Search

BPA is a plasticizer for the production of polycarbonate plastics and epoxy resins and is widely used in the production of household goods, including food packaging. Free BPA is known to migrate from packaging to food, and its uptake has been associated with adverse health effect, particularly the disruption of endocrine activity. The presence and migration of BPA from plastic consumer products are subject to strict regulation in the EU. The aim of this study is to analyse the migration of BPA from different packaging items and household products sold on the Croatian market. To simulate real life exposure, we treated samples with a food simulant. The analytical performance was confirmed with the EU requirements. BPA levels were assessed in 61 samples by HPLC-FLD and the LOQ of the method was 0.005 mg kg−1 for the food simulant. These results showed that the levels of BPA that migrated to the food simulant were below LOQ and in accordance with the specific migration limit into food, which was defined as 0.05 mg kg−1 for all samples. None of the analysed products presented a health hazard. However, these regulations do not refer to products intended for children’s use, in which BPA is banned. Furthermore, regulations require testing before putting products on the market, and previous research shows that possible BPA migration occurs due to various uses, along with a cumulative effect of exposure from even very small concentrations. Therefore, for accurate BPA consumer exposure evaluation and possible health risks, a comprehensive approach is needed.

The harmful effects of silver nanoparticles (AgNPs) have been confirmed in many organisms, but the mechanism of their toxicity is not yet fully understood. In biological systems, AgNPs tend to aggregate and dissolve, so they are often stabilized by coatings that influence their physico-chemical properties. In this study, the effects of AgNPs with different coatings [polyvinylpyrrolidone (PVP) and cetyltrimethylammonium bromide (CTAB)] on oxidative stress appearance and proteome changes in tobacco (

The Croatian National Health Care Act defines the areas of activities of the public health institute, including the activities of the epidemiology of infectious diseases and chronic non-communicable diseases, public health, health promotion, environmental health, microbiology, school and adolescent medicine, mental health and addiction prevention at Zagreb City level. This paper reviews the highly variable activities in the Andrija Štampar Teaching Institute of Public Health with the aim of promoting a comprehensive approach to the COVID-19 pandemic. Human and analytical resources in the Institute, activities and rapid implementation of innovations testify to the high capacities for adaptation to emerging risks. In the Institute, it is possible to carry out a whole range of tests and to monitor the environmental factors with predominant impact on human health and safety of the Zagreb environment. The supply of safe water for human consumption in the Republic of Croatia during the current COVID-19 crisis has been uninterrupted and in accordance with applicable legislation. Also, our laboratories have been developing and introducing a method for wastewater testing for SARS-CoV-2 presence. The sludge from wastewater treatment plants is used in agriculture, and potential risks associated with the COVID-19 outbreak should be assessed prior to each application on the soil. Increased use of disinfectants during the epidemic may present a higher risk to the aquatic environment. Air quality monitoring indicates a positive impact on air quality as result of isolation measures.

Extensive commercialization of silver nanoparticles (AgNPs) raises the risk of their accumulation in the soil-plant system. Once released into the environment, AgNPs are prone to chemical transformations, which make it hard to determine whether their phytotoxic effects are purely NP-related or a consequence of released Ag+ ions. In this study the effects of 25, 50, 75, 100 and 150 μM AgNPs and AgNO3 on seed germination and early growth of tobacco (Nicotiana tabacum L.) seedlings were compared. Additionally, the effects on photosynthetic performance and pigment content were investigated. Germination rate and index values indicated delayed and slower germination in some AgNP treatments. Lower AgNP concentrations stimulated root growth, but induced a prominent reduction in fresh weight. By contrast, all AgNO3 concentrations inhibited root growth but only the higher ones decreased fresh weight. Obtained results imply that the observed AgNP toxicity could be ascribed to NP form and can be correlated with high AgNP stability in the solid medium. On the other hand, the majority of AgNP and AgNO3 treatments induced an increase in chlorophyll content that was accompanied by significantly lower values of relative electron transport rate and coefficient of photochemical quenching, implying an inhibition of the electron transport chain. A similar impact of AgNPs and AgNO3 on photosynthesis can be correlated with lower stability of AgNPs in a liquid medium, resulting in AgNP aggregation and dissolution of Ag+ ions.

Widespread application of silver nanoparticles (AgNPs), due to their antibacterial and antifungal properties, increases their release into the environment and potential detrimental impact on living organisms. Plants may serve as a potential pathway for AgNPs bioaccumulation and a route into the food chain, hence investigation of AgNP phytotoxic effects are of particular importance. Since proteins are directly involved in stress response, studies of their abundance changes can help elucidate the mechanism of the AgNP-mediated phytotoxicity. In this study, we investigated proteomic changes in tobacco (Nicotiana tabacum) exposed to AgNPs and ionic silver (AgNO3). A high overlap of differently abundant proteins was found in root after exposure to both treatments, while in leaf, almost a half of the proteins exhibited different abundance level between treatments, indicating tissue-specific responses. Majority of the identified proteins were down-regulated in both tissues after exposure to either AgNPs or AgNO3; in roots, the most affected proteins were those involved in response to abiotic and biotic stimuli and oxidative stress, while in leaf, both treatments had the most prominent effect on photosynthesis-related proteins. However, since AgNPs induced higher suppression of protein abundance than AgNO3, we conclude that AgNP effects can, at least partially, be attributed to nanoparticle form.

