A. R. Yousuf

Assessment of endosulfan induced genotoxicity and mutagenicity manifested by oxidative stress pathways in freshwater cyprinid fish crucian carp (Carassius carassius L.)

Over the past few decades, endosulfan, one of the polychlorinated pesticides still in use, has received considerable attention of a number of international regulations and restriction action plans worldwide. This study aimed to evaluate the cytogenetic effects of endosulfan using robust genotoxicity assays, along with the oxidative stress pathways in order to understand biochemical mechanism, in Carassius carassius L. The LC50-96 h (95% confidence limits) value of endosulfan was 0.070 (0.046-0.093) ppm; and on its basis three test concentrations (sub-lethal I: 0.052, II: 0.035 and III: 0.017 ppm) were selected for 35 d in vivo exposure. The mean concentration of endosulfan in aquaria was always constant, when analyzed by dispersive liquid-liquid micro extraction (DLLME) followed by GC-MS. Autopsy was done on days 1, 2, 3, 4, 7, 14, 21, 28 and 35 of endosulfan exposure; the micronucleus formation (MN), authenticated by scanning electron microscopy, and chromosomal aberrations (CA), were induced significantly (p<0.05) in all the treated groups, including positive control cyclophosphamide (4 ppm), when compared to negative control. Similarly lipid peroxidation (LPO) was induced significantly with the maximal at higher concentration (SL-I) on 4th day (722.45%; p<0.01). Antioxidant biomarkers like glutathione reduced, superoxide dismutase and catalase also fluctuated significantly (p<0.01) in all treatment groups. Collective findings demonstrated that genotoxic effects were invariably accompanied and correlated with increased oxidative stress and disturbance of antioxidant enzymes; and the MN and CA assays are useful tools in determining potential genotoxicity of aquatic xenobiotics and might be appropriate as a part of monitoring program.

Pharmacological and toxicological evaluation of Urtica dioica.

Context: Medicinal plants are a largely unexplored source of drug repository. Urtica dioica L. (Urticaceae) is used in traditional medicine to treat diverse conditions. Objective: The present study describes the antidiabetic, antiinflammatory, antibacterial activity, and toxicological studies of Urtica dioica. Materials and methods: U. dioica leaves were subjected to solvent extraction with hexane, chloroform, ethyl acetate, methanol, and aqueous, respectively, and screened for antidiabetic (300 mg/kg bw by glucose tolerance test; GTT), antiinflammatory (200 mg/kg bw by rat paw edema assay) and antibacterial activities [by disc-diffusion and minimum inhibitory concentration (MIC) assays]. Toxicological studies were carried on Artemia salina and Wistar rats; phytochemical analyses were carried out, using chromatographic and spectroscopic techniques. Results: The aqueous extract of U. dioica (AEUD) significantly (p < 0.001; 67.92%) reduced the blood glucose level during GTT in Wistar rats with an effective dose of 300 mg/kg bw in dose-dependent studies. High-performance liquid chromatography with photodiode-array detection (HPLC-DAD) analysis showed the presence of hydroxycinnamic acid derivatives and flavonoids in AEUD. Hexane Fraction-2 (HF2) exhibited both antiinflammatory activity (48.83% after 3 h), comparable to that of indomethacin (53.48%), and potent antibacterial activity with MIC values ranging from 31.25–250 µg/mL against all the tested strains. Gas chromatography–mass spectrometry (GC–MS) analysis showed fatty acid esters and terpenes as the major constituents of HF2. Toxicity tests showed higher safety margin of all the solvent extracts with LC50 > 1000 μg/mL each on A. salina. Discussion and conclusion: Our results showed that the U. dioica leaves are an interesting source of bioactive compounds, justifying their use in folk medicine, to treat various diseases.

Cytotaxonomic status of Schizothoracine fishes of Kashmir Himalaya (Teleostei: Cyprinidae)

Abstract Karyotypic study of five Schizothorax species viz. S. niger, S. esocinus, S. labiatus, S. plagiostomus and S. curvifrons, belonging to subfamily Schizothoracinae, obtained from Dal lake and River Jhelum, Kashmir was carried out following the method of Thorgaard and Disney (1990).The diploid chromosome numbers recorded were 98 in S. niger (24m+32sm+22st+20t), 98 in S. esocinus (30m+22sm+10st+36t), 98 in S. labiatus (24m+20sm+2st+52t), 96 in S. plagiostomus (24m+18sm+54t) and 94 in S. curvifrons (26m+20sm+20st+28t).Pattern of chromosomal evolution and role of chromosomal rearrangements was discussed in addition to the cytotaxonomic status of these fishes. The study conclusively confirmed the specific status of five species of Schizothorax on the basis of their genetic material.

