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Grassland bird populations have decreased significantly due to environmental contaminants, evaluating their health status through biological indicators is a way to prevent population decline. The aim of the contribution was to examine the use of hematic biomarkers (Polychromatic erythrocytes; PCE, polychromatic erythrocytes with micronucleus; PCEMN, polychromatic erythrocytes with nuclear extensions; PCEPN, erythrocytes micronucleated; MNE, erythrocytes with nuclear extensions; PNE, leukocyte count and ratio H /L) and the possible exposure to xenobiotic substances of the grasshopper sparrow (Ammodramus savannarum) and Baird's sparrow (Centronyx bairdii). During the winter season of 2020 in Cuchillas de la Zarca, Durango, 57 specimens were collected (34 A. savannarum and 23 C. bairdii), the basal frequencies of the hematic biomarkers were obtained, whose values did not constitute evidence that the birds had been exposed to polluting substances such as agrochemicals, metals and metalloids. The basal frequencies of the blood biomarkers obtained in this study were similar to those reported in other apparently healthy bird species. The observed frequency of PCE and MNE suggests the possibility of using these bird species as biomonitor candidates when exposure to cytotoxic or genotoxic compounds is suspected., Las poblaciones de aves de pastizal han disminuido significativamente debido a los contaminantes ambientales, evaluar su estado de salud mediante indicadores biológicos es una forma de prevenir el decremento poblacional. El objetivo de la contribución fue examinar el uso de biomarcadores hemáticos (Eritrocitos policromáticos; EPC, eritocitos policromáticos con micronúcleos; EPCMN, erittrocitos policromáticos con prolongaciones nucleares; EPCPN, eritrocitos micronucleados; EMN, eritrocitos con prolongaciones nucleares; EPN, recuento leucocitario y relación H/L) y la posible exposición a sustancias xenobioticas del gorrión chapulinero (Ammodramus savannarum) y el gorrión de Baird (Centronyx bairdii). Durante la temporada invernal de 2020 en Cuchillas de la Zarca, Durango se colectaron 57 ejemplares (34 A. savannarum y 23 C. bairdii), se obtuvieron las frecuencias basales de los biomarcadores hemáticos cuyos valores no constituyeron evidencia de que las aves se hayan expuesto a sustancias contaminantes como agroquímicos, metales y metaloides. Las frecuencias basales de los biomarcadores hemáticos obtenidos en este estudio fueron similares a las reportadas en otras especies de aves aparentemente sanas. La frecuencia observada de EPC y EMN sugiere la posibilidad de usar a estas especies de aves como candidatas a biomonitoras cuando se sospecha de exposición a compuestos citotóxicos o genotóxicos.

Grassland bird populations have decreased significantly due to environmental contaminants, evaluating their health status through biological indicators is a way to prevent population decline. The aim of the contribution was to examine the use of hematic biomarkers (Polychromatic erythrocytes; PCE, polychromatic erythrocytes with micronucleus; PCEMN, polychromatic erythrocytes with nuclear extensions; PCEPN, erythrocytes micronucleated; MNE, erythrocytes with nuclear extensions; PNE, leukocyte count and ratio H /L) and the possible exposure to xenobiotic substances of the grasshopper sparrow (Ammodramus savannarum) and Baird's sparrow (Centronyx bairdii). During the winter season of 2020 in Cuchillas de la Zarca, Durango, 57 specimens were collected (34 A. savannarum and 23 C. bairdii), the basal frequencies of the hematic biomarkers were obtained, whose values did not constitute evidence that the birds had been exposed to polluting substances such as agrochemicals, metals and metalloids. The basal frequencies of the blood biomarkers obtained in this study were similar to those reported in other apparently healthy bird species. The observed frequency of PCE and MNE suggests the possibility of using these bird species as biomonitor candidates when exposure to cytotoxic or genotoxic compounds is suspected., Las poblaciones de aves de pastizal han disminuido significativamente debido a los contaminantes ambientales, evaluar su estado de salud mediante indicadores biológicos es una forma de prevenir el decremento poblacional. El objetivo de la contribución fue examinar el uso de biomarcadores hemáticos (Eritrocitos policromáticos; EPC, eritocitos policromáticos con micronúcleos; EPCMN, erittrocitos policromáticos con prolongaciones nucleares; EPCPN, eritrocitos micronucleados; EMN, eritrocitos con prolongaciones nucleares; EPN, recuento leucocitario y relación H/L) y la posible exposición a sustancias xenobioticas del gorrión chapulinero (Ammodramus savannarum) y el gorrión de Baird (Centronyx bairdii). Durante la temporada invernal de 2020 en Cuchillas de la Zarca, Durango se colectaron 57 ejemplares (34 A. savannarum y 23 C. bairdii), se obtuvieron las frecuencias basales de los biomarcadores hemáticos cuyos valores no constituyeron evidencia de que las aves se hayan expuesto a sustancias contaminantes como agroquímicos, metales y metaloides. Las frecuencias basales de los biomarcadores hemáticos obtenidos en este estudio fueron similares a las reportadas en otras especies de aves aparentemente sanas. La frecuencia observada de EPC y EMN sugiere la posibilidad de usar a estas especies de aves como candidatas a biomonitoras cuando se sospecha de exposición a compuestos citotóxicos o genotóxicos.

The EFSA Scientific Committee was asked to provide guidance on the most appropriate in vivo tests to follow up on positive in vitro results for aneugenicity, and on the approach to risk assessment for substances that are aneugenic but not clastogenic nor causing gene mutations. The Scientific Committee confirmed that the preferred approach is to perform an in vivo mammalian erythrocyte micronucleus test with a relevant route of administration. If this is positive, it demonstrates that the substance is aneugenic in vivo. A negative result with evidence that the bone marrow is exposed to the test substance supports a conclusion that aneugenic activity is not expressed in vivo. If there is no evidence of exposure to the bone marrow, a negative result is viewed as inconclusive and further studies are required. The liver micronucleus assay, even though not yet fully validated, can provide supporting information for substances that are aneugenic following metabolic activation. The gastrointestinal micronucleus test, conversely, to be further developed, may help to assess aneugenic potential at the initial site of contact for substances that are aneugenic in vitro without metabolic activation. Based on the evidence in relation to mechanisms of aneugenicity, the Scientific Committee concluded that, in principle, health-based guidance values can be established for substances that are aneugenic but not clastogenic nor causing gene mutations, provided that a comprehensive toxicological database is available. For situations in which the toxicological database is not sufficient to establish health-based guidance values, some approaches to risk assessment are proposed. The Scientific Committee recommends further development of the gastrointestinal micronucleus test, and research to improve the understanding of aneugenicity to support risk assessment.

The EFSA Scientific Committee was asked to provide guidance on the most appropriate in vivo tests to follow up on positive in vitro results for aneugenicity, and on the approach to risk assessment for substances that are aneugenic but not clastogenic nor causing gene mutations. The Scientific Committee confirmed that the preferred approach is to perform an in vivo mammalian erythrocyte micronucleus test with a relevant route of administration. If this is positive, it demonstrates that the substance is aneugenic in vivo. A negative result with evidence that the bone marrow is exposed to the test substance supports a conclusion that aneugenic activity is not expressed in vivo. If there is no evidence of exposure to the bone marrow, a negative result is viewed as inconclusive and further studies are required. The liver micronucleus assay, even though not yet fully validated, can provide supporting information for substances that are aneugenic following metabolic activation. The gastrointestinal micronucleus test, conversely, to be further developed, may help to assess aneugenic potential at the initial site of contact for substances that are aneugenic in vitro without metabolic activation. Based on the evidence in relation to mechanisms of aneugenicity, the Scientific Committee concluded that, in principle, health-based guidance values can be established for substances that are aneugenic but not clastogenic nor causing gene mutations, provided that a comprehensive toxicological database is available. For situations in which the toxicological database is not sufficient to establish health-based guidance values, some approaches to risk assessment are proposed. The Scientific Committee recommends further development of the gastrointestinal micronucleus test, and research to improve the understanding of aneugenicity to support risk assessment.

