7. General Framework
7.11. Ecotoxicologically Relevant Concentrations
Table21 lists the major exposure routes of key drivers, specifically for each organism group. For the single exposure routes, ecotoxicologically relevant types of concentrations (ERC) are listed in Table 23 below. The proposed ERC are based on the knowledge about the major exposure routes for in-soil organisms and the considerations about the temporal and spatial exposure profiles the organisms are exposed to.
From the data referred to above, it emerges that in-soil organisms inhabiting the upper soil layer or feeding there will be exposed to the highest concentration of the applied PPP after spraying. For these organism groups, the possible uncontaminated layers below the uppermost one do not deliver a definite shelter, since other vertical gradients (e.g. of food and water) attract the animals to the surface and in-soil organisms are not able to detect and to avoid all active substances.
Averaging of exposure between high concentrations in the top layer and the low concentrations in deeper soil layers would deliver a ‘space weighted average’, which, as the ‘time weighted average’
concentration, could be used only under specific preconditions (e.g. reciprocity) and only for specific measurement endpoints (see Section 7.10.3 for details).
Organisms with a greater mobility in the soil and having a geophagous feeding mode, although, will also dwell frequently in the uppermost soil layer, but not necessarily continuously and for longer periods. If the concentrations in the uppermost soil layers are sufficiently high, then a short permanence there will also elicit acute or chronic effects. If the concentrations in the uppermost soil layer are such that only a long permanence there would lead to acute or chronic effects, then moving to uncontaminated deeper layer and feeding on uncontaminated matter would decrease the animal chemical burden. For these organisms, no single soil depth, or in many cases type of exposure, would be relevant. The choice of one, single ecotoxicologically relevant type of concentration could possibly deliver unreliable exposure metrics. To deal with this situation, exposure and effects need to be integrated over time. This could be achieved by linking following components:
•
Reliable models of movement for endogeic earthworms, within the soil profile;•
Dynamic models of exposure providing soil and pore-water concentrations at all relevant soil depths and varying with time. Ideally, these would be linked to the systems models proposed for population assessment (see Section 7.3);•
Toxicokinetic–toxicodynamic models capable of integrating both internal concentrations and toxicological effects with time (see Section 9.9).Currently, there is no available systems model combining all three components, although technically this is considered to be feasible. Ideally, this combined system would be included in the systems model used to develop the population-modelling‘surrogate reference tier’ (see Section 7.3).
Since both exposure routes via total soil or pore-water concentration are considered to be relevant, and will have different relative importance for different substances and species, it is recommended to assess both exposure routes for in-soil organisms (see Table 21). Further research should clarify which exposure metrics are pertinent in different soil type/species/substance contexts.
Table 23: Ecotoxicologically relevant type of concentrations for in-soil organisms exposed to active substances in PPPs via different exposure routes. related to the (time course of) internal concentrations
7.11.1. Overall assessment of the exposure based on the different routes
As mentioned above, exposure by contact and oral uptake are both relevant. These two exposure routes require different exposure concentrations because the ecotoxicologically relevant concentration is different (Table23). Developing a worst-case exposure scenario that considers both routes would require a model that integrates both exposure routes, e.g. a TK/TD model (see also EFSA PPR Panel 2014b for deriving effects based environmental scenarios). Since such models are not yet available for regulatory purposes at the European level, it is not possible to assess the relative contributions of individual exposure routes to effects. Measured effects result from the combined exposure of both routes. Extrapolation from lower tier to the field situation is done by assessment factors (Section 7.6) which address uncertainties: this includes uncertainty about the relative contributions of different routes of exposure.
Exposure route related to the (time course of) internal concentrations
*–/–: The relevance of the impact of vertical movements for soil organisms living in the upper centimetres is deemed to be low.
Currently, available exposure data in test systems probably do not distinguish between the various exposure routes and the results are then to be seen as lumped over two routes (contact via pore water and via bulk soil and oral uptake). Available data should therefore be carefully checked and attributed to one of the two exposure routes or marked ‘lumped’. Appropriate values should be used for estimating exposure according to the two specified routes (see Section7.11.2).
7.11.2. Using consistent concentrations in the exposure and effects assessment For the scheme in Section7.10.1 to work, the same type of concentration should be used in the effect assessment and the exposure assessment. The type of concentration depends on the properties of the substance, the organism (e.g. soft-bodied or hard-bodied, see (EFSA, 2009c) and the intake route (Section 7.10.2) and is either the concentration in total soil or the concentration in pore water.
Both types of concentrations are delivered by the exposure assessment; however, different scenarios are used for the concentration in total soil and for the concentration in pore water (Section8).
