ORGANIC POLLUTANTS: An Ecotoxicological Perspective - Chapter 4 potx

The movement of chemicals along food chains and the fate of chemi-cals in the complex communities of sediments and soils are basic issues here.Ecotoxicology deals with the study of the h

of movement and distribution in the living environment—within individuals, munities, and ecosystems—where biological as well as physical and chemical factors come into play The movement of chemicals along food chains and the fate of chemi-cals in the complex communities of sediments and soils are basic issues here.Ecotoxicology deals with the study of the harmful effects of chemicals in ecosys-tems This includes harmful effects upon individuals, although the ultimate concern

com-is about how these are translated into changes at the levels of population, community, and ecosystem Thus, in the concluding sections of the chapter, emphasis will move from the distribution and environmental concentrations of pollutants to consequent effects at the levels of the individual, population, community, and ecosystem The relationship between environmental exposure (dose) and harmful effect (response) is fundamentally important here, and full consideration will be given to the concept of biomarkers, which is based on this relationship and which can provide the means of relating environmental levels of chemicals to consequent effects upon individuals, populations, communities, and ecosystems

4.2 MOVEMENT OF POLLUTANTS ALONG FOOD CHAINS

The pollutants of particular interest here are persistent organic pounds that have sufficiently long half-lives in living organisms for them to pass along food chains and to undergo biomagnification at higher trophic levels (see Box4.1) Some compounds of lesser persistence, such as polycyclic aromatic hydrocar-bons (PAHs) (Chapter 9), can be bioconcentrated/bioaccumulated at lower trophic levels but are rapidly metabolized by vertebrates at higher levels These will not be discussed further here, where the issue is biomagnification with movement along the

chemicals—com-entire food chain The best studied examples of this are the organochlorines (OCs)

dieldrin and p,pb-DDE (see Chapter 5), and the PCBs (see Chapter 6), where trations in the tissues of predators of the highest trophic levels can be 104–105-fold higher than in organisms at the lowest trophic levels Other examples include poly-chlorinated dibenzodioxin (PCDDs), polychlorinated dibenzofurans (PCDFs), and some organometallic compounds (e.g., methyl mercury)

concen-Biomagnification along terrestrial food chains is principally due to lation from food, the principal source of most pollutants (Walker 1990b) In a few instances, the major route of uptake may be from air, from contact with contaminated surfaces, or from drinking water The bioaccumulation factor (BAF) of a chemical is given by the following equation:

bioaccumu-Concentration in organism/concentration in food = BAF

Biomagnification along aquatic food chains may be the consequence of centration as well as bioaccumulation Aquatic vertebrates and invertebrates can absorb pollutants from ambient water; bottom feeders can take up pollutants from sediments The bioconcentration factor (BCF) of a chemical absorbed directly from water is defined as

biocon-Concentration in organism/concentration in ambient water = BCF

One of the challenges when studying biomagnification along aquatic food chains

is establishing the relative importance of bioaccumulation versus bioconcentration The processes that lead to biomagnification have been investigated with a view to developing predictive toxicokinetic models (Walker 1990b) When organisms are continuously exposed to pollutants maintained at a fairly constant level in food and/

or in ambient water/air, tissue concentrations will increase with time until either (1) a lethal concentration is reached and the organism dies or (2) a steady state is reached when the rate of uptake of the pollutant is balanced by the rate of loss The BCF or BAF at the steady state is of particular interest and importance because (A) it rep-resents the highest value that can be reached and therefore indicates the maximum risk, (B) it is not time dependent, and (C) the rates of uptake and loss are equal, thereby facilitating the calculation of the rate constants involved

BCFs and BAFs measured before the steady state is reached have little value because they are dependent on the period of exposure of the organism to the chemi-cal, and thus may greatly underestimate the degree of biomagnification that is possible This statement should be qualified by the reservation that there may be situations in which the duration of exposure cannot be long enough for the steady state to be reached, for example, where the life span of an insect is very short The principal processes of uptake and loss by different types of organisms are indicated

in Table 4.1 (see also Box 4.2)

A rough indication of the relative importance of different mechanisms of uptake and loss is given by a scoring system on the scale +−> ++++ Within each category of organ-ism there will be differences between compounds in the relative importance of differ-ent mechanisms, for example, due to differences in polarity and biodegradability

