin Chiocalteu's reagent. Results showed that JMSL has the highest total phenolic (108.359 ± 6.241 mg GAE/ G DWE) and flavonoid (32.080 ± 4.385 mg QE/ g DWE) contents whereas JMF has the highest tannin content (121.333 ± 17.889 mg TAE/ g DWE). In the light of these results, various parts of Jurinea species may be regarded as alternative sources for cytotoxic and/or antioxidant flavonoids, phenolics and unsaturated fatty acids that can arouse the interest of pharmaceutical, food and cosmetic industries.

Intranasal levosimendan prevents cognitive dysfunction and apoptotic response induced by repeated isoflurane exposure in newborn rats.

Anesthetic-induced toxicity in early life may lead to risk of cognitive decline at later ages. Notably, multiple exposures to isoflurane (ISO) cause acute apoptotic cell death in the developing brain and long-term cognitive dysfunction. This study is the first to investigate whether levosimendan (LVS), known for its protective myocardial properties, can prevent anesthesia-induced apoptotic response in brain cells and learning and memory impairment. Postnatal day (P)7 Wistar albino pups were randomly assigned to groups consisting of an equal number of males and females in this laboratory investigation. We treated rats with LVS (0.8 mg/kg/day) intranasally 30 min before each ISO exposure (1.5%, 3 h) at P7+9+11. We selected DMSO as the drug vehicle. Also, the control group at P7+9+11 received 50% O2 for 3 h instead of ISO. Neuroprotective activity of LVS against ISO-induced cognitive dysfunction was evaluated by Morris water maze. Expression of apoptotic-related proteins was detected in the whole brain using western blot. LVS pretreatment significantly prevented anesthesia-induced deficit in spatial learning (at P28-32) and memory (at P33, P60, and P90). No sex-dependent difference occurred on any day of the training and probe trial. Intranasal LVS was also found to significantly prevent the ISO-induced apoptosis by reducing Bax and cleaved caspase-3, and by increasing Bcl-2 and Bcl-xL. Our findings support pretreatment with intranasal LVS application as a simple strategy in daily clinical practice in pediatric anesthesia to protect infants and children from the risk of general anesthesia-induced cell death and cognitive declines.

Effects of microplastics and mercury on manila clam Ruditapes philippinarum: Feeding rate, immunomodulation, histopathology and oxidative stress.

Plastic pollution, which is one of the most important environmental problems at the present time, has been understood recently, and the effects of this pollution on ecosystem and biota are becoming a growing problem, especially in the aquatic ecosystems. Direct or indirect exposure to those particles leads to adverse effects on marine organisms. In the marine environment, plastic materials interact with other pollutants such as metals, thereby affecting the uptake levels of those pollutants in marine organisms. In the present study, the Manila clam Ruditapes philippinarum was exposed to polyethylene microbeads and mercury chloride in single, combined and incubated form at environmentally relative concentrations for one week in controlled laboratory conditions. The uptake and tissue distribution of both stressors as well as the vector role of microplastics on mercury uptake in the organisms were investigated. Filtration rates, biomarkers for immunomodulation and oxidative stress, and histological alterations were also evaluated. Microplastics were ingested by the clams, and translocated to the various tissues. However, contaminated microplastics displayed a negligible vector role in terms of mercury bioaccumulation in the clams. The single and interactive exposure of the stressors reduced the filtration rate in the clams. Both pollutants affected the immune system of the organisms. Histological alterations were determined in the gill and digestive gland tissues of the clams among the treatment groups, although oxidative stress biomarkers remained unchanged. This study suggests that the vector role of polyethylene microplastics in mercury uptake is negligible and reveals that the single and interactive one-week exposure of two pollutants induce toxicity in the manila clams.

Isoflurane exposure in infant rats acutely increases aquaporin 4 and does not cause neurocognitive impairment.