The EFSA Scientific Committee was asked to provide guidance on the most appropriate in vivo tests to follow up on positive in vitro results for aneugenicity, and on the approach to risk assessment for substances that are aneugenic but not clastogenic nor causing gene mutations. The Scientific Committee confirmed that the preferred approach is to perform an in vivo mammalian erythrocyte micronucleus test with a relevant route of administration. If this is positive, it demonstrates that the substance is aneugenic in vivo. A negative result with evidence that the bone marrow is exposed to the test substance supports a conclusion that aneugenic activity is not expressed in vivo. If there is no evidence of exposure to the bone marrow, a negative result is viewed as inconclusive and further studies are required. The liver micronucleus assay, even though not yet fully validated, can provide supporting information for substances that are aneugenic following metabolic activation. The gastrointestinal micronucleus test, conversely, to be further developed, may help to assess aneugenic potential at the initial site of contact for substances that are aneugenic in vitro without metabolic activation. Based on the evidence in relation to mechanisms of aneugenicity, the Scientific Committee concluded that, in principle, health-based guidance values can be established for substances that are aneugenic but not clastogenic nor causing gene mutations, provided that a comprehensive toxicological database is available. For situations in which the toxicological database is not sufficient to establish health-based guidance values, some approaches to risk assessment are proposed. The Scientific Committee recommends further development of the gastrointestinal micronucleus test, and research to improve the understanding of aneugenicity to support risk assessment.

Playa Penino is a natural reserve located on the south coast of Uruguay. It hosts 244 species of birds (more than 50% of the bird biodiversity recorded for Uruguay). The area is included in the International BirdLife Program for biodiversity preservation. Urbanization and pollution generally have affected the quality of the water and biodiversity in wetlands. The analysis of the major wetland of Playa Penino revealed high levels of organic compounds. The biomonitoring was done using the caviomorph rodent Ctenomys pearsoni (commonly known as tuco tuco) as an indicator species because they inhabit burrows around the wetland. Genetic effects were determined using the comet assay and micronucleus test. The significant correlations between chemical and microbiological parameters and genetic damage might indicate that macronutrients from sewage could be one of the causes of the genetic damage. There is an urgent need to conserve the biodiversity of this natural area by introducing sewage treatment, cesspool control and by controlling settlement in the area etc.

Potential health effects of extremely low-frequency (electro)magnetic fields (ELF-(E)MFs) have long been investigated, but the results are still inconclusive. With respect to genotoxicity, sound data related to the effects of long-term exposure to ELF-(E)MFs on the genetic material and the impact of long-term pre-exposure to ELF-(E)MFs on the sensitivity of cells to the damage induced by known mutagens are needed. In this manuscript, an optimized protocol for a combined in vitro comet/micronucleus study to investigate these effects in a human lymphoblastoid cell line (TK6) is provided including the description of a well-validated exposure system. Furthermore, the use of a shielding system to limit background ELF-MFs inside the incubator is described as well. • Optimized protocols for cytogenetic tests with ELF-MFs on TK6 cells ensure the reproducibility of test results. • Validation of exposure environment and exposure system are needed prior to performing tests with ELF-MFs. • A simple, but effective method to shield cells and reduce unintentional ELF-MF exposure consists of using the mu-metal cylinder. This is of particular interest when studying the effects of low exposure levels., SCOPUS: ar.j, info:eu-repo/semantics/published

Through this doctoral thesis chemical composition of acetone, ether, ethyl acetate and dichloromethane extracts and antioxidant, antibacterial and genotoxic activity of acetone extracts of the lichens Umbilicaria crustulosa (Аch.) Frey, Umbilicaria cylindrica (L.) Delise, Hypogimnia tubulosa (Schaer.) Hav. and Evernia prunastri (L.) Ach. were examined. Study on the content of the certain elements in these lichens was also carried out. Activity concentration of 40K, 137Cs, 226Ra and 232Th was determined only for lichen E. prunastri. Anticholinesterase activity of acetone extracts and isolated gyrophoric acid was also examined. The most abundant component of acetone extracts of lichen U. crustulosa and U. cylindrica was girophoric acid. The main components of acetone, ether and ethyl acetate extracts of H. tubulosa were physodic acid and the 3-hydroxyphysodic acid. The most abundant component of acetone extract of E. prunastri was evernic acid. The composition of dichloromethane extracts of all investigated lichens was different from the other extracts, atranorine was dominant component fallowed with small quantities of depsides and depsidons. Orcinol, atraric acid and orsellinic acid were the major volatile compounds in U. crustulosa, U. cylindrica and E. prunastri extracts, while olivetol, 3-hydroxyolivetonide and olivetonide were the main volatiles in H. tubulose extracts. Umbilicaria cylindrica has the highest content of Al, Co, Cr, Fe, Mn, V, Ba, Cd and Pb while the content of B, Si and Zn is the largest in U. crustulosa sample. The activity of 40K was 108 Bq kg-1 while the activities of 137Cs, 226Ra and 232Th were below minimal detectable activities. The results of antioxidant activity indicated a higher antioxidant capacity of depsidone compared to the depside. Gram-negative bacteria were resistant to the tested extracts. Extracts of U. cylindrica, E. prunastri and H. tubulosa had inhibition effect towards Gram-positive bacteria. The highest decreasing in the m

This work represents the study of quality of the Danube River, in Belgrade, at the Višnjica site, by analyzing the genotoxicity of water by using fish as bioindicators. With a population of about 2 million inhabitants (Belgrade) and poor legislation regarding the discharge of wastewaters, monitoring is the first step in approaching this serious problem. To obtain information on the genotoxicity of water as a measure of DNA damage, the comet assay, which detects the effects of exposure, and micronucleus test, which detects permanent effect, were used. For the comet assay we have used three types of cells: blood, liver, and gills, while the micronucleus test was done on blood cells. The fish used as bioindicators are the common perch (Perca fluviatilis), vimba bream (Vimba vimba), common barbell (Barbus barbus), and white bream (Blicca bjoerkna). Regarding the comet assay, significant difference in DNA damage was observed between species in all tested tissues. Vimba bream and common barbell had the highest level of DNA damage in gills, white bream in liver and common perch in blood. In the case of the micronucleus test, the highest number of micronuclei was detected in white bream blood cells. The obtained results showed that there is no correlation between comet assay and the micronucleus test suggesting the importance of performing different bioassays on multiple types of fish tissues in order to find the best biomarker for assessing the genotoxic potential of water.

The cylindrospermopsin (CYN) is one of the most important emerging cyanotoxins in fresh water due to its ubiquity, frequency and severity of the impacts causes in animal and human health.The liver seems to be its main target organ, but other several such as kidneys, lungs,heart, bone marrow, thymus,adrenal glands, immune system, intestinal tract and nervous system can also be affected. Different toxicity mechanisms of CYN have been described, but the most important are the inhibition of protein and glutathione synthesis, oxidative stressand, although genotoxicity has not been elucidated yet, there are some evidences about it.The aim of this work was to study the genotoxic potential of CYN for the first time in the cell line L5178Y/Tk+/-through the micronucleus (MN) assay in absence and presence of the metabolic fraction S9 mix. Our results showed a significant increase in the frecuency of MN in binucleated cells in the presence of S9 (0,25 – 2 μg CYN/mL), while there were not significantdifferences in absence of the metabolic fraction (0,08 – 1,35μg CYN/mL) compared to their respective control groups. For this reason, CYN does not seem to have genotoxic activity per se in the cell line L5178Y Tk+/- althoughits metabolites seem to cause clastogenic and/or aneugenic effects. Further research is required to progress in the knowledge of the genotoxic and carcinogenic potential of this cyanotoxin in order to avoidthose risks resulting in animal and humans from its exposure.

The objectives of the doctoral dissertation were achieved by determining the chemical composition of the extracts of H. physodes, isolation, structural determination and testing of the biological activity of physodalic acid, physodic acid, 3-hydroxyphysodic acid and atranorin. In this dissertation antioxidant activity, cytotoxic activity on cells of human cervix carcinoma and on rat thymocytes, anticholinesterase activity, antimicrobial activity and influence on the formation of micronuclei in lymphocytes of human blood plasma of physodalic acid, physodic acid, 3-hydroxyphysodic acid and atranorin were investigated The antioxidant activity and anticholinesterase activity were investigated by spectrophotometric methods. Cytotoxic activity on cells of human cervix carcinoma and rat thymocytes was assayed by a colorimetric MTT assay, and CCK-8 assay, respectively. The impact on the formation of micronuclei in lymphocytes of human blood plasma was investigated by micronucleus test. Antimicrobial activity was tested by using microdilution method. Comparing the literature available to us, and these results, it was found that for the first time was investigating the effect of Hypogymnia physodes methanol extract on cells of human cervical cancer and the number of micronuclei in lymphocytes of human blood plasma, impact of atranorin on the number of micronuclei in lymphocytes of human blood plasma and serum activity cholinesterase, antioxidant activity of 3-hydroxyphysodic acid and its effects on rat thymocytes, the number of micronuclei in lymphocytes of human blood plasma, the cells of the human cervical cancer and the activity of serum cholinesterase and the impact physodic acid on rat thymocytes, the number of micronuclei in human lymphocytes blood plasma, the cells of the human cervical cancer and serum cholinesterase activity and antioxidant activity of physodalic acid and its effect on rat thymocytes, the number of micronuclei in lymphocytes of human blood plasma, th

