Toxicological and ecological implications of biotransformation enzymes in the tropical teleost Chaetodon capistratus

Hepatic cytochromes P450 (phase I monooxygenases) and glutathione transferases (phase II conjugating enzymes) were investigated in Chaetodon capistratus (Linnaeus) collected in Florida and Belize in June and December 1991, respectively. These biotransformation enzymes play major roles in the detoxification of xenobiotics by converting lipophilic chemicals to more hydrophilic, readily excretable metabolites. Content of total microsomal P450 (0.501 to 0.821 nmol mg^-1 microsomal protein) and rates of NADPH-cytochrome c (P450) reductase (270.7 to 330.2 nmol min^-1 mg^-1 microsomal protein) and glutathione transferase (2.81 to 3.12 g min^-1 mg^-1 cytosolic protein) in these fish were greater than in most untreated fish species, i.e., fish that have not been exposed to PAHs (polycyclic aromatic hydrocarbons) or PCBs (polycyclic biphenyls). Ethoxyresorufin O-deethylase (EROD) rates (0.029 to 0.171 nmol min^-1 mg^-1) were also comparable to those in most untreated marine fish. Immunoblot analysis with monoclonal antibody (MAb) 1-12-3 to scup P450E (the EROD catalyst and a teleost representative of the PAH-inducible CYP1A gene subfamily) showed slight amounts of cross-reacting protein in C. capistratus liver microsomes. Hepatic CYP1A content and EROD activity did not differ significantly between fish collected in Florida and Belize, suggesting that the two sites differed little in contamination by CYP1A inducers. Immunochemical analyses with polyclonal antibodies to scup P450B (a teleost representative of the CYP2B subfamily) and human CYP3A4 cross-reacted strongly with C. capistratus hepatic proteins. The CYP2B and CYP3A subfamilies in mammals are believed to have partially evolved in response to toxic dietary allelochemicals. C. capistratus regularly feeds on terpenoid-rich gorgonian corals, suggesting that biotransformation enzymes may be involved in the metabolism of dietary allelochemicals as well as anthropogenic xenobiotics in this species.     

The effect of oxytetracycline on physiological and enzymatic defense responses in aquatic plant species Egeria densa,Azolla caroliniana,and Taxiphyllum barbieri

Oxytetracycline is an antibiotic widely employed in aquaculture to control and treat bacterial diseases of fish. Due to ineffective wastewater treatment, antibiotic residues from fish ponds are directly released into surface and groundwater affecting the environment. The Green Liver System® is a sustainable and cost-effective water treatment based on the ability of aquatic plants to biotransformation xenobiotics. To expand the application range of this system, Egeria densa, Azolla caroliniana, and Taxiphyllum barbieri were tested in response to oxytetracycline exposure. The aquatic plants were exposed to 0.5, 2.0, 5.0, and 25 μg/L oxytetracycline (n = 4) for 24 h in order to analyze the physiological responses (photosynthetic pigment contents and hydrogen peroxide formation -- H₂O₂), the biotransformation process (activity of glutathione S-transferase), and the antioxidant defense (glutathione reductase; peroxidase; and catalase) responses. There were no statistical differences in the photosynthetic pigment contents and H₂O₂ level between the treatment and control groups in the three aquatic plant species. It can be concluded that oxytetracycline does not provoke physiological damage to the tested species within 24 h of exposure. Nevertheless, the antioxidant defense mechanism was activated. Peroxidase and catalase were the most prominent antioxidant enzymes in the three plant species, whereas glutathione S-transferase activity was significantly increased in A. caroliniana and T. barbieri.     

Effects of nutrition and temperature on metabolic enzyme activities in larval and juvenile red drum,Sciaenops ocellatus,and lane snapper,Lutjanus synagris

The metabolic enzyme activities were determined in larvae of red drum, Sciaenops ocellatus, and lane snapper, Lutjanus synagris, to determine the effect of temperature and nutrition on metabolic enzyme activities and to evaluate if metabolic enzyme activities are useful in assessing the feeding condition of larval fish. During experiments conducted during the spring of 1990, lactate dehydrogenase (LDH) activities in both red drum and lane snapper were approximately an order of magnitude lower than values typical for adult fish; LDH and citrate synthase (CS) activities increased during early developmental stages, but nutritional effects were apparent. Clear differences (up to 4-fold) between well-fed and starving fish were evident in both LDH and CS activity in red drum. Differences between well-fed and poorly fed larvae were evident until 9 d after hatching. Lane snapper larvae reared at a 25°C had significantly lower LDH activities than larvae reared at 28°C.     