Isoflurane is commonly used in pediatric population, but its mechanism of action in cognition is unclear. Aquaporin 4 (AQP4) regulates water content in blood, brain, and cerebrospinal fluid. Various studies have provided evidence for the role of AQP4 in synaptic plasticity and neurocognition. In this study, we aimed to determine whether a prolonged exposure to isoflurane in infant rats is associated with cognition and what effect this exposure has on AQP4 expression. Ten-day-old [postnatal day (P) 10] Wistar albino rats were randomly allocated to isoflurane group (n = 32; 1.5% isoflurane in 50% oxygen for 6 hours) or control group (n = 32; only 50% oxygen for 6 hours). Acute (P11) and long-term (P33) effects of 6-hour anesthetic isoflurane exposure on AQP4 expression were analyzed in whole brains of P11 and P33 rats by RT-qPCR and Western blot. Spatial learning and memory were assessed on P28 to P33 days by Morris Water Maze (MWM) test. The analysis revealed that isoflurane increased acutely both mRNA (~4.5 fold) and protein (~90%) levels of AQP4 in P11 rats compared with control group. The increasing levels of AQP4 in P11 were not observed in P33 rats. Also, no statistically significant change between isoflurane and control groups was observed in the latency to find the platform during MWM training and probe trial. Our results indicate that a single exposure to isoflurane anesthesia does not influence cognition in infant rats. In this case, acutely increased AQP4 after isoflurane anesthesia may have a protective role in neurocognition.

Proteomic Identification of Allergenic Proteins of Morus alba L. Pollenexacerbation.

BACKGROUND: Tree pollens are well-known aeroallergens all over the world. Little is known about the allergenicity of Morus alba (white mulberry) pollen. OBJECIVE: We aimed to explore the potential allergens of this pollen and its clinical relevance in tree pollen allergic patients living in Istanbul, Turkey. METHODS: Twenty three seasonal allergic rhinitis patients with a confirmed tree pollen allergy and 5 healthy control subjects underwent skin prick and nasal provocation tests with M.alba pollen extract. The pollen extract was then resolved by gel electrophoresis, and immunoblotted with sera from patients/control individuals to detect the potential allergenic proteins. The prevalent IgE binding proteins from 1D-gel were analyzed by MALDI-TOF/TOF. RESULTS: Eleven out of 23 patients were reactive to the extract with skin prick tests. Seven of those patients also reacted positively to the nasal provocation tests. The most common IgE-binding pollen proteins were detected between 55-100 kDa, and also at molecular weights lower than 30 kDa for some patients. Mass spectrometry analyses revealed that the principal IgE-binding protein was methionine synthase (5-methyltetrahydropteroyltriglutamate homocysteine methyltransferase), which is then proposed as a novel allergen in M.alba pollen. CONCLUSION: This study provides the first detailed information for the potential allergens of Morus alba pollen of Istanbul. Methionine synthase with an apparent molecular weight of 80 to 85 kDa has been recognized as one of the allergens in Morus alba pollen for the first time.

Classical Galactosemia: Insight into Molecular Pathomechanisms by Differential Membrane Proteomics of Fibroblasts under Galactose Stress.

Classical galactosemia, a hereditary metabolic disease caused by the deficiency of galactose-1-phosphate uridyltransferase (GALT; EC 2.7.712), results in an impaired galactose metabolism and serious long-term developmental affection of the CNS and ovaries, potentially related in part to endogenous galactose-induced protein dysglycosylation. In search for galactose-induced changes in membrane raft proteomes of GALT-deficient cells, we performed differential analyses of lipid rafts from patient-derived (Q) and sex- and age-matched control fibroblasts (H) in the presence or absence of the stressor. Label-based proteomics revealed of the total 454 (female) or 678 (male) proteins a proportion of ∼12% in at least one of four relevant ratios as fold-changed. GALT(-) cell-specific effects in the absence of stressor revealed cell-model-dependent affection of biological processes related to protein targeting to the plasma membrane (female) or to cellular migration (male). However, a series of common galactose-induced effects were observed, among them the strongly increased ER-stress marker GRP78 and calreticulin involved in N-glycoprotein quality control. The membrane-anchored N-glycoprotein receptor CD109 was concertedly decreased under galactose-stress together with cadherin-13, GLIPR1, glypican-1, and semaphorin-7A. A series of proteins showed opposite fold-changes in the two cell models, whereas others fluctuated in only one of the two models.

Differential Proteomics of Urinary Exovesicles from Classical Galactosemic Patients Reveals Subclinical Kidney Insufficiency.

