Exposure to multiple “chemical mixtures”: methods for their human and ecological risk assessment

1. Introduction

Human and ecological risk assessments of combined exposure to several chemicals (“chemical mixtures”) pose some challenges to the scientists who perform the studies, to the risk assessors who have to interpret the data, as well as to the risk managers who have to determine the uses of chemicals and the limits of use and exposure to be defined to keep people and the environment safe. This is due in good part to the large number of chemicals involved, to the fact that people and the environment get exposed to them in different ways, and that they act and interact in many ways once they enter any organism.

In this context, it is essential to differentiate hazard identification, which refers to the detection of the intrinsic toxic properties of a chemical agent from risk assessment, which evaluates the probability of occurrence of (adverse) effects related to the level of exposure, both in intensity and in duration¹.

¹ About the fundamental distinction to be done between the notions of hazard and risk, see: www.youtube.com/watch?v=PZmNZi8bon8  Subtitles available in English, French, German, Dutch, Spanish, Chinese and Russian. A French speaking version is also available: https://youtu.be/wRmfvFYDNr8 

2. What is the method that is generally used to assess the risks resulting from exposure to chemical mixtures?

The conservative approach which is generally used is to simply add up all of the safety limits determined for the different components in a mixture. This means that for determining if a mixture is safe, one looks at the safety of all components separately, and if each of them is deemed safe, the same conclusion applies to the mixture.

When different components of a mixture provoke the same effect on the same endpoint, key studies have confirmed the validity of the estimation of effects by the mere addition of the various exposures to the different components of the mixture as a default (conservative) prediction approach for a simple risk assessment.

However, sometimes there may be a synergy between two components, for instance when two chemicals interact to produce, on the same endpoint, a combined effect, which is different from the one resulting from a simple additivity of their concentrations.

More tests than those considering them separately may thus be needed so that such interactions can be incorporated within specific risk assessments. This possibility can be relatively easily determined for substances for which we know the mode of action, for example the precise interaction of the substance with biological molecules that gives it its “toxic” properties. For example when a substance inhibits an enzyme that contributes to the elimination of another substance. This is fortunately not often the case, and more research is needed on that part.

For realistically occurring environmental mixtures, experimental evidence indicates that the number of chemicals that are relevant for the determination of a cumulative effect is indeed relatively low. According to the report, usually, no more than 3 or 4 individual chemicals determine 90% of the toxic potential of a complex mixture. This contributes to reduce the uncertainty between a simple concentrations addition and independent mode of actions, and this simplifies the prediction of the toxic response of the mixture.

3. What are the methods and tools that can be used to determine a safe level of exposure to chemical mixtures?

When there is insufficient understanding of mechanisms and/or of modes of action and of their relevance to joint effects, other approaches are needed. There are a number of potential tools that can be used for hazard identification:

• ‘In vivo’ studies, where the effects of chemicals are tested on laboratory animals. These are usually time consuming and expensive, have to be ethically justified, and are done only as a last resort;
• ‘In vitro’ studies, testing mixtures on isolated cells. Faster, more efficient but sometimes difficult to interpret;
• ‘In silico’² studies that use computer models to extrapolate the reaction of an organism to a given chemical or group of chemicals. These require important toxicological data banks;
• 'Omics’, a new approach looking either at a full set of the expression of interactions between the genes of an organism (genomics), at the full set of proteins expressed in a cell (proteomics), or at a number of other set of molecules in a biological system.
• Probabilistic methods³ which will also be pragmatically valuable for prospective assessments and the prediction of joint effects.

Overall, as the “omics” approaches are still under development, a joint, integrated approach making use of in vivo, in vitro and in silico methods is the most effective for the moment.

² in silico: literally ‘in silicon’, i.e. ‘in the computer’; referring to analysis or experimentation carried out in a computer environment, rather than in the laboratory
³ Probabilistic method or model: based on the theory of probability or the fact that randomness plays a role in predicting future events. The opposite is the deterministic approach which is the opposite of random and tells that something can be predicted exactly, without the added complication of randomness and uncertainty. www.statisticshowto.datasciencecentral.com/probabilistic/ 

4. What are the main conclusions on the « state of the art » in evaluating the risks of exposure to chemical mixtures?

Overall while the complexity of chemicals mixture toxicity assessment has always been a challenge, there have nonetheless been significant advances in the field. The interplays between initial biochemical interactions and the downstream nature of the physiological (and thus potentially toxic) response have been highlighted in a number of ways.

Although these issues related to exposure to chemical mixtures are widely recognised, current regulatory policies still generally remain focussed on the assessment of each individual chemical in a commercial mixture. However, while mixture approaches are not explicitly mentioned in all current regulatory contexts, they are increasingly appearing in both future regulations and retrospective adverse reaction assessment studies.

In the same time, in the key issues around the development of models for hazard identification and risk exposure assessment of chemical mixtures, the experimental tools and statistical and risk assessment methods can be used to provide a prediction of joint effects.

In this context, the major gaps in knowledge and challenges that remain to be addressed include:

• A common basis for the assessment of the mode of action;
• Information on the interactions (cross-talk) between these modes of action of different chemicals, also with respect to adverse effects outcome;
• The availability of data of the relation between exposure levels and adverse effects (dose-response data) in all single databases;
• Separating statistical uncertainty from variability which are both inherent within any risk assessment.  

Ultimately risk assessments may need to be supported by biomonitoring data which should include the specificity of responses to exposure to chemical mixtures.

In this context, one of the further major gaps in knowledge and challenge ahead is the need to establish a common database of reliable data on the modes of action of different chemicals and their interferences. This will make it possible to describe and integrate in a chain of causes, the successive biological reactions induced by the various constituents of a mixture finally leading to the triggering of an adverse effect, the so-called adverse outcome pathways (AOP)⁴.

⁴ Adverse outcome pathways (AOPs): depict the long, sometimes complexchain of processes and events from from the first interaction of any chemical with a molecular target (such as the interaction with a receptor or the inhibition of an enzyme) through to the perturbed function at cell and tissue level, leading finally to an appreciable disturbance of the organism’s function and/or structure causing health disorders in humans or animals.
www.openaccessgovernment.org/understanding-adverse-outcome-pathway-concept/36458/