The contents of phenols, flavonoids anthocyans and antioxidant activity of the water (WE), methanol (ME) and methanol-water (MWE) (50/50, v/v) extracts of M. sylvestris fresh and dry petals were determined. Further, for all above mentioned extracts, in vitro effect of the frequency of micronuclei (MN) on peripheral human lymphocytes was evaluated using cytochalasin-B micronucleus (CBMN) assay. Total phenol content (mg of gallic acid equivalent/g of petals) varied from 13.20 (WE) to 29.20 (ME) of fresh petals and for dry petals from 80 (WE) to 142 (ME). The content of flavonoids (mg of catechin equivalent /g of petals) ranged from 47.50 (WE) to 101.20 (ME) of fresh petals and for dry petals from 439 (WE) to 814 (ME). The anthocyans content (mg of cyanidin-3-O-glucoside/g of petals) ranged from 0.40 (WE) to 0.72 (ME) of fresh petals and for dry petals from 3.13 (WE) to 6.16 (ME). All extracts showed high antioxidant activity (more than 67%). ME of fresh petals showed a greater decrease in the frequency of MN (from 23.9 to 29.3%) in all tested concentrations (0.5; 1.0 and 2.0 µg/ml) compared to standard radio of protectant to Amifostine (20.8%). © 2017, Scibulcom Ltd. All rights reserved.

This paper reports for the first time the effect of an acetone extract of Umbilicaria crustulosa on the micronucleus distribution of human lymphocytes, and on the cholinesterase activity and antioxidant activity by the cupric ion reducing antioxidant capacity (CUPRAC) method. Additionally, the total phenolic compounds (TPC) and the antioxidant properties were estimated via DPPH, ABTS and TRP assays. Moreover, the antibacterial activity against two Gram-positive and three Gram-negative bacteria were determined. Acetone extract of U. crustulosa at concentration of 1 and 2 mu g mL(-1) decreased a frequency of micronuclei (MN) by 10.8 and 16.8 %, respectively, acting more or slightly less than the synthetic protector amifostine (AMF, WR-2721, 11.4 %, at concentration of 1 mu g mL(-1)). The tested extract did not inhibit cholinesterase activity nor did it exhibit activity toward the examined bacteria. The extract reduced the concentration of DPPH and ABTS radicals by 88.7 and 96.2 %, respectively. Values for total reducing power (TRP) and cupric reducing capacity (CUPRAC) were 0.6197+/-0.0166 mu g ascorbic acid equivalents (AAE) per mg of dry extract, and 19.7641+/-1.6546 mu g trolox equivalents (TE) per mg of dry extract, respectively. The total phenol content was 350.4188 +/-14.587 mu g gallic acid equivalents (GAE) per mg of dry extract. The results of the present study showed that U. crustulosa acetone extract is a promising candidate for in vivo experiments considering its antioxidant activity and protective effect on human lymphocytes.

In the present investigation, effects of Ramalina capitata acetone extract on micronucleus distribution on human lymphocytes, on cholinesterase activity and antioxidant activity (by the CUPRAC method) were examined, for the first time as well as its HPLC profile. Additionally, total phenolic compounds (TPC), antioxidant properties (estimated via DPPH, ABTS and TRP assays) and antibacterial activity were determined. The predominant phenolic compounds in this extract were evernic, everninic and obtusatic acids. Acetone extract of R. capitata at concentration of 2 mu g mL(-1) decreased a frequency of micronuclei (MN) for 14.8 %. The extract reduces the concentration of DPPH and ABTS radicals for 21.2 and 36.1 % (respectively). Values for total reducing power (TRP) and cupric reducing capacity (CUPRAC) were 0.4624 +/- 0.1064 mu g ascorbic acid equivalents (AAE) per mg of dry extract, and 6.1176 +/- 0.2964 mu g Trolox equivalents (TE) per mg of dry extract, respectively. The total phenol content was 670.6376 +/- 66.554 mu g galic acid equivalents (GAE) per mg of dry extract. Tested extract at concentration of 2 mg mL(-1) exhibited inhibition effect (5.2 %) on pooled human serum cholinesterase. The antimicrobial assay showed that acetone extract had inhibition effect towards Gram-positive strains. The results of manifested antioxidant activity, reducing the number of micronuclei in human lymphocytes, and antibacterial activity recommends R. capitata extract for further in vivo studies.

The contents of phenols, flavonoids anthocyans and antioxidant activity of the water (WE), methanol (ME) and methanol-water (MWE) (50/50, v/v) extracts of M. sylvestris fresh and dry petals were determined. Further, for all above mentioned extracts, in vitro effect of the frequency of micronuclei (MN) on peripheral human lymphocytes was evaluated using cytochalasin-B micronucleus (CBMN) assay. Total phenol content (mg of gallic acid equivalent/g of petals) varied from 13.20 (WE) to 29.20 (ME) of fresh petals and for dry petals from 80 (WE) to 142 (ME). The content of flavonoids (mg of catechin equivalent /g of petals) ranged from 47.50 (WE) to 101.20 (ME) of fresh petals and for dry petals from 439 (WE) to 814 (ME). The anthocyans content (mg of cyanidin-3-O-glucoside/g of petals) ranged from 0.40 (WE) to 0.72 (ME) of fresh petals and for dry petals from 3.13 (WE) to 6.16 (ME). All extracts showed high antioxidant activity (more than 67%). ME of fresh petals showed a greater decrease in the frequency of MN (from 23.9 to 29.3%) in all tested concentrations (0.5; 1.0 and 2.0 µg/ml) compared to standard radio of protectant to Amifostine (20.8%). © 2017, Scibulcom Ltd. All rights reserved.

In the present investigation, effects of Ramalina capitata acetone extract on micronucleus distribution on human lymphocytes, on cholinesterase activity and antioxidant activity (by the CUPRAC method) were examined, for the first time as well as its HPLC profile. Additionally, total phenolic compounds (TPC), antioxidant properties (estimated via DPPH, ABTS and TRP assays) and antibacterial activity were determined. The predominant phenolic compounds in this extract were evernic, everninic and obtusatic acids. Acetone extract of R. capitata at concentration of 2 mu g mL(-1) decreased a frequency of micronuclei (MN) for 14.8 %. The extract reduces the concentration of DPPH and ABTS radicals for 21.2 and 36.1 % (respectively). Values for total reducing power (TRP) and cupric reducing capacity (CUPRAC) were 0.4624 +/- 0.1064 mu g ascorbic acid equivalents (AAE) per mg of dry extract, and 6.1176 +/- 0.2964 mu g Trolox equivalents (TE) per mg of dry extract, respectively. The total phenol content was 670.6376 +/- 66.554 mu g galic acid equivalents (GAE) per mg of dry extract. Tested extract at concentration of 2 mg mL(-1) exhibited inhibition effect (5.2 %) on pooled human serum cholinesterase. The antimicrobial assay showed that acetone extract had inhibition effect towards Gram-positive strains. The results of manifested antioxidant activity, reducing the number of micronuclei in human lymphocytes, and antibacterial activity recommends R. capitata extract for further in vivo studies.

This paper reports for the first time the effect of an acetone extract of Umbilicaria crustulosa on the micronucleus distribution of human lymphocytes, and on the cholinesterase activity and antioxidant activity by the cupric ion reducing antioxidant capacity (CUPRAC) method. Additionally, the total phenolic compounds (TPC) and the antioxidant properties were estimated via DPPH, ABTS and TRP assays. Moreover, the antibacterial activity against two Gram-positive and three Gram-negative bacteria were determined. Acetone extract of U. crustulosa at concentration of 1 and 2 mu g mL(-1) decreased a frequency of micronuclei (MN) by 10.8 and 16.8 %, respectively, acting more or slightly less than the synthetic protector amifostine (AMF, WR-2721, 11.4 %, at concentration of 1 mu g mL(-1)). The tested extract did not inhibit cholinesterase activity nor did it exhibit activity toward the examined bacteria. The extract reduced the concentration of DPPH and ABTS radicals by 88.7 and 96.2 %, respectively. Values for total reducing power (TRP) and cupric reducing capacity (CUPRAC) were 0.6197+/-0.0166 mu g ascorbic acid equivalents (AAE) per mg of dry extract, and 19.7641+/-1.6546 mu g trolox equivalents (TE) per mg of dry extract, respectively. The total phenol content was 350.4188 +/-14.587 mu g gallic acid equivalents (GAE) per mg of dry extract. The results of the present study showed that U. crustulosa acetone extract is a promising candidate for in vivo experiments considering its antioxidant activity and protective effect on human lymphocytes.