The most commonly used tests in the effect assessment are the earthworm, collembolan and predatory mite reproduction tests (OECD 222, 232 and 226). The OECD guidelines recommend using artificial soil (70% sand, 20% kaolin clay, 10% or 5% coarse ground Sphagnum peat and pH adjusted to 6), but give also some indications to the use of natural soils. Artificial soil has the advantage that it is relatively well reproducible (Van Gestel, 1992). However, Sphagnum peat has different properties than organic matter in arable soils. The type of the organic matter influences sorption and hence bioavailability (EFSA PPR Panel, 2015c). Therefore, a standardised arable soil with properties closer to the scenarios in the exposure assessment would be preferred over an artificial soil with Sphagnum peat. However, developing a new standardised test would require extensive research to assess the suitability of the soil(s) selected to allow the survival and the reproduction of the test organisms in acceptable levels. For this reason, it is assumed that the OECD tests will be run with artificial soils in the nearest future until better alternatives are available and tested. The Panel recommends to investigate the use of feasible, alternative natural soils for standardised test systems and to perform a sensitivity analysis for a comparison of tests performed in natural soils to tests run with artificial soil.
7.11.3. Measuring exposure in test systems
In most test guidelines for in-soil organisms, the tested substance is incorporated into the soil;
either in a solution or with sand. The present test systems can be adapted in order to test effects of soil fumigants, treated seeds and granules. In the past, tests were also often performed with the substance applied to the soil surface. The advantage of spraying in test systems would be that it more realistically mimics the actual exposure of sprayed substances in the field (including a layer with high concentrations in the top centimetres of soil). Nevertheless, the Panel considers mixing through the soil a better option to avoid uncertainty about the actual exposure concentration of soil organisms in the test system.
A litter layer is not included in current, first-tier laboratory tests. Exposure via the litter layer is, however, a relevant route of exposure for some soil invertebrates, like macroarthropods, slugs and snails. Furthermore, exposure via food uptake is only partly included in the standard laboratory tests, since normally uncontaminated food is provided. This may lead to underestimation of internal exposure due to dilution.
As mentioned above, either the concentration in total soil or the concentration in pore water is to be used for linking exposure and effects. Total concentrations in laboratory tests are currently commonly expressed as nominal concentrations, i.e. the total mass of chemical added to a certain mass amount of dry or wet soil. During laboratory handling procedures of the spiked soils, however, possible losses of the pesticides due to volatilisation, degradation, and sorption to, e.g. the glass matrix of vessels used, may occur. The Panel therefore recommends that the exposure concentration be measured as a function of time regardless of the metric chosen (see also (EFSA, 2009c). Measuring the concentration increases the certainty about exposure, and could deliver information about formation of metabolites (ECHA, 2016).
Exposure could be measured using the two-step extraction procedure that is proposed in EFSA PPR Panel, (2015c). This consists of a 24-h extraction with a 0.01 M CaCl2 solution to characterise the pore-water concentration and a solvent extraction to characterise the total extractable mass (OECD, 2002). In principle, centrifugation would also deliver the pore-water concentration. It was, however, observed that variability was larger and therefore the CaCl2 extraction is preferred. Instead of the currently used harsh solvent extraction to determine the total extractable mass, a mild organic solvent
would be preferable because this fraction generally correlates better with ecologically relevant endpoints like uptake in organisms (see Appendix 1 in EFSA, 2009c for an overview). However, no standard protocol is currently available for mild chemical extractions in relation to bioavailability testing (see also EFSA, 2009c; EFSA PPR Panel, 2015c). Furthermore, there is no commonly agreed exposure assessment methodology to determine the fraction that is available for oral uptake by organisms. The Panel therefore recommends developing (i) a protocol for characterising the fraction that is available for uptake in organisms and (ii) an appropriate exposure assessment scheme for determining this fraction. As long as these protocols are not available, it is recommended to use the concentration in total soil as a proxy for the fraction available for oral uptake by organisms.
In order to compare the outcome of different tests – especially laboratory tests with field tests – the comparability of the time course of the concentrations in the different soils is essential. However, degradation, dissipation and movement of active substance in the field soils will differ from the degradation/dissipation in a standard artificial soil. Hence, it is recommended that the concentrations of active substances in soils is measured more than twice (see also the EFSA DegT50 EFSA Guidance), depending on the degradation of the active substances and the length of the test. It should be insured that the time course of exposure in the tests covers the predicted exposure profile in the field. The time course of the exposure profile could be used in models that consider toxicokinetics/
toxicodynamics (to be developed) to estimate toxicity based on internal body concentrations.
7.11.4. Calculating the exposure concentration in test systems
The Panel does not recommend calculating the pore-water concentration from the concentration in total soil using the partitioning coefficient. As indicated in EFSA PPR Panel, (2012), the coefficient for sorption on organic matter (Kom) is measured in at least four different soils and the variability of Kd-values between these four soils is generally more than 25%. This would give a high uncertainty of the calculated exposure concentration in the test system. However, in current ecotoxicological studies (legacy studies), only the initial nominal concentration is known. For such legacy studies, the initial pore-water concentration could be calculated when instantaneous sorption equilibrium is assumed and the water content and the sorption coefficient for the test soil are known. This can be done using the equation:
Mt
Vsoilþ MsoilmOMKOM ð1Þ
where c (kg/L) is the concentration in pore water, Vsoil (L) is the total volume of liquid in the system, Msoil(kg) is the total mass of soil in the system and Mt(kg) is the total amount of substance applied, mom (kg/kg) is the mass fraction of organic matter in the soil and KOM (L/kg) is the coefficient of equilibrium sorption on organic matter.
