Individual and combined ecotoxic effects of water-soluble polymers.

Water-soluble polymers (WSPs) are a class of high-molecular-weight compounds which are widely used in several applications, including water treatment, food processing, and pharmaceuticals. Therefore, they pose a potential threat for water resources and aquatic ecosystems. We assessed the ecotoxicity of four WSPs-non-ionic polyacrylamide (PAM) and polyethylene glycol (PEG-200), anionic homopolymer of acrylic acid (P-AA), and cationic polyquaternium-6 (PQ-6)-as single compounds and in mixture. For this purpose in vitro and in vivo assays were used to record baseline toxicity, mutagenic potential, endocrine effects, and growth inhibition in the freshwater alga Raphidocelis subcapitata. Furthermore, the mixture toxicity of the two polymers P-AA and PQ-6 which showed effects in the algae tests was evaluated with the concentration addition (CA), independent action (IA), and generalized concentration addition (GCA) model and compared with experimental data. No toxic effects were observed among the polymers and their mixtures in the in vitro assays. On the contrary, in the growth inhibition test with R. subcapitata the cationic PQ-6 caused high inhibition while the anionic P-AA and its mixture with the cationic polymer caused low inhibition. The non-ionic polymers PEG-200 and PAM showed no effect in R. subcapitata in the tested concentration range up to 100 mg/L. The IA model represented the mixture effect of the combination experiment better than the CA and GCA models. The results indicate (1) that the toxic effects of anionic and cationic polymers are most likely due to interactions of the polymers with the surfaces of organisms or with nutrients in the water and (2) that the polymers elicit their effects through different mechanisms of action that do not interact with each other.

The effects of elevated carbon dioxide and temperature levels on tilapia (Oreochromis mossambicus): Respiratory enzymes, blood pH and hematological parameters.

Oreochromis mossambicus were exposed to two different temperature and carbon dioxide partial pressure levels for about two weeks, as the ambient (Control; 25°C, 3.3mg/L CO2), high CO2 (25°C, 14mg/L CO2), high temperature (30°C, 3mg/L CO2) and combined (30°C, 14.1mg/L CO2) groups. No mortality was observed during the experiments. As a result of the study, elevated CO2 concentrations cause negative effects on the hematological parameters. At the end of the study, while the blood Carbonic Anhydrase (CA) activity, in the high CO2 group (25°C, 14mg/L CO2), statistically increased at the 7th day compared to the control group, it decreased at the 14th day (p<0.05). In addition, the blood CA activity, in the combined (30°C, 14.1mg/L CO2) group, showed a decrease at the 14th day compared to the control group (p<0.05). At the end of study, unlike the blood CA activity, gill, liver and kidney CA activity showed an increase in the tissues compared to the control groups (p<0.05). Furthermore, the Na(+), K(+)-ATPase activities were stimulated significantly in the gills in both high CO2 and temperature groups at day 7, but it showed a significant amount of inhibition at the 14th day compared to the control groups. Overall, increasing carbon dioxide concentration in different temperatures has negative effects on the hematological parameters and respiratory enzyme of the tilapia fish. In addition, it is observed that the fish survive at negative conditions with adaptation mechanisms.

Growth performance and antioxidant enzyme activities in rainbow trout (Oncorhynchus mykiss) juveniles fed diets supplemented with sage, mint and thyme oils.