Cilj predstavljenog istraživanja je da ispita uticaj malih doza jonizujućeg zračenja na zaposlene na različitim radnim mestima u zoni jonizujućeg zračenja. Materijal i metode: Za studiju retrospektivne prirode analizirani su rezultati dobijeni iz kartona preventivno-periodičnih medicinskih pregleda 148 zaposlenih na različitim radnim mestima u zoni jonizujućeg zračenja u Kliničkom Centru Srbije a koji obavljaju preglede u Institutu za medicinu rada Srbije. Ispitanici su podeljeni u tri velike grupe u odnosu na izvor zračenja-oblast rada na zaposlene u nuklearnoj medicini, intrerventnoj radiologiji i opštoj radiologiji. Podaci korišćeni za studiju su podaci sa prethodnog/prvog periodičnog pregleda koji je obavljan pre ulaska u zonu jonizujućeg zračenja ili u toku prve godine izlaganja (u toku obavljanja specijalizacije, u skladu sa odlukom poslodavca da pošalje zaposlenog na pregled) i podaci sa poslednjeg periodičnog pregleda. Uzimani su u obzir ukupni i ekspozicioni radni staž, vrsta izvora zračenja, hematološki parametri (eritrociti, retikulociti, leukociti sa leukocitarnom formulom, trombociti), frekvenca i broj nestabilnih hromozomskih aberacija (ishod testa nestabilnih hromozomskih aberacija) kao i ishod mikronukleusnog testa i njihov broj (značajnih za ocenu radne sposobnosti), primljena kumulativna petogodišnja doza zračenja, morbiditet ispitanika i prisutvo profesionalnih bolesti. Hematološki parametri (eritrociti-Er, retikulociti-Rtc, ukupni leukociti-Leu, monociti-Mo, neutrofili-Neut, eozinofili-Eo, bazofili-Ba, limfociti-Ly i trombociti-Tr) su dobijani ispitivanjem periferne venske krvi u aparatu Beckman Coulter HMX ( na poslednjem periodičnom pregledu) a na prethodnom su uglavnom ispitivani starom metodom ručnog brojanja ćelija (preračunatih za statistički obradu). Nestabilne hromozomske aberacije su dobijane modifikovanom Moorhead’s mikro metodom. Mikronukleusni test je rađen metodom po Fenech-u i Morley-u..., The aim of the research is to analyse the influence of the small doses of ionizing radiation on the employees from different workplaces in the zone of ionizing radiation. Material and methods: For this study of the retrospective nature we have analyzed results obtained from the medical records based on preventive-periodical examinations of 148 persons employed at different workplaces in the zone of ionizing radiation at the Clinical Center of the Republic of Serbia. All of the employees go to medical examinations at the Institute of Occupational Medicine of Serbia. The employees were divided into three large groups as related to the source of radiation – the field of work: the nuclear medicine employees, the interventional radiology employees and the general radiology employees. The data was taken from the previous/initial checkup which had been done before the entrance to the zone of ionizing radiation or during the first year of exposure to radiation (during the specialization period, in accordance with employer’s decision to send an employee to the examination) as well as the data from the last periodical examination. We also took into consideration total and exposition work periods, type of radiation source, haematological parameters (erythrocytes, reticulocytes, leukocytes with differential blood count, platelets), frequency and number of unstable chromosomal aberrations (the results of the test of the unstable chromosomal aberrations), the results of micronucleus test and their number (important for the assessment of working ability), as well as received cumulative five-year radiation dose, morbidity of patients and presence of occupational diseases. The hematological parameters (erythrocytes-RBC, reticulocytes-Rtc, total leucocytes- WBS, monocytes-Mo, neutrophils-Neu, eosinophils-Eo, basophils-Ba and platelets) were obtained through processing of the peripheral vein blood by Beckman Coulter HMX apparatus (at the periodical checkup), while at the previous / in

In this study we have performed a comprehensive genotoxicological survey along the 900 km of the Sava River - the major drainage basin of the Southeastern Europe and the greatest tributary to the Danube River with 945 km long and 97,713 km large catchment's area extending over Slovenia, Croatia, Bosnia and Herzegovina and Serbia. In total, 12 sites were chosen in compliance with routine monitoring program of the basin member states. The genotoxic potential was assessed by the complex battery of bioassays performed in prokaryotes and aquatic eukaryotes (freshwater fish). Battery comprised evaluation of mutagenicity by SOS/umuC test in Salmonella typhimurium TA1535/pSK1002. The level of DNA damage as biomarker of exposure (comet assay) and biomarker of effect (micronucleus assay) and the level of oxidative stress as well (Fpg-modified comet assay) was studied in blood cells of bleak Alburnus alburnus /Albumoides bipunctatus). We wanted to investigate whether the variation in genotoxic potential along the river can be linked to hotspots of faecal and industrial pollution. Hotspots of faecal pollution were identified by bacterial indicators (Escherichia coli numbers) while hotspots related to industry were identified by assessment of concentration of metals in tissue of bleak. The results indicated presence of genotoxic potential along the river resulted from recent or prolonged exposure to genotoxic pressure which can be traced to deterioration of quality of Sava River by urban/industrial wastewaters. The major highlight of the study is that we have provided complex set of data obtained from a single source (homogeneity of analyses for all samples).

Adverse effects of antifouling paints containing organotin compound tributylin (TBT) marked this agent as one of the most toxic substances released into the sea; therefore, its usage is banned. Considering the lack of TBT related data in the Boka Kotorska Bay, we decide to apply an ecotoxicological test battery of TBT influence on level of DNA damage in haemocytes and cardiac activity of the Mediterranean mussel Mytilus galloprovincialis L. During the 96 h treatment with 10, 100 and 1000 µg/l of TBT chloride and 50 µg/l of benzo(a)pyrene used as positive control, in static system, haemolymph samples were collected after 24, 48, 72 and 96 h from all aquaria. Results were done in duplicate and indicated on reduced cell viability compared to negative controls. The increase of DNA damage level in haemocytes obtained by comet assay and significant increase of micronucleus frequency after 72h of exposure in the first experiment and after 96 h in the second was occurred. By means of SOS/umuC assay, using special strains of Salmonella typhimurium, we did not detect mutagenic potency of TBT. However, dose depended decrease in cell biomass was observed. The heart rate recording was carried out by non-invasive method based on infrared sensor technology. The lowest TBT dose induced 10% increase of heart rate up on the treatment onset, while higher concentrations after the initial rise induced significant decline of cardiac activity. Heart rate baseline value was restored later during the treatment with 10 and 100 µg/l of TBT indicating on ability of mussels to maintain regular cardiac activity despite substantial amount of toxin.

Mikronukleus test (MN) se koristi u biomonitoring studijama za praćenje genetičkih efekata kod ljudi izloženih pesticidima. Cilj rada je bio da se, metodom mikronukleus testa, utvrdi oštećenje genetičkog materijala kod ispitanika koji su profesionalno izloženi pesticidima. Istraživanja su obuhvatila 119 ispitanika koji su podeljeni u tri grupe: u kontrolnoj grupi bilo je 39 ispitanika, u grupi izloženoj pesticidima u procesu proizvodnje (proizvođači) bilo je 40 ispitanika i 40 ispitanika koji rade u primeni na terenu. Mann-Whitney U-test je pokazao statistički značajnu razliku između srednjih vrednosti parametara svih ispitivanih varijabli u odnosu na kontrolnu grupu. Statistički značajne razlike konstatovane su između muškaraca u proizvodnji i primeni za parametar MN4, potom između nepušača u proizvodnji i primeni za parametere MN2, MN3, MN4 i NB, kao i u okviru cele posmatrane populacije između kontrolnih i radnika u proizvodnji za parametar MN2, i između proizvodnje i primene za parametar MN3. Spearman-ov test korelacije je pokazao pozitivnu korelaciju između učestalosti mikronukleusa i starosne dobi ispitanika, kao i navika vezanih za pušenje. Utvrđene su i statistički značajne razlike između ispitanika koji rade u proizvodnji i ispitanika koji rade u primeni u odnosu na praćene citogenetičke parametre. Svi dobijeni rezultati upućuju na zaključak da radnici koji rade na terenu ne koriste adekvatna sredstva lične zaštitite. Takođe, rezultati pokazuju da postoji potreba za kontinuiranim biomonitoringom radnika koji su u kontaktu sa pesticidima., Micronucleus (MN) is a biomarker widely used in biomonitoring studies for determining the genetic risk associated with exposure to pesticides. The purpose of this study was to assess damage to the genetic material of workers occupationally exposed to pesticides as detected in micronucleus tests. The research included 119 subjects divided into three groups: a control group of 39 subjects, a group of 40 subjects exposed to pesticides as producers and a group of 40 pesticide applicators in the field. A Mann-Whitney U-test displayed statistically significant differences between the parameter means of all variables, and the control group. Significant differences were observed between males involved in pesticide production and application for the parameter MN4, then between non-smoking producers and applicators regarding parameters MN2, MN3, MN4 and NB, as well as between the control and applicator groups for parameter MN2, and between producers and applicators for parameter MN3. Spearman's correlation test showed a positive correlation between the frequency of micronuclei and age of respondents, as well as their smoking habits. A statistically significant difference in relation to cytogenetic parameters was detected between the respondents working in pesticide production and those working in the field. The results suggest that applicators in the field do not use adequate personal protective equipment. Regular biological monitoring of workers exposed to pesticides is required.

