Evaluation of potential human health risks from exposure to volatile organic compounds in contaminated urban groundwater in the Sava river aquifer, Belgrade, Serbia.

The oil pollutant in the Sava River aquifer in the residential area of Belgrade, Serbia was investigated in order to analyze the extent, origin and spatial distribution of the pollution, with the aim to estimate potential human health risks from exposure to the compounds detected. Analytical methods indicated that the dominant compounds in this oil pollutant were gasoline range organic compounds. Benzene, toluene, ethylbenzene and xylenes (BTEX) were identified as compounds of concern and quantified by headspace gas chromatography. The concentrations of benzene measured at all sampling points were higher than the remediation value while the maximum concentrations of BTEX quantified were among the highest concentrations of these compounds reported in the petroleum-contaminated aquifers in the world. The assessment of the human health risks from exposure to BTEX-covered industrial scenario for adult receptors and residential scenario for adult receptors and children. The exposure routes analyzed were dermal contact with and ingestion of contaminated water, considering both cancer and non-cancer effects. The analysis of the lifetime incremental cancer risk indicated the potential for adverse health effects for human exposure at the investigated location, and because of that it was interpreted as an unacceptable risk level or risks of high priority which required immediate consideration for remedial measures at this location. A complete set of mitigation measures was proposed including: groundwater decontamination treatment, installation of filters for tap water, development of the system for monitoring of BTEX in the groundwater and development of the emergency response capacities at this location.

Study on the assessment of humification processes during biodegradation of heavy residual fuel oil.

The aim of this study was to investigate the creation of humic substances during biodegradation of heavy residual fuel oil, because there are indications that substances similar to humic substances are generated during biodegradation of polycyclic aromatic hydrocarbons. In the study, which lasted for 110 days, biodegradation of heavy residual fuel oil was carried out in a layer of artificial soil substrate. The initial concentration of the total petroleum hydrocarbon in the prepared artificial soil substrate (biopile) was 23.1 g kg-1 dry weight (d.w.). At the end of the process, the total petroleum hydrocarbons were reduced to 8.1 g kg-1 d.w. in the inoculated biopile, while the content of humic acids increased during bioremediation from 3.15 g kg-1 d.w. to 4.95 g kg-1 d.w. The humic acids extracted from biopile during the biodegradation process were characterized by various chemical techniques (elemental analysis, spectrofluorimetric analysis, electrochemical measurements, and size exclusion chromatography). The results showed that levels of C, H and the H/C ratio decreased as the biodegradation process progressed. This indicated that humic acids aromatization process took place and this was confirmed by the spectrofluorimetric analysis. The increase of oxygen percentage and the O/C ratio in the humic acids after the biodegradation treatment indicated an increase in functional oxygen groups. Additional analyses of humic acids from the inoculated biopile showed that they were transformed during the bioremediation process. They had greater redox and buffering capacities and a larger portion of the fractions had high molecular mass. Also, the humification parameters (the CHAs/CFAs ratio and CHAs/Corg ratio) increased during the biodegradation. This is one of the few studies that describes the generation of humic substances during the biodegradation of oil compounds.

Investigation of potentially toxic elements in urban sediments in Belgrade, Serbia.

Concentrations of 10 potentially toxic trace elements were measured in soil samples collected from 18 sites in urban areas of Belgrade, the capital and the largest city in Serbia. Samples were analysed in order to assess the extent of soil contamination and to distinguish natural and anthropogenic input. The results demonstrated a general Zn, Pb, and Cd enrichment in surface samples, hence, after comparison with guidelines, it has been determined that there is a potential risk for human health. In the surface and buried samples, traces of cooper were detected. The highest concentrations of Cr (121 ± 12.3 mg/kg) and Ni (94.9 ± 12.6 mg/kg) were found at a sediment depth of 2 m and should be ascribed to geogenic sources. The results of HCA and PCA analysis supported a natural origin of Co, Cu, Mn, Cr, and Ni, while Cd, Zn, and Pb originated from anthropogenic inputs. Based on the pollution indices, Cd made the most dominant contribution, posing a high contamination risk in the studied area. Results of Nemerow pollution index (PIN) and potential ecological risk index (PER) demonstrated that pollution by heavy metals in sediments on several sites is moderately intense.