BackgroundDuring the COVID-19 pandemic the Croatian Football Federation has launched a new model of pre-season systematic examination of football players, emphasizing the diagnosis of asymptomatic SARS-CoV-2 infection and preventing further spread among the players.ObjectivesThe aim of this study was to assess the prevalence and dynamics of SARS-CoV-2 IgA and IgG antibodies in the cohort of asymptomatic and SARS-CoV-2 PCR negative professional football players in the Croatian First Football League by using a commercial ELISA antibody assay in the paired serum samples taken 2 months apart.MethodsSerology testing was performed from May till July 2020 in a cohort of 305 asymptomatic football players and club staff members. RT-PCR for detection of SARS-CoV-2 from nasopharyngeal swabs was performed on three occasions, and Euroimmun ELISA for detection of IgA and IgG (S1 and NCP) antibodies was tested in paired serum samples in May and July.ResultsAll RT-PCR results were negative. Sixty-one (20%) participants were reactive in one or two classes of antibodies at baseline and/or follow-up serology testing. IgA reactivity was found in 41 (13.4% [95% CI=10.7-17.7]) baseline sera and 42 (13.8% [95% CI=10.3-18.9]) follow-up sera. IgG to S1 protein was found in 6 (2% [95% CI=0.9-4.2]) participants at baseline and 1 (0.33% [95% CI=0.0006-1.83]) at follow-up. IgG to NCP was found in 2 (0.7% [95% CI=0.2-2.4]) participants at baseline and 8 (2.6% (95% CI=1.3-5.1]) participants at follow-up. Noticeable dynamics in the paired sera was observed in 18 (5.9%) participants (excluding borderline IgA results) or 32 (10.5%) (including IgA borderline results).ConclusionVarious patterns of IgA and IgG reactivity were found in the paired serum samples. Based on serology dynamics we estimate that in 5.9%-10.5% of PCR negative football players asymptomatic exposure to SARS-CoV-2 during pandemics could not be excluded.

The aim of the study was to develop fast and accurate method for assessment of intracellular level of cadmium (Cd) and thallium (Tl), and to establish accumulation of the metals in the cells. HepG2 cells were treated with Cd or Tl (1.0 or 10.0 mg/L; 24 h) and level of Cd or Tl was assessed. ICP-MS was applied and the method was optimized and validated. Correlation coefficient (R2) for Cd was 0.9999 with intercept 0.0732 while for Tl was 1.00009 with intercept −0.1497, and limit of detection (LOD) for Cd was 0.020 μg/L and for Tl 0.097 μg/L. Both metals, Cd and Tl, accumulate in the cells in concentration-dependent manner. However, higher uptake of Cd in comparison to Tl was observed. Cells treated with the same concentration of the metal (1.0 mg/L) accumulated 10.0% of Cd and 1.0% of Tl. Higher uptake of Cd than Tl can explain higher toxicity of Cd toward HepG2 cells. Obtained results imply to the importance of monitoring the level of metals in the cells in order to connect changes at the molecular level with exposure to specific metal.

The small size of nanoparticles (NPs), with dimensions between 1 and 100 nm, results in unique chemical and physical characteristics, which is why they are implemented in various consumer products. Therefore, an important concern is the potential detrimental impact of NPs on the environment. As plants are a vital part of ecosystem, investigation of the phytotoxic effects of NPs is particularly interesting. This study investigated the potential phytotoxicity of silver nanoparticles (AgNPs) on tobacco (Nicotiana tabacum) plants and compared it with the effects of the same AgNO3 concentrations. Accumulation of silver in roots and leaves was equally efficient after both AgNP and AgNO3 treatment, with predominant Ag levels found in the roots. Exposure to AgNPs did not result in elevated values of oxidative stress parameters either in roots or in leaves, while AgNO3 induced oxidative stress in both plant tissues. In the presence of both AgNPs and AgNO3, root meristem cells became highly vacuolated, which indicates that vacuoles might be the primary storage target for accumulated Ag. Direct AgNP uptake by root cells was confirmed. Leaf ultrastructural studies revealed changes mainly in the size of chloroplasts of AgNP-treated and AgNO3- treated plants. All of these findings indicate that nano form of silver is less toxic to tobacco plants than silver ions.

Background and Purpose : Recombinant DNA technology has allowed expression of different heterologous proteins in many host systems, ranging from prokaryotic to eukaryotic organisms. Therapeutic properties of recombinant proteins are very often affected by the composition and heterogeneity of their glycans. Conventional expression systems for recombinant pharmaceuticals typically do not address this problem and result with products that contain a mixture of glycoforms that are neither identical to human glycans nor optimized for enhanced biological activity. Over the last decade plants have been developed as production platforms for recombinant proteins with pharmaceutical or industrial applications. Namely, plant expression systems contain very small differences in the post-translational modifications, mainly glycosylation, which can partly be overpowered by glycoengineering, whose goal is production of recombinant proteins with highly homogenous glycosylation that closely resembles the native system. This review attempts to present current accomplishments in the production of plant-derived glycoconjugates with humanized N- and O-glycans. Materials and Methods: Main goal of N-glycoengineering is to reduce or eliminate plant-specific N-glycans, and at the same time to introduce mammalian-specific N-glycans through the several approaches. The easiest way is to change intracellular targeting of plant-made recombinant proteins and to ensure their retention in the ER; next approach is to eliminate the addition of plant-specific glycans; while the final step is engineering the plant glycosylation pathway to introduce mammalian glycotransferases into plants with generation of biantennary and multi-antennary structures on complex N-glycans. Due to significant differences in O-glycosylation between humans and plants, different approaches to engineering of O-glycosylation have been taken. Besides having their typical O-glycoslyation on Hyp-residues, plants in general miss the machinery for production of mammalian-type O-glycosylation. Attempts have been made to mimic mammalian O-glycosylation in plants, specifically the mucin-type addition of GalNAc residues. Result: Efficient generation of bisected tetraantennary complex N-glycans without typical plant glycoepitopes on human erythropoietin (hEPO) and human transferrin (hTF) was obtained in Nicotiana benthamiana plants, thus demonstrating generation of recombinant proteins with human-type N-glycosylation at great uniformity. As for the O-glycosylation, attempts to produce mucin-type O-GalNAc and disialylated core 1 O-linked glycan structures on hEPO in N. benthamiana transgenic plants proved to be successful. Moreover, although small amounts of Hyp residues were found on recombinant EPO, no plant-specific O-glycans were detected, which demonstrates that plants are eligible candidates for production of recombinant therapeutics with fully humanized O- and N-glycans. Conclusion : Plants and methods of plant molecular farming offer a powerful expression platform for the production of a variety of recombinant proteins, which show similar, or even higher, biological activity then protein or native homologs in cultured mammalian cells currently used for large-scale production.

