Quantification of metallothionein in whole body gammarus locusta (crustacea: amphipoda) using differential pulse polarography

A polarographic method was optimised for the quantitative determination of metallothionein (MT) in whole body Gammarus locusta (Crustacea: Amphipoda). The identity of the MT-like proteins was examined using electrophoretic analysis (SDS-PAGE) and induction studies (water-borne exposures to copper). The MT-method was then used to examine changes in MT levels over the lifetime (juveniles, sub-adults and adults) of a laboratory population of G. locusta; copper (Cu) and zinc (Zn) body burden were also measured by flame atomic absorption spectrometry. MT was quantified by heat-denaturation of 30?000?g supernatants (to remove the bulk of non-MT proteins) followed by differential pulse polarography (DPP) to measure sulphydryl-group containing proteins. DPP proved to be a sensitive and reliable method to quantify heat-stable total sulphydryl-proteins. The majority of the latter was indicated to be MT from SDS-PAGE and induction studies. SDS-PAGE revealed a putative MT of 23?±?0.6?kDa app. mol. wt., consistent with the average size of dimeric MTs (10–20?kDa) found in several aquatic invertebrate species. Copper-exposure revealed parallel increases in the amounts of 23?kDa protein and total sulphydryl-proteins. Levels of whole body MT (rabbit pure MT as standard) range between 1.3 and 2.3?mg?g^?1 dry wt. over the lifetime of G. locusta. Sex-related changes in MT status were observed, the levels in females being 1.6 times higher than in males (p<0.05). Copper and Zn body content changed over the lifetime of G. locusta, the levels in juveniles (p>0.05) and sub-adults (p<0.05) being higher than in adults. It is concluded that the inclusion of MT in ecotoxicological studies with amphipod crustaceans require some caution with respect to the stages of moulting and/or sexual maturation of organisms, but this should not provide any major impediment in its use in metal pollution monitoring.     

Distribution and speciation of heavy metals in Apulian coastal sediments

In the framework of the Interreg II Project Italy–Greece, distribution and speciation of trace metals in surface coastal sediments of Apulian regions were studied. A sequential extraction technique was used to determine the chemical association of heavy metals (Zn, Pb, Ni, Fe, Cr, Mn, Cu and Cd) with major sedimentary phases (acid soluble phases, easily reducible fraction and oxidizable matter). The relationship between the metals' content and the grain-size composition of the sediments was also considered: generally the elements resulted associated with the fine fraction, even if in some cases (Cu in the oxidizable phase) the opposite behaviour was observed. The studied area is not particularly polluted, except for some defined zones, as for example Brindisi's harbour. Speciation analysis revealed that the major part of the heavy metals are not easily mobilizable and bioavailable. Even in this case, there are some exceptions to the general behaviour: Cu and Pb in several samples are largely present in the oxidizable phase, which even if not immediately available can become a source of associated elements when redox conditions change.     

Royal jelly inhibited N-acetylation and metabolism of 2-aminofluorene in human liver tumor cells (J5)

The present study was carried out to evaluate the question of whether or not royal jelly affects N-acetylation and metabolism of 2-aminofluorene (2-AF) in the human liver tumor cell line (J 5). N-acetylation and metabolism of 2-AF in intact J5 cells was determined by using high performance liquid chromatography for the amounts of acetylated and nonacetylated 2-AF and profile of 2-AF metabolism. The results indicated that royal jelly displayed a dose-dependent inhibition of N-acetylation of 2-AF in J5 cells. Royal jelly also decreased the profile of 2-AF metabolites in J5 cells. This report is the first demonstration which showed that royal jelly affects N-acetylation of 2-AF in human liver tumor cells (J5).     