High-quality draft genome sequence of Pseudomonas aeruginosa san ai, an environmental isolate resistant to heavy metals.

The strain Pseudomonas aeruginosa san ai, isolated from an extreme environment (industrial mineral cutting oil, pH 10), is able to survive and persist in the presence of a variety of pollutants such as heavy metals and organic chemicals. The genome of P. aeruginosa san ai is 6.98 Mbp long with a GC content of 66.08% and 6485 protein encoding genes. A large number of genes associated with proteins, responsible for microbial resistance to heavy metal ions and involved in catabolism of toxic aromatic organic compounds were identified. P. aeruginosa san ai is a highly cadmium-resistant strain. Proteome analysis of biomass after cadmium exposal confirmed a high tolerance to sublethal concentrations of cadmium (100 mg/L), based on: extracellular biosorption, bioaccumulation, biofilm formation, controlled siderophore production and a pronounced metalloprotein synthesis. Proteins responsible for survival in osmostress conditions during exposure to elevated concentrations of cadmium (200 mg/L) demonstrate a strong genetic potential of P. aeruginosa san ai for survival and adaptation. Sequencing of P. aeruginosa san ai genome provides valuable insights into the evolution and adaptation of this microbe to environmental extremes at the whole-genome level, as well as how to optimally use the strain in bioremediation of chemically polluted sites.

Phytochemical, Free Radical Scavenging and Antifungal Profile of Cuscuta campestris Yunck. Seeds.

This work was conceptualized with the goal to investigate the phytochemical, free radical scavenging and antifungal profile of Cuscuta campestrisYunck. seeds. Total phenolics, amino acid and carbohydrate contents were evaluated in ethanolic, acetone and chloroform extract. Effective antioxidant activity was evaluated throughout seven antioxidant methods. The antifungal activity was assessed against eight fungal strains and Candida albicans. The results showed total phenol, flavonoid, flavonols and phenolic acids contents in amount of 1.51 - 6.35 mg GAE/mL, 78 - 425 µg RU/mL, 1.04 - 2.98 mg QU/g and 12.01 - 30.58 µg CAE/mL, respectively. The total amino acids and carbohydrates content ranged from 8.29 to 185.45 µg Gly/mL and from 0.05 to 0.12 µg Glu/mL. The ethanolic extract showed the best antioxidant activity in phosphomolybdenum, DPPH free radical scavenging, ferric reducing power and lipid peroxidation assays. The best activity in ferrous ion chelating and H2 O2 assays had the acetone extract, whereas the best hydroxyl radical scavenging activity was observed with the chloroform extract. The ethanolic extract at a concentration of 6 mg/mL proved to be the most effective antimycotic, since it inhibited the growth of all tested fungi except Penicillium verrucosum. The obtained results indicate that C. campestris seeds could be attributed to a potential source of natural antioxidants in food and pharmaceutical products.

Potential of pure and mixed cultures of Cladosporium cladosporioides and Geotrichum candidum for application in bioremediation and detergent industry.

The effect of ethoxylated oleyl-cetyl alcohol (Henkel, "Merima", Serbia) on the growth and metabolic activity of Cladosporium cladosporioides, Geotrichum candidum and their mixed culture was in the focus of this paper. The cultures were grown in Czapek-Dox liquid nutrient medium with the addition of 0.5% pollutant and without it. The physico-chemical and biochemical changes of pH, the total biomass dry weight, the quantity of free and total organic acids, proteolytic activity and the quality of carbohydrates were evaluated from 4th to 19th day of fungal growth. The pollutant caused an inhibitory effect on biomass dry weight of C. cladosporioides and G. candidum for 10.36% and 4.65% respectively, and stimulatory effect on biomass of mixed culture for 3.80%. The pollutant had influence on the decrease in pH value of the media in the phase of culture growth, and pH changes were correlated with the amount of excreted total organic acids. The highest quantity of free and total organic acids was noted in media with pollutant of mixed culture and C. cladosporioides, respectively. The alkaline protease activities of C. cladosporioides, G. candidum and mixed culture were enhanced by addition of pollutant for 56.88%, 55.84% and 30.94% respectively. The obtained results indicate the potential of both pure and mixed cultures in mycoremediation environment contaminated by alcohol ethoxylated and detergent industry.