This study evaluated effects of dietary supplementation of sage (Salvia officinalis), mint (Mentha spicata) and thyme (Thymus vulgaris) oils on growth performance, lipid peroxidation level (melondialdehyde, MDA) and liver antioxidant enzyme activities (superoxide dismutase, SOD; catalase, CAT; glucose-6-phosphate dehydrogenase, G6PD; glutathione reductase, GR; glutathione-S-transferase, GST and glutathione peroxidase, GPx) in rainbow trout (Oncorhynchus mykiss) juveniles. For this purpose, triplicate groups of rainbow trout were fed daily ad libitum with diets containing sage, mint and thyme oils at 500, 1,000 and 1,500 mg kg(-1) for 60 days. While weight gain percentage of fish fed the diets containing sage and thyme oils was significantly higher than the control group, that of fish fed mint oil was the lowest. Similarly, specific growth rate was found to be the highest in all groups of the sage and thyme oil feeding and the lowest in the mint groups. Moreover, feed conversion ratio was significantly higher in the mint oil administered groups. Survival rate was also significantly reduced in the fish fed the diet containing mint oil. It was observed that SOD, G6PD and GPx activities were significantly increased in liver tissues of all the treated fish groups compared to that of control diet-fed group. However, CAT, GST and GR activities were significantly decreased in experimental diet-fed fish groups at the end of the experiment. On the other hand, a significant reduction was found in MDA levels in the fish fed the diets with sage and thyme oils compared to control and mint diets on the 30th and 60th days of experiment. Overall, dietary inclusion of sage and thyme oils is effective in enhancing rainbow trout growth, reduction in MDA and least changing antioxidant enzyme activities at a low level of 500 mg kg(-1) diet, and they can be used as important feed supplements for rainbow trout production.

The sexually dimorphic adipose fin is an androgen target tissue in the brown trout (Salmo trutta fario).

An investigation has been described on the relationship of body length, age and sex with adipose fin length and the number of androgen receptor (AR)-containing cells in the adipose fin as a secondary sexual characteristic for brown trout (Salmo trutta fario). Firstly, body and adipose fin lengths of 2- to 5-year-old brown trout were measured. Thereafter, these fish were killed by decapitation, then their sexes were determined, and adipose fins were excised. The cellular bases of AR binding activities in the adipose fins were analyzed with an antibody against human/rat AR peptide. Immunocytochemistry and western blotting techniques were performed with this antibody. Analysis of morphological measurements indicated that body length and age had a linear relationship with adipose fin length. The coefficients of determination for the body length and age were 0.92 and 0.85 in the male fish and 0.76 and 0.73 in the female fish against the adipose fin length, respectively. At 2 years of age, cells in the adipose fin did not exhibit AR immunoreactivity. However, AR-immunopositive cells were abundant in the adipose fin of 3- to 5-year-old fish. Moreover, the number of AR-immunopositive cells was significantly (P < 0.05) high in males and increased with age. These observations indicate that the adipose fin in the brown trout is a probable target for androgen action and that tissue function or development may to some extent be androgen dependent. In addition, it is likely that such an effect will be mediated by specific androgen receptors.

Metals in two species of fish in Karasu River.

In this study, cadmium (Cd), copper (Cu), iron (Fe), nickel (Ni), lead (Pb) and zinc (Zn) levels were determined in the tissues (muscle, liver and gills) of two fish species, Capoeta capoeta umbla and Chalcalburnus mosullensis, collected from three stations of the Karasu River. The lowest metal accumulation was detected in the muscle tissues. Moreover there was some variability in the metal concentrations measured in the same tissues from samples obtained from the three different stations. A positive correlation was observed for the concentration of metal pairs Fe-Cu, Fe-Zn, Ni-Pb, Pb-Zn. These findings were also compared with national and international food standards, and Pb and Cd concentrations were determined to be above the level set by the standards. In conclusion, it was supposed that excessive consumption of these two fish species, which already occurs in this region, might pose a public health risk.

Preliminary assessment of dietary mannanoligosaccharides on growth performance and health status of gilthead seabream Sparus auratus.