Since silver nanoparticles (AgNPs) are a dominant nanomaterial in consumer products, there is growing concern about their impact on the environment. Although numerous studies on the effects of AgNPs on living organisms have been conducted, the interaction of AgNPs with plants has not been fully clarified. To reveal the plant mechanisms activated after exposure to AgNPs and to differentiate between effects specific to nanoparticles and ionic silver, we investigated the physiological, ultrastructural and proteomic changes in seedlings of tobacco (Nicotiana tabacum) exposed to commercial AgNPs and ionic silver (AgNO3) from the seed stage. A higher Ag content was measured in seedlings exposed to AgNPs than in those exposed to the same concentration of AgNO3. However, the results on oxidative stress parameters obtained revealed that, in general, higher toxicity was recorded in AgNO3-treated seedlings than in those exposed to nanosilver. Ultrastructural analysis of root cells confirmed the presence of silver in the form of nanoparticles, which may explain the lower toxicity of AgNPs. However, the ultrastructural changes of chloroplasts as well as proteomic study showed that both AgNPs and AgNO3 can affect photosynthesis. Moreover, the majority of the proteins involved in the primary metabolism were up-regulated after both types of treatments, indicating that enhanced energy production, which can be used to reinforce defensive mechanisms, enables plants to cope with silver-induced toxicity.

Cilj: U ovom istraživanju uzorci cvjetnoga meda korišteni su kao bio-pokazatelji onečišćenja okoliša pesticidima. Materijal i metode: Ukupno je prikupljeno 40 (4x10) uzoraka sa 4 različite prostorne lokacije (subregije), u ukupno 16 općina, od proizvođača meda u Varaždinskoj županiji, kao i 20 (2x10) uzoraka s dvije različite lokacije proizvođača u susjednoj županiji, registriranih kao "ekoloških proizvođači", a udaljenih od poljoprivrednih površina pod konvencionalnom proizvodnjom. Analize su provedene korištenjem vezanog sustava plinske kromatografije i spektrometrije masa (GCMS) i tekućinskom kromatografijom visoke djelotvornosti (HPLC). Rezultati: Identificirano je i kvantificirano 78 aktivnih sastojaka pesticide, koji su dio od ukupno cca 400 pesticida koji se analiziraju u voću i povrću, a za koje je laboratorij raspolagao analitičkom opremom koja i dijelom određuje i vrstu i broj pesticida koje se detektiraju. Rezultati analiza ukazali su na mogući uzročno posljedični odnos između konvencionalnih poljoprivrednih aktivnosti u Varaždinskoj županiji s identificiranim i kvantificiranim aktivnim sastojcima pesticida. Iz rezultata dobivenih na udjelima identificiranih i kvantificiranih ostataka pesticida, s obzirom na dopustivost, vidljivo je da je udio dopuštenih pesticida u uzorcima cvjetnoga meda iznosio 55,2%, dok je udio ilegalnih pesticida iznosio 44,8%. Analizirajući broj otkrivenih insekticida i fungicida, ukupno na sve 4 lokacije utvrđeno je da cvjetni med sadrži 13 različitih vrsta insekticida i 3 vrste fungicida. Zaključak: Rezultati i statistička analiza podataka pokazuju da je cvjetni med dobar bio-indikator onečišćenja okoliša pesticidima, te stoga ova studija može poslužiti kao osnova za znanstvenike i stručnjake koji se bave istraživanjem štetnih učinaka pesticida na okoliš i ljudsko zdravlje., Aim: In this research samples of floral honey were used as bio-indicators of environmental pollution by pesticides. Material and methods: A total of 40 (4x10) samples from bee farms were collected at 4 different locations (sub-regions) including 16 municipalities in the Varaždin County, as well as 20 (2x10) samples from two locations – bee-farms registered as "ecological producers" in the neighborhood County, both located far away from conventional agricultural production. Analyses were carried out using the gas chromatography joint system with mass spectrometry (GCMS) and high-performance liquid chromatography (HPLC). Results: Identification and quantification of 78 pesticide active ingredients were performed which are part of a total of approximately 400 pesticides that are usually analyzed in fruits and vegetables whereby the laboratory had available analytical equipment that also determined the number of detected pesticides. Due to identified and quantified active ingredients of pesticides, the results of analyses might indicate a possible cause-effect relationship with conventional agricultural activities in the Varaždin County. It was evident from the obtained results on the shares of identified and quantified pesticide residues, and with respect to admissibility, that the share of permitted pesticides in the samples of floral honey accounted for 55.2%, while the share of illegal pesticides accounted for 44.8%. Analyzing the number of detected insecticides and fungicides, on a total of all 4 locations (sub-regions), it was found that floral honey contained 13 different types of insecticides and 3 types of fungicides. Conclusion: The results and statistical data analysis showed that floral honey is a good bio-indicator of environmental pollution by pesticides, therefore this study can serve as a basis for scientists and experts engaged in the research of harmful effects of pesticides on the environment and human health.

Acrylamide is a monomer widely used as an intermediate in the production of organic chemicals, e.g. polyacrylamides (PAMs). Since PAMs are low cost chemicals with applications in various industries and waste- and drinking water treatment, a certain amount of non-polymerised acrylamide is expected to end up in waterways. PAMs are non-toxic but acrylamide induces neurotoxic effects in humans and genotoxic, reproductive, and carcinogenic effects in laboratory animals. In order to evaluate the effect of acrylamide on freshwater organisms, bioassays were conducted on four species: algae Desmodesmus subspicatus and Pseudokirchneriella subcapitata, duckweed Lemna minor and water flea Daphnia magna according to ISO (International Organization for Standardisation) standardised methods. This approach ensures the evaluation of acrylamide toxicity on organisms with different levels of organisation and the comparability of results, and it examines the value of using a battery of low-cost standardised bioassays in the monitoring of pollution and contamination of aquatic ecosystems. These results showed that EC50 values were lower for Desmodesmus subspicatus and Pseudokirchneriella subcapitata than for Daphnia magna and Lemna minor, which suggests an increased sensitivity of algae to acrylamide. According to the toxic unit approach, the values estimated by the Lemna minor and Daphnia magna bioassays, classify acrylamide as slightly toxic (TU=0-1; Class 1). The results obtained from algal bioassays (Desmodesmus subspicatus and Pseudokirchneriella subcapitata) revealed the toxic effect of acrylamide (TU=1-10; Class 2) on these organisms.