BOX 4.1 PERSISTENT ORGANIC POLLUTANTS (POPS)

A list of hazardous environmental chemicals, sometimes referred to as the “dirty dozen,” has been drawn up by the United Nations Environment Programme (UNEP) These are:

POP Year of Introduction Classification

of plastics, PCBs

The selection of these compounds was made on the grounds of their icity, environmental stability, and tendency to undergo biomagnification; the intention was to move toward their removal from the natural environment In the REACH proposals of the European Commission (EC; published in 2003),

tox-a similtox-ar list of 12 POPs wtox-as drtox-awn up, the only differences being the inclusion

of hexachlorobiphenyl and chlordecone, and the exclusion of the by-products, dioxins, and furans The objective of the EC directive is to ban the manufac-ture or marketing of these substances It is interesting that no fewer than eight

of these compounds, which are featured on both lists, are insecticides

TABLE 4.1

Principal Mechanisms of Uptake and Loss for Lipophilic Compounds

Mechanisms of Uptake Mechanisms of Loss Habitat/Type of

From Ingested Water Diffusion Metabolism Aquatic

Terrestrial

BOX 4.2 MODELS FOR BIOCONCENTRATION

AND BIOACCUMULATION

As indicated in Table 4.1, aquatic mollusks present a relatively simple picture because they have little capacity for biotransformation of organic pollutants, the principal mechanism of both uptake loss being diffusion It is not sur-prising, therefore, that bioconcentration factors (BCFs) for diverse lipophilic

compounds, measured at the steady state, are related linearly to log Kow ues (Figure 4.1) Thus, the more hydrophobic a compound is, the greater the tendency to partition from water into the lipids of the mollusk The relation-ship shown in Figure 4.1 has been demonstrated in several species of aquatic

val-mollusks, including the edible mussel (Mytilus edulis), the oyster (Crassostrea virginica), and soft clams (Mya arenaria) (Ernst 1977) A similar relationship

has also been found with rainbow trout and other fish for some pollutants On the other hand, some organic pollutants do not fit the model well (Connor 1983)

It seems probable that some compounds that are metabolized relatively rapidly

by fish will be eliminated faster than would be expected if diffusion were the only process involved (Walker 1987) Such compounds would not be expected

to follow closely a model for BCF based on Kow alone This point aside, Kow

values can give a useful prediction of BCF values at the steady state for philic pollutants in aquatic invertebrates A great virtue of the approach is that

lipo-Kow values are easy and inexpensive to measure or predict (Connell 1994) Other more complex and sophisticated models have been developed for fish (see, for example, Norstrom et al 1976) but are too time-consuming/expensive

to be used widely in environmental risk assessment where cost-effectiveness

is critically important Modeling for bioaccumulation by terrestrial animals

presents greater problems, and BAFs cannot be reliably predicted from Kowvalues (Walker 1987) For example, benzo[a]pyrene and dieldrin have log Kow

values of 6.50 and 5.48, respectively, but their biological half-lives range from

a few hours in the case of the former to 10 –369 days for the latter Endrin

is a stereoisomer of dieldrin with a similar Kow, but has a half-life of only 1 day in humans, compared with 369 days in the case of dieldrin These large differences in persistence have been attributed to differences in the rate of metabolism by P450-based monooxygenases (Walker 1981) Effective predic-tive models for bioaccumulation of strongly lipophilic compounds by terrestrial animals need to take account of rates of metabolic degradation This is not a straightforward task and would require the sophisticated use of enzyme kinet-ics to be successful In one model, it has been suggested that Lineweaver–Burke plots for microsomal metabolism might be used to predict BAF values in the steady state (Walker 1987) (Figure 4.2) In principle, when an animal ingests a lipophilic compound at a constant rate in its food, a steady state will eventually

be reached where the rate of intake of the compound is balanced by the rate of its metabolism It is assumed that the rate of loss of the unchanged compound

by direct excretion is negligible Primary metabolic attack upon many highly

The main points to bring out are as follows:

1 The uptake and loss by exchange diffusion is important for aquatic isms but not for terrestrial ones

organ-2 Metabolism is the main mechanism of loss in terrestrial vertebrates, but is less important in fish, which can achieve excretion by diffusion into ambi-ent water

