Mineral Content in Honeybee Wax Combs as a Measurement of the Impact of Environmental Factors.

Environmental pollution from metals needs to be constantly monitored due to their predominantly negative impacts on living organisms. As apian products stored in hives are considered useful bioindicators, the objective of this study was to: (a) investigate and compare the essential and toxic metal concentrations in freshly constructed combs (light combs, LC) and old combs (dark combs, DC) in use for two to three beekeeping seasons, and (b) compare the mineral content of beeswax combs from apiaries exposed to different levels of environmental pollution using the energy dispersive x-ray fluorescence method. Concentrations of ten elements (Cr, Pb, Cu, Ni, Fe, Zn, Mn, Sr, Rb, Ca) were determined in 18 honeybee wax comb samples from three apiaries in continental Croatia. The results showed that the influence of comb age and/or geographical origin (representing varying levels of environmental pollution exposure) on the elemental composition of beeswax was evident for the toxic elements Cr, Pb, Cu and Ni, and for the essential elements Fe, Zn, Mn and Sr, but not Rb. In addition to monitoring the environmental element content, wax combs can be used to determine contamination levels. Additionally, in-time analysis results can enable beekeepers to adjust management practices, such as moving apiaries to better positions. They can also be useful in the creation of policies on acceptable limits for toxic metal levels in particular geographical areas.

Seasonal variations as predictive factors of the comet assay parameters: a retrospective study.

Since there are several predicting factors associated with the comet assay parameters, we have decided to assess the impact of seasonal variations on the comet assay results. A total of 162 volunteers were retrospectively studied, based on the date when blood donations were made. The groups (winter, spring, summer and autumn) were matched in terms of age, gender, smoking status, body mass index and medical diagnostic exposure in order to minimise the impact of other possible predictors. Means and medians of the comet assay parameters were higher when blood was sampled in the warmer period of the year, the values of parameters being the highest during summer. Correlation of meteorological data (air temperature, sun radiation and sun insolation) was observed when data were presented as the median per person. Using multivariate analysis, sampling season and exposure to medical radiation were proved to be the most influential predictors for the comet assay parameters. Taken together, seasonal variation is another variable that needs to be accounted for when conducting a cohort study. Further studies are needed in order to improve the statistical power of the results related to the impact of sun radiation, air temperature and sun insolation on the comet assay parameters.

Cytokinesis-block micronucleus cytome assay parameters in peripheral blood lymphocytes of the general population: Contribution of age, sex, seasonal variations and lifestyle factors.

The cytokinesis-block micronucleus cytome (CBMN Cyt) assay was used to evaluate the baseline frequency of cytogenetic damage in peripheral blood lymphocytes of the general population (average age, 38.28 ± 12.83 years) in relation to age, sex, body mass index, seasonal variations (season of sampling, period of sampling and different meteorological parameters) and lifestyle factors (smoking habit, alcohol consumption, exposure to medications and diagnostic radiation, physical activity, and family history of cancer). The background frequency of micronuclei (MNi) for the 200 subjects assayed was 5.06 ± 3.11 per 1000 binucleated cells, while the mean frequency of nucleoplasmic bridges (NPBs) was 1.21 ± 1.46 and of nuclear buds (NBUDs) 3.48 ± 2.14. The background frequency of apoptosis and necrosis was 1.58 ± 1.50 and 1.39 ± 1.56, respectively, while the mean nuclear division index (NDI) was 1.99 ± 0.14. The cut-off value, which corresponds to the 95th percentile of the distribution of 200 individual values, was 11 MNi, 4 NPBs and 7 NBUDs. The study also confirmed an association of the above mentioned parameters with age, sex and several lifestyle factors. Moreover, significant confounders based on our results are also sampling season, sampling period and different meteorological parameters that were dependent on the CBMN Cyt assay parameters. In line with the above mentioned, several factors should be taken into account when it comes to the monitoring of exposed populations using cytogenetic biomarkers. Moreover, the normal and cut-off values obtained in this study present background data for the general population, and can later serve as baseline values for further biomonitoring studies.

Detoxification of polluted marine sediments using water treatment sludge.