The proliferation of silver nanoparticle (AgNP) production and use owing to their antimicrobial properties justifies the need to examine the resulting environmental impacts. The discharge of biocidal nanoparticles to water bodies may pose a threat to aquatic species. This study evaluated the effects of citrate-coated AgNPs on the standardized test organism Daphnia magna Straus clone MBP996 by means of biochemical biomarker response. AgNP toxicity was compared against the toxic effect of Ag(+). The toxicity endpoints were calculated based upon measured Ag concentrations in exposure media. For AgNPs, the NOAEC and LOAEC values at 48 h were 5 and 7 μg Ag/L, respectively, while these values were 0.5 and 1 μg Ag/L, respectively, for Ag(+). The EC50 at 48 h was computed to be 12.4 ± 0.6 and 2.6 ± 0.1 μg Ag/L for AgNPs and Ag(+), respectively, with 95 % confidence intervals of 12.1-12.8 and 2.3-2.8 μg Ag/L, respectively. These results indicate significant less toxicity of AgNP compared to free Ag(+) ions. Five biomarkers were evaluated in Daphnia magna neonates after acute exposure to Ag(+) or AgNPs, including glutathione (GSH) level, reactive oxygen species (ROS) content, and catalase (CAT), acetylcholinesterase (AChE), and superoxide dismutase (SOD) activity. AgNPs induced toxicity and oxidative stress responses in D. magna neonates at tenfold higher concentrations than Ag. Biochemical methods revealed a clear increase in AChE activity, decreased ROS level, increased GSH level and CAT activity, but no significant changes in SOD activity. As Ag(+) may dissolve from AgNPs, these two types of Ag could act synergistically and produce a greater toxic response. The observed remarkably high toxicity of AgNPs (in the parts-per-billion range) to crustaceans indicates that these organisms are a vulnerable link in the aquatic food chain with regard to contamination by nanosilver. Graphical Abstract ᅟ.

Metals have a variety of negative outcomes on plants, essential components of any ecosystem. The effects of CdCl2 (5 μmol L-1), ZnCl2 (25 or 50 μmol L-1), and CuCl2 (2.5 or 5 μmol L-1) and combinations of CdCl2 with either ZnCl2 or CuCl2 on the growth, photosynthetic pigments, and photosystem II (PSII) efficiency of duckweed (Lemna minor L.) were investigated. All of the treatments caused growth inhibition and remarkable metal accumulation in plant tissue after 4 and 7 days. In the combined treatments, the accumulation of each metal applied was lesser in comparison to treatments with single metals. After 4 days, all of the treatments generally diminished chlorophyll a content and decreased the maximum quantum yield (Fv/Fm) and effective quantum yield (ΔF/F’m) of PSII. However, after 7 days of exposure to a combination of Cd and Zn, pigment content and PSII activity recovered to control levels. A higher concentration of Cu (5 μmol L-1) as well as Cd in combination with Cu had a prolonged inhibitory effect on photosynthetic features. Our results suggest that growth inhibition was due to the toxic effect of absolute metal quantity in plant tissue. Zn counteracted Cd uptake, as seen from the recovery of pigment content and PSII efficiency in plants exposed for 7 days to the Cd and Zn combination. Cu-induced oxidative stress led to a prolonged inhibitory effect in plants treated both with a higher concentration of Cu (5 μmol L-1) and simultaneously with Cd and Cu. Our findings could contribute to general knowledge on anthropogenic and environmental contaminants that endanger plant communities and significantly disrupt the sensitive balance of an ecosystem by influencing photosynthetic mechanisms.

Sulfonamides, macrolides, torasemide, fumagillin and chloramphenicol were simultaneously analyzed in surface water samples by using solid-phase extraction (SPE) and reversed phase (RP) liquid chromatography- electrospray tandem mass spectrometry (LC-ESI- MS/MS). In the pre-concentration and clean-up process the pH value of samples and volume of the solvent for extraction of analytes from cartridge were optimized. Extraction recoveries were high with values in the range from 62 to 115 %. Limits of quantification (LoQ) were in the range from 0.02 g L–1 to 0.2 g L–1. Repeatability of the method was evaluated at LoQ and expressed as relative standard deviation (RSD). Calculated RSDs were low with values in the range from 2.4 to 14.5 %. The method was successfully applied for analysis of surface water real samples. Samples were collected along the rivers in Croatia on 4 sampling sites in 2012 in Danube catchment areas, 19 sampling sites in Danube and Adriatic catchment areas in 2013, and another 19 places in 2014. Altogether, 20 target compounds were analysed in 164 water samples and detected in 24 samples in range (0.1–5.3) g L–1 or in 14.6 % of samples. The most frequent and highest concentrations were detected for macrolide antibiotics. This is the first attempt of such monitoring in surface waters in Croatia.