Role of bacteria with regard to chitin degradation in the digestive tract of the cod Gadus morhua

Numbers of aerobic, heterotrophic and chitinolytic bacteria in stomach, pyloric caeca and intestine of eight specimens of freshly caught cod (Gadus morhua L.) were determined by the spread plate method (medium: sea water agar, including 1% chitin). In crude enzyme extracts, chitinase activity was assayed according to the end product measurement. Results revealed no correlation between the number of chitinoclasts and the level of chitinase activity in any part of the digestive tract of cod. In another series of studies, antibiotics (Ampicillin/Sisomicin, or Chloramphenicol) were applied via tank water and feed. In most cases, antibiotic treatment resulted in the complete elimination of bacteria from the digestive tract. In contrast, up to 1.0×10⁹ chitinoclasts per g wet wt were determined in control fish. Chitinase did not reflect the absence or presence of bacteria: no significant difference in enzyme activities of cod treated with Ampicillin/Sisomicin and non-treated fish could be demonstrated. A decrease of chitinase activities of cod exposed to Chloramphenicol is interpreted as a toxic effect of this antibiotic on the fish. It is concluded that chitinase activities measured in the digestive tract of G. morhua primarily originate from fish tissues.Results presented in this paper are part of the author's Ph.D. dissertation. Authorization for pre-publication was given by the Graduation Committee of the Biology Department, University of Hamburg     

Influence of temperature and pH on the effects of zinc and copper on proteolytic activities of the intestinal mucosa of planktivorous and benthophagous fishes and their potential preys

Influences of temperature (0 and 20°C) and pH (3.0, 5.0, and 7.4) on the effect of zinc (Zn) and copper (Cu) on proteolytic activities of intestinal mucosa in planktivorous (blue bream, bleak), bentophagous (bream, roach) fishes and their potential preys (pond snail Limnaea stagnalis, planorbid Planorbarius purpura, dreissena Dreissena polymorpha, midge larvae Chironomus sp., water flea Daphnia longispina and total zooplankton) are revealed in this article. Cu decreases the caseinolytic and hemoglobinolytic activities in both fish and their preys more than Zn at temperature 20°C and pH 7.4. Low temperature intensifies the negative effect of the metals on the protease activity in fish (in the case of Zn 5–10 times, in the case of Cu 5–30 times). In fish prey species, the negative effect of the metals on the proteinase activity may be more significant. The influence of pH on Zn and Cu effects is less pronounced than that of low temperature. Maximum reduction of enzyme activities is observed for the combined action of low temperature and pH as well as of the studied metals.     

Mercuric chloride induced proteotoxicity and structural destabilization in marine fish (Therapon jarbua)

The reaction byproducts derived from lipid peroxidation (LPO), as well as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) enzyme activities were measured in tissues of marine fish (Therapon jarbua) exposed to mercuric chloride (HgCl) in water dispersions of 0.125 or 0.25 ppm. LPO was significantly increased in various tissues relative to control values after 96-h exposure. SOD and GPx activities significantly decreased after exposure to first two doses but significantly elevated CAT in Dose II kidney and liver tissues. The elevated levels of LPO, decreased activities of SOD and GPx, and increased CAT activities in all tissues examined in T. jarbua are an index of oxidative stress in fish. Structural analysis by scanning electron microscopy studies revealed the structural deformation in HgCl₂-exposed animals. These observations suggest that HgCl acts as a mediator in free radical generation. The increase in CAT and decrease in SOD and GPx activities in these tissues may be an adaptive response.     

