Ecological risk of mixtures
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1 VALUTAZIONE DEL RISCHIO PER ESPOSIZIONE COMBINATA A SOSTANZE CHIMICHE MILANO, 7 LUGLIO 2015 Ecological risk of mixtures Marco Vighi Department of Earth and Environmental Sciences University of Milano Bicocca, Milano, Italy DEPARTMENT OF EARTH AND ENVIRONMENTAL SCIENCES
2 What an environmentally relevant mixture is? Ecosystems are never exposed to individual chemicals. They are always exposed to more or less complex mixtures of different origin. a. Chemicals used as technical mixtures Commercial formulations containing several active ingredients (plant protection, biocidal, cosmetics or pharmaceutical products) industrial technical mixtures of congeneric chemicals (PCBs and PBDEs). b. Chemicals emitted by a human activity Wastes from industrial typologies or production processes, combination of pesticides applied on an agricultural crop, etc. c. Chemicals likely to be present in the environment as the result of multiple emissions Mixtures of chemicals resulting from the combination of all the emissions of human activities in a given territory (e.g. mixture present in a river as a result of the emissions in the hydrographic basin).
3 What an environmentally relevant mixture is? The example of pesticides Many active ingredients may be present in the same formulation Many formulations may be applied on the same crop Many crops may be present on the same agricultural area }Final mixture
4 Mixtures in surface water Urban settlements (organic matter, detergents, pharmaceuticals, personal care products) Agriculture (fertilizers, pesticides) Industry (industrial chemicals) Predicting the composition of an ecologically relevant mixture requires detalied knowledge of land use.
5 Assessing exposure to mixtures Terrestrial ecosystems: epigean communities (birds and beneficial insects) Foraging area The community potentially exposed is moving within the area. Exposure is a function of the variable concentrations on different systems in the area (e.g.: treated crops, non crop vegetation, etc) as well as of the behaviour and the ecological role and niche of the organism.
6 ytu (B) Isobologram describing possible toxicological responses to a binary mixture of toxic chemicals TUi = Ci/ECi xtua+ytub= 1TU(AB) 1 additive response: x+y=1 2 less than additive: x +y>1 (x and y <1) 3 antagonism: x+y>1 (x or y >1) 4 synergism: x+y< TU (A) x and y are the toxic units (TU) of the two chemicals (A and B). The four lines represent the loci where the response of the mixture corresponds to 1 TU x
7 Approaches for predicting mixture response No interactions among chemicals The Dose/Concentration Addition (CA) approach. It applies to chemicals with the same mode of action. The Independent Action (IA) or Effect Addition approach. It applies to chemicals with different mode of action Interactions among chemicals Synergism and antagonism. The response is higher or lower than predicted respectively
8 The Concentration Addition (CA) approach Two reference models are generally used for predicting the effects of mixtures in ecotoxicology: The Concentration Addition (CA) model, applicable to chemicals with the same mode of action TUm = n i= 1 TUi = n C EC i= 1 x, i i ECx Mix 1 n p i = i= 1 ECxi n is the number of mixture components pi is the relative fraction of chemical i in the mixture x is a common effect level, which is provoked by an exposure to a substance or mixture concentration ECx Mix resp. ECxi. single
9 The Independent Action (IA) approach The Independent Action (IA) model, applicable to chemicals with different mode of action For a binary mixture f: is the fraction of the toxicological end point produced by the mixture and by individual chemicals For a mixture of n components f 1+2 = f 1 + f 2 - f 1 f 2 f Mix = 1 [1 fi] n i= 1 or
10 Triazine mixture Observed and predicted algal toxicity of a mixture of 18 s-triazines. Components mixed in the ratio of their individual EC1 values. ( ) experimentally observed toxicity; (ο) controls; (-----) prediction according to CA ( ) prediction according to IA Comparison of the effects of a s- triazine mixture with the effects of individual components. Concentrations of individual mixture components equal individual EC1 values. CA, IA - predictions of the total effect according to CA and IA, respectively.
11 Eterogeneous mixture % Inhibition B 36 c o n t r o l s c m i x = Σ E C 1 i T o t a l C o n c e n t r a t i o n ( µ m o l / L ) Observed and predicted algal toxicity of mixtures of 16 structurally and toxicologically different components mixed in the ratio of their individual EC1 values. ( ) experimentally observed toxicity; (ο) controls; (-----) prediction according to CA ( ) prediction according to IA Comparison of the effect of a mixture of heterogeneous chemicals with the effects of all individual components. Concentrations of individual mixture components equal individual EC1 values. CA, IA - predictions of the total effect according to CA and IA respectively.
12 Two Stage Calculation Naturally occurring mixtures are rarely composed by «all similar» or «all dissimilar» chemicals. In case of more complex mixtures the two stage approach (TSA) may be applied.
13 How interactions may occur Chemical interaction is a complex issue that may occur in different phases of the toxic effect, outside or inside of the organism. 1. Chemicals may interact in the environment and the resulting complex may increase bioavailability and the capability to enter into the organism. 2. Chemicals may interact into the organism and the resulting complex may be more active to the receptor. 3. Chemicals may act to the same receptor and chemical A may increase the sensitivity of the receptor to chemical B.