The purpose of this study was development of the optimal conditions for the inertization of the polluted marine sediments using groundwater treatment sludge highly enriched in iron and aluminum. For that purpose fine-grained sediment (>85% clay and silt fraction) highly enriched in copper and zinc was amended with the waste sludge (from 10% to 50%). The sample with the optimum percentage of the waste sludge was further subjected the thermal treatment at 200-800°C. The efficiency of the treatment was determined by the leaching tests and toxicity testing using Hordeum vulgare L. as biosystem. The percentage of the seed germination, the root elongation, and the germination index before and following the treatment were determined. Untreated sediment leachate caused significant arrest in the percentage of the seed germination (6.7 ± 6.7%), the root elongation (1.9 ± 2.2%), and the germination index (0.2 ± 0.3%) with EC50 of 24.9%, suggesting a significant toxic potential of the sediments. After amendment with 30% of the waste sludge the concentrations of the considered micropollutants decreased below regulated values while all three toxicological parameters showed no significant difference compared to the negative control. The toxicity of the sediment was completely removed following the thermal treatment at 400°C.

Toxicity assessment of untreated/treated electroplating sludge using human and plant bioassay.

The purpose of this work was to assess the risk to the environment arising from the electroplating sludge from both chemical and toxicological point of view. Both approaches were used for the assessment of the treatment efficiency which consisted of CaO based solidification followed by thermal treatment at 400°C. The elemental composition was determined in the bulk samples and the leachates of untreated sludge. The toxicity of the leachate was determined using two human colorectal adenocarcinoma cell lines (Caco-2 and SW 480) and Hordeum vulgare L. based plant bioassay. The same toxicity tests were employed to the leachate of the treated sludge. Untreated sludge showed extremely high cytotoxic effect to both human and plant bio-system in dose-dependent manner. The percentages higher than 0.5% and 0.05% of the leachate caused significant cytotoxic effect on Caco-2 and SW 480 cells, respectively. The percentages of the leachate higher than 0.05% also showed significant toxic effect to H. vulgare L. bio-system with complete arrest of seed germination following the treatment with 100% to 5% of the leachate. The leachate of the treated sludge showed no toxicity to any of the test systems confirming the efficiency and justification of the employed procedures for the detoxification of electroplating sludge.

Physico-chemical characterization and the in vitro genotoxicity of medical implants metal alloy (TiAlV and CoCrMo) and polyethylene particles in human lymphocytes.

BACKGROUND: The main objective of the present study was to investigate chemical composition and possible cyto/genotoxic potential of several medical implant materials commonly used in total hip joint replacement. METHODS: Medical implant metal alloy (Ti6Al4V and CoCrMo) and high density polyethylene particles were analyzed by energy dispersive X-ray spectrometry while toxicological characterization was done on human lymphocytes using multi-biomarker approach. RESULTS: Energy dispersive X-ray spectrometry showed that none of the elements identified deviate from the chemical composition defined by appropriate ISO standard. Toxicological characterization showed that the tested materials were non-cyto/genotoxic as determined by the comet and cytokinesis-block micronucleus (CBMN) assay. Particle morphology was found (by using scanning electron and optical microscope) as flat, sharp-edged, irregularly shaped fiber-like grains with the mean particle size less than 10µm; this corresponds to the so-called "submicron wear". The very large surface area per wear volume enables high reactivity with surrounding media and cellular elements. CONCLUSIONS: Although orthopedic implants proved to be non-cyto/genotoxic, in tested concentration (10µg/ml) there is a constant need for monitoring of patients that have implanted artificial hips or other joints, to minimize the risks of any unwanted health effects. GENERAL SIGNIFICANCE: The fractal and multifractal analyses, performed in order to evaluate the degree of particle shape effect, showed that the fractal and multifractal terms are related to the "remnant" level of the particles' toxicity especially with the cell viability (trypan blue method) and total number of nucleoplasmic bridges and nuclear buds as CBMN assay parameters.

Toxicological assessment and management options for boat pressure-washing wastewater.

Boats are washed periodically for maintenance in order to remove biofoulants from hulls, which results in the generation of wastewater. This study aimed at evaluating the cyto/genotoxic and mutagenic properties of wastewater produced by pressure washing of boats. The chemical characterisation of this wastewater showed that Cu, Zn, V, Cr, Fe, Pb, and select organic contaminants exceeded the maximum allowable values from 1.7 up to 96 times. The wastewater produced negative effects on human lymphocytes resulting in decreased cell viability after 4 and 24h of exposure. Chromosome aberration, micronucleus, and comet assay parameters were significantly higher after 24h of exposure. At the same time, the Salmonella typhimurium test showed negative for both TA98 and TA100 strains at all of the concentrations tested. After the treatment of wastewater using electrochemical methods/ozonation during real scale treatment plant, removal rates of colour, turbidity and heavy metals ranged from 99.4% to 99.9%, while the removal of total organic carbon (TOC) and chemical oxygen demand (COD) was above 85%. This was reflected in the removal of the wastewater's cyto/genotoxicity, which was comparable to negative controls in all of the conducted tests, suggesting that such plants could be implemented in marinas to minimise human impact on marine systems.

