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Reactive nitrogen and public policies for environmental protection.

A core goal of both US and European pollution control policies has been to establish rules and regulations pertaining to the movement of reactive-nitrogen (Nr) through the environment. This is manifest in US federal legislation such as the Clean Air Act Amendments and the Clean Water Act Amendments and in various protocols of the United Nations Economic Commission for Europe (UNECE) and its Convention on Long-Range Transboundary Air Pollution (CLRTAP). In this paper, we begin by reviewing the two US laws and their effectiveness and make some comparisons with the approaches used mainly in Europe by the UNECE and CLRTAP. Next we use the Mississippi drainage/Gulf of Mexico hypoxia case study to highlight the importance of applying a "systems approach" to address the reactive nitrogen problem at the regional scale. After briefly posing a number of unanswered questions related to nitrogen control policies, we conclude by sketching a blueprint for future actions related to the development of improved policies to regulate reactive nitrogen.

Environment↗

Soft ligands--biodegradable complexants in plant cultivation and environmental protection.

In this paper, the place and role of biodegradable complexants in light of modern plant cultivation criteria is discussed. From this point of view, micro- and macroelement (metal) chelators play a remarkable role. They are widely used in a variety of consumer products and processes, especially in agriculture. The great majority of the traditional chelators (e.g., EDTA) are, however, essentially non-biodegradable. We have systematically investigated some new biodegradable ligands with polyaminopolycarboxylic structure. We conclude that these compounds may be considered as a special group of the soft (safer) chemicals proposed first by Bodor based on retrometabolic concepts. We studied the structural conditions of biodegradability, especially regarding the new ligands, by the physicochemical characterization of their complexes formed with micro- and macroelements. The study of the complicated equilibrium conditions required the prior determination of the stability constants of the metal aqua/hydroxo complexes and the protonation constants of the ligands as well. Knowledge of the pH-dependence of the proper competitive equilibriums allowed the optimization of the investigated complexation reactions. The protonation and stability constants obtained were compared with the adequate values of the traditional, but weakly biodegradable EDTA. Excellent or very good linear correlations have been found between the logarithms of the different ligands' protonation constants, but the slopes of the straight lines were different and generally smaller than EDTA's corresponding values. Presumably, the log stability constants (logK(st)) of complexes formed by different ligands (X(m), X(n)) having analogue structures with a given metal-ion series (M(i), i = 1, 2, ...) are linearly correlated [i.e., log K(X(m)M(i)) = a log K(X(n)M(i)) + b], just as the stability constants of complexes formed by different ions (M(k), M(l)) with a given ligand series (X(j), j = 1, 2, ...). Based on the rich data basis of EDTA, these regression equations allow the calculation of some still unmeasured stability constants of the new biodegradable complexes. In the analysis of structural effects we would like to especially highlight the role of stereochemical relations in addition to the usual structural elements. From this point of view, the eventual presence of chiral nitrogen atoms is of particular importance. The stability of these compounds increases in their complexed form. As a special research field, we have examined the possibility of obtaining metal ion buffers. These systems may get an important role both from theoretical and practical points of view.

Biodegradation, Environmental↗

Radiolytic degradation of the herbicide dicamba for environmental protection.

The radiolytic degradation of the widely used herbicide dicamba (3,6-dichloro-2-methoxybenzoic acid), employing gamma irradiation in laboratory batch conditions and with a beam of accelerated electrons in flow-through installation, was investigated. The effects of dose magnitude, ozone or hydrogen peroxide in irradiated solution, and scavengers such as nitrate and hydrogen carbonate on the effectiveness of dicamba decomposition and the products formed were investigated. Changes in the toxicity of irradiated solutions were measured with the Microtox and Spirotox toxicity tests. The application of radiolytic degradation was also examined for decomposition of herbicides in commercial agrochemical preparations.

Aliivibrio fischeri↗

Biotechnological development of effective phytases for mineral nutrition and environmental protection.

Phytases are hydrolytic enzymes that initiate the release of phosphate from phytate (myo-inositol hexakisphosphate), the major phosphorus (P) form in animal feeds of plant origin. These enzymes can be supplemented in diets for food animals to improve P nutrition and to reduce P pollution of animal excreta. This mini-review provides a synopsis of the concept of "ideal phytase" and the biotechnological approaches for developing such an enzyme. Examples of Escherichia coli AppA and Aspergillus fumigatus PhyA are presented to illustrate how new phytases are identified from microorganisms and developed by genetic engineering based on the gene sequences and protein structures of these enzymes. We also discuss the characteristics of different heterologous phytase expression systems, including those of plants, bacteria, fungi, and yeast.

6-Phytase↗

An evaluation of the host-mediated assay and body fluid analysis. A report of the U.S. Environmental Protection Agency Gene-Tox Program.