Acrylamide is a monomer widely used as an intermediate in the production of organic chemicals, e.g. polyacrylamides (PAMs). Since PAMs are low cost chemicals with applications in various industries and waste- and drinking water treatment, a certain amount of non-polymerised acrylamide is expected to end up in waterways. PAMs are non-toxic but acrylamide induces neurotoxic effects in humans and genotoxic, reproductive, and carcinogenic effects in laboratory animals. In order to evaluate the effect of acrylamide on freshwater organisms, bioassays were conducted on four species: algae Desmodesmus subspicatus and Pseudokirchneriella subcapitata, duckweed Lemna minor and water flea Daphnia magna according to ISO (International Organization for Standardisation) standardised methods. This approach ensures the evaluation of acrylamide toxicity on organisms with different levels of organisation and the comparability of results, and it examines the value of using a battery of low-cost standardised bioassays in the monitoring of pollution and contamination of aquatic ecosystems. These results showed that EC50 values were lower for Desmodesmus subspicatus and Pseudokirchneriella subcapitata than for Daphnia magna and Lemna minor, which suggests an increased sensitivity of algae to acrylamide. According to the toxic unit approach, the values estimated by the Lemna minor and Daphnia magna bioassays, classify acrylamide as slightly toxic (TU=0-1; Class 1). The results obtained from algal bioassays (Desmodesmus subspicatus and Pseudokirchneriella subcapitata) revealed the toxic effect of acrylamide (TU=1-10; Class 2) on these organisms.

Metals have a variety of negative outcomes on plants, essential components of any ecosystem. The effects of CdCl2 (5 μmol L-1), ZnCl2 (25 or 50 μmol L-1), and CuCl2 (2.5 or 5 μmol L-1) and combinations of CdCl2 with either ZnCl2 or CuCl2 on the growth, photosynthetic pigments, and photosystem II (PSII) efficiency of duckweed (Lemna minor L.) were investigated. All of the treatments caused growth inhibition and remarkable metal accumulation in plant tissue after 4 and 7 days. In the combined treatments, the accumulation of each metal applied was lesser in comparison to treatments with single metals. After 4 days, all of the treatments generally diminished chlorophyll a content and decreased the maximum quantum yield (Fv/Fm) and effective quantum yield (ΔF/F’m) of PSII. However, after 7 days of exposure to a combination of Cd and Zn, pigment content and PSII activity recovered to control levels. A higher concentration of Cu (5 μmol L-1) as well as Cd in combination with Cu had a prolonged inhibitory effect on photosynthetic features. Our results suggest that growth inhibition was due to the toxic effect of absolute metal quantity in plant tissue. Zn counteracted Cd uptake, as seen from the recovery of pigment content and PSII efficiency in plants exposed for 7 days to the Cd and Zn combination. Cu-induced oxidative stress led to a prolonged inhibitory effect in plants treated both with a higher concentration of Cu (5 μmol L-1) and simultaneously with Cd and Cu. Our findings could contribute to general knowledge on anthropogenic and environmental contaminants that endanger plant communities and significantly disrupt the sensitive balance of an ecosystem by influencing photosynthetic mechanisms., Izloženost metalima može izazvati različite štetne učinke u biljaka. Vodene leće izložili smo solima teških metala CdCl2 (5 μmol L 1), ZnCl2 (25 μmol L 1 ili 50 μmol L 1) i CuCl2 (2,5 μmol L 1 ili 5 μmol L 1) te kombinaciji CdCl2 sa svakom od navedenih koncentracija ZnCl2 i CuCl2. Rast biljaka, količina fotosintetskih pigmenata i učinkovitost fotosistema II (PSII) mjereni su nakon četiri i sedam dana tretmana. Utvrđeno je da su svi tretmani uzrokovali značajnu inhibiciju rasta te akumulaciju metala u biljci. U biljaka koje su bile izložene kombinacijama teških metala količina pojedinog metala u tkivu bila je niža u odnosu na količinu istog metala u biljaka izloženih samo tom metalu. Nakon četiri dana tretmana sva su tri metala, neovisno o tome jesu li bila primijenjena zasebno ili u kombinacijama, uzrokovala smanjenje količine klorofila a i pad vrijednosti maksimalnog (Fv/Fm) i efektivnog (ΔF/F’m) prinosa PSII. Međutim, u biljaka koje su bile istovremeno izlagane kadmiju i cinku, vrijednosti količine pigmenata i učinkovitost PSII vratile su se nakon sedam dana na kontrolnu razinu, a bakar u koncentraciji 5 μmol L 1 te kombinacija kadmija i bakra i dalje su imali inhibitorni učinak. Budući da smanjeno primanje pojedinog metala uočeno u biljaka izloženih kombiniranim tretmanima nije ublažilo inhibitorni učinak na rast, možemo zaključiti da je inhibicija rasta uzrokovana apsolutnom količinom metala primljenog u tkivo. Povećanje količine fotosintetskih pigmenata i učinkovitosti PSII nakon sedam dana tretmana kadmijem i cinkom upućuje na oporavak biljaka, što se može objasniti ublažavajućim djelovanjem cinka na učinak kadmija uslijed smanjenog primanja kadmija u biljku. Suprotno tome, dugotrajni inhibitorni učinak istovremenog tretmana biljaka kadmijem i bakrom te samim bakrom u koncentraciji 5 μmol L 1 može se objasniti oksidacijskim stresom uzrokovanim bakrom. Rezultati ovoga istraživanja pridonose saznanjima o štetnim učincima antropogenih i okolišnih onečišćivača, koji narušavanjem mehanizama fotosinteze ugrožavaju ne samo biljke i njihove zajednice nego i osjetljivu ravnotežu ekosustava.