Oxidative stress and genotoxicity of 1-methyl-3-octylimidazolium chloride on loach (Misgurnus anguillicaudatus)

This study was a preliminary step to evaluate the acute toxicity of 1-methyl-3-octylimidazolium chloride ([C₈mim]Cl) on loach (Misgurnus anguillicaudatus) by determining the effects on hepatic antioxidant enzyme activities and by the comet assay. The results showed that [C₈mim]Cl had acute toxicity at concentrations above 20 mg L^?1, inducing oxidative stress and genotoxicity on fish liver cells. In respect to enzyme activities, [C₈mim]Cl induced changes in the activities of superoxide dismutase, catalase, and glutathione content the livers of fish exposed at 20–80 mg L^?1. [C₈mim]Cl at the same exposure level caused a remarkable increase in malondialdehyde level. The comet assay indicated that [C₈mim]Cl at 20–80 mg L^?1 induced genotoxicity in liver cells. With increased exposure concentration and time, the two comet parameters trailing rate and tail moment were significantly increased, with significant differences (P < 0.05) observed between control group and each treatment group. The present study shows that ionic liquids can be a threat to the health of aquatic organism when accidentally released to aquatic ecosystems.     

Wirkung subakuter PCB-Exposition (Aroclor 1254) auf Sauerstoffverbrauch, Schwimmbewegung und Biotransformation (GST-Aktivität) des Karpfens (Cyprinus carpio)

Objective and Background

PCB (polychlorinated biphenyle) are typical man-made environmental pollutants that cause a broad spectrum of effects in vertebrates. Although a lot has been studied about the toxic effects of xenobiotic substances on aquatic organisms, their subacute effects are not yet well known. Fish occupies a central position in freshwater food webs and therefore, carp was chosen to be our test organism. The aim of our study was the investigation of subacute effects of Aroclor 1254 (22μg ^l?1) on respiration, swimming activity, and biotransformation, allowing discussion of changed interaction between the available energy resources.

Methods

The respiration experiments under controlled laboratory conditions were run for 29 days (5 d before, 16 d with and 8 d after chemical exposure) with exposed carp compared to the non-exposed fish. Furthermore, the BehavioQuant system quantitatively monitors the positions of each individual fish before (8 d) and during (21 d) PCB exposure and swimming activity (number of horizontal turnings s^?1) of animals was calculated. Thereafter, liver samples were taken from animals for analysis of the phase 2 enzyme activity (glutathione-S-transferase). In the respiration experiments were 6 exposure, 6 controll, and 3 solubilizer controll groups (12 animals each group); in the behavioral experiments were 9 exposure, 6 controll- und 3 solubilizer controll groups (6 animals each group), and in the enzyme activity measurements were 9 exposure, 6 controll and 3 solubilizer controll groups (6 animals each group) examined.

Results and Discussion

A clearly increased (33%) oxygen consumption due to the PCB-exposure is found to be reversible; after the exposure period the respiration of carp shows a level comparable to the oxygen consumption during non-exposure time. By evaluating quantitative behavioral parameters of animals, it becomes evident that the exposure to PCB also causes a reversible change in their swimming behavior. Chemical stress leads to a decrease of mean daily swimming activity combined with an increase of the mean number of turnings during the night. Thereafter, the swimming activity shifts to the circadian swimming behavior under control conditions. Furthermore, our measurement of soluble and microsomal glutathione-S-transferase activity of fish liver shows a significant elevation after exposure period.

Conclusion

Our results prove that it was feasible to detect sublethal effects of PCB-mixture on all parameters under these conditions successfully. Our findings imply that fish are able to cope with the chemicals and we purse the hypothesis that the higher physiological energy demand caused by increased activity of enzymes, which are involved in biotransformation of foregin substances, may be related to a higher respiration of fish stressed by xenobiotics.

Recommendation and Outlook

Further experiments should investigate the effects of PCB on ecophysiological parameters of carp fed with exposed natural food allowing discussion of ecological consequences.     