A feeding trial was performed to assess the potential beneficial effect of two levels of mannanoligosaccarides (MOS) on the growth performance, feed utilization, hematological parameters, and liver histopathology of gilthead seabream Sparus auratus (also known as gilthead bream). Mannanoligosaccarides were added at the rates of 2 and 4 g/kg to a fish-meal-based control diet, and each diet was given (twice daily [midmorning and midafternoon] to apparent satiation) to triplicate groups of gilthead seabream growers (mean weight = approximately 170 g) in sea cages. The trial lasted 12 weeks, and the average ambient water temperature ranged from 19.6 degrees C to 24.7 degrees C during the experimental period. At the end of the experiment, fish attained market size (350-450 g) and their health status was evaluated by blood analysis and liver histology. There were no differences in survival rates among fish fed experimental diets. However, there were significant improvements in both growth and feed utilization among fish fed diets supplemented with MOS. Hemoglobin (Hb) and hematocrit (Ht) levels and erythrocyte, leukocyte, and thrombocyte (Thr) counts were unaffected by any dietary MOS. The levels of Hb (g/dL; mean +/- SD) and Ht (%; mean +/- SD) were 11.0 +/- 2.5 and 45.6 +/- 6.7 for the control group, 11.1 +/- 1.7 and 39.3 +/- 8.0 for the 2-g/kg group, and 11.2 +/- 1.9 and 40.2 +/- 8.4 for the 4-g/kg group. The mean Thr count ranged from 47.6 to 53.8 x 10(3)/mm3. Despite the apparently higher Thr counts for fish fed diets supplemented with MOS, these differences were not significant. Moreover, no histopathological differences were observed in liver tissue cross sections between control and treatment groups. These results suggest that supplementation of diets with MOS had no significant effects on general fish health.

Carbonic anhydrase activities from the rainbow trout lens correspond to the development of acute gas bubble disease.

Dissolved gas supersaturation is hazardous to fish and can result in gas bubble disease (GBD). Signs of GBD typically include bubbles in the eyes, fins, skin, lateral line, and gill filaments. Ocular abnormalities in diseased salmonids typically occur after aberrant gas production in the eyes. In this study, freshwater rainbow trout Oncorhynchus mykiss were exposed experimentally to percent total gas pressure (TGP%) levels of 104% (control) and 115%. No mortalities occurred during the 7-d experimental period. Effects of GBD were observed externally as a darkened skin, exophthalmia, localized hemorrhage in the eye, and gas bubbles on the operculum. Additional signs included increased swimming activity and, more frequently, panic episodes. Carbonic anhydrase (CA) enzyme activities from the lens and retina were determined at days 0, 1, 3, 5, and 7 of the study. Venous blood gases were also measured on day 7. Retinal pH did not differ between normal and affected fish, but blood characteristics such as the partial pressure of O2, partial pressure of CO2, carboxyhemoglobin level, and bicarbonate ion concentration were significantly elevated in affected fish relative to normal fish. Venous blood pH and oxyhemoglobin levels were not significantly different between affected and normal fish. Patterns of response to total dissolved gas levels differed between the lens and the retina. Mean CA activities in the lenses of fish exposed to a TGP% level of 115% were significantly below those of control fish. However, retinal CA activities did not significantly differ between the two groups over the course of the experiment. These findings show that dissolved gas supersaturation reduces CA activity in the rainbow trout lens.

Effects of some metals on carbonic anhydrase from brains of rainbow trout.

Carbonic anhydrase (CA) enzyme was purified from rainbow trout brain by Sepharose-4B-L: -tyrosine-sulfanilamide affinity chromatography. The enzyme was obtained with a specific activity of 2,275 EU mg(-1) and a yield of 22.5%. The sample obtained from the affinity column was used for kinetic properties and inhibition studies. Both optimum and stable pH were found as 9.0 in 1 M Tris-SO(4) at 4 degrees C, respectively. To check the purity and subunit molecular weight of enzyme, sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis was performed, and MW was found as approximately 29.0 kDa. The molecular weight of native enzyme was estimated to be approximately 27.3 kDa by gel filtration chromatography. The purified enzyme had apparent K (m),V (max), and k (cat) as follows: 0.92 mM, 0.207 micromol.min(-1) and 43.6 s(-1) for p-nitrophenylacetate. The inhibitory effects of Co(II), Cu(II), Zn(II), Ag(I), and Cd(II) on CA enzyme activity were determined using the esterase method under in vitro conditions at low concentrations of the corresponding metals. The obtained IC(50) values, which cause 50% inhibition on in vitro enzyme activity, were 0.05, 30, 0.31, 159, and 82.5 mM for cobalt, copper, zinc, silver, and cadmium, respectively. K ( i ) values were also calculated from Linewaever-Burk plots for these substances as 0.014, 27.68, 2.15, 193.86, and 94.18 for cobalt, copper, zinc, silver, and cadmium, respectively; it was determined that cobalt, silver and cadmium inhibited the enzyme competitively, copper inhibited noncompetitively while zinc inhibited the enzyme uncompetitively.