3 Most aquatic invertebrates have very little capacity for metabolism; this is particularly true of mollusks Crustaceans (e.g., crabs and lobsters) appear

to have greater metabolic capability than mollusks (see Livingstone andStegeman 1998; Walker and Livingstone 1992)

The balance between competing mechanisms of loss in the same organism depends

on the compound and the species in question In fish, for example, some compounds

lipophilic compounds (e.g., polyhalogenated aromatic compounds and PAHs) takes place predominantly in the endoplasmic reticulum, particularly that of the liver in vertebrates Thus, microsomes (especially hepatic microsomes of verte-brates) can serve as model systems for measuring rates of enzymic detoxication Lineweaver–Burke and similar metabolic plots can relate concentrations of pol-lutants in microsomal membranes to rates of metabolism In the steady state, rate of intake of chemical should equal rate of metabolism in the membranes

of the endoplasmic reticulum The concentration of the chemical required in the membranes to give this balancing metabolic rate can be estimated from the Lineweaver–Burke plot The necessary balancing metabolic rate can be calcu-lated from the defined rate of intake in food, and then the microsomal concen-tration that will give this rate can be read from the plot Thus, the concentration

in endoplasmic reticulum can be compared to the dietary concentration to give

an estimate of BAF Estimates can also be made of BAF for the liver or the whole body if approximate ratios of concentrations of chemical in different compartments of the body when at the steady state are known

that are good substrates for monooxygenases, hydrolases, etc., can be metabolized relatively rapidly even though they, as a group, have relatively low metabolic capac-ity (Chapter 2) So, in this case metabolism as well as diffusion is an important fac-tor determining rate of loss By contrast, many polyhalogenated compounds are only metabolized very slowly by fish, so metabolism does not make a significant contribu-tion to detoxication, and loss by diffusion is the dominant mechanism of elimination.Some further aspects of detoxication by fish need to be briefly mentioned When fish inhabit polluted waters, exchange diffusion occurs until a steady state is reached, and no net loss will occur by this mechanism unless the concentration in water falls When a recalcitrant pollutant is acquired from prey, digestion can lead to the tissue levels of that pollutant temporally exceeding those originally existing while in the steady state Here, diffusion into the ambient water may provide an effective excre-tion mechanism in the absence of effective metabolic detoxication Seen from an evolutionary point of view, the requirements of fish for metabolic detoxication would appear to have been limited on the grounds that loss by diffusion would often have prevented tissue levels becoming too high The poor metabolic detoxication sys-tems of fish relative to those of terrestrial omnivores and herbivores are explicable

on these grounds (Chapter 2) However, the advent of refractory organic pollutants, which combine high toxicity with high lipophilicity, has exposed the limitations of existing detoxication systems of fish The very high toxicity of compounds such as dieldrin and other cyclodiene insecticides to fish was soon apparent, with fish kills occurring at very low concentrations in water (see Chapter 5) and metabolically resistant strains of fish being reported in polluted rivers such as the Mississippi

Gut

Liver

Peripheral tissues Redistribution

Metabolism and excretion

FIGURE 4.2 (a) A bioaccumulation model for terrestrial organisms A kinetic model for

liver RU, rate of uptake from the gut; RM, rate of metabolism in liver; CL, concentration of pollutant in liver The arrows indicate the routes of transfer of pollutant within the animal The rates of uptake and metabolism are expressed in terms of kilograms of body weight The final elimination of water-soluble products (metabolites and conjugates) is in the urine (b)

Lineweaver–Burke plot to estimate the bioaccumulation factor; Vmax and v are expressed as milligrams of pollutant metabolized per kilogram of body weight per day; S is expressed as

the concentration of pollutant, ppm by weight (either in terms of grams of liver or milligrams

of hepatic microsomal protein) (from Walker 1987).

More rapid elimination was needed than could be provided by passive diffusion in order to prevent tissue concentrations reaching toxic levels.