Neutralization/purification of the wastewaters from printed circuit boards production using waste by-products.

The purpose of this work was development of a new method for the neutralization/purification of printed circuit boards wastewater (PCBW) originating from Zagreb, Croatia, using two industrial by-products. PCBW was characterized with low pH value (2.11) and high concentration of TDS (50190 mg L(-1)), copper (4190 mg L(-1)) and iron (2660 mg L(-1)). Waste base (WB), by-product of the alumina production, and waste sludge, by-product of the electrochemical treatment of groundwater, were employed as neutralization/adsorption agents. Due to its high neutralization capacity WB was used for pH adjustment to pH 8 and heavy metals removal from both effluents, yet the final removal of the contaminants down to the regulated values was assessed by adsorption/coagulation with the iron and aluminum rich waste sludge. Following the combined treatment the removal efficiency of iron and copper was higher than 99.99% with their final concentration in the treated water of 0.151 mg L(-1) and 0.129 mg L(-1), respectively. Following the ozone base oxidation the removal efficiency of the organic contaminants was more than 83%. The successful application of the industrial waste by-products for neutralization/purification of the PCBW with the removal efficiencies of the contaminants comparable or better than those obtained with conventional treatment represented the main advantage of our presented method.

Removal of arsenic, phosphates and ammonia from well water using electrochemical/chemical methods and advanced oxidation: a pilot plant approach.

The purpose of this work was to develop a pilot plant purification system and apply it to groundwater used for human consumption, containing high concentrations of arsenic and increased levels of phosphates, ammonia, mercury and color. The groundwater used was obtained from the production well in the Vinkovci County (Eastern Croatia). Due to a complex composition of the treated water, the purification system involved a combined electrochemical treatment, using iron and aluminum electrode plates with simultaneous ozonation, followed by a post-treatment with UV, ozone and hydrogen peroxide. The removal of the contaminant with the waste sludge collected during the electrochemical treatment was also tested. The combined electrochemical and advanced oxidation treatment resulted in the complete removal of arsenic, phosphates, color, turbidity, suspended solids and ammonia, while the removal of other contaminants of interest was up to 96.7%. Comparable removal efficiencies were obtained by using waste sludge as a coagulant.

Cytogenetic status of healthy children assessed with the alkaline comet assay and the cytokinesis-block micronucleus cytome assay.

In the present study the alkaline comet assay and the cytokinesis-block micronucleus cytome (CBMN Cyt) assay were used to evaluate the baseline frequency of cytogenetic damage in peripheral blood lymphocytes (PBLs) of 50 healthy children from the general population in Croatia (age, 11.62±1.81 years). Mean values of tail length, tail intensity and tail moment, as comet assay parameters, were 12.92±0.10, 0.73±0.06 and 0.08±0.01, respectively. The mean frequency of micronuclei (MN) for all subjects was 2.32±0.28 per 1000 bi-nucleated cells, while the mean frequency of nucleoplasmic bridges (NPBs) was 1.72±0.24 and of nuclear buds (NBUDs) 1.44±0.19. The mean nuclear division index (NDI) was 1.70±0.05. When comet-assay parameters were considered, higher mean values for all three were found for the female population. According to the Mann-Whitney U test applied on the results of the comet assay, the only statistically significant difference between the male and female populations was found for tail length. Similar to the results obtained by the comet assay, girls showed higher mean values of all three measured parameters of the CBMN Cyt assay. This difference was statistically significant for total number of NPBs only. In the case of the NDI, a higher mean value was also obtained in girls, but this difference was not statistically significant. The results obtained present background data that could be considered as normal values for healthy children living in urban areas, and can later on serve as baseline values for further toxicological monitoring. Additionally, the usefulness of both techniques in measuring cytogenetic damage during bio-monitoring of children is confirmed.