Classical galactosemia is caused by a nearly complete deficiency of galactose-1-phosphate uridyltransferase (GALT; EC 2.7.712), resulting in a severely impaired galactose metabolism with galactose-1-phosphate and galactitol accumulation. Even on a galactose-restricted diet, patients develop serious long-term complications of the central nervous system and ovaries that may result from chronic cell-toxic effects exerted by endogenous galactose. To address the question of whether disease-associated cellular perturbations could affect the kidney function of the patients, we performed differential proteomics of detergent-resistant membranes from urinary exovesicles. Galactosemic samples (showing drastic shifts from high-mannose to complex-type N-glycosylation on exosomal N-glycoproteins) and healthy, sex-matched controls were analyzed in quadruplex iTRAQ experiments performed in biological and technical replicates. Particularly in the female patient group, the most striking finding was a drastic increase of abundant serum (glyco)proteins, like albumin, leucine-rich α-2-glycoprotein, fetuin, immunoglobulins, prostaglandin H2 d-isomerase, and α-1-microglobulin protein (AMBP), pointing to a subclinical failure of kidney filter function in galactosemic patients and resulting in a heavy overload of exosomal membranes with adsorbed serum (glyco)proteins. Several of these proteins are connected to TBMN and IgAN, proteinuria, and renal damage. The impairment of renal protein filtration was also indicated by increased protein contents derived from extracellular matrices and lysosomes.

Effect of α-tocopheryl succinate on the molecular damage induced by indomethacin in C6 glioma cells.

Indomethacin is a member of the non-steroidal anti-inflammatory drug (NSAID) class, which has great potential for use in the treatment of glioma. However, it induces the generation of reactive oxygen species (ROS) and causes molecular damage while inducing its effects. Vitamin E is widely used in the complementary therapy of cancers. The main goal of the present study was to investigate the effects of α-tocopheryl succinate (α-TOS) against the oxidative damage induced by indomethacin in C6 glioma cells. Cells were treated with 10 µM α-TOS alone or in combination with 200 µM indomethacin for two days. The intracellular ROS level, molecular damage as revealed by lipid peroxidation and protein carbonyl formation, and the COX activity in C6 glioma cells were measured. Treatment of the cells with α-TOS and indomethacin, alone or in combination, caused the levels of ROS generation and protein damage to increase, but protected against lipid peroxidation and reduced COX activity.

Evaluation of biological activities of Physalis peruviana ethanol extracts and expression of Bcl-2 genes in HeLa cells

Physalis species are used in folk medicine for phytotherapeutic properties. The extracts of medicinal plants are known to possesscytotoxic and chemopreventative compounds. In this study we investigated antibacterial, antioxidant, DNA damage preventativeproperties of Physalis peruviana (golden berry) on leaf and shoot ethanol extracts and their effects on cytotoxicity of HeLa cellsand expression of apoptotic pathway genes. Among the tested bacteria for antibacterial activity, maximum inhibition zone wasdetermined in Lactococcus lactis. The phenolic content was found higher in leaf extracts than shoot extracts. The antioxidantactivity showed the highest TEAC values of the leaf (2 mg/mL) and the shoot (0.5 mg/mL) extracts as 0.291±0.04 and 0.192±0.015,respectively. In DNA damage prevention assay both leaf and shoot extracts, especially 30 and 20 µg/mL concentrations, exhibitedsignificant protection against DNA damage-induced by hydroxyl radical generated by Fenton reaction. Our results suggest thatleaf and shoot extracts possess cytotoxic effect on HeLa cells when applied as 100 µg/mL concentration. Also mRNA expressionanalysis showed the alteration of antiapoptotic genes, so the results suggest that P. peruviana ethanol extracts induce apoptoticcell death and should be investigated for identification of active compounds and their mechanisms of action.

Comparison of antioxidant capacity, protein profile and carbohydrate content of whey protein fractions.