Effects of light on trace elements and triiodothyronine levels in plasma of mirror carp (Cyprinus carpio).

The changes of copper, zinc, iron, magnesium, manganese, and triiodothyronine as a result of short- and long-term isothermal exposure to light were determined in the plasma of mirror carp (Cyprinus carpio). Two groups of fish were subject to a 3-mo treatment that consisted of (1) 8 h of light followed by 16 h of darkness and (2) 16 h of light followed by 8 h of darkness at 25 degrees C. No significant changes were recorded between the two groups at the beginning of the study. After 3 mo, the triiodothyronine and elements levels as well as the growth rates of fish in the long-time exposure period were significantly higher than those in the short-time exposure (p<0.05).

Sublethal ammonia and urea concentrations inhibit rainbow trout (Oncorhynchus mykiss) erythrocyte glucose-6-phosphate dehydrogenase.

In vitro and in vivo effects of sublethal ammonia and urea concentrations were assayed on glucose-6-phosphate dehydrogenase (G6PD) of rainbow trout (Oncorhynchus mykiss) erythrocyte. G6PD was purified from erythrocytes with a specific activity of 16.7 EU (mmol NADP+/min)/mg protein and approximately 1600-fold in a yield of approximately 60% by ammonium sulphate precipitation and 2',5'-ADP Sepharose 4B affinity chromatography. The purity of the enzyme was confirmed using SDS polyacrylamide gel electrophoresis. Experiments with ammonia (2.2-5.5 microM) and urea (20-50 microM) showed the inhibitory effects on the enzyme, in vitro. Inhibition effects were determined in vitro by Lineweaver-Burk and regression graphs. The dissociation constant of the enzyme inhibitor complex (Ki) and 50% inhibitory values were 2.26+/-1.21 and 2.86+/-3.51 microM for ammonia and 18.69+/-6.75 and 23.77+/-4.58 microM for urea, respectively. In vivo studies in rainbow trout erythrocytes showed significant (p < 0.01) inhibition of G6PD by ammonia and urea. However, ammonia inhibited more than urea since there were significant differences between the final values of erythrocyte G6PD activities.

Effects of low molecular weight plasma inhibitors of rainbow trout (Oncorhynchus mykiss) on human erythrocyte carbonic anhydrase-II isozyme activity in vitro and rat erythrocytes in vivo.

The effects of low molecular weight plasma inhibitors from rainbow trout (Oncorhynchus mykiss) (RT) were investigated on the carbonic anhydrase enzyme (CA) activities in in vitro human and in in vivo Sprague-Dawley rat erythrocytes. The RT blood was used as extracellular fluid (plasma) source and plasma inhibitors were obtained by dialysis of the plasma. For the in vitro study, human carbonic anhydrase-II (HCA-II) isozyme was obtained by Sepharose 4B-L-tyrosine-sulfanylamide affinity chromatography with an overall purification of about 646-fold. The enzyme (specific activity of 7750 EU/mg protein) was obtained with a yield of 71.1% and SDS-PAGE showed a single band. From in vitro studies, the I50 value for RT plasma inhibitors obtained was 0.37 mg/ml. From in vivo studies on rat erythrocytes, CA activity was significantly inhibited by the inhibitors from the extracellular fluid of RT for up to 3 h (p < 0.05) following intraperitoneal administration.