Effects of environmental hypercapnia on hemato-immunological parameters,carbonic anhydrase,and Na+, K+-ATPase enzyme activities in rainbow trout (Oncorhynchus mykiss) tissues

In this study, the effects of environmental hypercapnia on hemato-immunological parameters and the activities of respiratory enzymes such as carbonic anhydrase (CA) and Na⁺, K⁺-ATPase were investigated in rainbow trout (Oncorhynchus mykiss) tissues (gill, liver and kidney). Batches of 12 fish were exposed to 4.5 mg L^?1 (control) and 14 mg L^?1 CO₂. No mortalities occurred during the 14 days of the experimental period. Red blood cell (RBC), hemoglobin (Hb), and hematocrit (Ht) levels, and innate immune parameters such as nitro blue tetrazolium (NBT), lysozyme, and myeloperoxidase activities, and the melano-macrophage frequency were negatively affected by elevated CO₂ levels. Patterns of change in CA activity differed among the gill, liver, and kidney. Compared with the activities of CA in the control group, the CA enzyme was significantly stimulated at day 7 in the gill tissue, whereas it was stimulated at day 14 of the experiment in the liver tissue of fish exposed to 14 mg L^?1 CO₂ (P < 0.05). In contrast to the pattern of CA enzyme activities, the Na⁺, K⁺-ATPase enzymes were stimulated significantly in the liver after day 7 but inhibited in the kidney and gill (P < 0.05). These results suggest that a subchronic exposure to hypercapnia of rainbow trout tissues may lead to adaptive changes in the respiratory enzymes and negatively affects hemato-immunological parameters.     

Biomarkers of oxidative stress as indicators of water pollution in Nile tilapia (Oreochromis niloticus) from a water reservoir in Riyadh,Saudi Arabia

Antioxidant enzyme activities of fish (Oreochromis niloticus) were determined in order to assess the status of pollution in the Wadi Namar (WN), near Riyadh, Saudi Arabia. Activities of four antioxidant enzymes as superoxide dismutase (SOD), catalase (CAT), glutathione S-transferase (GST), and concentrations of glutathione (GSH) and oxidant malondialdehyde (MDA) were selected as bioindicators. Fish (n = 14) were sampled in the month of April 2013 from WN and a control site (CS). SOD activity was increased by 37.9%, 47%, and 29% in kidney, liver, and heart, respectively, while a significant decrease (36.4%) was observed in gills of O. niloticus from WN as compared to fish from CS. CAT activity was reduced by 51%, 55%, 47%, and 35% in kidney, liver, heart, and gills of O. niloticus from WN. The GST activities in kidney, liver, and heart of O. niloticus from WN were elevated by 34%, 48%, and 32%, respectively. However, significant fall (49%) in gills of fish was noted from WN compared to fish from CS. GSH levels were increased by 44%, 36%, and 38% in kidney, liver, and heart, respectively, but decreased by 30% in gills. MDA levels of O. niloticus were significantly increased in kidney, liver, and heart in fish from WN. Data indicated that WN is polluted mainly by industrial and urban discharge of liquid waste products.     

Effect of cobalt-supplemented diets on bioaccumulation,digestive enzyme activities and growth of Cyprinus carpio

Experiments were conducted to determine the effects of excess dietary cobalt (Co) on bioaccumulation, digestive enzyme activities, and growth in freshwater Cyprinus carpio. Four isonitrogenous diets (average crude protein: 35%) were formulated to prepare a control diet (T₁) with no Co, and three Co-supplemented diets with 1 (T₂), 1.5 (T₃), or 2 (T₄)% Co. The results showed that C. carpio fed with T₃ diet showed maximal apparent protein digestibility, feed conversion, protein utilization, and growth. Protease and lipase activities were maximal in the fish group given T₃ diet. Accumulation of Co in different soft and hard tissues of fish did not correlate with levels of Co in the diet. The concentration of Co in the water increased with dietary level, being at a disturbing adverse effect level at 2% dietary Co. Evidence indicates that an additional supply of dietary Co up to a level of 1.5% is a viable option to augment growth of C. carpio, but at higher levels of Co it may be detrimental to fish health and the aquatic ecosystem.     