Some models for predicting bioconcentration and biomagnification are presented

in Box 4.1

4.3 FATE OF POLLUTANTS IN SOILS AND SEDIMENTS

Regarding soils, a central issue is the persistence and movement of pesticides that are widely used in agriculture Many different insecticides, fungicides, herbicides, and molluscicides are applied to agricultural soils, and there is concern not only about effects that they may have on nontarget species residing in soil, but also on the pos-sibility of the chemicals finding their way into adjacent water courses

Soils are complex associations between living organisms and mineral particles Decomposition of organic residues by soil microorganisms generates complex organic polymers (“humic substances” or simply “soil organic matter”) that bind together mineral particles to form aggregates that give the soil its structure Soil organic matter and clay minerals constitute the colloidal fraction of soil; because

of their small size, they present a large surface area in relation to their volume Consequently, they have a large capacity to adsorb the organic pollutants that con-taminate soil Within a freely draining soil there are air channels and soil water, the latter being closely associated with solid surfaces Depending on their physical properties, organic compounds become differentially distributed between the three phases of the soil, soil water, and soil air

Hydrophobic compounds of high Kow become very strongly adsorbed to soil loids (Chapter 3, Section 3.1), and consequently tend to be immobile and persistent

col-OC insecticides such as DDT and dieldrin are good examples of hydrophobic pounds of rather low vapor pressure that have long half-lives, sometimes running into years, in temperate soils (Chapter 5) Because of their low water solubility and their refractory nature, the main mechanism of loss from most soils is by volatiliza-tion Metabolism is limited by two factors: (1) being tightly bound, they are not freely available to enzymes of soil organisms, which can degrade them, and (2) they are, at best, only slowly metabolized by enzyme systems Because of strong adsorption and low water solubility, there is little tendency for them to be leached down the soil pro-file by percolating water The degree of adsorption, and consequently the persistence and mobility, is also dependent on soil type Heavy soils, high in organic matter and/

com-or clay, adscom-orb hydrophobic compounds mcom-ore strongly than light sandy soils, which are low in organic matter Strongly lipophilic compounds are most persistent in heavy soils When OC insecticides are first incorporated into soil, they are lost relatively rapidly, mainly due to volatilization, before they become extensively adsorbed to soil colloids (Figure 4.3) With time, however, most residual OC insecticide becomes adsorbed, and subsequently there is a period of very slow exponential loss

In marked contrast to hydrophobic compounds, more polar ones tend to be less adsorbed and to reach relatively high concentrations in soil water Phenoxyalkanoic acids such as 2,4-D and MCPA are good examples (Figure 4.3) Their half-lives in soil are measured in weeks rather than years, and they are more mobile than OC insec-ticides in soils When first applied they are lost only slowly After a lag period of a

few days, however, they disappear very rapidly as a consequence of metabolism by soil microorganisms This has been explained on the grounds that it takes time for

a buildup in numbers of strains of microorganisms that can metabolize them; these microorganisms use the herbicides as an energy source It has also been suggested that the lag period relates to the time it takes for enzyme induction to occur Whatever the explanation, soils treated with these compounds stay enriched for a period, and further additions of the original compounds will be followed by rapid metabolism without

a lag phase If, however, the soils are untreated for a long period, they will revert to their original state and not show any enhanced capacity for degrading the herbicides

An important difference from the OC insecticides and related hydrophobic pollutants

is that, because of their polarity and water solubility, they are freely available to the microorganisms that can degrade them Interestingly, the phenoxyalkanoic acid 2,4,5-T

is more persistent than either 2,4-D or MCPA With three substituted chlorines in its phenyl ring, it is metabolized less rapidly than the other two compounds, and it would appear that metabolism is a rate-limiting factor determining rate of loss from soil

It was long assumed that there is little tendency for most pesticides or other organic pollutants to move through soil into drainage water Indeed, this is to be expected with intact soil profiles Hydrophobic compounds will be held back by adsorption, whereas water soluble ones will be degraded by soil organisms Some soils, how-ever, depart from this simple model Soils high in clay can crack and develop deep fissures during dry weather If rain then follows, pesticides, in solution or adsorbed

to mobile colloids, can be washed down through the fissures, to appear in ing drainage ditches and streams This was found to happen with pesticides such as carbofuran, isoproturon, and chlorpyrifos in the Rosemaund experiment conducted

neighbor-in England durneighbor-ing the period 1987–1993 (Williams et al 1996)

0

0 10 20 30 40 50 60 70 80 90 100

Days after Application

Time Following Application

(b) (a)

Concentration that would have been found if all applied material were retained by soil

Period of slow exponential loss

FIGURE 4.3 Loss of pesticides from soil (a) Breakdown of herbicides in soil (b)

Dis-appearance of persistent organochlorine insecticides from soils (from Walker et al 2000).