Whey is used as an additive in food industry and a dietary supplement in nutrition. Here we report a comparative analysis of antioxidant potential of whey and its fractions. Fractions were obtained by size exclusion chromatography, before and after enzymatic digestion with pepsin or trypsin. Superoxide radical scavenging, lipid peroxidation inhibition and cupric ion reducing activities of different fractions were checked. Peptides were detected by SDS-PAGE and GC-MS was used to determine carbohydrate content of the fractions. All samples showed antioxidant activity and the second fraction of the trypsin hydrolysate showed the highest superoxide radical scavenging activity. CUPRAC value of this fraction was two-times higher than that of whey filtrate. The first fraction of the pepsin hydrolysate was the most effective inhibitor of lipid peroxidation. Each sample exhibited a different polypeptide profile. Different percentages of carbohydrates were identified in whey filtrate and in all second fractions, where galactose was the major component.

Investigation of the relationship between oxidative stress and glucose signaling in Schizosaccharomyces pombe.

The invertase mutant defective in the glucose signaling pathway of Schizosaccharomyces pombe (ird11) is resistant to glucose repression. This mutant is able to consume sucrose alongside glucose and grows in glucose-containing media with a generation time close to that of the wild type. Intracellular oxidation, protein carbonyl, and reduced glutathione levels and catalase, superoxide dismutase, and glutathione peroxidase activity were investigated in ird11, to determine the relationship between oxidative stress response and glucose signaling. The expression profiles of some genes involved in regulation of glucose repression (fbp1, fructose-1,6-bis-phosphatase; hxk2, hexokinase) and stress response (atf1 and pap1 transcription factors; ctt1, catalase; sod1, Cu,Zn superoxide dismutase) were analyzed using the quantitative real-time PCR technique. Oxidative stress response in ird11 seems to be affected by glucose signaling in a manner different from that caused by glucose deprivation.

Expression of human A4V mutant Cu,Zn superoxide dismutase in Schizosaccharomyces pombe: investigations of its toxic properties.

Cu,Zn superoxide dismutase (SOD1) is an antioxidant enzyme that catalyzes the removal of superoxide radicals generated in various biological oxidations. Amyotrophic lateral sclerosis (ALS) is one of the most common neurodegenerative disorders, occurring in families (FALS) and sporadically (SALS). FALS and SALS are distinguishable genetically but not clinically. More than 100 point mutations in the human SOD 1 gene have been identified that cause FALS. In order to determine the effects of mutant SOD protein, we first cloned wild-type and A4V mutant human SOD1 into Schizosaccharomyces pombe. This study shows viabilities and some antioxidant properties including SOD, catalase, proteasomal activity, and protein carbonyl levels of transformants in SOD1 deleted strain (MN415); and its parental strain (JY741) at different stress conditions. There was no more oxidative damage in the human mutant SOD carrying the transformant strain compared with other strains. These results may help to explain whether ALS progresses as a consequence of cellular oxidative damage.

Antioxidant and cytotoxic activities of Aphanes arvensis extracts.

The objectives of this study were to examine the free radical scavenging activity and the protective effects against macromolecular oxidation as well as the cytotoxic activity of Aphanes arvensis aqueous and methanolic extracts. Free radical scavenging activity was determined by DPPH method. The methanolic extract showed a scavenging activity nearly equivalent to Trolox and Vitamin C and has an IC(50) value of 4.54 microg/mL. Total antioxidant capacity was determined by CUPRAC method. The antioxidant capacity of aqueous and methanolic extract was 0.792 and 1.550 mmol TE/g DWE, respectively. The protective effect of A. arvensis extracts against lipid peroxidation was evaluated using a liposome oxidation system. The methanolic extract was more active than the aqueous extract. The aqueous extract possessed protective effect against protein oxidation in a dose dependent manner. Both extracts showed inhibitory effect on DNA oxidation as measured by plasmid relaxation assay. Results presented here indicate that A. arvensis possess strong antioxidant activity and protective effects with very little cytotoxic effect, and they can therefore be used as a natural additive in food, cosmetic and pharmaceutical industries.

Intracellular scavenging activity of Trolox (6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid) in the fission yeast, Schizosaccharomyces pombe.