Natural attenuation of petroleum hydrocarbons-a study of biodegradation effects in groundwater (Vitanovac, Serbia).

The role of natural attenuation processes in groundwater contamination by petroleum hydrocarbons is of intense scientific and practical interest. This study provides insight into the biodegradation effects in groundwater at a site contaminated by kerosene (jet fuel) in 1993 (Vitanovac, Serbia). Total petroleum hydrocarbons (TPH), hydrochemical indicators (O2, NO3-, Mn, Fe, SO42-, HCO3-), δ13C of dissolved inorganic carbon (DIC), and other parameters were measured to demonstrate biodegradation effects in groundwater at the contaminated site. Due to different biodegradation mechanisms, the zone of the lowest concentrations of electron acceptors and the zone of the highest concentrations of metabolic products of biodegradation overlap. Based on the analysis of redox-sensitive compounds in groundwater samples, redox processes ranged from strictly anoxic (methanogenesis) to oxic (oxygen reduction) within a short distance. The dependence of groundwater redox conditions on the distance from the source of contamination was observed. δ13C values of DIC ranged from - 15.83 to - 2.75‰, and the most positive values correspond to the zone under anaerobic and methanogenic conditions. Overall, results obtained provide clear evidence on the effects of natural attenuation processes-the activity of biodegradation mechanisms in field conditions.

Redox properties of transitional milk from mothers of preterm infants.

AIM: There is a discrepancy between the amount of transitional milk produced by mothers of preterm infants and the low capacity of premature infants to consume it. This milk can be used in milk banks, but previous studies found that there are large variations in the level of host-defence proteins in individual samples of milk from mothers of premature infants, which implies that large individual variations in antioxidative defence composition are also possible. METHODS: Milk samples were collected from 20 healthy mothers of preterm infants. We determined the values for non-enzymatic antioxidative capacity parameters (oxygen radical absorbance capacity (ORAC)), static oxidation-reduction potential (ORP), activities of antioxidant defence enzymes and the amount of vitamin C in whole milk, skim and whey fractions of transitional milk. RESULTS: The main low-molecular-weight antioxidant in transitional milk is vitamin C and most of it is contained in whey. ORAC is higher in whole transitional milk than in skim milk and whey, and ORP is lower in whole transitional milk than that in skim milk and whey. Antioxidative enzyme activities are similar in all individual samples of transitional milk from mothers of preterm infants. CONCLUSIONS: Our results indicate that transitional milk of mothers of preterm infants shows slow individual variations in antioxidative defence composition; therefore, it can be used in human milk banks.

Brachybacterium sp. CH-KOV3 isolated from an oil-polluted environment-a new producer of levan.

Various microorganisms isolated from polluted environments, such as Pseudomonas sp. and Micrococcus sp. can synthesize exopolysaccharides (EPSs) which are natural, non-toxic and biodegradable polymers. EPSs play a key role in protection of microbial cells under various external influences. For humans, these substances have potential use in many industries. EPSs can be applied as a flavor or a fragrance carrier, an emulsifier, a stabilizer, a prebiotic, an antioxidant or an antitumor agent. In this study, we characterized an environmental microorganism that produces EPS, optimized EPS production by this strain and characterized the EPS produced. Isolate CH-KOV3 was identified as Brachybacterium paraconglomeratum. The sucrose level in the growth medium greatly influenced EPS production, and the highest yield was when the microorganism was incubated in media with 500g/L of sucrose. The optimal temperature and pH were 28°C and 7.0, respectively. The nuclear magnetic resonance (NMR) results and GC-MS analysis confirmed that the residues were d-fructofuranosyl residues with ß-configuration, where fructose units are linked by ß-2,6-glycosidic bonds, with ß-2,1-linked branches. All these data indicate that the investigated EPS is a levan-type polysaccharide. Thus, it was concluded that Brachybacterium sp. CH-KOV3 could constitute a new source for production of the bioactive polysaccharide, levan.

Effect of the edaphic factors and metal content in soil on the diversity of Trichoderma spp.