The methodologies and status of the Host-Mediated Assay were reviewed using the published literature available up to June 1980. The Working Group reviewed 274 documents, including abstracts, research articles, review articles, and publicly available contracts and grant final reports. From this group, abstracts and reviews were rejected from critical evaluation. 77 documents were accepted and reviewed by the Working Group and the test results summarized. These selected documents yielded 208 chemicals that were evaluated in th host-mediated assay. Of these chemicals, 133 were mutagenic in this assay with one or more indicators. 76 chemicals, several of which are not considered to be carcinogenic, were not detected by any of the indicators. Of the 208 chemicals, 125 had been tested in carcinogenicity assay in rodents. 90, or 71%, of the carcinogens were detected as mutagens in the Host-Mediated Assay. In several cases, those carcinogens not detected may have been negative because of improper selection of the indicator. The Working Group concluded that the Host-Mediated Assay is an important test in mutagenicity/carcinogenicity research and that, by proper selection of protocols and indicators, valuable information can be gained that otherwise would be overlooked strict, in vitro assays.

Animals↗

Chromosome aberration assays in Allium. A report of the U.S. Environmental Protection Agency Gene-Tox Program.

The common onion (Allium cepa) is an excellent plant for the assay of chromosome aberrations after chemical treatment. Other species of allium (A. cepa var. proliferum, A. carinatum, A. fistulosum and A. sativum) have also been used but to a much lesser extent. Protocols have been given for using root tips from either bulbs or seeds of Allium cepa to study the cytological end-points, such as chromosome breaks and exchanges, which follow the testing of chemicals in somatic cells. It is considered that both mitotic and meiotic end-points should be used to a greater extent in assaying the cytogenetic effects of a chemical. From a literature survey, 148 chemicals are tabulated that have been assayed in 164 Allium tests for the clastogenic effect. Of the 164 assays which have been carried out, 75 are reported as giving a positive reaction (i.e., causing chromosome aberrations), 49 positive and with a dose response, 1 positive and temperature-related, 9 borderline positive, and 30 negative; 76% of the chemicals gave a definite positive response. It is proposed that the Allium test be included among those tests routinely used for assessing chromosomal damage induced by chemicals.

Chromosome Aberrations↗

Specific-locus mutation assays in Zea mays. A report of the U.S. Environmental Protection Agency Gene-Tox Program.

The use of the angiosperm Zea mays in assays for the induction of mutations at specific genetic loci is discussed. The effect of chemical mutagens on 3 genetic end points is evaluated. The end points surveyed were the loss of the phenotype of dominant alleles in heterozygotes, the induction of segregating point mutations in M2 plants, and the induction of forward and reverse mutations in pollen grains. Maize is sensitive to a wide range of mutagens and has the capacity to activate promutagens. This plant has been used to study mutagenesis under both laboratory and in situ conditions.

Genes↗

An evaluation of the mouse sperm morphology test and other sperm tests in nonhuman mammals. A report of the U.S. Environmental Protection Agency Gene-Tox Program.

The literature on the mouse sperm morphology test and on other sperm tests in nonhuman mammals was reviewed (a) to evaluate the relationship of these tests to chemically induced spermatogenic dysfunction, germ-cell mutagenicity, and carcinogenicity, and (b) to make an interspecies comparison to chemicals. A total of 71 papers were reviewed. The mouse sperm morphology test was used to assess the effects of 154 of the 182 chemical agents covered. 4 other murine sperm tests were also used: the induction of acrosomal abnormalities (4 agents), reduction in sperm counts, (6 agents), motility (5 agents), and F1 sperm morphology (7 agents)). In addition, sperm tests for the spermatogenic effects of 35 agents were done in 9 nonmurine mammalian species; these included analyses for sperm count, motility, and morphology, using a large variety of study designs. For the mouse sperm morphology test, 41 agents were judged by the reviewing committee to be positive inducers of sperm-head shape abnormalities, 103 were negative, and 10 were inconclusive. To evaluate the relationship between changes in sperm morphology and germ cell mutagenicity, the effects of 41 agents on mouse sperm shape were compared to available data from 3 different mammalian germ-cell mutational tests (specific locus, heritable translocation, and dominant lethal). The mouse sperm morphology test was found to be highly sensitive to germ-cell mutagens; 100% of the known mutagens were correctly identified as positives in the sperm morphology test. Data are insufficient at present to access the rate of false positives. Although it is biologically unclear why one might expect changes in sperm morphology to be related to carcinogenesis, we found that (a) a positive response in the mouse sperm morphology test is highly specific for carcinogenic potential (100% for the agents surveyed), and (b) overall, only 50% of carcinogens were positive in the test (i.e., sensitivity approximately equal to 50%). Since many carcinogens do not produce abnormally shaped sperm even at lethal doses, negative findings with the sperm test cannot be used to classify agents as noncarcinogens. We conclude that the mouse sperm morphology test has potential use for identifying chemicals that induce spermatogenic dysfunction and perhaps heritable mutations. Insufficient numbers of chemicals agents have been studied by the other sperm tests to permit similar comparisons. A comparison of 25 chemicals tested with sperm counts, motility, and morphology in at least 2 species (including man, mouse and 9 other mammals) demonstrated good agreement in response among species. With further study, interspecies comparisons of chemically induced sperm changes may be useful for predicting and evaluating human effects.