Cloning and expression of high choriolytic enzyme,a component of the hatching enzyme system,during embryonic development of the marine ornamental fish<Emphasis Type="Italic"> Chrysiptera parasema</Emphasis>

In the present study, a cDNA for the hatching enzyme of a marine tropical fish, Chysiptera parasema, was cloned. This is the first demonstration of hatching enzyme cDNA from a marine tropical fish. The amino acid (aa) sequence deduced from the cDNA consisted of an 18-aa signal sequence, a 53-aa propeptide sequence and a 196-aa mature enzyme portion, having a consensus active site sequence for astacin family proteases. Phylogenetic analysis showed that the C. parasema enzyme was included in the clade of HCEs (high choriolytic enzymes), one of the hatching enzymes of freshwater fishes such as medaka (Oryzias latipes), masu salmon (Oncorhynchus masou) and zebrafish (Danio rerio), but not in the group of LCEs (low choriolytic enzymes), another type of hatching enzymes identified in the medaka. The developmental expression patterns of the C. parasema HCE gene were highly similar to that of the medaka HCE gene. The results suggested that the hatching enzyme system is highly conserved between these marine and freshwater fish species.Communicated by R. Cattaneo-Vietti, Genova     

离体模型预测鱼体肝脏清除速率：鲑鱼肝细胞和肝脏S9组分的比较

将离体肝细胞和肝脏S9组分用于获取鱼类的体外生物转化数据可以优化模拟评估对化学物质的生物富集作用。然而涉及2种方式之间的直接对比的研究却几乎没有。本研究采用冷藏保存的鲑鱼肝细胞来测定对于6种多环芳烃(PAHs)的体外本征清除速率。我们运用测定结果推测体内本征清除速率,并将其作为输入值输入一种充分搅匀的肝脏模型中来预测肝清除速率。将事先由体外灌流肝脏测定的速率作为参考来评价预测结果。在2种竞争结合的假说前提下,由鲑鱼肝细胞测出的肝清除速率与实现的测定结果基本一致(6种多环芳烃中的5种都保持在2.1倍差异以下)。尽管多环芳烃的高代谢率是可能的原因之一,这些发现与之前由肝脏S9组分得出的结果相似。对苯并芘这一种化合物而言,由S9数据得出的体内本征清除速率是由肝细胞得出结果的10倍左右,这一结果可能是由细胞吸收速率造成的传播限制引起的。尽管苯并芘的结果差异较大,由任何一种体外测试方法得出的体内本征清除速率结果通常是一致的。这些结果显示离体肝细胞和肝脏S9组分2种系统均可用于优化鱼类的生物富集评估,尤其对于体外反应速率较高的情况。不同系统在化工领域的应用性是否相同则需要进一步的研究工作。
精选自Kellie A. Fay, Patrick N. Fitzsimmons, Alex D. Hoffman, John W. Nichols. Comparison of trout hepatocytes and liver S9 fractions as in vitro models for predicting hepatic clearance in fish. Environmental Toxicology and Chemistry: Volume 36, Issue 2, pages 463–471, July 2017. DOI: 10.1002/etc.3572
详情请见http://onlinelibrary.wiley.com/wol1/doi/10.1002/etc.3572/full
     

Metabolic constraints on burst-swimming in the Antarctic teleost Notothenia neglecta

Immediate-energy-supply pathways were analysed during burst-swimming in white muscle of the Antarctic fish Notothenia neglecta, (collected during Austral summer 1984 near Signy Island, South Orkneys). Selected enzyme activities were determined, and tissue metabolites were measured, before and after 3 min of burst-work. The activity of glycolytic enzymes is very low compared to that in other fish, while the activities of creatine phosphokinase, adenylate kinase and AMPdeaminase are relatively high. Concentrations of metabolites associated with cellular energy status declined with exercise. At the same time, there was no change in the concentrations of most of the glycolytic metabolites measured (including glycogen, glucose and lactate), while [creatine phosphate] declined and [inosine monophosphate] increased. These observations were used to propose that the potential for ATP generation via creatine-phosphate hydrolysis is enhanced in N. neglecta, while glycolytic capacity is relatively reduced. This is an unusual pattern of ATP production and places severe constraints upon the behavior of the fish. While the capacity for very short-term, high-power-output swimming is increased, the capacity for medium-duration swimming is reduced. Ecological repercussions are discussed.     

