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«: AGROCHEMICALS: FATE IN FOOD AND THE ENVIRONMENT PROCEEDINGS OF A SYMPOSIUM, ROME, 7 - 1 1 JUNE 1982 JOINTLY ORGANIZED BY IAEA AND FAO l^J I N T E R ...»

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Sources of pesticides are runoff, leaching, adverse deposition of pesticides from agriculture and, in certain areas, industrial sources. Several bioassays for toxicity tests are described. For photoautotrophic activity (algae, particularly Scenedesmus quadricauda) growth is monitored by: turbidity, cell counting, chlorophyll determination, oxygen production or 14 C incorporation;

for heterotrophic activity: oxygen consumption, dehydrogenase activity, grazing capacity of protozoa, acidification by Pseudomonas, ATP, production of НТО or 14 C-dioxide from 14 C- or 3H-acetate or glucose; for higher animals: adverse behaviour or death. Radioenzymatic and radioimmunoassays have also been used. Often adverse effects, e.g. on fish (taste, poor health), are observed, but unequivocal causative relationships to particular pesticides or polluters can seldom be established. Metabolic transformations of pesticides in organisms are very involved;

they include: oxidation, hydrolysis, reduction, conjugation, dehydrohalogenation, ring scission, hydration and chelation (metals). Some guidelines are established and experiments, particularly with 14C label, are reported. Bioaccumulation occurs mainly in fat and bone. Concentrations take place in the food chain. Research methods are similar to those for degradation. In water, pesticides may be transported as dissolved material or by adhering to suspended sediments.

Some studies on isotopic and non-isotopic sediment tagging are reported.

–  –  –

1. INTRODUCTION Although extended use of pesticides has proved beneficial, as in the production o f f o o d, it has also developed into a major threat to the environment. Pesticides will either accumulate in the soil, evaporate, degrade or enter fresh water bodies.

These events will be discussed with respect to the application o f radionuclides in

the context o f the following properties:

–  –  –

Radiotracers as well as stable isotopic tracers are powerful tools in experiments on these properties.

A short primer on limnic metabolism will be given and the numerous applications o f nuclear and some non-nuclear methods for pesticide research will be described.

Limnic metabolism Some aspects are unique to water ecosystems compared with land systems.

Water supports the organisms mechanically and supplies dissolved nutrients.

These are the properties that allow development o f plankton, a form o f life unknown to land systems. Concentration o f dissolved oxygen and carbon dioxide or carbonate is not constant, as it is in air. The hypolimneon o f deep lakes, or the whole pelagic o f shallow lakes at night, may be depleted o f oxygen and enriched in carbon dioxide.

Primary production is restricted to a lighted, warm, oxygen-rich surface layer (euphotic zone) where it exceeds community respiration. When dead organic material sinks to the bottom it causes oxygen consumption and carbon dioxide production, which may use up all the oxygen in the depths and cause strongly reducing anoxic conditions. Oxygen is supplied either from photosynthesis (biogenic aeration) or from the atmosphere (atmospheric aeration).

Thermal stratification divides a lake into three layers o f different temperature and density: epilimneon (warm), metalimneon (transient zone) and hypolimneon IAEA-SM-263/32 (cold), if it is deep enough. In moderate climates this stratification is so stable during the summer that oxygen supply to the hypolimneon is prevented completely.

Tropical lakes may even show permanent stratification. Detailed discussions o f these relations are given in Goltermann's text b o o k [1 ] and in Ref. [2]. Lake metabolism under oxygen-depleted conditions is described in Refs [3—5].

Almost all water quality problems centre around the balance or imbalance between production and mineralization, and oxygen supply and oxygen consumption. In shallow lakes wind forces are usually strong enough to prevent stratification during the summer. However, lack o f biogenic aeration during the night may also cause oxygen depletion. In rivers turbulence prevents stratification.

Atmospheric aeration can support a remarkable respiratory turnover, which mineralizes planktic detritus or sewage (self-purification). However, under heavy organic charge large, slow-flowing lowland rivers may be depleted o f oxygen.