Acrylamide is a monomer widely used as an intermediate in the production of organic chemicals, e.g. polyacrylamides (PAMs). Since PAMs are low cost chemicals with applications in various industries and waste- and drinking water treatment, a certain amount of non-polymerised acrylamide is expected to end up in waterways. PAMs are non-toxic but acrylamide induces neurotoxic effects in humans and genotoxic, reproductive, and carcinogenic effects in laboratory animals. In order to evaluate the effect of acrylamide on freshwater organisms, bioassays were conducted on four species: algae Desmodesmus subspicatus and Pseudokirchneriella subcapitata, duckweed Lemna minor and water flea Daphnia magna according to ISO (International Organization for Standardisation) standardised methods. This approach ensures the evaluation of acrylamide toxicity on organisms with different levels of organisation and the comparability of results, and it examines the value of using a battery of low-cost standardised bioassays in the monitoring of pollution and contamination of aquatic ecosystems. These results showed that EC50 values were lower for Desmodesmus subspicatus and Pseudokirchneriella subcapitata than for Daphnia magna and Lemna minor, which suggests an increased sensitivity of algae to acrylamide. According to the toxic unit approach, the values estimated by the Lemna minor and Daphnia magna bioassays, classify acrylamide as slightly toxic (TU=0-1; Class 1). The results obtained from algal bioassays (Desmodesmus subspicatus and Pseudokirchneriella subcapitata) revealed the toxic effect of acrylamide (TU=1-10; Class 2) on these organisms., Akrilamid je monomer koji se koristi kao intermedijer u proizvodnji organskih kemikalija, npr. poliakrilamida (PAM-a). PAM se primjenjuje u različitim industrijama te u obradi otpadnih voda i tretmanu vode za piće, a prihvatljiv je i zbog niske cijene. Široka upotreba PAM-a u spomenute svrhe predstavlja rizik od toga da određena količina nepolimeriziranog akrilamida dospije u vodene ekosustave. PAM je netoksičan, ali akrilamid ima neurotoksične učinke u ljudi te je također dokazana njegova genotoksičnost, kancerogenost i štetan utjecaj na reprodukcijski sustav u laboratorijskih životinja. U cilju procjene toksičnosti akrilamida provedeni su biotestovi na četirima vrstama slatkovodnih organizama: na zelenim algama Desmodesmus subspicatus i Pseudokirchneriella subcapitata, vodenoj leći Lemna minor i vodenbuhi Daphnia magna. Biotestovi su provedeni prema standardiziranim ISO metodama, što osigurava procjenu toksičnosti akrilamida na organizme različitog stupnja organizacije, usporedivost rezultata i procjenu pogodnosti korištenja niza standardiziranih biotestova prihvatljive cijene za praćenje zagađenja i onečišćenja vodnih ekosustava. Rezultati su pokazali da su EC50 vrijednosti niže nakon izlaganja zelenih algi Desmodesmus subspicatus i Pseudokirchneriella subcapitata akrilamidu u odnosu na vrijednosti dobivene za vodenu leću Lemna minor i vodenbuhu Daphnia magna, što pokazuje veću osjetljivost algi. S obzirom na klasifikacijski sustav za izražavanje stupnja toksičnosti, a na temelju vrijednosti utvrđenih za vodenu leću Lemna minor i vodenbuhu Daphnia magna, akrilamid se može klasificirati kao slabo toksičan (TU=0-1; razred 1). Rezultati dobiveni testovima na algama (Desmodesmus subspicatus i Pseudokirchneriella subspicata) dokazuju toksičan učinak akrilamida na tim organizmima (TU=1-10; razred 2).

Metals have a variety of negative outcomes on plants, essential components of any ecosystem. The effects of CdCl2 (5 μmol L-1), ZnCl2 (25 or 50 μmol L-1), and CuCl2 (2.5 or 5 μmol L-1) and combinations of CdCl2 with either ZnCl2 or CuCl2 on the growth, photosynthetic pigments, and photosystem II (PSII) efficiency of duckweed (Lemna minor L.) were investigated. All of the treatments caused growth inhibition and remarkable metal accumulation in plant tissue after 4 and 7 days. In the combined treatments, the accumulation of each metal applied was lesser in comparison to treatments with single metals. After 4 days, all of the treatments generally diminished chlorophyll a content and decreased the maximum quantum yield (Fv/Fm) and effective quantum yield (ΔF/F'm) of PSII. However, after 7 days of exposure to a combination of Cd and Zn, pigment content and PSII activity recovered to control levels. A higher concentration of Cu (5 μmol L-1) as well as Cd in combination with Cu had a prolonged inhibitory effect on photosynthetic features. Our results suggest that growth inhibition was due to the toxic effect of absolute metal quantity in plant tissue. Zn counteracted Cd uptake, as seen from the recovery of pigment content and PSII efficiency in plants exposed for 7 days to the Cd and Zn combination. Cu-induced oxidative stress led to a prolonged inhibitory effect in plants treated both with a higher concentration of Cu (5 μmol L-1) and simultaneously with Cd and Cu. Our findings could contribute to general knowledge on anthropogenic and environmental contaminants that endanger plant communities and significantly disrupt the sensitive balance of an ecosystem by influencing photosynthetic mechanisms.

Silver nanoparticles (AgNPs) are the dominating nanomaterial in various consumer products. To enhance nanoparticle useful properties, different surface coatings are used, which affect physico-chemical characteristics of AgNPs. As plants can have a significant role in accumulation and biodistribution of many environmentally released substances, they are also very likely to be influenced by AgNPs, serving as a potential pathway for transport and bioaccumulation of AgNPs into food chains. In this study we investigated the phytotoxicity of AgNO3 and three types of laboratory-synthesized AgNPs with different surface coatings [citrate, polyvinylpyrrolidone (PVP) and cetyltrimethylammonium bromide (CTAB)] on tobacco (Nicotiana tabacum L.), agriculturally interesting and frequently used model plant. In vitro grown 3-months old plants were treated with 25, 50, 75, 100 and 150 microM of AgNO3, AgNPs-citrate, AgNPs-PVP and AgNPs-CTAB. After seven days chlorophyll fluorescence parameters, concentration of photosynthetic pigments and Ag uptake were determined. Treatments with AgNO3 and all types of AgNPs resulted with the increased Ag uptake in the leaves. Positively charged AgNPs-CTAB lowered most parameters of chlorophyll fluorescence, while at higher concentrations decreased the concentration of photosynthetic pigments, including chlorophylls, thus indicating strong negative influence on photosynthetic apparatus. Treatments with AgNO3 and AgNPs-PVP, NPs with the low negative charge, decreased the non-photochemical quenching and negatively influenced majority of photosynthetic pigments, but only at higher concentrations. Negatively charged AgNPs-citrate exhibited the weakest impact by only increasing the non-photochemical quenching and concentration of lutein. We can conclude that the phytotoxicity of AgNPs is correlated with their surface coating and overall surface charge.

