Determination and multivariate statistical analysis of biochemical responses to environmental contaminants in feral freshwater fish Leuciscus cephalus L.

Modulations of 11 prospective biochemical markers of impacts of aquatic pollutants in liver tissue of chub (Leuciscus cephalus), caught at several sampling sites of a river with various pollution types and rates, were matched against analytical data of concentrations of organochlorine compounds, polycyclic aromatic hydrocarbons (PAHs), and heavy metals. Multivariate principal component analysis (PCA) of the field data showed general patterns of biochemical responses to different types of pollutants and relationships among the biomarkers. Cytochrome P4501A-dependent 7-ethoxyresorufin O-deethylase (EROD) activity, inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin and structurally related planar compounds, was strongly enhanced in the more contaminated areas. Compared with polychlorinated aromatic hydrocarbons, PAHs did not contribute so significantly to EROD induction. Testosterone 6 beta- and 16 alpha-hydroxylase activities, as an expression of the cytochrome P4503A27, were slightly increased at several sites but were significantly decreased in samples from some heavily polluted areas. Recently, these activities have been suggested as potential biomarkers of exposure to contaminants that do not induce cytochrome P4501A. In this study, their inhibition or induction was not associated with a specific class of monitored contaminants, and selectivities of these modulations are still to be investigated. Similar modulations of the prospective biochemical indicators of oxidative stress, including microsomal glutathione S-transferase activity, cytosolic glutathione S-transferase with ethacrynic acid, and glutathione reductase, were demonstrated by PCA. The pattern of the modulations of the microsomal nicotinamide adenine dinucleotide phosphate (NADPH)-dependent lipid peroxidation in vitro differed from the responses of the rest of oxidative stress parameters at some sampling sites. Further biochemical markers of oxidative stress under study, including in vivo lipid peroxidation, in vitro production of reactive oxygen species, and the concentration of metallothioneins did not correlate well with the concentrations of the contaminants. Principal component analysis demonstrated that the EROD activity, glutathione-dependent enzymes, and Fe(II)-enhanced lipid peroxidation formed a suitable battery of biomarkers of exposure.

[1]  C. V. Gestel,et al.  Incorporation of the biomarker concept in ecotoxicology calls for a redefinition of terms , 1996 .

[2]  D. Siebers,et al.  Biochemical responses and environmental contaminants in breams (Abramis brama L.) caught in the river Elbe. , 1995, Ecotoxicology and environmental safety.

[3]  M. Machala,et al.  Biochemical markers for differentiation of exposures to nonplanar polychlorinated biphenyls, organochlorine pesticides, or 2,3,7, 8-tetrachlorodibenzo-p-dioxin in trout liver. , 1998, Ecotoxicology and environmental safety.

[4]  E. Jansen,et al.  Measurement of enzyme activities of cytochrome P-450 isoenzymes by high-performance liquid chromatographic analysis of products. , 1991, Journal of chromatography.

[5]  J. H. Zar,et al.  Biostatistical Analysis (5th Edition) , 1984 .

[6]  M. Goldstein,et al.  Multivariate Analysis: Methods and Applications , 1984 .

[7]  G. Roesijadi.  Metallothioneins in metal regulation and toxicity in aquatic animals , 1992 .

[8]  C. Walker,et al.  The role of biomarkers in environmental assessment (3). Vertebrates , 1994, Ecotoxicology.

[9]  C. Weihs,et al.  Multivariate exploratory data analysis and graphics: A tutorial , 1993 .

[10]  R W Olafson,et al.  An electrochemical approach to quantitation and characterization of metallothioneins. , 1979, Analytical biochemistry.

[11]  N. Vermeulen,et al.  Biomonitoring of aquatic pollution with feral eel (Anguilla anguilla). II. Biomarkers: pollution-induced biochemical responses. , 1996 .

[12]  W B Jakoby,et al.  Glutathione S-transferases. The first enzymatic step in mercapturic acid formation. , 1974, The Journal of biological chemistry.

[13]  A. Arukwe,et al.  Xenobiotic and steroid biotransformation enzymes in Atlantic salmon (Salmo salar) liver treated with an estrogenic compound, 4‐nonylphenol , 1997 .

[14]  Y. Shen,et al.  Detection of elevated reactive oxygen species level in cultured rat hepatocytes treated with aflatoxin B1. , 1996, Free radical biology & medicine.

[15]  T. W. Moon,et al.  3,3',4,4'-tetrachlorobiphenyl effects on antioxidant enzymes and glutathione status in different tissues of rainbow trout. , 1995, Pharmacology & toxicology.