Influence of edaphic factors and metal content on diversity of Trichoderma species at 14 different soil sampling locations, on two depths, was examined. Forty-one Trichoderma isolates from 14 sampling sites were determined as nine species based on their internal transcribed spacer (ITS) sequences. Our results indicate that weakly alkaline soils are rich sources of Trichoderma strains. Also, higher contents of available K and P are connected with higher Trichoderma diversity. Increased metal content in soil was not inhibiting factor for Trichoderma species occurrence. Relationship between these factors was confirmed by locally weighted sequential smoothing (LOESS) nonparametric smoothing analysis. Trichoderma strain (Szeged Microbiology Collection (SZMC) 22669) from soil with concentrations of Cr and Ni above remediation values should be tested for its potential for bioremediation of these metals in polluted soils.

Synthesis and characterization of a new type of levan-graft-polystyrene copolymer.

Novel macromolecular graft copolymers were synthesized by reaction of the hydroxyl groups of the microbial polysaccharide levan, produced using Bacillus licheniformis, with polystyrene (Lev-g-PS). Synthesis was performed by the free radical reaction using potassium persulfate (PPS) as initiator. The prepared copolymer was characterized by FTIR, SEM, TG/DTA, XRD and (13)C NMR. The influence of the different conditions (reaction temperature, air or nitrogen atmosphere, reaction time, type of amines and ascorbic acid (AA) concentration) on the grafting reaction was investigated. Results showed that maximum percentage of grafting (58.1%) was achieved at a reaction temperature 70°C, in a nitrogen atmosphere and using dimethylethanolamine (DMEA) as the amine activator. On the basis of the obtained results, the likely reaction mechanism was proposed. Synthesized copolymers have better thermal stability in comparison with their initial components. Copolymers such as Lev-g-PS could potentially have many applications, such as compatibilizers and material for membranes.

Treatment of a mud pit by bioremediation.

The mud generated from oil and natural gas drilling, presents a considerable ecological problem. There are still insufficient remedies for the removal and minimization of these very stable emulsions. Existing technologies that are in use, more or less successfully, treat about 20% of generated waste drilling mud, while the rest is temporarily deposited in so-called mud pits. This study investigated in situ bioremediation of a mud pit. The bioremediation technology used in this case was based on the use of naturally occurring microorganisms, isolated from the contaminated site, which were capable of using the contaminating substances as nutrients. The bioremediation was stimulated through repeated inoculation with a zymogenous microbial consortium, along with mixing, watering and biostimulation. Application of these bioremediation techniques reduced the concentration of total petroleum hydrocarbons from 32.2 to 1.5 g kg(-1) (95% degradation) during six months of treatment.

Interactions of the metal tolerant heterotrophic microorganisms and iron oxidizing autotrophic bacteria from sulphidic mine environment during bioleaching experiments.

Iron and sulfur oxidizing chemolithoautotrophic acidophilic bacteria, such as Acidithiobacillus species, hold the dominant role in mine environments characterized by low pH values and high concentrations of reduced sulfur and iron compounds, such as ores, rocks and acid drainage waters from mines. On the other hand, heterotrophic microorganisms, especially their biofilms, from these specific niches are receiving increased attention, but their potential eco-physiological roles have not been fully understood. Biofilms are considered a threat to human health, but biofilms also have beneficial properties as they are deployed in waste recycling and bioremediation systems. We have analyzed interactions of the metal tolerant heterotrophic microorganisms in biofilms with iron oxidizing autotrophic bacteria both from the sulphidic mine environment (copper mine Bor, Serbia). High tolerance to Cu(2+), Cd(2+) and Cr(6+) and the presence of genetic determinants for the respective metal tolerance and biofilm-forming ability was shown for indigenous heterotrophic bacteria that included strains of Staphylococcus and Rhodococcus. Two well characterized bacteria- Pseudomonas aeruginosa PAO1 (known biofilm former) and Cupriavidus metallidurans CH34 (known metal resistant representative) were also included in the study. The interaction and survivability of autotrophic iron oxidizing Acidithiobacillus bacteria and biofilms of heterotrophic bacteria during co-cultivation was revealed. Finally, the effect of heterotrophic biofilms on bioleaching process with indigenous iron oxidizing Acidithiobacillus species was shown not to be inhibitory under in vitro conditions.

Antioxidative Activity of Colostrum and Human Milk: Effects of Pasteurization and Storage.