Honey is used for nutritional, medicinal and industrial purposes and antibiotic residues may harm its quality and constitute a danger to human health. The broad spectrum antibiotic chloramphenicol (CAP) was used for curative purposes in veterinary medicine, but is now forbidden in European Union (EU) because of its many serious side effects (e.g. aplastic anaemia, grey syndrome, severe bone marrow depression and hypersensitivity). The aim of this study was to facilitate analyses of the quality and safety of Croatian honey distributed to whole European Union market; an assessment that has not previously been made.CAP in honey was qualifying and quantifying by validated liquid chromatography tandem mass spectrometry with negative electrospray ionisation method (LC-MS/MS). The target antibiotic was separated on chromatographic column Zorbax SB C18 (150 mm x 2.1 mm, 3.5 μm) with a gradient elution using acetonitrile - 0.1% formic acid mobile phase at a flow rate of 0.3 mL/min, with column temperature 35°C for CAP and 5D-CAP as internal standard. Homogenised honey samples were diluted with acetate buffer solution and extracted on Oasis Hydrophilic-Lipophilic-Balanced (HLB) sorbents. The method was used to analyse 280 domestic honey samples collected throughout Croatia between 2005.-2013.Recoveries of the method for real (acacia, chestnut, linden and flower) honey samples were 102% with RSD 8.4%. The value CCα and CCβ were 0.09 and 0.12 μg/kg, respectively. Results showed only three subsequent positive detections (1.1%) of CAP in honey.Analysed honey samples from Croatia showed good quality and safety what is the one of the main objective in consumer health policy in EU.

Discharge of silver nanoparticles (AgNP) from textiles and cosmetics, todays major application areas for metallic AgNP, into wastewater is inevitable. Transformation in wastewater treatment plants (WWTP) will determine the impact of AgNP on aquatic and terrestrial environments. This study aimed to determine the behavior of AgNP in the sludge at the WWTP by adressing the stability of citrate- coated AgNPs and their interactions with laundry-relevant surfactants (sodium dodecyl sulphate (SDS) and dodecylammonium chloride (DDACl)). The stability of the AgNPs was shown to depend on the charge and concentration of the adsorbed surfactants. The presence of DDACl prevented the formation of AgNP agglomerates even at higher ionic strength. In order to determine the effect of AgNP on biodegradation of laundry surfactants, we employed standardized method for water quality and HRN- EN-ISO-10707:2000 (Evaluation in an aqueous medium of the "ultimate" aerobic biodegradability of organic compounds). Additionally, we evaluated the inhibitory effect of AgNPs on the growth of activated sludge microorganisms (ASMs)). Results showed that AgNPs were toxic to ASMs at very high concentration (>50 mg/L). However, biodegradability of SDS, DDACl and commercial laundry detergent were inhibited by more than 5% and 20% in the presence of 0.1 and 1 mg/L AgNP, respectively. It can be concluded that AgNP discharged to the wastewater stream will influenced overall biodegradability of organic matter at various degrees in the sewer system. Such knowledge is important as it may influence the subsequent transport of AgNPs through different chemical transients and thus their potential bioavailability and toxicity.

The objective of the present study was to investigate the effects of cadmium-zinc (Cd-Zn) interactions on their uptake, oxidative damage of cell macromolecules (lipids, proteins, DNA) and activities of antioxidative enzymes in tobacco seedlings as well as roots and leaves of adult plants. Seedlings and plants were exposed to Cd (10 µM and 15 µM) and Zn (25 µM and 50 µM) as well as their combinations (10 µM or 15 µM Cd with either 25 µM or 50 µM Zn). Measurement of metal accumulation exhibited that Zn had mostly positive effect on Cd uptake in roots and seedlings, while Cd had antagonistic effect on Zn uptake in leaves and roots. According to examined oxidative stress parameters, in seedlings and roots individual Cd treatments induced oxidative damage, which was less prominent in combined treatments, indicating that the presence of Zn alleviates oxidative stress. However, DNA damage found in seedlings, and lower glutathione reductase (GR) and superoxide dismutase (SOD) activity recorded in both seedlings and roots, after individual Zn treatments, indicate that Zn accumulation could impose toxic effects. In leaves, oxidative stress was found after exposure to Cd either alone or in combination with Zn, thus implying that in this tissue Zn did not have alleviating effects. In conclusion, results obtained in different tobacco tissues suggest tissue-dependent Cd-Zn interactions, which resulted in activation of different mechanisms involved in the protection against metal stress.

Food supplements adulterated with analogues pose a pharmacological and toxicological health risk. The scale at which erectile dysfunction (ED) medicines are obtained outside of the official health system rivals and possibly exceeds legitimate sales. The legitimate health systems were dominated by the EMA and FDA approved ED medicines ; Viagra® (sildenafil), Cialis® (tadalafil) and Stendra® (avanafil). According to literature a high-risk segment of this market is occupied by adulterated food supplements. We studied food supplements (capsules, pastes and drinks) purchased from Croatian market or as a row material on sildenafil, tadalafil and avanafil content. Substances were determined by high-pressure liquid chromatography-electrospray tandem mass spectrometry (LC-ESI-MS-MS). For each substance two transitions was monitored, using protonated molecules as precursor ions and sildenafil D8 as internal standard. The drugs were isolated from food remedies using simple methanol extraction. The target drug was separated using reversed-phase liquid chromatography on chromatographic column Zorbax SB C18 (150 mm × 2.1 mm, 3.5 μm) with an isocratic elution using acetonitrile – 0.1% formic acid mobile phase at a flow rate of 0.2 mL/min. 59 samples were analysed in period from 2012 till 2014. 46% of samples were collected by sanitary inspection and others by private companies. The study showed that sildenafil and tadalafil were detected in 12 samples in range (