Cytogenotoxicity and histopathological assessment of Lekki Lagoon and Ogun River in Synodontis clarias (Linnaeus, 1758)

The cytogenotoxicity and histopathological alterations induced by xenobiotics in Lekki Lagoon and Ogun River on Synodontis clarias were investigated. Fish from these water bodies and a fish farm (control) were examined for micronucleated, binucleated, and immature erythrocytes in both gill and peripheral blood. Also gill, liver, kidney, and ovary were processed for histopathology using hematoxylin-eosin staining. Concentrations of cadmium, zinc, lead and copper in the water were determined. There was significant (p < 0.05) increase in micronucleated, binucleated, and immature erythrocytes in both gill and peripheral blood of S. clarias from the lagoon and river compared to the reference site. Loss and disorganization of the primary and secondary lamellae, multifocal degeneration, hemorrhages, cellular infiltration, congestions, vacuolations, atresia, and necrosis were common lesions in the examined tissues of fish from the lagoon and river. Cd, Zn, Pb, and Cu in water samples from the lagoon and river were higher than the reference site. Xenobiotics in Lekki Lagoon and Ogun River, mostly metals, induced deoxyribonucleic acid (DNA) and pathological damage in S. clarias.     

Feeding energetics and metabolism in demersal fish species from Antarctic,temperate and tropical environments

The energetics of feeding has been investigated in demersal fish with similar sedentary lifestyles from the Antarctic (Notothenia neglecta Nybelin), North Sea (Myoxocephalus scorpius L.) and Indian Ocean (Cirrhitichys bleekeri Bleeker). In general, the metabolic rates of fasting individuals were positively correlated with adaptation temperature: values for a standard 100 g fish (mg O₂/h) were 3.3 for N. neglecta at around 0 °C, 2.7 for winter-acclimatized M. scorpius at 5 °C, 4.3 for summer-acclimatized M. scorpius at 15 °C, and 7.0 for C. bleekeri at 25 °C. In all species, following a single satiating meal, oxygen consumption increased to a peak of 2 to 3.5 times the fasting values. Maximum rates of oxygen consumption after feeding were several-fold higher in the warm-than in the cold-water species. After controlling for the effects of body mass and energy intake by analysis of covariance, the duration of the increase in metabolic rate, referred to as specific dynamic action (SDA), was found to be 3 to 4 times shorter in the warm- than in the cold-water fish, ranging from 57 h in C. bleekeri to 208 h in N. neglecta. In contrast, the SDA was not significantly different in the various species, corresponding to 15 to 23% of the energy ingested. Seasonal influences on metabolism and feeding were also studied in N. neglecta acclimated to simulated winter (-1.0 to-0.5 °C; 3 h light:21 h dark) or summer (0 to 0.9 °C; 21 h light:3 h dark) conditions. The metabolic rates of fasting and fed individuals, and the characteristics of the SDA were found to be independent of acclimation conditions. This suggests that N. neglecta is capable of processing food at similar rates throughout the year. Energy stores and enzyme activities were measured in the swimming muscles and liver of fish fed ad libitum. Summer-acclimated fish had higher concentrations of liver triglyceride stores and elevated activities of some enzymes of intermediary metabolism relative to winter-acclimated fish. The observed changes in intermdiary metabolism are probably related to annual cycles of growth and reproduction. It is suggested that the low aerobic scope for physiological performance in Antarctic fish may necessitate the seasonal switching of energy allocation between growth and reproduction.     

Bioaccumulation of cobalt,digestive enzyme activities,and growth of freshwater catfish,Heteropneustes fossilis (Bloch), fed cobalt-supplemented diets

Experiments were conducted to determine the effects of excess dietary cobalt (Co) on its bioaccumulation, on digestive enzyme activities, and on the growth of freshwater catfish Heteropneustes fossilis (Bloch). Four isonitrogenous diets (average crude protein: 32.7%) were formulated to prepare a control diet (T1) with no Co, and three Co-supplemented diets with 0.05 (T2), 0.1 (T3), and 1.0 (T4)% Co. The results showed that H. fossilis fed with T3 diet showed maximum apparent protein digestibility, feed conversion, protein utilization, and growth. Protease and lipase activities were also maximum in T3 diet. Accumulation of Co in different soft and hard tissues of the fish did not show any correlation with the level of Co in the diet. The concentration of Co in the water increased with its dietary level, being at an alarming level at 1.0% dietary Co. It is concluded that an additional supply of dietary Co up to a level of 0.1% is a viable option to augment growth of the catfish H. fossilis, but at higher levels of Co it may be detrimental to the fish and the aquatic ecosystem.     