Bioassay guided isolation and identification of anti-inflammatory and anti-microbial compounds from Urtica dioica L. (Urticaceae) leaves

The present study describes the anti-inflammatory, anti-microbial activity and lipophilic profile with acute toxicological studies of Urtica dioica. Successive extraction of the leaves with organic solvents of increasing polarity and their screening for anti-inflammatory and anti-microbial activity was assessed. Hexane extract showed good anti-inflammatory and anti-microbial activity; hence it was further fractionated using open silica gel column chromatography into 19 sub fractions which were pooled together according to their thin layer chromatography (TLC) profile to give an overall 5 fractions. Among the 5 fractions, fraction-II (FII) at a dose of 200 mg/kg body-weight (bw) exhibits equipotent antiinflammatory activity (48.83% after 3 h) as that of the standard drug indomethacin (53.48%) in Wistar rats. FII also showed a potent anti-microbial activity against all the tested bacterial strains and its minimum inhibitory concentration (MIC) value was 125, 15.62, 31.25, 250, 31.25, 125 and 7.81 µg/ml against Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus, Shigella flexneri and Salmonella typhi, respectively which was determined by serial tube dilution method. FII was subjected to gas chromatography-mass spectrometry (GC-MS) analysis in search of potent anti-inflammatory and anti-microbial compound(s). 2,4-Di-t-butylphenol (4.56%), neophytadiene (26.97%), butyl tetradecyl ester (9.53%), dibutyl phthalate (7.45%), bis(2-ethyl hexyl) maleate (8.80%), 1,2-benzenedicarboxylic acid (9.89%) and 2-tert-butyl-4,6-bis(3,5-di-tert-butyl-4hydroxybenzyl)phenol (3.19%) were the major constituents responsible for both anti-inflammatory and anti-microbial activity of hexane extract of U. dioica. Sub-acute oral toxicity of crude n-hexane extract of U. dioica was carried out in Wistar rats at doses of 250, 500, 1000 and 2000 mg/kg bw to assess the safety index. Hematological parameters from blood and other biochemical parameters from serum confirmed its safety at tested concentrations. Our results corroborate the anti-inflammatory and antimicrobial activity of U. dioica, and could justify its use in folk medicine for the treatment of rheumatic arthritis and other infectious diseases.

An introduction about Genotoxicology Methods as Tools for Monitoring Aquatic Ecosystem: Present status and Future perspectives

Pesticide residues with genotoxic potential reach the aquatic environment and constitute a major issue that give rise to concerns at local, regional, national and global scales. Fish serves as an excellent genetic model for the genetic hazard assessment as they are very sensitive to changes in their environment. As there is a close consortium of DNA damage, mutation and induction of various types of genetic disorders, genotoxicity tests like chromosomal aberration test, micronuclei and comet assay are gaining credence and since past few decades many tests have been developed for evaluating genetic alterations in aquatic organisms. These tests rely on the premise that any change to DNA may have enduring and ardent consequences. Thus, the first aim of the genotoxicology is to describe the outcome produced by toxic substances in various test species, but only from the genetic point of view and to draw conclusions that can be extrapolated to man. This review summarizes the genotoxicity tests developed till date and the role of piscine model in genotoxicology. The text also delves the latest knowledge and thinking on these cardinal approaches for the assessment of aquatic environmental health, management and conservation, besides providing useful repository for the researchers especially dealing with aquatic genotoxicity tests. The perspectives for further research on the use of genotoxicology tests were also highlighted.

Oxidative stress in the freshwater cyprinid crucian carp (Carassius carassius L.) upon chronic exposure to endosulfan

Oxidative stress and antioxidant responses of crucian carp, upon chronic exposure to endosulfan, were evaluated in vivo. The lethal concentration (LC50–96 h) was 70 μg L−1; on its basis, the fish were exposed to endosulfan at 20, 35, and 50 μg L−1 and autopsy was done on days 1, 2, 3, 4, 7, 14, 21, 28, and 35. Lipid peroxidation was induced in a concentration-dependent manner, being highest at 50 μg L−1 (3/4 LC50–96 h, sub-lethal concentration-I, SL-I) on day 4 (720% versus control), followed in its extent (490%) at 30 μg L−1 (1/2 LC50–96 h, sub-lethal concentration-II, SL-II) on day 7 and lowest (260%) at 10 μg L−1 (1/4 LC50–96 h, sub-lethal concentration-III, SL-III) on day 14. Glutathione showed a concentration- and time-dependent elevation in the initial phase, with highest level on day 4 (180%) at SL-I, but showed significant reduction in all test concentrations from day 21 of post-exposure. Superoxide dismutase was decreased significantly throughout the study, with highest reduction (63%) on day 4 at SL-I; catalase increased in all test concentrations up to day 14 but showed a significant decrease from the day 28 of post-exposure. The potential role of these parameters as indicators of pesticide pollution in aquatic systems is discussed.