[16]  D. Slone,et al.  Channel catfish glutathione S-transferase isoenzyme activity toward (±)-anti-benzo[a]pyrene-trans-7,8-dihydrodiol-9, 10-epoxide , 1996 .

[17]  C. V. Van Gestel,et al.  Incorporation of the biomarker concept in ecotoxicology calls for a redefinition of terms. , 1996, Ecotoxicology.

[18]  V. Palace,et al.  VARIATION OF HEPATIC ENZYMES IN THREE SPECIES OF FRESHWATER FISH FROM PRECAMBRIAN SHIELD LAKES AND THE EFFECT OF CADMIUM EXPOSURE , 1993 .

[19]  P. Thomas,et al.  Effects of cadmium and Aroclor 1254 on lipid peroxidation, glutathione peroxidase activity, and selected antioxidants in Atlantic croaker tissues , 1993 .

[20]  B. Mannervik,et al.  Glutathione transferases--structure and catalytic activity. , 1988, CRC critical reviews in biochemistry.

[21]  N. Vermeulen,et al.  Biochemical markers in feral roach (Rutilus rutilus) in relation to the bioaccumulation of organic trace pollutants , 1994 .

[22]  L. Viganò,et al.  Biomarker responses in cyprinids of the middle stretch of the River Po, Italy , 1998 .

[23]  E. Levin,et al.  Oxidative stress in toxicology: established mammalian and emerging piscine model systems. , 1998, Environmental health perspectives.

[24]  P. K. Smith,et al.  Measurement of protein using bicinchoninic acid. , 1985, Analytical biochemistry.

[25]  M. Machala,et al.  Glutathione‐dependent detoxifying enzymes in rainbow trout liver: Search for specific biochemical markers of chemical stress , 1997 .

[26]  K. Kolaja,et al.  The role of oxidative stress in chemical carcinogenesis. , 1998, Environmental health perspectives.

[27]  C. Miranda,et al.  Cloning, sequencing, and tissue expression of CYP3A27, a new member of the CYP3A subfamily from embryonic and adult rainbow trout livers. , 1998, Archives of biochemistry and biophysics.

[28]  R. Cattell.  The Scree Test For The Number Of Factors. , 1966, Multivariate behavioral research.

[29]  P. Namour,et al.  In situ pollution induced cytochrome P450 activity of freshwater fish: barbel (Barbus barbus), chub (Leuciscus cephalus) and nase (Chondrostoma nasus) , 1991 .

[30]  C. Pueyo,et al.  Biochemical Indicators of Oxidative Stress in Fish from Polluted Littoral Areas , 1993 .

[31]  A. Bainy,et al.  Oxidative stress in gill, erythrocytes, liver and kidney of Nile tilapia (Oreochromis niloticus) from a polluted site , 1996 .

[32]  V. Palace,et al.  Oxidative stress in Lake Sturgeon (Acipenser fulvescens) orally exposed to 2,3,7,8-tetrachlorodibenzofuran , 1996 .

[33]  A. J. Collins,et al.  Interpretation of transformed axes in multivariate analysis , 1993 .

[34]  L. Dušek,et al.  Responses of carp hepatopancreatic 7‐ethoxyresorufin‐O‐deethylase and glutathione‐dependent enzymes to organic pollutants—a field study , 1997 .

[35]  R. Mayer,et al.  Direct fluorometric methods for measuring mixed function oxidase activity. , 1978, Methods in enzymology.

[36]  H. Ischiropoulos,et al.  Evaluation of the probe 2',7'-dichlorofluorescin as an indicator of reactive oxygen species formation and oxidative stress. , 1992, Chemical research in toxicology.

[37]  T. Collier,et al.  Hepatic xenobiotic metabolizing enzymes in two species of benthic fish showing different prevalences of contaminant-associated liver neoplasms. , 1992, Toxicology and applied pharmacology.

[38]  M. Uchiyama,et al.  Determination of malonaldehyde precursor in tissues by thiobarbituric acid test. , 1978, Analytical biochemistry.

[39]  H. Kappus.  12 – Lipid Peroxidation: Mechanisms, Analysis, Enzymology and Biological Relevance , 1985 .

[40]  L. Förlin,et al.  Temporal aspects of induction of hepatic cytochrome P450 1A and conjugating enzymes in the viviparous blenny (Zoarces viviparus) treated with petroleum hydrocarbons , 1994 .