Animals↗

Testing of chemicals for genetic activity with Saccharomyces cerevisiae: a report of the U.S. Environmental Protection Agency Gene-Tox Program.

The yeast Saccharomyces cerevisiae is a unicellular fungus that can be cultured as a stable haploid or a stable diploid . Diploid cultures can be induced to undergo meiosis in a synchronous fashion under well-defined conditions. Consequently, yeasts can be used to study genetic effects both in mitotic and in meiotic cells. Haploid strains have been used to study the induction of point mutations. In addition to point mutation induction, diploid strains have been used for studying mitotic recombination, which is the expression of the cellular repair activities induced by inflicted damage. Chromosomal malsegregation in mitotic and meiotic cells can also be studied in appropriately marked strains. Yeast has a considerable potential for endogenous activation, provided the tests are performed with appropriate cells. Exogenous activation has been achieved with S9 rodent liver in test tubes as well as in the host-mediated assay, where cells are injected into rodents. Yeast cells can be recovered from various organs and tested for induced genetic effects. The most commonly used genetic end point has been mitotic recombination either as mitotic crossing-over or mitotic gene conversion. A number of different strains are used by different authors. This also applies to haploid strains used for monitoring induction of point mutations. Mitotic chromosome malsegregation has been studied mainly with strain D6 and meiotic malsegregation with strain DIS13 . Data were available on tests with 492 chemicals, of which 249 were positive, as reported in 173 articles or reports. The genetic test/carcinogenicity accuracy was 0.74, based on the carcinogen listing established in the Gene-Tox Program. The yeast tests supplement the bacterial tests for detecting agents that act via radical formation, antibacterial drugs, and other chemicals interfering with chromosome segregation and recombination processes.

Aneuploidy↗

Radiolytic degradation of herbicide 4-chloro-2-methyl phenoxyacetic acid (MCPA) by gamma-radiation for environmental protection.

Reversed-phase HPLC determination of the herbicide MCPA and its products of radiolytic degradation has been optimized. The radiolytic degradation was carried out using gamma-irradiation and was optimized in terms of irradiation dose and pH of irradiated MCPA solution. Decomposition of 100 ppm MCPA in pure solutions required irradiation with a 3 kGy dose. The main products of irradiation in the dose range up to 10-kGy were various phenolic compounds and carboxylic acids. The developed method was applied for treatment of industrial waste from production of MCPA. The 10-kGy dose was needed for decomposition of 500 ppm of MCPA in the industrial waste samples; however, the presence of stoichiometric amount of hydrogen peroxide in the irradiated waste allowed a 50% reduction of the gamma-irradiation dose. Despite complete decomposition of MCPA in the industrial waste, in order to reduce the toxicity of irradiated waste, measured by the Microtox bioluminescence test, higher than a 10 kGy irradiation dose was needed.

2-Methyl-4-chlorophenoxyacetic Acid↗

The potential of cost-effective coconut husk for the removal of toxic metal ions for environmental protection.

Coconut (Cocos nucifera) husk, an agricultural waste, has been thoroughly investigated for the removal of toxic Cd(II), Cr(III) and Hg(II) ions from aqueous media. The parameters like nature and composition of electrolyte, concentration of toxic ions, dosage of coconut husk, and equilibration time between the two phases were optimized for their maximum accumulation onto the solid surface. The effect of common ions on the uptake of metal ions has been monitored under optimal conditions. The variation of retention of each metal ion with temperature was used to compute the thermodynamic quantities DeltaH, DeltaS and DeltaG. The values 18.1+/-0.6 kJmol(-1), 74+/-2 Jmol(-1)K(-1), and -3.8+/-0.04 kJmol(-1) at 298 K; 10.8+/-0.8 kJmol(-1), 48.8+/-2.7 Jmol(-1)K(-1), and -4.6+/-0.3 kJmol(-1) at 298 K; and -37.4+/-2k Jmol(-1), 105+/-7 Jmol(-1)K(-1) and -2.58+/-0.5 kJmol(-1) at 298 K were obtained for Cd(II), Cr(III) and Hg(II) ions, respectively. The sorption data were analysed by applying different sorption isotherms. The sorption capacity and energy were evaluated for each metal ion. The values of the Freundlich constants 1/n and C(m) were 0.92+/-0.04 and 52.6+/-22.2 mmolg(-1); 0.85+/-0.05 and 56.0+/-0.03 mmolg(-1); and 0.88+/-0.03 and 6.84+/-0.45 mmolg(-1) for Cd(II) Cr(III) and Hg(II) ions, respectively. Similarly, the Dubinin-Radushkevich (D-R) constants beta, X(m,) and E were evaluated for the three metal ions. To check the selectivity of the sorbent, sorption of a number of elements was measured under similar conditions. Separation of Zn(II) from Cd(II); Cr(III) from I(I), Zr(IV), Se(IV), and Hg(II) from Se(IV) and Zn(II) can be achieved using this sorbent. This cheap material has potential applications in analytical chemistry, water decontamination, industrial effluent treatment and in pollution abatement.

Adsorption↗