The equation above is based on the assumption that sorption is linear. In reality, sorption is non-linear and because the exponent is usually < 1, the concentration in pore water will be overestimated when linear sorption is assumed. Overestimation of the pore-water concentration in the effects study would underestimate toxicity, and therefore, the Panel recommends using the Freundlich equation for calculating the pore-water concentration (in line with current practice in exposure modelling):
ct ¼Vsoil
Msoilcþ Kommomcref c cref
1=n
ð2Þ
where ct(mg kg) is the concentration in total soil, cref is the reference concentration (usually 1 mg/kg) and 1/n is the Freundlich exponent. This equation requires an iterative solution, which could be easily implemented using, e.g. standard spreadsheet software.
Notice further that the equations above only give the initial concentration of a parent substance. If a time-weighted average concentration is needed, it could be calculated assuming first-order degradation kinetics. For this purpose, the Panel recommends developing a dedicated version of the PERSAM model (EFSA PPR Panel, 2012). This tool could also be helpful for characterising the exposure concentration of metabolites in the test system. Specific attention is needed for parameterisation of this model so that it will lead to conservative estimates of the pore-water concentration (Van der Linden et al., 2006, 2008).
Time-dependent or aged sorption is a generally accepted phenomenon, which reduces the concentration in pore water (see e.g. EFSA PPR Panel, 2015c). Consideration of this process is not necessary if measured pore water concentrations are used in the effects study, because this process is then implicitly included in the measurements.
7.11.5. Scaling of the toxicity endpoint to account for bioavailability
It is now common practice to divide the obtained toxicity endpoint (LC50 or NOEC) of lipophilic substances (log(Kow) > 2) for in-soil organisms (except microorganisms) by a factor of two. This is done because the organic matter content of the artificial substrate of the earthworm laboratory tests is higher than that of many natural soils (Van Gestel, 1992). The scaling factor was needed because concentrations were only available as nominal concentrations (Section 7.11.5), while toxicity endpoints generally correlated better with pore-water concentrations. This was demonstrated because differences between soils almost completely disappeared when LC50 values in mg/kg were recalculated towards values in mg/l pore water concentrations (Van Gestel and Ma, 1988, 1990; see also EFSA, 2009c).
Scaling of the toxicity endpoint is no longer necessary (and even not justified) if the appropriate exposure concentration in the test system is either measured (Section7.11.3) or calculated (Section 7.11.4). When only the nominal concentration is available (legacy studies) and when the ERC is expressed in terms of a pore-water concentration, scaling of the toxicity endpoint is justified (Table 23). Notice, however, that the factor of two has no scientific basis for assessments at the European level because it is based on the ratio of the organic matter content of the test medium (10%) and the organic matter content of the old Dutch standard soil (4.7%) and not on that of the proposed exposure scenarios described in Section 8. So, instead of the default factor of two, scaling should be done based on the following equation:
Sf¼ Vsoilþ KomOMtestMsoil
Vsoilþ KomOMscenarioMsoil ð3Þ
If in the equation Vsoilis neglected, equation 3 reduces to:
Sf¼ OMtest
OMscenario ð4Þ
Figure21 gives the scaling factor calculated with equation 3 for a range of values of Kom. In this example, the mass fraction of organic matter in the test medium is 0.1 g/g (or 10%). Thefigure indicates that the scaling factor is substance-dependent. For substances with a Kom > 10, the equation yields the traditional factor of two when the organic matter content of the test medium is 10% and the organic matter content of the scenario is 5%. For lower values of Kom, ignoring Vsoil would overestimate the scaling factor, and would therefore give a conservative estimate of the toxicity endpoint.
Figure 21: Scaling factor of the toxicity endpoint as a function of the mass fraction of organic matter of the exposure scenarios for a range of values of Kom. In this example, the mass fraction of organic matter in the test medium is 0.1 g/g (or 10%)
The implicit assumption of this scaling is that organic matter is the main adsorbent (Van Gestel, 1992). The latter is true for the majority of PPPs; however, in some cases, PPPs show affinity for other soil constituents such as clay minerals or sesquioxides (see EFSA PPR Panel, 2015c) for an overview of sorption processes). Clearly, in those cases, the scaling procedure is not valid and the pore-water concentration should be calculated or measured. Van Gestel (1992) further mentions that for substances with a log(Kow) < 2 differences in soil moisture content should be taken into account when extrapolating toxicity data. However, current practice at the EU level is not to scale the toxicity endpoint at all for such substances. This is not in line with the original description given by Van Gestel (1992).
If the PEC is derived from a Tier 1 or 2A exposure assessment (see Section8), the organic matter content of one of the standard concentrations in total soil scenarios should be selected. If the PEC is derived from a Tier 3 assessment (substance or crop specific exposure scenarios), the organic matter of this Tier 3 scenario should be used. Notice that organic matter content is not delivered when running a Tier 2B/C exposure assessment. It can, however, easily be obtained by running Tier 3B. It has to be noticed that the scaling factor is less than one when the organic matter content of the selected exposure scenario is higher than the organic matter content of the test system.