Effects of modified atmosphere and vacuum packaging on microbiological and chemical properties of rainbow trout (Oncorynchus mykiss) fillets.

Microbial (psychrotrophic, mesophilic aerobic bacteria and Enterobacteriacae counts), and chemical analysis [pH, total volatile bases nitrogen (TVB-N), lipid oxidation (Thiobarbituric acid reactive substance, TBARS)] of rainbow trout (Oncorynchus mykiss) fillets in air (control), vacuum and modified atmosphere packaging (MAP) with various gas mixtures conditions at 4+/-1 degrees C were determined. The gas mixtures evaluated were 100% CO2, 2.5% O2+7.5% N2+90% CO2 and 30% O2+30% N2+40% CO2. Psychrotrophic bacteria count was above 1 x 10(7) cfu/g on the 12th day in 100% CO2. However; mesophilic bacteria count was below 1 x 10(6) cfu/g at the end of the 14-day storage period. Enterobacteriaceae count was significantly lower in samples packaged with MAP. Lipid oxidation increased rapidly after 6 days of storage in the samples containing 30% O2. While minimum TBARS values were recorded in fillets containing 100% CO2 and vacuumed fillets, the lowest TVB-N values were obtained in fillets with 100% CO2.

Inhibitory effects of ammonia and urea on gill carbonic anhydrase enzyme activity of rainbow trout (Oncorhynchus mykiss).

The effects of ammonia and urea on branchial carbonic anhydrase (CA) enzyme which plays a key role in ionoregulation, osmoregulation and acid-base balance of rainbow trout (Oncorhynchus mykiss) were investigated. CA activity of the control group for ammonia and urea was determined as 1285.7 ± 67.9 and 1261.7 ± 60.8EU/mg protein, respectively. The CA enzyme activities of the other groups were measured at 1, 2 and 3h after ammonia and urea applications. The corresponding activities of ammonia were 774.9 ± 68.8, 732.1 ± 48.6 and 768.1 ± 59.5EU/mg protein, respectively and that of urea were 769.3 ± 58.9, 638.2 ± 47.7 and 1108.1 ± 61.1EU/mg protein, respectively. The differences between the initial CA activities for the controls was not significantly (P > 0.01). The CA activities were significantly (P < 0.01) inhibited both in ammonia and urea group. However, the ammonia inhibited more than urea since there was significant differences between final values of gill CA activities.

The in vitro and in vivo inhibitory effects of some sulfonamide derivatives on rainbow trout (Oncorhynchus mykiss) erythrocyte carbonic anhydrase activity.

The in vitro and in vivo inhibitory effects of 5-(3alpha, 12alpha-dihydroxy-5-beta-cholanamido)-1,3,4-thiadiazole-2-sulfonamide (1), 5-(3alpha, 7alpha, 12alpha-trihydroxy-5-beta-cholanamido)-1,3,4-thiadiazole-2-sulfonamide (2), 5-(3alpha, 7alpha, 12alpha-triacetoxy-5-beta-cholanamido)-1,3,4-thiadiazole-2-sulfonamide (3) and acetazolamide on rainbow trout (Oncorhynchus mykiss) (RT) erythrocyte carbonic anhydrase (CA) were investigated. The RT erythrocyte CA was obtained by affinity chromatography with a yield of 20.9%, a specific activity of 422.5EU/mg protein and a purification of 222.4-fold. The purity of the enzyme was confirmed by SDS-PAGE. Inhibitory effects of the sulfonamides and acetazolamide on the RT erythrocyte CA were determined using the CO2-Hydratase method in vitro and in vivo studies. From in vitro studies, it was found that all the compounds inhibited CA. The obtained I50 value for the sulfonamides (1), (2) and (3) and acetazolamide were 0.83, 0.049, 0.82 and 0.052 microM, respectively. From in vivo studies, it was observed that CA was inhibited by the sulfonamides (1), (2) and (3) and acetazolamide.