OBJECTIVES: Milk banks collect, pasteurize, and freeze/store human milk. The processing may alter redox properties of milk, but the effects have not been fully examined. METHODS: We collected 10 mature milk and 10 colostrum samples and applied a battery of biochemical assays and electron paramagnetic resonance spectroscopy to inspect changes that milk undergoes with pasteurization and 30 days storage at -20°C. RESULTS: Pasteurization and storage of raw milk did not affect total nonenzymatic antioxidative capacity, but specific components and features were altered. Urate radical and ascorbyl radical emerge as products of exposure of milk to hydroxyl radical-generating system. Processing shifted the load of antioxidative activity from ascorbate to urate and lowered the capacity of milk to diminish hydroxyl radical. Pasteurization caused a significant drop in the activity of 2 major antioxidative enzymes-superoxide dismutase and glutathione peroxidase, whereas freezing/storage of raw milk affected only superoxide dismutase. Colostrum showed drastically higher total nonenzymatic antioxidative capacity, hydroxyl radical scavenging ability, and glutathione reductase activity compared with mature milk. CONCLUSIONS: Pasteurization and storage affect nonenzymatic and enzymatic antioxidative agents in human milk. It appears that nonenzymatic antioxidative systems in colostrum and milk are different. The effects of processing may be partially compensated by fortification/spiking with ascorbate before use.

Antioxidants of Edible Mushrooms.

Oxidative stress caused by an imbalanced metabolism and an excess of reactive oxygen species (ROS) lead to a range of health disorders in humans. Our endogenous antioxidant defense mechanisms and our dietary intake of antioxidants potentially regulate our oxidative homeostasis. Numerous synthetic antioxidants can effectively improve defense mechanisms, but because of their adverse toxic effects under certain conditions, preference is given to natural compounds. Consequently, the requirements for natural, alternative sources of antioxidant foods identified in edible mushrooms, as well as the mechanistic action involved in their antioxidant properties, have increased rapidly. Chemical composition and antioxidant potential of mushrooms have been intensively studied. Edible mushrooms might be used directly in enhancement of antioxidant defenses through dietary supplementation to reduce the level of oxidative stress. Wild or cultivated, they have been related to significant antioxidant properties due to their bioactive compounds, such as polyphenols, polysaccharides, vitamins, carotenoids and minerals. Antioxidant and health benefits, observed in edible mushrooms, seem an additional reason for their traditional use as a popular delicacy food. This review discusses the consumption of edible mushrooms as a powerful instrument in maintaining health, longevity and life quality.

Dropwort (Filipendula hexapetala Gilib.): potential role as antioxidant and antimicrobial agent.

The aim of this study was to investigate the antioxidant activity of the methanolic extracts of Filipendula hexapetala Gilib. aerial parts (FHA) and roots (FHR) and their potential in different model systems, as well as antimicrobial activity. According to this, a number of assays were employed to evaluate the antioxidant and antimicrobial potential of F. hexapetala extracts. In addition, the antioxidant activity assays in different model systems were carried out, as well as pH, thermal and gastrointestinal stability studies. The phenolic compounds contents in FHA and FHR were also determined. The results showed that F. hexapetala extracts had considerable antioxidant activity in vitro and a great stability in different conditions. The extracts exhibited antimicrobial activity against most of the tested bacterial and fungal species. Also, the extracts contain high level of phenolic compounds, especially aerial parts extract.

Nutraceutical properties of the methanolic extract of edible mushroom Cantharellus cibarius (Fries): primary mechanisms.

The methanolic extract of the wild edible mushroom Cantharellus cibarius Fr. (chanterelle) was analyzed for in vitro antioxidative, cytotoxic, antihypertensive and antibacterial activities. Various primary and secondary metabolites were found. Phenols were the major antioxidant components found in the extract (49.8 mg g(-1)), followed by flavonoids, whose content was approximately 86% of the total phenol content. Antioxidant activity, measured by four different methods, was high for inhibition of lipid peroxidation (EC50 = 1.21 mg mL(-1)) and chelating ability (EC50 = 0.64 mg mL(-1)). The antioxidant activity of the C. cibarius methanol extract was achieved through chelating iron compared to hydrogen atom and/or electron transfer. The extract showed good selectivity in cytotoxicity on human cervix adenocarcinoma HeLa, breast carcinoma MDA-MB-453 and human myelogenous leukemia K562, compared to normal control human fetal lung fibroblasts MRC-5 and human lung bronchial epithelial cells BEAS-2B. The extract had inhibitory activity against angiotensin converting I enzyme (ACE) (IC50 = 0.063 mg mL(-1)). The extract revealed selective antimicrobial activity against Gram-positive bacteria with the highest potential against E. faecalis. The medicinal and health benefits, observed in wild C. cibarius mushroom, seem an additional reason for its traditional use as a popular delicacy food.