The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed. (C) 2015 The Authors. Published by Elsevier Ltd.

The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed. (C) 2015 The Authors. Published by Elsevier Ltd.

The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed. (C) 2015 The Authors. Published by Elsevier Ltd.

The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed.

The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed. (C) 2015 The Authors. Published by Elsevier Ltd.

The induction of micronucleated reticulocytes in the bone marrow is a sensitive indicator of chromosomal damage. Therefore, the micronucleus assay in rodents is widely used in genotoxicity and carcinogenicity testing. A test system based on cultured human primary cells could potentially provide better prediction compared to animal tests, increasing patient safety while also implementing the 3Rs principle, i.e. replace, reduce and refine. Hereby, we describe the development of an in vitro micronucleus assay based on animal-free ex vivo culture of human red blood cells from hematopoietic stem cells. To validate the method, five clastogens with direct action, three clastogens requiring metabolic activation, four aneugenic and three non-genotoxic compounds have been tested. Also, different metabolic systems have been applied. Flow cytometry was used for detection and enumeration of micronuclei. Altogether, the results were in agreement with the published data and indicated that a sensitive and cost effective in vitro assay to assess genotoxicity with a potential to high-throughput screening has been developed. (C) 2015 The Authors. Published by Elsevier Ltd.

Three lichen secondary metabolites atranorin (1), evernic acid (2), and usnic acid (3), were evaluated for their in vitro clastogenic and antiproliferative effects on human lymphocytes using the cytochalasin-B blocked micronucleus (CBMN) assay at concentrations of 2 mu g/mL, 4 mu g/mL and 6 mu g/mL of final culture solution. The frequency of micronucleus (MN) was scored in binucleated cells, and cytokinesis-block proliferation index (CBPI) was calculated. Among the tested compounds, 3 exhibited the most prominent effect decreasing the frequency of MN in the range of 42.5% - 48.9%, that is about double of the positive control amifostin WR-2721 that reduces MN frequency for 22.0%. The effect of evernic acid was approximately equal to action of amifostin (23.2% -32.9%). Atranorin at concentrations of 2 mu g/mL and 4 mu g/mL decreasing the frequency of MN only for 11.1% and 1.8%, while in concentration of 6 mu g/mL increases the frequency of MN for 9.6 %. The comparable CBPI values of the investigated compounds and control suggested that they did not show a statistically significant inhibitory effect on lymphocyte cell proliferation at applied concentrations.

Three lichen secondary metabolites atranorin (1), evernic acid (2), and usnic acid (3), were evaluated for their in vitro clastogenic and antiproliferative effects on human lymphocytes using the cytochalasin-B blocked micronucleus (CBMN) assay at concentrations of 2 mu g/mL, 4 mu g/mL and 6 mu g/mL of final culture solution. The frequency of micronucleus (MN) was scored in binucleated cells, and cytokinesis-block proliferation index (CBPI) was calculated. Among the tested compounds, 3 exhibited the most prominent effect decreasing the frequency of MN in the range of 42.5% - 48.9%, that is about double of the positive control amifostin WR-2721 that reduces MN frequency for 22.0%. The effect of evernic acid was approximately equal to action of amifostin (23.2% -32.9%). Atranorin at concentrations of 2 mu g/mL and 4 mu g/mL decreasing the frequency of MN only for 11.1% and 1.8%, while in concentration of 6 mu g/mL increases the frequency of MN for 9.6 %. The comparable CBPI values of the investigated compounds and control suggested that they did not show a statistically significant inhibitory effect on lymphocyte cell proliferation at applied concentrations.

The safety of several azo colouring agents, used as food additives, has during the years been questioned. Allura Red AC (E129) has in some publications been classified as genotoxic. In fact, in the European Union, Allura Red is permitted as a food additive in human food, but, surprisingly, it was not acceptable as an additive for use in animal feed. In this study we have evaluated whether Allura Red is genotoxic using a flow cytometer-based micronucleus assay in peripheral blood of mice. Male FVB mice were given a single intra-peritoneal injection of various doses of Allura Red and sacrificed at 46 h after treatment. The tested doses were 0, 100, 200, 400, 600, 800, 1000, 1500, and 2000 mg/kg body weight (b.w.). Each dose group constituted three mice, except for in the dose group of 1000 mg/kg b.w., which constituted four mice. Blood samples were collected and the frequency of micronucleated polychromatic erythrocytes (fMNPCE) and the cell proliferation (%PCE) was determined. The analyses did not show any significant difference in the %PCE or in the fMNPCE. Consequently, under the testing circumstances one can conclude that Allura Red is not genotoxic.

The safety of several azo colouring agents, used as food additives, has during the years been questioned. Allura Red AC (E129) has in some publications been classified as genotoxic. In fact, in the European Union, Allura Red is permitted as a food additive in human food, but, surprisingly, it was not acceptable as an additive for use in animal feed. In this study we have evaluated whether Allura Red is genotoxic using a flow cytometer-based micronucleus assay in peripheral blood of mice. Male FVB mice were given a single intra-peritoneal injection of various doses of Allura Red and sacrificed at 46 h after treatment. The tested doses were 0, 100, 200, 400, 600, 800, 1000, 1500, and 2000 mg/kg body weight (b.w.). Each dose group constituted three mice, except for in the dose group of 1000 mg/kg b.w., which constituted four mice. Blood samples were collected and the frequency of micronucleated polychromatic erythrocytes (fMNPCE) and the cell proliferation (%PCE) was determined. The analyses did not show any significant difference in the %PCE or in the fMNPCE. Consequently, under the testing circumstances one can conclude that Allura Red is not genotoxic., AuthorCount:2

Cadmium (Cd) is highly toxic heavy metal which may cause severe biological effects in vivo and in vitro. In this study, an evaluation of the acute Cd ability to trigger micronuclei (MNi) formation was carried out on 3-monthold male and female Albino Oxford (AO) rats using micronucleus (MN) test. Experimental animals were treated intraperitoneally with three different concentrations of cadmium chloride (CdCl2): 0.5, 1, and 2 mg CdCl2 per kg of body weight. Control animals received equal volume of sterile phosphate buffered saline. The results showed that 2 mg CdCl2 per kg b.w. concentration caused a highly statistically significant (P lt 0.001) increase in MNi formation in the bone marrow polychromatic erythrocytes (PCEs), exerting a clear-cut concentration-dependent effect. Lower concentrations of CdCl2 used (0.5 and 1 mg/kg b.w.) also caused MNi formation, but with lower statistical significance. Sex differences in MNi production in bone marrow PCEs after acute exposure to different experimental concentrations of CdCl2 were not observed in our study. Our results indicate the ability of CdCl2 to exerts genotoxic effects in bone marrow of AO rats, and complement previous data on the genotoxicity of this important environmental contaminant, burdening the body from different sources - major being industrial exposure, food and cigarette smoking., Kadmijum (Cd) je veoma toksičan teški metal koji može da prouzrokuje ozbiljne biološke efekte in vivo i in vitro. U ovom istraživanju, procena potencijala Cd da izazove produkciju mikronukleusa (MNA) u polihromatskim eritrocitima (PHE) košatne srži, sprovedena je na tri meseca starim mužjacima i ženkama AO (Albino Oxford) soja pacova, primenom mikronukleus testa. Eksperimentalne životinje tretirane su intraperitonealno sa tri različite koncentracije kadmijum-hlorida (CdCl2) i to: 0,5, 1 i 2 mg CdCl2 po kg telesne mase. Kontrolne životinje primile su istu količinu sterilnog fiziološkog rastvora sa fosfatnim puferom. Rezultati su pokazali da koncentracija CdCl2 od 2 mg/kg t.m. prouzrokuje statistički visoko značajno (P lt 0,001) povećanje produkcije MNA u PHE koštane srži, i taj efekat je koncentracijski zavisan. Niže koncentracija CdCl2 koje su korišćene u ovom istraživanju (0,5 i 1 mg/kg t.m.) takođe su prouzrokovale produkciju MNA, ali efekat nije bio statistički visoko značajan. Polne razlike u produkciji MNA u PHE koštane srži, nakon akutnog izlaganja različitim eksperimentalnim koncentracijama CdCl2, nisu uočene u našem istraživanju. Dobijeni rezultati ukazuju na potencijal CdCl2 da prouzrokuje genotoksične efekte u koštanoj srži pacova AO soja i upotpunjuju bazu podataka o genotoksičnosti ovog zagađivača životne sredine, koji je najčešće uzročnik bolesti ljudi izloženih Cd u industrijskoj proizvodnji i onih koji konzumiranju cigarete i hranu kontaminiranu Cd.

