2.2 Test Compounds
The drug acetylsalicylic acid of 99.0% purity (CAS Number: 50-78-2) was purchased from Sigma-Aldrich. AST and ALT kits were purchased from Coral Clinical
Systems, India. All the other chemicals and reagents (of analytical grade) used in the
present investigation were purchased from HiMedia Laboratories Pvt. Ltd., India.
1 g of acetylsalicylic acid (ASA) was dissolved in 0.9 mL of dimethyl sulfoxide
(DMSO) was used to prepare stock solution (1,000 mg L
À1 ) of ASA. From this stock
solution, appropriate quantity was taken and dissolved in experimental tanks to get
the desired concentration of toxicity solution.
2.3 Chronic Toxicity Assay
Two different concentrations were selected for testing the chronic toxicity of
acetylsalicylic acid such as 100 and 200 mg L
À1 , and they were grouped as
Treatment I and Treatment II, respectively. Three glass aquaria of 100 L capacity
were taken and filled with 80 L of water in which one is maintained as control and
the other two as Treatment I and Treatment II. The glass tanks were aerated. From
the stocking tank, 40 healthy fish fingerlings were randomly selected and introduced
into each tank. The toxicant was renewed and the water in the tanks was changed
daily, in order to prevent accumulation of faecal matter.
2.4 Blood and Organs Sampling
Fish from control and ASA-treated groups were sacrificed without anaesthetizing,
and blood was collected in a heparinized syringe by puncturing dorsal aorta. The
collected blood was maintained in sterilized plastic vials. A freshly pooled whole
blood sample was used for haematological analysis. The remainder of the blood were
centrifuged at 10,000 rpm for 20 min to separate the plasma and transferred to clean
vials for biochemical (plasma glucose and protein) and enzymological (AST and
ALT) analysis. Subsequently gill and liver tissues were excised out for the study of
antioxidant parameters (SOD, LPO, CAT, GST and GSH). The collected tissue
samples (liver and gill) were washed with 0.9% NaCl solution dried with filter paper
and maintained at À80
C.
196
S. Siddeswaran et al.
The drug acetylsalicylic acid of 99.0% purity (CAS Number: 50-78-2) was purchased from Sigma-Aldrich. AST and ALT kits were purchased from Coral Clinical
Systems, India. All the other chemicals and reagents (of analytical grade) used in the
present investigation were purchased from HiMedia Laboratories Pvt. Ltd., India.
1 g of acetylsalicylic acid (ASA) was dissolved in 0.9 mL of dimethyl sulfoxide
(DMSO) was used to prepare stock solution (1,000 mg L
À1 ) of ASA. From this stock
solution, appropriate quantity was taken and dissolved in experimental tanks to get
the desired concentration of toxicity solution.
2.3 Chronic Toxicity Assay
Two different concentrations were selected for testing the chronic toxicity of
acetylsalicylic acid such as 100 and 200 mg L
À1 , and they were grouped as
Treatment I and Treatment II, respectively. Three glass aquaria of 100 L capacity
were taken and filled with 80 L of water in which one is maintained as control and
the other two as Treatment I and Treatment II. The glass tanks were aerated. From
the stocking tank, 40 healthy fish fingerlings were randomly selected and introduced
into each tank. The toxicant was renewed and the water in the tanks was changed
daily, in order to prevent accumulation of faecal matter.
2.4 Blood and Organs Sampling
Fish from control and ASA-treated groups were sacrificed without anaesthetizing,
and blood was collected in a heparinized syringe by puncturing dorsal aorta. The
collected blood was maintained in sterilized plastic vials. A freshly pooled whole
blood sample was used for haematological analysis. The remainder of the blood were
centrifuged at 10,000 rpm for 20 min to separate the plasma and transferred to clean
vials for biochemical (plasma glucose and protein) and enzymological (AST and
ALT) analysis. Subsequently gill and liver tissues were excised out for the study of
antioxidant parameters (SOD, LPO, CAT, GST and GSH). The collected tissue
samples (liver and gill) were washed with 0.9% NaCl solution dried with filter paper
and maintained at À80
C.
196
S. Siddeswaran et al.