Assessment of surface water in the vicinity of fertilizer factory using fish and plants.

The genotoxic and toxic potential of polluted surface water exposed to a fertilizer factory effluent was evaluated using assays with fish (Cyprinus carpio) and plant (Lemna minor) model organisms. Beside classical physicochemical parameters, the contents of fluorides, some heavy metals and polycyclic aromatic hydrocarbons were analyzed as well. Surface water caused inhibition of plant growth and decrease of photosynthetic pigment content. Regarding DNA damage and oxidative stress parameters, both fish and plants showed similar response to the surface water. In confirmation to biochemical markers, histopathological analysis of gill and liver tissues revealed a higher incidence of lesions in fish exposed to polluted surface water. Generally, results obtained by biological monitoring were mostly in agreement with chemical analysis of the surface water, although several discrepancies were observed which might be due to difference in sensitivity of model organisms or in experimental conditions (laboratory and field exposure). The results imply that conventional chemical analysis should be extended to genotoxicity/toxicity assays as measured biological effects and the potential health hazard cannot be predicted based on the physicochemical characteristics of water samples alone.

Preparation of drinking water used in water supply systems of the towns Zrenjanin and Temerin by electrochemical methods.

The aim of this work was the development and application of the pilot plant with the capacity of 1000 L/day for the purification of groundwater used for human consumption characterized with high concentration of arsenic and increased values of organic pollutants, ammonia, nitrites, color and turbidity. For that purpose, groundwater from the production wells supplying the towns Zrenjanin and Temerin (Vojvodina, Serbia) was used. Due to its complex composition, the purification system required the combination of the electroreduction/electrocoagulation, using iron and aluminum electrode plates with/without ozonation, followed by the electromagnetic treatment and the finally by the simultaneous ozonation/UV treatment. The electroreduction was used for the removal of nitrates, nitrites, and Cr(VI), while the removal of arsenic, heavy metals, suspended solids, color and turbidity required the application of the electrocoagulation with simultaneous ozonation. Organic contaminants and ammonia were removed completely in the last treatment step by applying the simultaneous ozonation/UV treatment. All measured parameters in the purified water were significantly lower compared to the regulated values. Under the optimum treatment conditions, the removal efficiencies for color, turbidity, suspended solids, total arsenic, total chromium, Ni(II), total copper, sulfates, fluorides, chemical oxygen demand, ammonia, nitrates, and nitrites were 100%. The removal efficiencies of the total manganese and iron were 85.19% and 97.44%, respectively, whilst the final concentrations were 4 and 7 µg/L, respectively.

Electroplating wastewater treatment by the combined electrochemical and ozonation methods.

This article presents a pilot-plant study of the electroplating wastewater treatment by the processes of electroreduction with iron electrode plates, and electrocoagulation/ozonation with aluminum electrode set, followed by the process of ozonation. The initial effluent was found to be highly enriched in heavy metals and to possess the elevated levels of organic contaminants. The values of Cr(VI), Fe, Ni, Cu, Zn, Pb, TOC, and COD exceeded the upper permissible limits of 63, 220.2, 1.1, 7, 131.3, 1.7, 12.3, and 11.4 times, respectively. The heavy metal removal was forced either by the coagulation/flocculation using Fe(II), Fe(III), and Al(III) ions released into the treated solution by the electrochemical corrosion of the sacrificial iron and aluminum electrodes, or the precipitation of the metal hydroxides as well as co-precipitation with iron and aluminum hydroxides. The principle organic matter destruction mechanisms were ozone oxidation and the indirect oxidation with chlorine/hypochlorite formed by the anodic oxidation of chloride already present in the wastewater. Following the combined treatment, the removal efficiencies of Cr(VI), Fe, Ni, Cu, Zn, Pb, TOC, and COD were 99.94%, 100.00%, 95.86%, 98.66%, 99.97%, 96.81%, 93.24%, and 93.43%, respectively, thus complying with the regulated values.

Treatment of winery wastewater by electrochemical methods and advanced oxidation processes.

