Combined Actions and Interactions of Chemicals in Mixtures

interaction of TCDD and retinoic acid also revealed a synergistic effect on the
percent of mouse foetuses with cleft palate (Birnbaum et al. 1989). The action is
interpreted as a dissimilar action i.e. same target organ but different mode of
action.
Besides laboratory animal models to clarify human exposure, models representing
wild life exposure are used in order to evaluate the ecological consequences of
chemical mixtures in the environment. Length and weight of Zebra fish were used
as growth parameters in a combination study of 3,4-dichloroaniline and lindane.
When testing the mixtures in a complete life cycle model a significant reduction in
length and weight of fish was observed. None of the chemicals tested individually
showed any effect on growth not even in much higher doses than in the mixture
(Ensenbach et al. 1997). This effect reflects a synergistic interaction between the
two chemicals on growth. However, the study did not include investigation of the
mode of action.
7.4.4 Evaluation of the in vivo studies
One of the major problems when evaluating the literature regarding combination
toxicology is the lack of a consistent use of terms and definitions. On the surface
these definitions appear to be relatively straightforward. However, there is not
uniform agreement in the use of terms like synergism and potentiation. For
instance, the interaction between TCDD and glycocorticoids was described as a
synergistic effect (Birnbaum et al. 1986). However, according to other definitions
this effect is better defined as a potentiation. TCDD alone at this concentration did
not induce cleft palate, but increased the effect of glycocorticoids. If TCDD
induced cleft palate at the selected dose, the interaction would be defined as
synergistic. Another problem is that some authors do not define the interactions
they observe. In some cases the term ‘cumulative’ has been applied in order to
define the phenomenon observed, when an effect of two or more chemicals were
observed at their individual No Observed Effect Level (NOEL).
Another important issue when investigating chemical interaction is the selection of
appropriate dose levels. It is our impression that in most studies of mixtures the
low observed effect level (LOEL) is used as a minimum dose. The advantage of
using LOEL is that one has the opportunity to characterise the observed interaction.
The problem is the extrapolation to the actual human exposure level. However,
selecting dose levels, corresponding to human exposure has the limitation that the
sensitivity of the study in most cases will be too low to identify any effect i.e. a
precise description of the toxicological effect and the possible interaction of the
tested compounds.
Time of exposure according to ‘critical windows’ is a significant issue in
reproductive and developmental combination toxicology. As the chemical agents
often have different modes of action, the exposed animals may also have different
periods of sensitivity during embryo and foetal life. Should the two or more
chemicals be administrated at the same time or by the same route? Exposing
animals to a chemical before pregnancy may antagonise the effect of another
toxicant given during gestation. Exposing animals to two chemicals within an 8-hr
interval or at the same time was shown to result in an interaction between the
chemicals. However, if the order of exposure of the two chemicals was reversed,
no interaction was observed (Hassoun & Dencker 1982).
In a literature review on the effect of interaction of chemicals on reproductive and
developmental toxicology in vivo, Nelson (1994) has gone through and evaluated
160 published studies. The conclusions were that:
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