The influence of polarity on movement of chemicals down through the soil profile has been exploited in the selective control of weeds using soil herbicides (Hassall 1990) In general, the more polar and water soluble the herbicide, the further it will

be taken down into the soil by percolating water Insoluble herbicides such as the triazine compound simazine (water solubility, 3.5 ppm), remain in the first few cen-timeters of soil when applied to the surface More water-soluble compounds such

as the urea herbicides diuron and monuron (water solubilities 42 ppm and 230 ppm, respectively) are more mobile, and can move farther down the soil profile Selective weed control can be achieved in some deep-rooting crops by judicious selection from this range of herbicides, so that the herbicide will only percolate far enough down the soil profile to control surface rooting weeds without reaching the main part of the root system of the crop (depth selection) Thus, when applied to the soil surface, simazine should only be toxic to shallow rooting weeds and should not affect crops that root farther down Other more water-soluble herbicides can give weed control

to greater depths in situations where the rooting systems of the crops are sufficiently deep When attempting depth selection in weed control, account needs to be taken

of soil type Herbicides will move farther down the profile in the case of light sandy soils than they will in heavy clays or organic soils

Although the major concern about the fate of organic pollutants in soil has been about pesticides in agricultural soils, other scenarios are also important The dis-posal of wastes on land (e.g., at landfill sites) has raised questions about movement

of pollutants contained in them into the air or neighboring rivers or water courses The presence of polychlorinated biphenyls (PCBs) or PAHs in such wastes can be a significant source of pollution Likewise, the disposal of some industrial wastes in landfill sites (e.g., by the chemical industry) raises questions about movement into air

or water and needs to be carefully controlled and monitored

In certain respects, sediments resemble soils Sediments also represent an ciation between mineral particles, organic matter, and resident organisms The main difference is that they are situated underwater and are, in varying degrees, anaerobic The oxygen level influences the type of organisms and the nature of biotransformations that occur in sediments A feature with sediments, as with soils, is the limited availability of chemicals that are strongly adsorbed Again,

asso-compounds with high Kow tend to be strongly adsorbed, relatively unavailable, and highly persistent There is much interest in the question of sediment toxicity and the availability to bottom-dwelling organisms of compounds adsorbed by sedi-ments (Hill et al 1993) One case in point is pyrethroid insecticides (see Chapter

12), which are strongly retained in sediments on account of their high Kow values Because of their ready biodegradability, they are not usually biomagnified in the higher trophic levels of aquatic food chains

However, they are available to bottom-dwelling organisms low in the food chain Questions are asked about the possible long-term buildup of pyrethroids in sedi-ments and their effects on organisms in lower trophic levels

4.4 EFFECTS OF CHEMICALS UPON INDIVIDUALS—

THE BIOMARKER APPROACH

Until now this narrative has been concerned with questions about the movement and distribution of chemicals in the living environment, a topic that relates to the field of toxicokinetics in classical toxicology, although on a much larger scale It is now time

to move to a consideration of the effects that chemicals may have upon living isms, which relates to the area of toxicodynamics in classical toxicology Effects upon individuals will be discussed before dealing with consequent changes at the higher levels of biological organization—population, community, and ecosystem.Measuring effects of chemicals upon free-living individuals in the natural envi-ronment is not an easy matter Mobile animals need to be captured so that samples

organ-of tissues can be taken for analysis, but this is organ-often difficult to do in a properly controlled way in the field It is easier to obtain samples from sedentary species (e.g., mollusks or plants) or of eggs in the case of birds, reptiles, and some invertebrates All too often sampling is destructive, which raises problems of experimental design and statistical evaluation of results In principle, measuring behavioral effects of chemicals is an attractive option, but this can be hard to achieve in practice because

of the difficulty of making reliable measurements in the field The problems of pling can, to some extent, be circumvented by deploying indicator species that have been maintained in a “clean” environment into the field Thus, control fish from the laboratory can be held in cages in contaminated waters and samples taken from them after periods of exposure to pollutants Uncontaminated birds’ eggs can be intro-duced into the nests of birds of the same species that are breeding in a polluted area

sam-In this way, changes caused by pollutants can be measured and evaluated

Problems of sampling aside, the success of any strategy of this kind depends on the availability of reliable tests that can measure harmful effects of chemicals under field conditions Reference has already been made to biomarkers (Chapter 2, Box 2.2) In the following account they are defined as “biological responses to environ-mental chemicals at the individual level or below, which demonstrate a departure from normal status.” This definition includes biochemical, physiological, histologi-cal, morphological, and behavioral changes, but does not extend to changes at higher levels of organization Changes at population, community, or ecosystem level are regarded instead as bioindicators