Distribution Pattern of Rooted Floating Leaf Type Macrophytes in Response to Water Depth in a Fresh Water Lake of Kashmir Himalaya

The distribution of rooted floating-leaf type macrophytes were studied in response to water depth in Manasbal Lake of Kashmir Himalaya. Seven aquatic plant species viz., Potamogeton natans, Trapa natans, Nelumbo nucifera, Nymphoides peltatum, Nymphaea alba, Hydrocharis dubia and Euryale ferox were observed along three different depth zones in the lake. The depth zones D1 (0-100 cm) and D2 (101-200 cm) were noticed to be suitable for the growth of macrophytes in terms of their community features while as less values of the same for plant species were observed at a depth range of 201-300 cm (D3). Thus, a certain effect was depicted by water depth on the community of rooted floating aquatic plant species, which could be used as a potential management strategy to control their infestation in the Lake ecosystem.

Oxidative stress-inducing potential of butachlor in a freshwater fish, Cyprinus carpio (L)

Oxidative stress-inducing potential of butachlor was studied in fresh water fish, Cyprinus carpio. Fish was exposed to butachlor (0.17 ppm in acetone) for a period of 24, 48, or 72 h. Various markers of oxidative stress, including catalase, glutathione S-transferase, glutathione reductase, levels of reduced glutathione (GSH) and lipid peroxidation (LPO), were estimated in liver, kidneys, and gills of fish. Butachlor-induced LPO and antioxidant enzyme activity and depleted levels of GSH in all organs examined. Fish exposed to butachlor experienced oxidative stress which was time dependent. The tissue antioxidants seemed to be induced in order to counteract the impact of free radicals, denoting an adaptation of fish to overcome chemical-induced stress.

Coupled biochemical genetic and karyomorphological analyses for taxonomic classification - A case study of Schizothorax species complex (Teleostei: Cyprinidae)

Biochemical genetic and karyomorphological studies were evaluated for taxonomic importance. Five Schizothorax species namely: Schizothorax niger , Schizothorax curvifrons , Schizothorax esocinus , Schizothorax labiatus and Schizothorax plagiostomus were studied in this experiment as a model for serum proteins and chromosomal analysis. Serum protein band patterns of Schizothorax species complex were studied by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Their electrophoretograms revealed similarities as well as differences in the number and molecular weight of protein bands. S. esocinus showed five bands, S. curvifrons five, S. niger seven, S. labiatus and S. plagiostomus each showed six bands; they also showed species characteristic bands. Karyotypic study of these was carried out. The diploid chromosome numbers recorded were 98 in S. niger (24 m + 32 sm + 22 st + 20 t), 98 in S. esocinus (30 m + 22 sm + 10 st + 36 t), 98 in S. labiatus (24 m + 20 sm + 2 st + 52 t), 96 in S. plagiostomus (24 m + 18 sm + 5 4t) and 94 in S. curvifrons (26 m + 20 sm + 20 st + 28 t). Coupled biochemical genetic and karyomorphological analysis proved a good taxonomic tool as the results were decisive in establishing the species status of these species despite their overlapping morphological characters. Keywords: Schizothorax , serum proteins, karyotype, taxonomy, biochemical African Journal of Biotechnology , Vol 13(14), 1623-1630

Karyoevolutionary and karyosystematic considerations on Schizothorax curvifrons and Schizothorax niger (Teleostei: Cyprinidae): Important hill-stream food fishes of Kashmir Himalaya

Cytogenetic studies have helped in clarifying the problem of disagreement amongst taxonomists on the identity of a given species. Cytogenetic studies were performed on two fishes of the genus Schizothorax viz. Schizothorax curvifrons Heckel and Schizothorax niger Heckel (Cyprinidae: Schizothoracinae) obtained from Sindh Stream and Dal Lake Srinagar Kashmir, respectively. These fishes are considered to be the subspecies of the same species. The two species showed a diploid number of 98 in S. niger and 94 in S. curvifrons . The karyological data are analyzed in terms of the taxonomic aspects within this genus, and the validity of their existence as species chromosomally distinct from each other is emphasized. Keywords: Sindh stream, karyotype, cytotaxonomy, Kashmir Himalaya, chromosome.