Speciation of heavy metals in Albanian coastal sediments

This article presents results on heavy metal (Fe, Mn, Cr, Ni, Cu, Cd, Pb and Zn) speciation in the coastal sediments of Albania. Sediment samples were collected within the framework of the activities of the Interreg Italia–Albania Project, carried out in the Southern Adriatic Sea in 2000–01. This study shows that Albanian coastal sediments are highly influenced by river input, as the decreasing concentrations from coast to offshore confirm. Pb represents an exception of the general trend; meteoric inputs are prevalent for this metal. Ni, Cr, Fe, Zn and Cu correlate significantly and positively in the entire area, indicating a common origin for these metals in the analyzed sediments. All metals result mostly associated to the most refractory phases, which constitute up to 80% of the total concentration. Nevertheless a recent input of Cr, Ni, Cu and Mn can be observed, as the result of mining and industrial activities, in some selected areas, such as the Drin Bay and Durres. Their high concentration in the bioavailable phase can determine toxic effect upon biota.     

In vitro and in vivo study of neutrophil functions after acute methanol intoxication in albino rats

Toxicity from methanol (MeOH), a potentially significant problem due to occupational, accidental, or intentional, as well as daily ingestion of small amounts of the agent, only receives considerable attention after severe signs of intoxication have set in or death is imminent. While accidental and intentional exposures usually involve very high doses, the occupational and ingestion forms more often reflect small daily intakes. Still, even at the low levels, little is known about the potential immunotoxic implications (and less so in regard to mechanisms) from these ongoing exposures. This study has been attempted to focus the effects of acute methanol exposure on blood neutrophil functions in both in vitro and in vivo conditions at 30 and 60?min time intervals. Innate immunity of the host defense relies mostly on neutrophils. The blood MeOH levels were determined upon exposing the experimental animals to ¼ of LD₅₀ MeOH. No significant change in the blood methanol level was observed at 30 and 60?min of methanol exposure. The in vitro results showed a decrease in the neutrophil functions of adherence and phagocytic index without affecting the avidity index, followed by an increased nitroblue tetrazolium (NBT) reduction. The in vivo exposures of MeOH showed a decline in neutrophil adherence. However, the other neutrophil functions were found to be significantly increased, which indicates that in vivo results are different from that of the in vitro studies. The present study concludes that MeOH exposure can reconstitute the innate immune status by altering the neutrophil functions. Moreover, the in vivo effects are much pronounced compared to the in vitro effects, which may be due to the formation of the metabolites under in vivo conditions.     

Removal of selenite using ‘waste’ Fe(III)/Cr(III) hydroxide: Adsorption kinetics and isotherms

Removal of selenite [Se (IV)] from aqueous solution on to industrial solid ‘waste’ Fe(III)/Cr(III) hydroxide as adsorbent was investigated in the present article. Maximum adsorption was found to be at pH 4.0. Pretreated Fe(III)/Cr(III) hydroxide was found to be more efficient for the removal of selenite compared to untreated adsorbent. Langmuir and Freundlich isotherms have been studied. The Langmuir adsorption capacity (Q ₀) of the pretreated and untreated adsorbents was found to be 15.63 and 6.04?mg?g^?1, respectively. The adsorption process fit into the second-order kinetics. Thermodynamic parameters show that the adsorption process is spontaneous and endothermic in the temperature range 32 to 60°C. Coexisting anions vanadate and phosphate significantly affect the adsorption of selenite for both the pretreated and untreated adsorbents. Molybdate, thiocyanate, sulphate, nitrate and chloride do not significantly affect the removal of selenite for pretreated adsorbent.     