14 May interactions be predicted? At present models capable to predict chemical interactions are not available. Therefore, the possibility of synergism must be assessed case-by case. However, experimental evidence indicates that toxic interactions are relatively infrequent. Moreover, interactions usually occur at medium or high dose levels (relative to the lowest effect levels). At low exposure levels they are either not occurring or toxicologically insignificant.
15 The problem of modes of action The selection of the most appropriate approach between CA and IA (or TSA) requires the knowledge of the mode of action of all the component of a mixture. The major knowledge gap at present is the limited number of chemicals for which information on mode of action is available. Currently there is neither an agreed inventory of modes of action, nor a defined set of criteria to characterise a mode of action for data poor chemicals. In ecotoxicology, the problem is particularly complex. A knowledge of all possible modes of actions that may occur in the different types of organisms of a complex biological community is difficult (if not impossible) to be attained.
16 The problem of modes of action The complexity of ecotoxicology The mode of action of chemicals may be fully different on different taxonomic groups of organisms. Chemicals with specific toxic effect on animals (organophosphate and chlorinated insecticides) behave as baseline, narcotic-type, toxicants on algae and plants. 6 substituted benzenes organophosphorus triazines chlorinated ins 5 Relationship between toxicity on algae (Log 1/EC50) and Log Kow for chemicals with specific (triazines) and non-specific (all others) toxicological mode of action on algae log 1/c m mol/l log Kow
17 The Meolo river case study Meolo River flows in an intensive agricultural area in the province of Treviso. The River Meolo Basin Major crops are vinejard, maize soybean and sugar beet
18 List of active ingredients applied on the Meolo basin 54 different active ingredients (herbicides, insecticides and fungicides) are applied on the four main crops (maize, soybean, grapevines and sugar beet). The total amount used in the basin is extremely variable. Chemicals kg Chemicals kg Chemicals kg 1 Mancozeb Ethofumesate Cyprodinil Terbuthylazine Fenpropidin Penconazole Metolachlor Dicamba Clopyralid Fosetyl-Al Dimethenamide Fludioxonil Dimethomorph Chlorpyriphos Nicosolfuron Procymidone Imazethapyr Azoxystrobin Pirimethanil Cymoxanil Flufenoxuron Glyphosate Metalaxyl Trifloxistrobin Flufenacet Mesotrione Tetraconazole Metamitron Metribuzin Desmedipham Folpet Propargite Cyproconazole Metiram Difenoconazole Linuron Fenitrothion Quizalofop-ethyl Propaquizafop Imazamox Lenacil Thifensulfuron-met Isoxaflutole Oxasulfuron Deltamethrin Chloridazon Phenmedipham Propiconazole Chlorothalonil Cycloxydim Triflusulfuron-met Glufosinate-Ammoniun Clethodim Cyhalothrin 0.31
19 The seasonal trend of mixture risk The time course of the potency of the total mixtures (as TUs) present in Meolo River has been calculated for algae, Daphnia and fish. 1 0,1 Toxic Units 0,01 ALGAE DAPHNIA FISH 0,001 0, /4 7/5 14/ / 5 28/5 4/6 11/ / 6 25/6 2 7/ 9 / 7 16/7 Time 23/7 30/ 7 6 / 8 13/8 20/ 8 27/ 8 3 / 9 10/9 17/ 9 24/ 9
20 The problem of modes of action Differences between human toxicology and ecotoxicology The goal of human toxicology is the protection of individuals. The goal of ecotoxicology is protecting structure and functions of biological communities and ecosystems. Therefore relevant end-points may be different in human toxicology and in ecotoxicology. Ecotoxicological end-points are relatively broad, related to ecologically-relevant parameters ( massive mortality, reduction of fertility and any other effect affecting reproductive capability). End-points, that in human toxicology are often referred to a specific target organ, are meaningless in ecotoxicology.
21 The IA/CA ratio The CA approach generally overestimated the response in comparison to IA. Therefore it may represent an overprotective approach if applied by default. The key factors affecting the IA/CA ratio are: The number of component n CA IA The effect ratio of individual chemicals The effect level The slope of the concentration-response curve of individual components
22 The role of slope High slopes are relatively infrequent in concentrationresponse curves
23 How many chemicals are relevant? Theoretically, the number of chemicals that may be present in the real environment (e.g. in a large river, alimented by a big hydrographic basin) may be extremely high. However, the idea of considering all chemicals potentially present is unrealistic. A treshold of concern should be established in order to consider only significant chemicals. 3 1 Toxic Units 0,3 0,1 0,03 Treshold of concern Relationship between the number of chemicals in a mixture and the total mixture toxic potency. 0,01 0,003 0, Number of chemicals in the mixture It has been empirically observed that in mixtures likely to occur in the real environment only a few chemicals explain a high percentage of the toxic potency.
