Enzymatic changes Mean±SE (N=6) of the fish
The elemental lead (Pb) occurs naturally in the environment as well as being produced by mining and manufacturing activities . Lead and its compounds are serious pollutants of the aquatic environment. Moreover, several authors also agree that toxic and non-biodegradable metals such as lead accumulate in many fish species, causing various diseases such as renal [2, 3], hepatic lesions , endocrine impairment  and effect of cell membrane lipids in cells of the central nervous system.
Zeolites are used in industry, agriculture, environment protection and even in medicine. Zeolites have a relatively high Si/Al compositional ratio which gives it is special ion-exchange selectivity for large monovalent cations. Natural or synthetic zeolites (sodium aluminum silicates) are known to easy adsorb metal ions by exchange reactions .
Hematological indices such as hematocrit (Hct), hemoglobin (Hb), red blood cells (RBCs), white blood cells (WBCs) and plasma enzyme activities such as AST (aspartate aminotransferase), ALT (alanine aminotransferase), stress hormone cortisol and choline esterase have been used as an indicator of metal pollution in the aquatic environment [9-11].
There is a strong correlation between lead and zeolite that have been found in aquatic organisms and several authors have demonstrated that zeolite protects against lead and also other heavy metals in the experiment [12, 13]. There are numerous biological mechanisms between lead and zeolite. These mechanisms are (I) ion-exchange and (II) adsorb metal ions by exchange reactions [6, 14]. This effects decreases homeostatic mechanisms on fish.
The aim of this study was to determine the effects on hematological and some biochemical parameters on adult
2. Materials and methods
Water quality characteristics in tanks;
Dissolved Oxygen: 7.6±0.5 mg/L
Total Hardness: 184.5±4.38 CaCO3 mg/L
Total Alkalinity: 278.2±8.7 CaCO3mg/L
The effects of lead and zeolite were shown as 0.1 mg/L Lead (Pb), 0.1 mg/L Lead+0.1 g/L Zeolite (PbZ1), 0.1 mg/L Lead+0.2 g/L Zeolite (PbZ2) and Control (C). A total of 4 aquariums sized 40x100x40 cm in height were divided into two groups. These were filled with 100 L. aerated class aquarium tanks. Nine fish were put in each aquarium (3 repetition x 3 fish). One aquariums of the first group contained 0.1 mg Pb/L (PbCl2, 2 H2O) solutions and two aquariums of the second group contained 0.1 mg/L Lead+0.1 g/L Zeolite and 0.1 mg/L Lead+0.2 g/L Zeolite solutions and one aquarium was used as a control. All fish were fed with Pinar Yem at a concentration of 1 % of their body mass per day.
Fish were anaesthetized with MS-222 blood was collected from each fish by cutting the caudal peduncle. Fish blood was collected for hematological parameters. The blood was centrifuged at 4000 rpm over 10 min at 15 0C to obtain the serum. Blood samples were sent to Cukurova University (Balcalı Hastahanesi, Merkez laboratory) for hematological analysis. The serum was divided in two portions from the ephandof tubes, first portion for cortisol and cholineesterase, second portion for ALT and for AST, ALT and AST activities. Those analyses were determined using UV test technique . The serum samples were frozen and stored -200C until required for assays. Cortisol, ALT, AST and cholinesterase were determined by ROCHE Hitachi E-170 and DPP.
Data are presented as mean ± standard error. For the statistical analysis, it was used one-way analysis of variance (ANOVA) followed by Student Newman–Keul’s test using SPSS 10.0 statistical software (SPSS Inc., Chicago, IL). Differences were considered signiﬁcant if
3. Results and discussion
No mortality was observed at concentrations of the lead and its zeolite mixtures studied during the experiments. The statistical analysis was done with “SNK” differences among groups were measured to be significant at p<0.05 and showed Table 1 and 2.
Zeolite may decrease the toxicity of lead in water and Pb may form a complex with Zeolite. Hb, Hct, RBCc and WBCc levels decreased Pb, PbZ1 and PbZ2 exposed fish at both exposure periods (Fig 1-4). The exposures of Pb, PbZ1 and PbZ2 did not cause any significant changes in RBCc and WBCc levels of fish at 10 days while they caused a decrease in its levels at the end of the exposure period.
Serum Cortisol, ALT, AST and Cholinesterase activities of
4. Hematological parameters
Fig. 1-4 show distribution of hematological (Hb, Hct, RBCc and WBCc) parameters Pb and Pb+Ze mixtures exposed to
Lead was classified as a toxic substance for fish. Changes in the erythrocyte profile suggest a compensation of oxygen deficit in the body due to gill damage. Hematological indices are commonly used as indicators of metal pollution in fish [17, 18]. This index reflects respiratory status of animals. In addition to these status, infectious and stress have been shown to influence the fish hematology.
Metals in hematological parameters of fish generally occur due to the osmotic changes resulting in hemodilution (an increase in the volume of plasma, resulting in a reduced concentration of red blood cells in blood) or hemoconcentrations (an increase in the concentration of blood cells resulting from the loss of plasma or water from the bloodstream) . Generally, spleen is responsible for this change. Because, spleen, serving as a potent blood storage organ in some teleost, sequestering blood cell under resting conditions and releasing them to circulating blood associated with various stress . In  have been reported that histopathological lesions occur in spleen and intestine lead exposure on
5. Enzyme activity
In this study, at 10 and 20 days, compared with controls, ALT, AST, Cortisol and Cholinesterase were decreased PbZ1 and PbZ2 concentrations while ALT, AST and Cortisol increased only Pb concentrations. However, Cholinesterase decreased at not Pb, but PbZ1 and PbZ2 combinations (P<0.005) during 10 and 20 days.
Plasma cortisol level is widely used as a general indicator of stressful conditions in fish [21, 22]. Despite the interest in plasma cortisol measurement as an indicator of stress, few studies have actually measured the kinetics of cortisol in fish. A study , reported that during chronic stress, plasma cortisol back to the resting levels on
Serum ALT and AST activities used in diagnosis of damage fish tissues (i.e. gill, muscle, liver) . Determinations of transaminases (AST and ALT) have been useful in the diagnosis of liver and kidney diseases in fish . These enzymes of
There are multiple forms of esterase in vertebrates’ blood plasma . However, acetyl cholinesterase content of fish blood is present in low concentration compared with other vertebrate . One of the biomarkers most frequently used in fish for the diagnosis of exposure to pollutants is the measurement of the inhibition of the enzyme cholinesterase (ChE) . Table 1 and 2 presents ChE activity in plasma of
In conclusion, the data from this investigation which is the blood-based enzymatic and hematological parameters responded to relatively Pb and its mixtures concentrations show useful for monitoring on fish [33, 34]. The hematological data, as well as gross observations from sample handing and fish necropsy, suggest that this data may have been related to erythrocyte fragility (erythrocyte easily broken, damaged, or destroyed) and hemorrhaging exposed to metals [20, 35]. It is possible mechanisms that the decrease in blood parameters may be hemolysis and damage to hematopoietic tissues by lead and its zeolite mixtures. Further, the decrease of serum enzymatic mechanisms may be indicated liver damage [25, 28 and 36] and may be occurring from Pb and Ze mixtures form.
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