The correlation of total and extractable heavy metals from soil and domestic sewage sludge and their transfer to maize ( Zea mays L.) plants

A study to understand the mobility and transport of heavy metals (HMs) from soil and soil amended with sewage sludge to maize plants was carried out. The total and ethylenediaminetetraaceticacid (EDTA)-extractable HMs in agricultural soil and untreated domestic sewage sludge samples, and the correlation between the total and extractable metals in soil and sewage sludge were carried out. Pot experiments were performed to study the transfer of HMs to maize grains, grown in soil (control) and in soil amended with sewage sludge (test samples). The total and extractable HMs in soil, sewage sludge, and maize grains were analysed by FAAS/ETAAS (flame atomic absorption spectrometer/electrothermal atomic absorption spectrometer) after digestion in microwave oven. Statistically significant correlations were obtained between the total contents of Cu, Cd, As and their respective extractable fractions in soil, while in domestic wastewater sludge (DWS) the better correlation was observed only for Ni and Cd. The edible part of maize plants (grains) from test samples presented high concentration of Zn, Pb, Ni, Cd, Cu, As, and Cr concentrations (80.7–85.6, 3.8–3.95, 2.35–2.5, 0.75–0.82, 3.21–3.29, 0.23–0.27, and 0.22–0.29?mg?kg^?1, respectively). Good correlations were found between metals in exchangeable fractions of both soil and DWS and total metals in control and test samples of maize grains. The transfer factor of all HMs from DWS to maize grains was also determined.     

Fatty acid composition of marine macroalgae from the Black Sea and Dardanelles

The fatty acid (FA) composition was determined in 14 species of marine macroalgae belonging to the Chlorophyta, Phaeophyta and Rhodophyta, which were collected from ?ile in the Black Sea and Kepez in the Dardanelles. Generally, polyunsaturated FAs and monounsaturated FAs were major components (50–77%). Total saturated FAs ranged from 22% to 50%, with 16?:?0 as the most abundant saturate (32–38%). Two samples of Cystoseira barbata collected from a different station had some differences from each other in their contents of 18?:?2n-6 and 18?:?3n-3 and in the 18?:?2n-6/20?:?4n-6 ratios. Green algal species had a significantly higher proportion of unsaturated FAs and a significantly lower proportion of saturated FAs than the red and brown algae. The amount of n-3 FAs was significantly higher in Ulva rigita, Chaetomorpha linum, Enteromorpha linza and Gracilaria verrucosa (8.88, 6.44, 5.31 and 5.24, respectively).     

Determination of 4-nonylphenol in carp blood samples by solid phase extraction and chromatographic techniques

In this study, selective methods were developed for isolation, purification, separation and determination of 4-nonylphenol (4NP) in biological samples. Several methods such as high-performance liquid chromatography (HPLC) with fluorescence and diode array detection and gas chromatography coupled to mass spectromotry (GC-MS) are described for the simultaneous determination of 4NP. The nonylphenols in the plasma of fish were extracted through solid phase process by using octadecyl, polymeric and octadecyl endcapping sorbents. The average recoveries in plasma samples spiked with 4NP of levels 1?µg?mL^?1 were between 80–95% using added surrogate standards. The aim of this study was to determine whether trace amounts of 4NP in fish plasma samples could be detected by solid phase extraction and chromatographic methods (HPLC, GC-MS). This technique of monitoring the levels of endocrine-disruptors in plasma samples is consistent, reliable as well as inexpensive.     

In vitro genotoxicity of fly ash leachate in earthworm coelomocytes

Fly ash is the major coal combustion byproduct from thermal power plants. Considering its plant–nourishing constituents, its soil amendment in farmland is one of its promoted disposal methods. A substantial amount of heavy metals present in fly ash, which may leach out due to rainwater or irrigation water, may cause serious problem with long term use, especially to soil organisms. These metals may cause DNA damage through Reactive Oxygen Species (ROS) generation. In the present study, single cell gel electrophoresis [(SCGE) i.e., comet assay] was used to detect DNA damage in earthworm (Dichogaster curgensis) coelomocytes, following an in vitro exposure. Significant DNA damage was observed at the lowest concentration of fly ash leachate (6.25%) examined. DNA damage by all the tested concentrations (6.25%, 12.5%, 25%, 50%) differed significantly (p?<?0.001) from that of the negative control. Hence, long-term application of fly ash might prove harmful for earthworm populations.