Three lichen depsidones, physodalic acid (1), physodic acid (2), and 3-hydroxy physodic acid (3), were isolated from Hypogymnia physodes diethyl ether extract using column chromatography, and their structures determined by comparing their UV, H-1 and C-13 NMR spectroscopic and MS data with those given in the literature, as well as with data computed by CHEM draw ultra 11 software. The contents of 1, 2 and 3 were determined in the methanol (ME), acetone (AE), and diethyl ether (EE) extracts using reversed-phase high performance liquid chromatography with photodiode array detection. The extracts, isolated depsidones 1-3 and EE fraction F23 (consisting of 90% 2 and 3, in the ratio 5.5: 1) were evaluated for their in vitro effects on chromosome aberrations in peripheral human lymphocytes using the cytochalasin-B blocked micronucleus (CBMN) assay in doses of 1 mu g/mL and 2 mu g/mL of final culture solution. The frequency of MN was scored in binucleated cells, and nuclear proliferation index was calculated. It was found that 1, 2, 3, F23, and EE at 1.0 mu g/mL exerted a beneficial effect on lymphocyte cells giving a significant decrease of the frequency of MN in comparison with the positive control Amifostin WR-2721. Among the tested depsidones at a concentration of 1 mu g/mL, 3 exhibited the most prominent effect decreasing the frequency of MN by 30.3%, followed by 2 (28.2%) and 1 (22.0%). The extracts were less effective than the isolated depsidones.

Cadmium (Cd) is highly toxic heavy metal which may cause severe biological effects in vivo and in vitro. In this study, an evaluation of the acute Cd ability to trigger micronuclei (MNi) formation was carried out on 3-month-old male and female Albino Oxford (AO) rats using micronucleus (MN) test. Experimental animals were treated intraperitoneally with three different concentrations of cadmium chloride (CdCl2): 0.5, 1, and 2 mg CdCl2 per kg of body weight. Control animals received equal volume of sterile phosphate buffered saline. The results showed that 2 mg CdCl2 per kg b.w. concentration caused a highly statistically significant (P LT 0.001) increase in MNi formation in the bone marrow polychromatic erythrocytes (PCEs), exerting a clear-cut concentration-dependent effect. Lower concentrations of CdCl2 used (0.5 and 1 mg/kg b.w.) also caused MNi formation, but with lower statistical significance. Sex differences in MNi production in bone marrow PCEs after acute exposure to different experimental concentrations of CdCl2 were not observed in our study. Our results indicate the ability of CdCl2 to exerts genotoxic effects in bone marrow of AO rats, and complement previous data on the genotoxicity of this important environmental contaminant, burdening the body from different sources-major being industrial exposure, food and cigarette smoking.

Cadmium (Cd) is highly toxic heavy metal which may cause severe biological effects in vivo and in vitro. In this study, an evaluation of the acute Cd ability to trigger micronuclei (MNi) formation was carried out on 3-monthold male and female Albino Oxford (AO) rats using micronucleus (MN) test. Experimental animals were treated intraperitoneally with three different concentrations of cadmium chloride (CdCl2): 0.5, 1, and 2 mg CdCl2 per kg of body weight. Control animals received equal volume of sterile phosphate buffered saline. The results showed that 2 mg CdCl2 per kg b.w. concentration caused a highly statistically significant (P lt 0.001) increase in MNi formation in the bone marrow polychromatic erythrocytes (PCEs), exerting a clear-cut concentration-dependent effect. Lower concentrations of CdCl2 used (0.5 and 1 mg/kg b.w.) also caused MNi formation, but with lower statistical significance. Sex differences in MNi production in bone marrow PCEs after acute exposure to different experimental concentrations of CdCl2 were not observed in our study. Our results indicate the ability of CdCl2 to exerts genotoxic effects in bone marrow of AO rats, and complement previous data on the genotoxicity of this important environmental contaminant, burdening the body from different sources - major being industrial exposure, food and cigarette smoking., Kadmijum (Cd) je veoma toksičan teški metal koji može da prouzrokuje ozbiljne biološke efekte in vivo i in vitro. U ovom istraživanju, procena potencijala Cd da izazove produkciju mikronukleusa (MNA) u polihromatskim eritrocitima (PHE) košatne srži, sprovedena je na tri meseca starim mužjacima i ženkama AO (Albino Oxford) soja pacova, primenom mikronukleus testa. Eksperimentalne životinje tretirane su intraperitonealno sa tri različite koncentracije kadmijum-hlorida (CdCl2) i to: 0,5, 1 i 2 mg CdCl2 po kg telesne mase. Kontrolne životinje primile su istu količinu sterilnog fiziološkog rastvora sa fosfatnim puferom. Rezultati su pokazali da koncentracija CdCl2 od 2 mg/kg t.m. prouzrokuje statistički visoko značajno (P lt 0,001) povećanje produkcije MNA u PHE koštane srži, i taj efekat je koncentracijski zavisan. Niže koncentracija CdCl2 koje su korišćene u ovom istraživanju (0,5 i 1 mg/kg t.m.) takođe su prouzrokovale produkciju MNA, ali efekat nije bio statistički visoko značajan. Polne razlike u produkciji MNA u PHE koštane srži, nakon akutnog izlaganja različitim eksperimentalnim koncentracijama CdCl2, nisu uočene u našem istraživanju. Dobijeni rezultati ukazuju na potencijal CdCl2 da prouzrokuje genotoksične efekte u koštanoj srži pacova AO soja i upotpunjuju bazu podataka o genotoksičnosti ovog zagađivača životne sredine, koji je najčešće uzročnik bolesti ljudi izloženih Cd u industrijskoj proizvodnji i onih koji konzumiranju cigarete i hranu kontaminiranu Cd.

Three lichen depsidones, physodalic acid (1), physodic acid (2), and 3-hydroxy physodic acid (3), were isolated from Hypogymnia physodes diethyl ether extract using column chromatography, and their structures determined by comparing their UV, H-1 and C-13 NMR spectroscopic and MS data with those given in the literature, as well as with data computed by CHEM draw ultra 11 software. The contents of 1, 2 and 3 were determined in the methanol (ME), acetone (AE), and diethyl ether (EE) extracts using reversed-phase high performance liquid chromatography with photodiode array detection. The extracts, isolated depsidones 1-3 and EE fraction F23 (consisting of 90% 2 and 3, in the ratio 5.5: 1) were evaluated for their in vitro effects on chromosome aberrations in peripheral human lymphocytes using the cytochalasin-B blocked micronucleus (CBMN) assay in doses of 1 mu g/mL and 2 mu g/mL of final culture solution. The frequency of MN was scored in binucleated cells, and nuclear proliferation index was calculated. It was found that 1, 2, 3, F23, and EE at 1.0 mu g/mL exerted a beneficial effect on lymphocyte cells giving a significant decrease of the frequency of MN in comparison with the positive control Amifostin WR-2721. Among the tested depsidones at a concentration of 1 mu g/mL, 3 exhibited the most prominent effect decreasing the frequency of MN by 30.3%, followed by 2 (28.2%) and 1 (22.0%). The extracts were less effective than the isolated depsidones.

Cadmium (Cd) is highly toxic heavy metal which may cause severe biological effects in vivo and in vitro. In this study, an evaluation of the acute Cd ability to trigger micronuclei (MNi) formation was carried out on 3-month-old male and female Albino Oxford (AO) rats using micronucleus (MN) test. Experimental animals were treated intraperitoneally with three different concentrations of cadmium chloride (CdCl2): 0.5, 1, and 2 mg CdCl2 per kg of body weight. Control animals received equal volume of sterile phosphate buffered saline. The results showed that 2 mg CdCl2 per kg b.w. concentration caused a highly statistically significant (P LT 0.001) increase in MNi formation in the bone marrow polychromatic erythrocytes (PCEs), exerting a clear-cut concentration-dependent effect. Lower concentrations of CdCl2 used (0.5 and 1 mg/kg b.w.) also caused MNi formation, but with lower statistical significance. Sex differences in MNi production in bone marrow PCEs after acute exposure to different experimental concentrations of CdCl2 were not observed in our study. Our results indicate the ability of CdCl2 to exerts genotoxic effects in bone marrow of AO rats, and complement previous data on the genotoxicity of this important environmental contaminant, burdening the body from different sources-major being industrial exposure, food and cigarette smoking.