The aim of this research was development of new system for the treatment of highly polluted wastewater (COD = 10240 mg/L; SS = 2860 mg/L) originating from vine-making industry. The system consisted of the main treatment that included electrochemical methods (electro oxidation, electrocoagulation using stainless steel, iron and aluminum electrode sets) with simultaneous sonication and recirculation in strong electromagnetic field. Ozonation combined with UV irradiation in the presence of added hydrogen peroxide was applied for the post-treatment of the effluent. Following the combined treatment, the final removal efficiencies of the parameters color, turbidity, suspended solids and phosphates were over 99%, Fe, Cu and ammonia approximately 98%, while the removal of COD and sulfates was 77% and 62%, respectively. A new approach combining electrochemical methods with ultrasound in the strong electromagnetic field resulted in significantly better removal efficiencies for majority of the measured parameters compared to the biological methods, advanced oxidation processes or electrocoagulation. Reduction of the treatment time represents another advantage of this new approach.

Relationships between radionuclides, metals, and sediment properties in sediment of a bay exposed to anthropogenic pressure and mixed sediment sources (Kastela Bay, Adriatic Sea, Croatia).

Natural and anthropogenic radionuclides, metals, organic matter, sediment grain size, mineral composition, and sediment sources were studied in marine sediment of Kastela Bay up to a depth of 0.5 m. Deposition of man-modified material into the sea was evidenced in sediment mineral composition. Presence of pyrite and hematite in this sediment may pose an environmental concern. Metals, radionuclides, and organic matter were grouped in three groups: (i) variables under no anthropogenic influence and preferentially associated with carbonates (Ca, Sr); (ii) variables under no or weak anthropogenic influence and preferentially associated with aluminosilicates (Al, K, Ti, V, Cr, Mn, Fe, Co, Ni, Ga, Rb, Y, 40K, 232Th); (iii) variables under notable anthropogenic influence and/or natural processes of separation (Cu, Zn, Pb, As, 226Ra, 238U, 137Cs, organic matter). Predominant influencing parameters change with sediment depth for some variables. Anthropogenic influence was the most emphasised for Cu, Zn, and Pb, followed by 137Cs.

Chemical composition and genotoxicity assessment of sanitary landfill leachate from Rovinj, Croatia.

Chemical analysis and an in vitro approach were performed to assess elemental composition and genotoxic effects of the samples of landfill leachate taken from Lokva Vidotto sanitary landfill the official landfill for Rovinj town, Croatia. Two samples of landfill leachate were collected and analyzed in order to evaluate macro, micro and trace elements by atomic absorption spectroscopy, energy dispersive X-ray spectrometry and colorimetry. Genotoxicity of sanitary landfill leachate was evaluated in human lymphocytes by the use of the micronucleus test and comet assay. Samples were characterized with relatively low concentrations of heavy metals while organic component level exceeded upper permissible limit up to 39 times. Observed genotoxic effects should be connected with high concentrations of ammonia nitrogen, which exceeded permissible limit up to 180 times. Leachate samples of both sanitary landfills increased the frequency of micronuclei, nucleoplasmic bridges and nuclear buds. Increase of DNA damage in human lymphocytes was also detected by virtue of measuring comet assay parameters. All parameters showed statistically significant difference compared to negative control. Increased micronucleus and comet assay parameters indicate that both samples of sanitary landfill leachate are genotoxic and could pose environmental and human health risk if discharged to an aquatic environment.

The correlation of the lifestyle and medical conditions with the vaginal infections and production of 2-phenylethanol.

OBJECTIVE: The objective of this study was determination of causative factors of the genital infections and their correlation with various predictor variables. Secondary objectives included: (1) determination of the presence and the type of low molecular weight metabolites in the samples of vaginal secretion formed in vivo, (2) determination of the concentration of 2-phenylethanol formed in vitro for each Candida species, (3) determination of the relationship between fungal/bacterial/viral infections with the metabolites formed in vivo using multivariate analysis. METHODS: One hundred and ninety-seven women in the age range from 18 to 65 years were included in the study. After the completion of questionnaire, all the patients were subjected to Pap test, cervical swabs for the presence of aerobic bacteria, yeasts, Ureaplasma urealyticum, Chlamydia trachomatis, Mycoplasma, and hrHPV DNA. The presence and the concentration of low-molecular weight metabolites in vitro and in vivo were determined by gas chromatography-mass spectrometry (GC-MS) method. Multivariate analysis methods were used for statistical evaluation. RESULTS: The most important risk factors of fungal/bacterial/viral infections were determined. The presence of 2-phenylethanol in vivo was confirmed in 14 of 74 tested samples and connected with the Candida species. The presence of symptoms, hrHPV DNA and Ureaplasma urealyticum are the predictor variables with the highest influence on the formation of the metabolite in vivo. The results in vitro confirmed that various Candida species produced 2-phenylethanol with the concentrations ranging from 0.6 to 4.64 µg/mL. CONCLUSION: The medical exposure to irradiation, marital status, and number of partners as well as stress factors (miscarriages, chronic, viral, or tumor illnesses) had the highest influence on the development of the bacterial/fungal/viral infections. The formation of 2-phenylethanol, both in vivo and in vitro, was confirmed and connected with Candida species. Besides, according to statistical tests, it seems that presence of symptoms, hrHPV DNA, and Ureaplasma urealyticum had also significant role on the formation of 2-phenylethanol in vivo.