The concept of biomarkers is illustrated in Figure 4.4 As the dose of a chemical increases, the organism moves from a state of homeostasis to a state of stress With fur-ther increases in dose, the organism enters first the state of reversible disease, and even-tually the state of irreversible disease, which will lead to death In concept, all of these stages can be monitored by biomarker assays (lower part of conceptual diagram).Some biomarker responses provide evidence only of exposure and do not give any reliable measure of toxic effect Other biomarkers, however, provide a measure

of toxic effects, and these will be referred to as mechanistic biomarkers Ideally, marker assays of this latter type monitor the primary interaction between a chemical and its site of action However, other biomarkers operating “down stream” from the original toxic lesion also provide a measure of toxic action (see Figure 14.3 in Chapter 14), as, for instance, in the case of changes in the transmission of action potential

bio-following the interaction of DDT or pyrethroids with Na+ channels, although these are changes that may also be caused by the operation of other toxic mechanisms These will also be treated as mechanistic biomarkers in order to distinguish them from other responses that are only biomarkers of exposure (e.g., the induction of certain enzymes that can occur at low levels of exposure before any toxic effects are manifest) Mechanistic biomarkers (see Table 4.2 for examples) have potential for measuring adverse effects of chemicals in the field—effects that may be translated into changes at the population level and above.

Physiological Condition Pollutant Concentration

Homeostasis Compensation Death

FIGURE 4.4 Relationship between exposure to pollutant, health status, and biomarker

responses Upper curve shows the progression of the health status of an individual as sure to pollutant increases; h, the point at which departure from the normal homeostatic response range is initiated; c, the limit at which compensatory responses can prevent develop- ment of overt disease; r, the limit beyond which pathological damage is irreversible by repair mechanisms The lower graph shows the response of five different hypothetical biomarkers used to assess the health of the individual (Reproduced from Depledge et al 1993 With permission from Springer-Verlag.)

expo-Of the examples given, brain cholinesterase inhibition has been frequently sured in field studies involving OP insecticides and when investigating cases of poi-soning on agricultural land (see Chapter 10) A number of studies on fish, rodents, and birds in the field and/or in the laboratory give evidence for a range of sublethal neurotoxic and behavioral effects of OP insecticides when brain acetylcholinesterase inhibition is in the range 40–50%—before the appearance of severe toxic manifesta-tions and death (see Chapters 10 and 16) A few OP compounds cause delayed neu-ropathy in mammals and birds, and this has been related to inhibition of neuropathy target esterase (NTE) Symptoms of this type of poisoning appear after aging of the inhibited enzyme occurs (Chapter 10) Warfarin and related anticoagulant rodenti-cides act as competitive antagonists of vitamin K at binding sites for this cofactor in the liver and consequently inhibit the carboxylation of precursors of blood-clotting proteins (Chapter 11) Thus, undercarboxylated Gla proteins are released into the blood After a period of time (usually 5 days or more) the blood becomes depleted

mea-of normal clotting proteins and loses its capacity to coagulate A valuable biomarker assay involves the measurement of levels of undercarboxylated clotting protein in the blood by immunochemical determination An increase of this nonfunctional protein

in the blood provides a measure of the toxic process that leads to hemorrhaging and death—although it does not measure the primary interaction between rodenticide and the vitamin K binding site in the liver Certain metabolites of coplanar PCBs such as 4-OH, 3,3b,4,4b-TCB can compete with thyroxine (T4) for binding sites on the protein transthyretin in blood This interaction leads to the breaking apart of

TABLE 4.2

Some Mechanistic Biomarker Assays

Acetylcholine buildup in synapse and synaptic block

Reduced levels of retinol and thyroxine (T4) in blood

6