24 How many chemicals are relevant? Examples of mixtures realistically occurring in surface water. A: mixture of herbicides emitted from a typical intensive agricultural area (maize crop) in Northern Italy; 24 individual chemicals; TU=PEC/EC50 algae. B: effluent of a typical petrochemical industry; 26 chemicals; TU=PEC/PNEC. In both cases only three chemicals represent more than 80 and 93% respectively of the total mixture potency.
25 Example of mixture risk assessment at continental level (Fraunhofer priority) The assessment has been performed on the 140 substances selected as priority chemicals within the Water Framework Directive. Continental PECs have been calculated according to EUSES. The mixture may represent a background general mixture for European surface waters PECcont/PNEC Substances fluorantene pentaclorofenolo 1-cloro-3-nitrobenzene Benzo-a-antracene Indeno(1,2,3-cd)pyrene benzo-a-pirene benzo-g-h-i-perylene benzo-b-fluorantene benzo-k-fluorantene esaclorobenzene antracene esaclorobutadiene naftalene 1,2,3-TCB 1,2,4-TCB 1,3,5-TCB acenaftene 1,2-dicloroetano triclorometano diclorometano Mixture potency has been calculated as PEC/PNEC. Total mixture potency is 9.9. Only 7 chemicals over 140 represent 90% of the total mixture potency.
26 Maximum Cumulative Ratio (MCR)
27 Examples of MCR MCR for Case 1 is: MCR = 3.5/0.8 = 4.4 MCR for Case 2 is: MCR = 3.5/3.2 = 1.1 The need for a cumulative risk assessment increases with the size of MCR
28 MCR values are constrained Values can approach 1 when one chemical completely dominates the toxicity Values can approach n (where n is the number of chemicals in the mixture) when each chemical is present in equitoxic concentrations Thus the MCR value of cumulative exposure to 30 chemicals will fall in a range from 1 to 30
29 MCR values for chronic effect in fish MCR closer to 1 than n: Most values of MCR ranged from 1-6 MCR declined with toxicity: Average for mixtures with HI values <1 is 1.9 Average value for mixtures with HI values >1 is 1.3
30 Back to Meolo River Relationship between toxic potency on algae Daphnia and fish and the MCRs for the pesticide mixtures emitted in the RiverMeolo basin.
31 Options for a Predictive Hazard Assessment of Chemical Mixtures 1: Case by case selection of the most appropriate concept: Independent Action (IA) OR Concentration Addition (CA) OR Two Stage Calculation (TSC) that is a combination of both Prerequisite: Problem: Sound criteria for classifying chemicals into groups of similar or dissimilar action Need for knowledge on the mechanisms of action of chemicals 2:IA by Default Problems: Underestimations of mixture toxicity Extensive data requirements 3:CA by Default Problem: Overestimations of mixture toxicity
32 Differences between CA and IA models % E f f e c t CA IA L o g T o t a l C o n c e n t r a t i o n All available evidence (theoretical and experimental) indicates that quantitative differences between predictions of effect concentrations for multi-component mixture derived from the competing concepts of Independent Action and Concentration Addition usually are relatively small (< or 1 order of magnitude). CA as an acceptable worst case Thus, our current status of knowledge may justify the general use of Concentration Addition as a pragmatic default approach to the predictive hazard assessment of chemical mixtures.
33 Advantages of the CA model It is a conservative approach If applied by default, it does not require knowledge on mode of action It requires only knowledge on simple toxicological end-point (e.g. EC50) It is a powerfool tool for screening-level mixture assessment Drawbacks of the CA model It does not allow predicting synergistic effects. At the present level of our knowledge, predictive approaches for synergistic and antagonostic effects are not available
34 CA as an acceptable default approach A precise knowledge of modes of action is rarely available. The complexity of toxicological modes of action of POPs makes the problem even more difficult. The CA approach overestimates the toxicological response, so it may be assumed as a conservative worst case. It may be thoeretically demonstrated and experimentally supported that the ratio CA/IA is generally relatively low. Therefore, if the available information is not sufficent for a scientifically based selection between the CA and IA models, there is large consensus on accepting by default the CA approach as a realistically acceptable conservative approach
35 Proposal for a Decision Tree for mixture RA Information on the Mixture composition available NO YES Data available on the mixture as a whole Information on the MoA for each component available NO Interaction suspected YES Similar MoA NO YES NO YES NO YES No RA possible RA based on Mixture Independent action or TSA Concentration addition Case by case RA
36 Conclusions Predicting mixture composition is a key issue for assessing the actual pesticide risk for ecosystems. The composition of pesticide mixtures is highly variable in time, both qualitatively (number and type of chemicals involved) and quantitatively (concentrations of individual chemicals). The CA approach is a practically applicable compromise for predicting mixture response if information for a more precise assessment is lacking.
37 Risk for baby bears in the Arctic The Hazard Index of the mixture of POPs present in the polar bear milk is three orders of magnitude higher than the treshold of 1. The risk for baby bears is likely to be very high. Several individual chemicals may be hazardous. However, in this case too, about 80% of the total mixture potency is covered by three chemicals (or congeneric groups): PFOS, PCBs, chlordane.
38 Thanks for your attention
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