The effects of 10 μmol L-1 and 15 μmol L-1 cadmium (Cd), a nonessential toxic element and 25 μmol L-1 and 50 μmol L-1 zinc (Zn), an essential micronutrient, on proteins and glycoproteins of Nicotiana tabacum L. seedlings and plants were investigated after exposure to each metal alone or to their combinations. Changes in only few polypeptides related to heavy metal treatments were observed in tobacco seedlings and leaves of adult plants, while the greatest change in total soluble protein pattern was observed in plant roots. Differences between control and treated tobacco tissues were more pronounced in the glycoprotein pattern, which was analysed by application of different lectins. The majority of the detected glycoproteins in leaves and roots of adult plants can be considered as a result of enhanced glycosylation due to heavy metal stress. The difference in glycoproteins between Cd and Zn application on tobacco seedlings and adult plants could not be determined since enhanced glycosylation was noticed after treatment with either metal alone or in combination. Therefore, it can be concluded that both metals induced N- and Oglycosylation as a result of changed environmental conditions., Ispitivali smo učinke 10 μmol L-1 i 15 μmol L-1 kadmija (Cd), neesencijalnoga toksičnog elementa i 25 μmol L-1 i 50 μmol L-1 cinka (Zn), esencijalnog mikronutrijenta, na proteine i glikoproteine u klijancima i odraslim biljkama Nicotiana tabacum L. nakon izlaganja svakomu metalu posebno ili njihovim kombinacijama. Promjene kod nekoliko polipeptida koje su uočene u klijancima i listovima odraslih biljaka nisu bile brojne, dok su one najvažnije zabilježene u uzorku ukupnih topljivih proteina u korijenu biljke. Razlike između kontrole i tretiranog tkiva duhana bile su izraženije kod glikoproteina koji su analizirani primjenom različitih lektina. Većina glikoproteina uočenih u listovima i korijenu odraslih biljaka može se smatrati rezultatom povećane glikozilacije zbog stresa koji uzrokuju teški metali. Nije bilo moguće utvrditi razliku u glikoproteinima između tretiranja klijanaca i odraslih biljaka duhana kadmijem i cinkom jer je povećana glikozilacija utvrđena i nakon tretmana svakim metalom posebno i nakon tretmana njihovim kombinacijama. Stoga se može zaključiti da su, kao rezultat promijenjenih uvjeta u okolišu, oba metala potaknula N- i O-glikozilaciju.

The effects of 10 μmol L-1 and 15 μmol L-1 cadmium (Cd), a nonessential toxic element and 25 μmol L-1 and 50 μmol L-1 zinc (Zn), an essential micronutrient, on proteins and glycoproteins of Nicotiana tabacum L. seedlings and plants were investigated after exposure to each metal alone or to their combinations. Changes in only few polypeptides related to heavy metal treatments were observed in tobacco seedlings and leaves of adult plants, while the greatest change in total soluble protein pattern was observed in plant roots. Differences between control and treated tobacco tissues were more pronounced in the glycoprotein pattern, which was analysed by application of different lectins. The majority of the detected glycoproteins in leaves and roots of adult plants can be considered as a result of enhanced glycosylation due to heavy metal stress. The difference in glycoproteins between Cd and Zn application on tobacco seedlings and adult plants could not be determined since enhanced glycosylation was noticed after treatment with either metal alone or in combination. Therefore, it can be concluded that both metals induced N- and Oglycosylation as a result of changed environmental conditions.

The effects of 5 μM cadmium (Cd), a non-essential toxic element and 25 and 50 μM zinc (Zn), an essential micronutrient, were investigated in aquatic plant Lemna minor L. after 4 and 7 days of exposure to each metal alone or to their combinations. Both metals showed tendency to accumulate with time, but when present in combination, they reduced uptake of each other. Cd treatment increased the lipid peroxidation and protein oxidation indicating appearance of oxidative stress. However, Zn supplementation in either concentration reduced values of both parameters, while exposure to Zn alone resulted in elevated level of lipid peroxidation and protein oxidation but only on the 7th day. Enhanced DNA damage, which was found on the 4th day in plants treated with Cd alone or in combination with Zn, was reduced on the 7th day in combined treatments. Higher catalase activity obtained in all treated plants on the 4th day of experiment was reduced in Zn-treated plants, but remained high in plants exposed to Cd alone or in combination with Zn after 7 days. Cd exposure resulted in higher peroxidase activity, while Zn addition prominently reduced peroxidase activity in the plants subjected to Cd stress. In conclusion, Cd induced more pronounced oxidative stress and DNA damage than Zn in applied concentrations. Combined treatments showed lower values of oxidative stress parameters— lipid peroxidation, protein oxidation and peroxidase activity as well as lower DNA damage, which indicates alleviating effect of Zn on oxidative stress in Cd-treated plants.

Effect of Copper on the Toxicity and Genotoxicity of Cadmium in Duckweed (Lemna Minor L.)We investigated interactions between copper (in the concentrations of 2.5 μmol L-1 and 5 μmol L-1) and cadmium (5 μmol L-1) in common duckweed (Lemna minor L.) by exposing it to either metal or to their combinations for four or seven days. Their uptake increased with time, but it was lower in plants treated with combinations of metals than in plants treated with either metal given alone. In separate treatments, either metal increased malondialdehyde (MDA) level and catalase and peroxidase activity. Both induced DNA damage, but copper did it only after 7 days of treatment. On day 4, the combination of cadmium and 5 μmol L-1 copper additionally increased MDA as well as catalase and peroxidase activity. In contrast, on day 7, MDA dropped in plants treated with combinations of metals, and especially with 2.5 μmol L-1 copper plus cadmium. In these plants, catalase activity was higher than in copper treated plants. Peroxidase activity increased after treatment with cadmium and 2.5 μmol L-1 copper but decreased in plants treated with cadmium and 5 μmol L-1 copper. Compared to copper alone, combinations of metals enhanced DNA damage after 4 days of treatment but it dropped on day 7. In conclusion, either metal given alone was toxic/genotoxic and caused oxidative stress. On day 4 of combined treatment, the higher copper concentration was more toxic than either metal alone. In contrast, on day 7 of combined treatment, the lower copper concentration showed lower oxidative and DNA damage. These complex interactions can not be explained by simple antagonism and/or synergism. Further studies should go in that direction.