Genotoxic effects on fauna after waterborne pollutant exposure have been demonstrated by numerous research programmes. Less effort has been focused on establishing relationship between genotoxicity and long-term responses at higher levels of biological organization. Taking into account that embryos may be more sensitive indicators of reproductive impairment than alterations in fertility, we have developed two assays in multiwell plates to address correlations between embryo toxicity and genotoxicity. The potential teratogenicity was assessed by analyzing abnormal development and mortality of Physa acuta at embryonic stage. Genotoxicity was measured by the micronucleus (MN) test using embryonic cells. Our results showed that linkage between genotoxicity and embryo toxicity depends on mechanisms of action of compounds under study. Embryo toxic responses showed a clear dose-related tendency whereas no clear dose-dependent effect was observed in micronucleus induction. The higher embryo toxicity was produced by benzo(a)pyrene exposure followed by fluoxetine and bisphenol A. Vinclozolin was the lower embryo toxic compound. Binary mixtures with BaP always resulted in higher embryo toxicity than single exposures but antagonistic effects were observed for MN induction. Benzo(a)pyrene produced the higher MN induction at 0.04. mg/L, which also produced clear embryo toxic effects. Fluoxetine did not induce cytogenetic effects but 0.25. mg/L altered embryonic development. Bisphenol A significantly reduced hatchability at 0.5. mg/L while MN induction appeared with higher treatments than those that start causing teratogenicity. Much higher concentration of vinclozolin (5. mg/L) reduced hatchability and induced maximum MN formation. In conclusion, while validating one biomarker of genotoxicity and employing one ecologically relevant effect, we have evaluated the relative sensitivity of a freshwater mollusc for a range of chemicals. The embryo toxicity test is a starting point for

Although alcohol is known to be a carcinogen for humans, ethanol-genotoxicity studies are incomplete. Ethanol seems not to be a bacterial mutagen, but the results are conflicting in rodent assays. We investigate the genotoxicity in the bone marrow micronucleus (MN) test and in the dominant lethal mutation (DLM) assay using two long-term ethanol exposure protocols. In the MN test, mice consumed three doses (5, 10 and 15% v/v) for 32 weeks. MN induction was compared to two control groups of 5- and 38-week-old mice (the ages of the treated mice when the treatment was initiated and when they were killed, respectively). For the three groups treated with ethanol there was no significant increase in MN induction as compared to the first control group, but observed MN frequencies were significantly lower than in the 38-week-old control group. This suggests a protective effect against genotoxic damage caused by aging, probably due to ethanol action as a hydroxyl radical scavenger. In the DLM assay, male mice drank ethanol at 15% or 30% (v/v) for 20 weeks. In both groups the number of dead implants was similar to the control, but there was a significant reduction in total implants, indicating a pre-implantation loss.

The aim of the present study was to investigate the water quality and the ecosystem health of a segment of Pescara river (Abruzzo-Italy) with the aid of an integrated approach. Sixteen priority polycyclic aromatic hydrocarbons and heavy metals were determinated in water and chub samples in order to calculate the bioconcentration factor. Biochemical, physical and microbiological characterisation were also carried out on water samples. Moreover, real and synergic effects of water pollution on biota were investigated by genotoxicological tests. Water samples were tested by Salmonella mutagenicity assay, whereas micronucleus assay on chub was used to evaluate environmental genotoxic effects on fish. The characterisation of water samples did not reveal any relevant contamination of Pescara river, but some analytical results obtained for biota were remarkable. In particular, the comparison between the site downstream from the industrial area of Chieti and the control site showed significant differences in chub micronuclei frequencies., Obiettivo del lavoro è stato indagare alcuni aspetti della qualità delle acque e della salute ecosistemica di un segmento del fiume Pescara (Abruzzo). E’stata dapprima effettuata una caratterizzazione dei siti mediante l’analisi di parametri chimico-fisici e microbiologici delle acque, quindi è stata determinata la bioconcentrazione nel cavedano di idrocarburi policiclici aromatici e metalli pesanti. L’analisi chimica è stata accompagnata da test genotossicologici per valutare gli effetti sinergici sul biota. In particolare, è stato eseguito il test del micronucleo su L. cephalus come biomarker e il test di mutagenesi su Salmonella. Dallo studio è emerso che la qualità delle acque del fiume Pescara non è ancora compromessa; indicazioni su un’alterazione dei siti di studio risultano invece dall’analisi delle componenti biologiche. In particolare, il campione di cavedani raccolto a valle della zona industriale di Chieti presentava una maggiore incidenza di cellule micronucleate rispetto a quello raccolto nel sito controllo.

Mikrojaderný test zastupuje širokou skupinu genotoxických testů, které slouží k detekci genotoxických látek. V dnešní době tyto testy využíváme k monitorování nových látek, ale i běžně používaných látek v chemickém, farmaceutickém, zemědělském a potravinářském průmyslu. Slouží i k identifikaci přítomnosti genotoxických faktorů v životním prostředí. V důsledku interakce genotoxické látky s DNA (deoxyribonukleová kyselina) dochází k bodovým i strukturním změnám v DNA, které zvyšují strukturní labilitu a narušují integritu v místech poškození DNA. Za určitých podmínek mohou dát vznik mutacím, chromozomálním aberacím, karcinogenezi. Tato práce je zaměřena na detekci genotoxických účinků u vybraných látek mikrojaderným testem in vitro, in vivo. Mezi testované látky byl zařazen cyklofosfamid, kolchicin, formaldehyd, methylcelulosa, sulfid cínatý. Výsledná data testu ukázaly, že sulfid cínatý testovaný metodou in vivo mikrojaderného testu, neprokazuje genotoxicitu. Stejného výsledku jsme dosáhli u mikrojaderného testu in vitro s methylcelulosou. Pozitivní výsledky mikrojaderného testu in vitro byly u cyklofosfamidu, kolchicinu, formaldehydu. U všech testovaných látek byla sledována závislost míry cytotoxicity na změně koncentrace jednotlivých testovaných látek. Tato cytotoxicita se hodnotila mikroskopickým hodnocením a následně hodnocením dle indexu CBPI (proliferační index cytokinetického bloku)., Micronucleus test represents a broad group of genotoxicity tests used to detect genotoxic substances. Today we use these tests to monitor new substances, but substance commonly used in chemical, pharmaceutical, agricultural and food industries. Serves for identifying the presence of genotoxic factors in the environment. As a result of interaction of genotoxic agents with DNA (deoxyribonucleic acid) leads to a point and structural changes in DNA that increase the structural instability and undermine the integrity of the sites of DNA damage. Under certain conditions, may give rise to mutations, chromosomal aberrations, carcinogenesis. This work is focused on the detection of genotoxic effects of selected substance in the micronucleus test in vitro and in vivo. Among the tested substances was included cyclophosphamide, colchicine, formaldehyde, methyl cellulose, Stannous sulphide. The resulting test data show that Stannous sulphide tested using in vivo micronucleus test does not establish the probability or possibility of confirmation of genotoxicity. The same result was obtained in the micronucleus test in vitro with, Katedra biologických a biochemických věd, 1. Prezentace výsledků diplomové práce 2. Diskuze k posudkům vedoucího a oponenta diplomové práce 3. Diplomantka zodpověděla všechny dotazy a přípomínky k DP, Dokončená práce s úspěšnou obhajobou

When food is heated, a vast number of compounds are formed. Some of these are known to be toxic. Among these are furan, HMF, PhIP, IQ, and MeIQx, the subjects of this thesis. All these compounds are known or suspected carcinogens but the detailed mechanisms behind their carcinogenicity have not yet been fully examined. The aim of this thesis was to study genotoxic properties of these compounds using both in vitro and in vivo methods. Clastogenic effects of all five compounds were assessed with the flow cytometer-based micronucleus assay in vivo and for furan also with the micronucleus assay in vitro. DNA-damaging effects of HMF were studied using the comet assay. No induction of micronuclei was obtained after exposure to IQ, MeIQx or furan. Hence, it can be argued that non-genotoxic mechanisms are partly responsible for the carcinogenic properties of these compounds. PhIP, on the other hand, generated a clear response in the in vivo test. Comparing these result with previous results on acrylamide indicates that PhIP is much more potent. However, acrylamide probably poses a higher risk for humans as the intake is considerably higher. For HMF no effects were seen using the in vivo setup. To further investigate the influence of bioactivation of HMF by sulfotransferases (SULTs) the comet assay was performed in cell lines expressing different levels of SULT. However, no correlation between SULT-expression and DNA-damage was observed. Thus, the DNA-damaging effects found in our experimental setup is probably due to other factors than SULT mediated effects. Furthermore, in this thesis the effects of folic acid on chromosomal stability in healthy people were studied. A negative correlation was found between micronucleus frequency and folate status. The results gained within this thesis will hopefully contribute to the risk assessment of compounds present in our diet.