A combined CaO/electrochemical treatment of the acid mine drainage from the "Robule" Lake.

The purpose of this work was development and application of the purification system suitable for the treatment of the acid mine drainage (AMD) accumulated in the "Robule" Lake, which represents the part of the Bor copper mining and smelting complex, Serbia. The study was undertaken in order to minimize adverse effect on the environment caused by the discharge of untreated AMD, which was characterized with low pH value (2.63) and high concentration of heavy metals (up to 610 mg/L) and sulfates (up to 12,000 mg/L). The treatment of the effluent included pretreatment/pH adjustment with CaO followed by electrocoagulation using iron and aluminum electrode sets. Following the final treatment, the decrease in the concentration of heavy metals ranged from 40 up to 61000 times depending on the metal and its initial concentration. The parameters, color and turbidity were removed completely in the pretreatment step, while the removal efficiencies for other considered parameters were as follows: EC = 55.48%, SO(4) (2-) = 70.83%, Hg = 98.36%, Pb = 97.50%, V = 98.43%, Cr = 99.86%, Mn = 97.96%, Fe = 100.00%, Co = 99.96%, Ni = 99.78%, Cu = 99.99% and Zn = 99.94%. Because the concentrations of heavy metals in the electrochemically treated AMD (ranging from 0.001 to 0.336 mg/L) are very low, the negative impact of this effluent on the aquatic life and humans is not expected. The sludge generated during the treatment of AMD is suitable for reuse for at least two purposes (pretreatment of AMD and covering of the flotation waste heap). From the presented results, it could be concluded that electrochemical treatment is a suitable approach for the treatment of AMD.

Characterization and electrochemical treatment of landfill leachate.

A combined treatment approach using advanced oxidation, electrochemical methods and microwaves was developed and applied for the treatment of landfill leachate taken from Piskornica (Koprivnica, Croatia) sanitary landfill. Due to the complex nature of the effluent and extremely low bio-degradability (BOD(5)/COD ratio = 0.01) the purification of the leachate started with pre-treatment with ozone followed by simultaneous ozonation and electrocoagulation/electrooxidation using the set of iron and aluminum electrodes, and finally, the degradation of organic residue and ammonia with microwaves. Applied treatment approach resulted in clear, colorless and odorless effluent with the values of all measured parameters significantly lower compared to the upper permissible limit for discharge into the environment. The removal percentages of the parameters: color, turbidity, suspended solids, ammonia, COD and iron following the combined treatment were 98.43%, 99.48%, 98.96%, 98.80%, 94.17% and 98.56%, respectively.

Chemical and toxicological characterization of the bricks produced from clay/sewage sludge mixture.

The present study aimed to characterize chemical properties of clay bricks containing 20 % of sewage sludge. After detection of potentially hazardous metals, we simulated precipitation exposure of such material to determine the amount of heavy metals that could leach out of the bricks. Metals, such as copper, zinc, nickel, cobalt, chromium, etc., were detected in leachate in low concentrations. Moreover, human peripheral blood lymphocytes were exposed to brick leachate for 24 h in order to evaluate its possible negative impact on human cells and genome in vitro. Cytotoxicity tests showed no effect on human peripheral blood lymphocytes viability after exposure to brick's leachate. On the contrary, the alkaline comet assay showed slight but significant increase in DNA damage with all three parameters tested. As we might predict, interactions of several heavy metals in low concentrations could be responsible for DNA damaging effect. In that manner, our findings suggest that leachates from sewage sludge-produced bricks may lead to adverse effects on the exposed human population, and that more stabile bricks should be developed to minimize leaching of heavy metals into the environment. Bricks with lower percentage of the sludge may be one of the solutions to reduce the toxic effect of the final product.