Primary productivity, if it is as warm and bright as in summer, is limited by the availability o f plant nutrients. Increased supply (eutrophication) leads to a series o f largely undesirable events, such as intense algal growth affecting water-work operation and/or production of toxines [6], overcharge o f the self-purification capacity, killing o f fish, damage to reed belts and less species diversity. Lake trophic indices have been compiled by Shapiro [7]; for detailed discussions o f eutrophication see Refs [1, 8, 9].

In most cases phosphorus is the first limiting factor for primary productivity.

Hence, the phosphorus balance o f lakes and their watersheds deserves particular attention. Once an algal cell dies most o f its phosphorus content leaves the cell at a much faster rate than other cell material (particularly the cellulose walls) is mineralized. Thus, long before the cell detritus has disappeared the phosphorus again helps to build up new organic material (short-circuited phosphorus metabolism [8]).





Biomass is reduced in each step o f the f o o d chain, often by a factor o f 10 (details are given in Réf. [1 ]). This means that the biomass of plants is much greater than that o f all animals. In spite o f this animals play an important role in accumulation problems. They contain more lipids than plants [9] and hence they accumulate hydrophobic xenobiotics, e.g. many insecticides.

2. ECOTOXICITY

Discussion o f ecotoxicity often causes a dilemma. Frequently ecosystems are observed that show effects like changes in species composition, adverse behaviour o f animals or increased occurrence o f sickness, and it is generally known that such systems suffer from pollution by a complex mixture o f eutrophicating, toxic and other, probably not dangerous, chemicals. However, it is not always possible to establish a causal relationship between pollution and its effect [10].

130 ERNST On the other hand, there are many ways of determining the toxic levels o f selected pesticides or certain mixtures on a well-defined biological property, such as respiratory turnover, photosynthetic activity, etc., some of which are discussed later in this section. Such results establish a stringent relationship between pollution and one o f its effects, sometimes called 'hard' methods in contrast to the 'soft' information described above, but the concentrations causing significant effects are usually much higher than those observed in the abovementioned complex cases (see Refs [ 1 1 - 1 3 ] ). Radionuclide methods are usually typical examples o f 'hard' experiments. Turnover rates o f tagged metabolites are accurately measured under well-defined laboratory conditions. Several authors have pointed out that such experiments should be carried out under conditions as near to nature as possible [14, 15]. Armstrong [16] reports an interesting relation between a 'hard' result (the toxicity of waters o f Galveston Bay, Texas, was checked in a bioassay measuring the growth depression o f blue green algae) and its general ecological relevance (growth depression is correlated to species diversity).

The essential process o f self-purification o f water bodies is the heterotrophic activity of aquatic bacteria. It is usually limited by oxygen depletion or it may be 'self-limited', i.e. the bacterial activity cannot compensate for the influx o f organic material. One o f the major concerns o f ecotoxicity is the question o f whether pesticides reduce self-purification in rivers or whether they affect the proper operation of sewage treatment plants.

Several reviews exist on pesticide interaction on the microflora o f soils and water; they report on the negligible influence of pesticides on community gross respiration provided they are applied in reasonably low concentrations [13, 15, 17].

Sewage plants may be affected by intense pulses o f poison, although such events seldom occur in urban sewage. A remarkable easing o f the organochlorine charge in rivers has been observed since the DDT ban. In the Federal Republic o f Germany, for example, lindane and HCB are now o f major concern, but these pesticides are controlled more because o f their importance to the human f o o d chain than their environmental toxicity. However, there are regionally different situations in particular cases. In the lower Rhine self-purification is assumed to be reduced by one-third owing to a complex pattern of environmental chemicals, including pesticides [18]. Such a reduction may be caused either by damage to the heterotrophic activity which reduces the degradation o f organic burden, or by lower primary production which reduces biogenic aeration; this may play an important role in large, slow-flowing lowland rivers. Such a burden is more remarkable, both from the economical and ecological standpoint, than occasional killing o f fish, although it is less visible to the public.

A case of damage to primary productivity (measured by the 14C method) by papermill effluents is reported in Ref. [19]. Butler's review [14] compiles the effects o f numerous herbicides, insecticides and fungicides on algae. Usually, 1 4 C-dioxide uptake or growth has been used to establish the toxicity levels of IAEA-SM-263/34 131 phytoplanktic organisms. Particularly dangerous are the phenylurea derivatives [14, 17, 20], whereas 2,4D is tolerated to much higher levels (approximately 100 p p m ) [21].