Physiological responses of Cladophora glomerata to cyanotoxins: a potential new phytoremediation species for the Green Liver Systems

Due to increased pollution of potable water sources as a consequence of eutrophication and anthropogenic xenobiotics, sustainable water purification is an essential concern. Therefore, the Green Liver System, a natural, economic and sustainable water purification system employing the biotransformation capabilities of aquatic plants, was developed. To expand the capacities and applications of this system, new aquatic plants are continually evaluated for their potential to remediate various aquatic pollutants. In this study, the potential of Cladophora glomerata to internalize cyanotoxins, microcystins (MCs) and anatoxin-a, and consequently its ability to cope with the subsequent oxidative stress associated with toxin-uptake were investigated. C. glomerata was able to take up all three of the tested MC congeners as well as anatoxin-a, similarly to previous toxin internalizations reported for aquatic plants such as Ceratophyllum demersum, Myriophyllum spicatum and Hydrilla versiculata. The antioxidative stress defense of C. glomerata proved to efficiently endure the toxin-uptake with no adverse effects. Subsequently, the uptake potential of C. glomerata was investigated at lab-scale by exposure to the three MC congeners and anatoxin-a collectively. After a period of seven days, 95–97% of the MCs and 100% of anatoxin-a were removed from the exposure media. C. glomerata therefore, is a suitable candidate to be incorporated in future Green Liver Systems.     

Enzymatic data as a criterion for characterizing two populations of Disostichus eleginoides (Notothenioidei)

An enzymatic parameter was examined with a view to providing a supplementary criterion to individualize separate geographical populations of a single fish species, Disostichus eleginoides (Notothenioidei). An acid phosphatase of specimens fished from the Crozet and Kerguelen islands in February 1982 and February 1983 was studied and compared between populations. The enzyme was extracted from the white back-muscle of fresh fish and proved to be very similar in both groups, except that their K _m s differed slightly. The optimum temperature for in vitro enzyme activity was 30°C, optimal pH was 5; the active purified polypeptides had a molecular mass of 55 kdaltons and exhibited very low substrate-specificity.     

Characterisation of cysteine proteinase activities in the digestive tract of juvenile <Emphasis Type="Italic">Paragnathia formica</Emphasis> isopods,ectoparasites of estuarine fish

Juveniles of Paragnathia formica Hesse (Isopoda; Gnathiidae) are haematophagous ectoparasites, feeding on fish blood which supplies the nutrients for their development through growth and moulting, and the subsequent survival and reproduction of the free-living adults. Little is known of the mechanisms of digestion in juvenile gnathiids, and biochemical studies on the digestive tract of these interesting estuarine isopods have not been undertaken previously. Here, functionally active cathepsin-like cysteine proteinases are identified in the digestive system of juvenile praniza (fed) and zuphea (unfed) forms. The physiological pH of the digestive tract and the optimum proteolytic activities detected in praniza 3 homogenates using the cathepsin B/L, cathepsin B, and cathepsin H fluorogenic substrates, N-Carbobenzoxy-Phe-Arg-4-methoxy-2-naphthylamine (Z-phe-arg-MNA), N-Carbobenzoxy-Arg-Arg-4-methoxy-2-naphthylamine (Z-arg-arg-MNA) and Arg-4-methoxy-2-naphthylamine (H-arg-MNA), respectively, are in the acidic range (pH 5.8–6.7). Inhibition profiles against Z-phe-arg-MNA and Z-arg-arg-MNA using the cathepsin B inactivator urea, and cysteine proteinase inhibitors, support the presence of cathepsin L- and B-like enzymes. These proteolytic activities are 10–50 times higher in homogenates of praniza 3 compared with zuphea 3 forms. Histochemistry of praniza 3 sections reveals that the predominant enzyme activity towards Z-phe-arg-MNA is limited to the digestive glands during early and mid stages of digestion; later, this activity appears in the lining of the anterior hindgut. Moreover, activity towards Z-arg-arg-MNA is generally restricted to the digestive glands, and only occasionally present in the anterior hindgut. In conclusion, the digestive glands are the main site of cathepsin-like cysteine proteinase activities in P. formica juveniles; these enzymes appear to be important to the digestion of host fish blood enabling development through to the free-living adult.