When food is heated, a vast number of compounds are formed. Some of these are known to be toxic. Among these are furan, HMF, PhIP, IQ, and MeIQx, the subjects of this thesis. All these compounds are known or suspected carcinogens but the detailed mechanisms behind their carcinogenicity have not yet been fully examined. The aim of this thesis was to study genotoxic properties of these compounds using both in vitro and in vivo methods. Clastogenic effects of all five compounds were assessed with the flow cytometer-based micronucleus assay in vivo and for furan also with the micronucleus assay in vitro. DNA-damaging effects of HMF were studied using the comet assay. No induction of micronuclei was obtained after exposure to IQ, MeIQx or furan. Hence, it can be argued that non-genotoxic mechanisms are partly responsible for the carcinogenic properties of these compounds. PhIP, on the other hand, generated a clear response in the in vivo test. Comparing these result with previous results on acrylamide indicates that PhIP is much more potent. However, acrylamide probably poses a higher risk for humans as the intake is considerably higher. For HMF no effects were seen using the in vivo setup. To further investigate the influence of bioactivation of HMF by sulfotransferases (SULTs) the comet assay was performed in cell lines expressing different levels of SULT. However, no correlation between SULT-expression and DNA-damage was observed. Thus, the DNA-damaging effects found in our experimental setup is probably due to other factors than SULT mediated effects. Furthermore, in this thesis the effects of folic acid on chromosomal stability in healthy people were studied. A negative correlation was found between micronucleus frequency and folate status. The results gained within this thesis will hopefully contribute to the risk assessment of compounds present in our diet.

The distribution of polyphenolic compounds in a grape (Vitis vinifera) seed extract (GSE) was determined using LC/ESI-TOF MS, HPLC/DAD, and C-13-NMR. The 17 identified compounds comprised gallic and protocatechuic acid, catetchin and epicatechin monomers, procyanidin oligomers, and procyanidin gallates. This study addresses the in vitro effects of grape seed extract (GSE) on the frequency of micronuclei with reference to the antioxidant status in human lymphocytes. To establish the most effective protective support, we used four different concentrations of GSE, in the range 1-6 mu g/mL. Treatment of lymphocytes with GSE at a concentration of 2.5 mu g/mL induced a significant decrease in the frequency of micronuclei by 40%, reduction of malonyldialdehyde production by 30%, while a concentration of 5 mu g/mL increased catalase and glutathione S-transferase activity by 10 % and 15 %, respectively. These results demonstrate that GSE may be effective in the prevention of oxidative lymphocyte damage by ROS.

The distribution of polyphenolic compounds in a grape (Vitis vinifera) seed extract (GSE) was determined using LC/ESI-TOF MS, HPLC/DAD, and (13)C-NMR. The 17 identified compounds comprised gallic and protocatechuic acid, catetchin and epicatechin monomers, procyanidin oligomers, and procyanidin gallates. This study addresses the in vitro effects of grape seed extract (GSE) on the frequency of micronuclei with reference to the antioxidant status in human lymphocytes. To establish the most effective protective support, we used four different concentrations of GSE, in the range 1-6 mu g/mL. Treatment of lymphocytes with GSE at a concentration of 2.5 mu g/mL induced a significant decrease in the frequency of micronuclei by 40%, reduction of malonyldialdehyde production by 30%, while a concentration of 5 mu g/mL increased catalase and glutathione S-transferase activity by 10 % and 15 %, respectively. These results demonstrate that GSE may be effective in the prevention of oxidative lymphocyte damage by ROS.

This study addresses the in vitro protective effects of proanthocyanidins and quercetin on the frequency of micronuclei with reference to the antioxidant status of cultured human lymphocytes also subjected to 2 Gy of gamma-radiation. Treatment of lymphocytes with proanthocyanidins induced a significant decrease in the frequency of micronuclei and reduction of malonyldialdehyde production, as well as increased catalase and glutathione S-transferase activity Quercetin induces a higher incidence of micronuclei and production of malonyldialdehyde. The seven-fold increase of micronuclei frequency induced by a therapeutic dose of gamma-radiation was decreased in the presence of proanthocyanidins. These results demonstrate that proanthocyanidins may be important in the prevention of ROS-induced oxidative lymphocyte damage by decreasing DNA damage, lowering membrane lipid peroxidation, and increasing the activity AOP enzymes, as well as by reducing the level of gamma-radiation-induced DNA damage. Our results support the potential benefits of proanthocyanidins as efficient antioxidants and radioprotectors.

The distribution of polyphenolic compounds in a grape (Vitis vinifera) seed extract (GSE) was determined using LC/ESI-TOF MS, HPLC/DAD, and (13)C-NMR. The 17 identified compounds comprised gallic and protocatechuic acid, catetchin and epicatechin monomers, procyanidin oligomers, and procyanidin gallates. This study addresses the in vitro effects of grape seed extract (GSE) on the frequency of micronuclei with reference to the antioxidant status in human lymphocytes. To establish the most effective protective support, we used four different concentrations of GSE, in the range 1-6 mu g/mL. Treatment of lymphocytes with GSE at a concentration of 2.5 mu g/mL induced a significant decrease in the frequency of micronuclei by 40%, reduction of malonyldialdehyde production by 30%, while a concentration of 5 mu g/mL increased catalase and glutathione S-transferase activity by 10 % and 15 %, respectively. These results demonstrate that GSE may be effective in the prevention of oxidative lymphocyte damage by ROS.

When food is heated, a vast number of compounds are formed. Some of these are known to be toxic. Among these are furan, HMF, PhIP, IQ, and MeIQx, the subjects of this thesis. All these compounds are known or suspected carcinogens but the detailed mechanisms behind their carcinogenicity have not yet been fully examined. The aim of this thesis was to study genotoxic properties of these compounds using both in vitro and in vivo methods. Clastogenic effects of all five compounds were assessed with the flow cytometer-based micronucleus assay in vivo and for furan also with the micronucleus assay in vitro. DNA-damaging effects of HMF were studied using the comet assay. No induction of micronuclei was obtained after exposure to IQ, MeIQx or furan. Hence, it can be argued that non-genotoxic mechanisms are partly responsible for the carcinogenic properties of these compounds. PhIP, on the other hand, generated a clear response in the in vivo test. Comparing these result with previous results on acrylamide indicates that PhIP is much more potent. However, acrylamide probably poses a higher risk for humans as the intake is considerably higher. For HMF no effects were seen using the in vivo setup. To further investigate the influence of bioactivation of HMF by sulfotransferases (SULTs) the comet assay was performed in cell lines expressing different levels of SULT. However, no correlation between SULT-expression and DNA-damage was observed. Thus, the DNA-damaging effects found in our experimental setup is probably due to other factors than SULT mediated effects. Furthermore, in this thesis the effects of folic acid on chromosomal stability in healthy people were studied. A negative correlation was found between micronucleus frequency and folate status. The results gained within this thesis will hopefully contribute to the risk assessment of compounds present in our diet.

[Abstract] The spill from the oil tanker Prestige (NW Spain, November 2002) was perhaps the biggest ecological disaster that happened worldwide in the last decades. As a consequence of this catastrophe a general concern led to a huge mobilization of human and technical resources. Given that no information was reported in the scientific literature regarding to the chronic repercussions to human health of exposure to oil spills, a pilot study was performed by our group revealing some increased genotoxic effects in the subjects exposed to the oil during cleaning activities. Due to the seriousness of the results, we extended our study comprising a larger population and including an extensive evaluation of the main polymorphic sites in metabolizing and DNA-repair genes. General increases in micronucleus (MN) frequency and decreases in the proliferation index were observed in individuals with longer time of exposure. Age was a significant predictor of MN frequency. CYP1A1 3′-UTR, EPHX1 codons 113 and 139, GSTP1, GSTM1 and GSTT1 metabolic polymorphisms, and XRCC3 codon 241 and XPD codon 751 repair polymorphisms influenced cytogenetic damage levels. In view of these results, it seems essential to pay more attention to the chronic human health effects of exposure to oil and to focus new studies on such a relevant but overlooked public health field that involves a large number of people all over the world.