A study o f nonachlor and chlordane effects in the marine fauna o f f the Canadian east coast has not yet revealed toxic effects [22]. The bulk o f the pesticides used are either herbicides acting on phytoplankton or insecticides acting on the nerves o f higher animals. An example o f the latter is given in Ref. [23], where the insecticide affected fish and zooplankton, whereas it caused a b l o o m in hydra, which does not have nerves. In experiments with Chlorella pyrenoidosa it has been shown [ 2 4 ] that exposure to HCB at levels o f 0.1, 1.0 and 5 ppm inhibited photosynthesis but not, or hardly ever, respiration.

In bioassays for toxicity tests one may differentiate according to:

(1 ) If the time o f incubation is short compared with the Time of incubation.

life span of the test organism the test is called acute. Mortality, metabolic irregularities and adverse behaviour may be observed. In chronic tests the exposure time is comparatively long. Long-term metabolic and reproductive damage is checked.

(2) If the pesticide is administered once and the water is not changed Test form.

the test is called static. Pesticide concentration will decrease during exposure.

If it is to be kept constant automated flow-through methods have to be used.

(3) Primary productivity tests require illuminated assays for algal Trophic level.

growth. Test parameters may be: oxygen evolution, 1 4 C-dioxide incorporation, chlorophyll measurements or cell counts. The first two parameters are well known in limnology for the measurement o f primary productivity; for details see Refs [25—30]. Chlorophyll may be measured photometrically after extraction, if necessary even after chromatographic separation. All these methods require some care concerning incomplete extraction and proper discrimination between the different photosynthetic pigments, particularly as far as the unintentional degradation to phaeopigments is concerned; details are reported in Refs [28] and [31].

These problems are circumvented by in vivo fluorometric methods, which also save much work. However, in living algae the quantum efficiency may vary with physiological conditions. This can be avoided by blocking the electron transport chain with DCMU or CMU [ 3 1 - 3 4 ].

Pesticide concentrations preventing growth in phytoplankton are listed in Ref. [17]; they range from 0.02 ppb (urea derivatives) to 1000 ppm (trichlorphon).

Fresh water algae that are frequently used for such tests are: Chlorella sp., which are distinguished by their storage capacity o f lipophilic substances [35], aeruginosa and Scenedesmus quadricaudae, Skeletonema costatum, Microcystis Anabaena flos-aque.

ERNST For high trophic levels Daphnia and Guppy, Golden Orfe which have some central position in j3-mesosaprobic waters [35], or Rainbow trout, as well as the mussel Dreissena polymorpha, are standard organisms. The relative toxicity o f several pesticides to Daphnia are given in Ref. [17].

The heterotrophic activity o f microorganisms has frequently been determined by measuring the mineralization o f 14 C-glucose, 14 C-acetate and corresponding 3 H-labelled substances. ATP measurement by the luziferine-luziferase method [36] has also been used. Techniques to trap the 1 4 C-dioxide are described in Refs [ 3 7 - 3 9 ].

Albright has used heterotrophic activity methods with 14 C- or 3 H-labelled organic substrates (glucose, acetate or amino acids) to determine the effect o f 11 heavy metals; 3 H-substrates can be achieved at higher specific activities. Since natural substrate concentrations are very low, such a high specific activity may be necessary not to disturb the natural system by t o o high a substrate addition.

Michaelis-Menthen-kinetics, and deviations from it, have been found [37].

Further details o f heterotrophic radiotracer techniques are described in Refs [38—40].

Since most insecticides act via nerve intoxication, anti-cholinesteratic activity is an important property. Horváth [41 ] and Horva'th and Forster [42] have described an extremely sensitive method using carboxyacetylcholine as substrate and the H liquid scintillation technique.

3. D E G R A D ABILITY

Metabolic transformations o f the numerous pesticides in numerous organisms modified by numerous different conditions are very involved. It is impossible to establish simple unequivocal guidelines.

The major types o f reactions are: oxidation, hydrolysis, reduction, conjugation, dehydrohalogenation, ring scission and, important for heavy metals, chelation.

Despite these difficulties it is reasonable to establish a few guidelines which, however, should be applied carefully.



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