Bioactivity, stability and phenolic characterization of Filipendula ulmaria (L.) Maxim.

The purpose of the present study was to evaluate the antioxidant and antimicrobial potential of Filipendula ulmaria (L.) Maxim. methanolic extracts, their stabilities under different pH and thermal conditions and in vitro digestibility. The results showed a considerable content of phenolic compounds in the extracts, especially total phenolic acids (47.47 mg CAE g(-1)) and flavonoids (45.47 mg RUE g(-1)) in aerial parts. HPLC analysis indicated the presence of spiraeoside in the aerial part extract. The extracts revealed an interesting antimicrobial effect against the tested microorganisms, especially bacteria E. coli and E. faecalis (MIC 0.156-0.625 mg mL(-1)), and fungi P. cyclopium and F. oxysporum (MIC 2.5-5 mg mL(-1)). The extracts exerted high antioxidant activities, particularly the root extract, paralleled by their considerable activities against the lipid oxidation process. The results of this study suggest that both extracts potentially could be functional food ingredients considering their good antioxidant and antimicrobial activities, and stability under different conditions.

Dietary polysaccharide extracts of Agaricus brasiliensis fruiting bodies: chemical characterization and bioactivities at different levels of purification.

Polysaccharides of the European strain of A. brasiliensis were obtained by hot water extraction and ethanol precipitation (HWPE I) of fruiting bodies, and further purified by dialysis (HWPE II) and pronase incubation (PPE). These polysaccharides consisted mainly of (1→6)-ß-d-glucans. PPE was free of proteins and polyphenols as demonstrated by quantitative assays and NMR profiling. They showed a clear IFN-γ inducing activity in human PBMCs, which suggests these polysaccharides to have proinflammatory effects. Treatment by ß-glucosidase caused the polysaccharides to be degraded into smaller fragments and at the same time increased their IFN-γ inducing activity in PBMCs fourfold. In vitro, PPE showed a dose-dependent inhibition of the proliferation of the human leukemia Jurkat cell. At 100µg/mL the cells' viability was decreased by appr. 51% compared to the control. EPR spin trapping demonstrated a high antioxidative activity against •OH and •O2- radicals of HWPE I and PPE. Further, the results of the antioxidant assays indicated that antioxidant activity against •OH radicals in the Fenton system was achieved through scavenging or through chelating iron mechanisms. The good immunomodulating and antioxidative properties of A. brasiliensis polysaccharide extract obtained by hot water extraction and ethanol precipitation make it suitable for everyday use as an inexpensive dietary supplement.

Perfluorinated compounds in sediment samples from the wastewater canal of Pancevo (Serbia) industrial area.

Perfluoroalkyl sulfonates (PFSAs) and perfluoroalkyl carboxylates (PFCAs) were analyzed in sediment samples from the wastewater canal draining the industrial complex of Pancevo, Serbia (oil refinery, petrochemical plant, and fertilizer factory). The canal is directly connected to Europe's second largest river, the Danube, which drains its water into the Black Sea. Perfluorooctane sulfonate (PFOS) up to 5.7ngg(-1) dry weight (dw) and total Perfluorinated compounds (PFCs) up to 6.3ngg(-1) dw were detected. Compared to other reports, high levels of PFOS were found, even though PFCs are not used in the industrial production associated with this canal. The PFOS concentration in water was recalculated using the adsorption coefficient, KOC from literature. Using the average output of wastewater from the canal, a mass load of 1.38kg PFOS per year discharged in the Danube River has been calculated, which undoubtedly points to the contribution to global persistent organic pollution of surface waters originating from this industrial place.