The ability of Trolox (6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid), a water-soluble vitamin E analogue, to prevent oxidative damages is well characterized, but the mechanisms underlying it remain unclear. The protective effect of Trolox pre-treatment on H(2)O(2)-induced toxicity might be attributed to the decreased cellular permeability to H(2)O(2) or in vitro scavenging activity of Trolox, induction of antioxidant enzymes or the direct scavenging activity of Trolox. The results obtained rule out the first and second possibilities and intracellular scavenging activity was found to be the mechanism whereby Trolox confers protection. This was confirmed by measuring protein oxidation (levels), and the observed decrease in proteasomal activity indicated that the decrease in protein carbonyls was due to Trolox scavenging activity rather than proteasome activation. In conclusion, the intracellular scavenging activity of Trolox is a key protective mechanism against H(2)O(2). These findings obtained in Schizosaccharomyces pombe, a good model organism for eukaryotic cells, can be used as standard protocols for investigating the antioxidant activity of pure or complex potential antioxidants.

Fatty acid profile and in vitro antioxidant and antibacterial activities of red grape (Vitis vinifera L. cvs. Oküzgözü and Bogazkere) Marc extracts.

The marcs of two red grape (V. vinifera L.) varieties (Bogazkere and Oküzgözü), grown in eastern Anatolia (Elazig), were evaluated for their fatty acid composition, and antioxidant and antibacterial activities. The hexane extracts of both varieties were found to contain linoleic, palmitic, stearic and oleic acids by GC-MS analyses. The major fatty acid (linoleic acid) was detected relatively as 57.13% in Oküzgözü and 59.07% in Bogazkere in methylated hexane extracts. In addition, myristic and palmitoleic acids were observed in Bogazkere as minor components. The free radical (DPPH) scavenging activity of Oküzgözü was higher than that of Bogazkere. The IC50 values were calculated as 403.0 +/- 7.8 microg/mL for Oküzgozü and 552.0 +/- 23.6 microg/mL for Bogazkere. The extracts were found to be effective on the four gram (+) and four gram (-) test bacteria, but not as good as standard antibiotic, gentamycine by MIC method.

Antioxidant activity of whey protein fractions isolated by gel exclusion chromatography and protease treatment.

Whey proteins were isolated from whey powder by a combination of gel exclusion chromatography and protease (pepsin or trypsin) treatment. Whey solution (6g/dl) was applied to Sephadex G-200 column chromatography and three fractions were obtained. Gel electrophoresis (SDS-PAGE) was used to identify the fractions; the first one contained immunoglobulins and bovine serum albumin, the second contained beta-lactoglobulin and alpha-lactalbumin whereas the third fraction contained small peptides. We have also subjected the whey filtrate to proteases (pepsin and trypsin). Treatment with proteases showed that beta-lactoglobulin can be obtained after hydrolysis of the second fraction with pepsin. When the whey filtrate was treated with pepsin and then applied to Sephadex G-200 column chromatography three fractions were obtained; the first one was bovine serum albumin, the second was beta-lactoglobulin and the third fraction contained small peptides. After trypsin treatment only two fractions were obtained; the first one was serum albumin and the second fraction was an alpha-lactalbumin rich fraction. We have determined the antioxidant activity of the fractions using an assay based on the measurement of superoxide radical scavenging activity. Our results showed that among the three fractions, the first fraction had the highest superoxide radical scavenging activity. Also, protease treatment of the second fraction resulted in an increase in the antioxidant activity.

The effect of superoxide dismutase deficiency on zinc toxicity in Schizosaccharomyces pombe.

Zinc is a metal which is a cofactor in many enzymes and a structural element in zinc finger motifs those are important in relation between DNA and regulator proteins. Little is known about uptake, distribution, toxicity and detoxification of zinc ions in cells. In this study, zinc toxicity and detoxification levels have been compared in wild type and Cu/Zn superoxide dismutase mutant (sod1Delta) cells of the fission yeast Schizosaccharomyces pombe. We evaluated the toxic levels of zinc, total zinc content, lipid peroxidation levels and catalase activities for both strains which were grown in medium containing different concentrations of zinc. sod1Delta mutant showed important growth retardation and has higher lipid peroxidation and catalase activities than wild type. Cu/Zn superoxide dismutase (SOD1) activity of wild type cells was markedly increased when they were treated with elevated levels of zinc. SOD1 mRNA level also significantly increased when the cells treated with higher concentrations of zinc. These results indicate that the mutant cells were more sensitive to zinc stress and seemed to have more oxidative intracellular environment than wild type cells. Our results support the idea that superoxide dismutase is an important factor for zinc detoxification in eukaryotes.