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EC number: 200-087-7
CAS number: 51-28-5
Dinoseb, the pesticide used in this study, is a herbicide and insecticide belonging to the dinitrophenol family. Structural formulas of 2,4-dinitrophenol and Dinoseb are similar.Dinitrophenols may also originate as impurities in certain pesticides such as Dinoseb (Wegman RCC, Wammes JI. 1983. Determination of nitrophenols in water and sediment samples by high-performance liquid chromatography. Med Fat Landbouwwet Rijksuniv Gent 48:961-969).
No effect on adult survival was observed after 28 days at the hightest tested concentration of 20 µg/kg soil dw (NOEC);
The most conservative data obtained from reproduction test is the EC50 of 14.4 µg dinoseb/kg soil dw.
the pesticide used in this study, is a herbicide and insecticide
belonging to the dinitrophenol family. Structural formulas of
2,4-dinitrophenol and Dinoseb are similar. Dinitrophenols
may also originate as impurities in certain pesticides such as Dinoseb
(Wegman RCC, Wammes JI. 1983. Determination of nitrophenols in water and
sediment samples by high-performance liquid chromatography. Med Fat
Landbouwwet Rijksuniv Gent 48:961-969).
The effect of dinoseb on the energy
reserves (lipid and protein content) of the Collembola Folsomia candida
was investigated to test the hypothesis that dinoseb exposure reduces
energy reserves before an effect can be detected on life cycle
parameters (growth, reproduction and survival).
Folsomia candida was exposed to dinoseb in
artificial soil, while survival, reproduction, weight, lipid and protein
content were determined at different time intervals in order to
determine the relative sensitivity of those parameters. The observed
effects at different levels of biological organization were analyzed and
compared over time (temporal analysis) as well as after certain time
intervals (fixed time analysis).
reproduction test was conducted according to the ISO 11267 standard
Results are explained here below:
significant effect on survival was recorded after 28 days at the highest
tested concentration of 20 µg/g soil dry soil. In chronic toxicity test
conducted on a compressed soil with 30 Collembola per mini Petri dish a
highly significant decrease of survival was observed after 21 days of
exposure at 20 µg/g dry soil. However, as an important mortality was
observed in the control at the beginning of the experiment, no LC50
could be estimated. This mortality was probably due to overpopulation.
In the second test conducted with one Collembola per box, the toxicity
is higher than for the experiment conducted according to the ISO
standard (LC50 17 µg/g dry soil).
of soil seems to have an impact on the sensitivity of Folsomia candida
(Rundgren and Van Gestel, 1998). On a compressed soil, a water film
appears on the soil surface. The ventral tube of Folsomia candida that
enables them to regulate their water content (Rundgren and Van Gestel,
1998) is in these conditions constantly in contact with the water
surface. As soil pore water is probably the most important route of
uptake of dinoseb, it can be can hypothized that the degree of exposure
is more important in this test that in the ISO standard test with loose
soil. Furthermore, as Collembola are aggregative organisms, contact
between individuals is necessary (Green, 1964; Verhoef and Nagelkerke,
1977; Joosse and Veltkamp, 1970). Thus, isolation could also be
responsible for the observed increase of sensitivity
because it can provoke additional stress. According to Green (1964),
optimal conditions correspond to 1.2 cm2 per individual. Thus, in these
experiments we should work with approximately four organisms per mini
analysis of dinoseb concentrations in artificial soil from a previous
study (Staempfli et al., 2002) showed that degradation is observed
during the first 14 days. This phase is followed by a stationary phase,
showing a relative stability of the compound. The quantity of dinoseb
degraded over time (4 µg/g dry soil in 21 days) was not correlated with
the initial concentrations (10 and 20 µg/g dry soil), which indicates
that degradation is probably due to microbial processes.
mode of action
final weight, length and the total lipid content of adults after 21 days
was affected by dinoseb concentrations of 25 and 30 µg/g dry soil. These
effects can be linked to the mode of action of dinoseb. Dinoseb
uncouples ATPsynthesis by canceling the membrane potential, thus
disturbing the transformation of nutritive macromolecules such as
glucose or triglycerides into useful energy. The energy, instead of
being used to form ATP, is dissipated as heat (Escher, 1995). The
decrease of energy available for maintenance could be responsible for
the lower size and weight reached by the organisms after 3 weeks of
exposure. The energy left for storage also decreases, which can explain
the decrease of the total lipid content (lipids being the main form of
energy reserves in invertebrates). Jager et al. (2004), who tested the
toxicity of triphenyltin, which is also an ATP synthesis inhibitor, onFolsomia
candidafound that this compound decreased the growth and
reproduction rate of the exposed organisms over time but increased their
lifespan. It has been observed such a lifespan increase, but our results
on weight, length, and lipid content are in agreement with their results.
and growth stimulation
length and total lipid content a slightly stimulating effect on the
growth rate coefficient b could be observed for all the treatments. It
has been observed that longevity decreased with increasing dinoseb
concentrations. This observation is in agreement with Ernsting and
coworkers observation of the growth pattern of the springtailOrchesella
cincta. They found that longevity was negatively related to the Von
Bertalanffy growth coefficient (Ernsting et al., 1993; Stam et al.,
1996). Results in this study indicate that shortly after exposure to
dinoseb, the Collembola adopted a strategy of growth stimulation in
which they reach sexual maturity more quickly and reproduce more
efficiently than unexposed organisms. This phenomenon involves a
decrease in longevity. The hypothesis stated above, that stress induced
by dinoseb exposure can increase the reproduction rate, is confirmed by
the fact that the number of eggs counted at 20µg/gdry soil was 1.5 times
greater than the number of juveniles hatched in the control.
direct comparison of the number of eggs laid between exposed and
unexposed organisms is difficult because different adult Collembola
behave differently. It has been observed that despite the compression of
the substrate, unexposed organisms succeeded in laying their eggs at the
bottom of the boxes. With increasing dinoseb concentrations the
organisms made less effort to lay eggs at the bottom of the boxes.
reduction of 50% in the number of living juveniles was observed at 15
µg/g dry soil, indicating that dinoseb affects the hatching rate or the
survival of juveniles. This assumption is confirmed by the observation
of deadjuveniles (93%) at a concentration of 20 µg/g dry soil.
higher sensitivity of juveniles to dinoseb can be explained by their
thinner cuticle and higher surface/volume ratio (Holmstrup and Krogh,
1996). These results are in accordance with the EC50 of 14µg/g dry soil
obtained with the reproduction tests following the ISO standard (ISO,
of protein synthesis
increase of the protein synthesis observed in exposed organisms after 6
days may reflect the fact that Collembola have begun to respond to
stress. This hypothesis is strengthened by the induction of the heat
shock protein Hsp70 involved in repairing of protein unfolding observed
in previous work (Staempfli et al., 2002). The highest induction was
found after 5 days of exposure in organisms exposed to 20 mg dinoseb/g
dry soil. The decrease of induced Hsp70 after this time speaks for a
strong stress reaction, meaning that the optimum of the Hsp70 response
has been surpassed and that cell damage leads to decreasing values of
Hsp70 (Staempfli et al., 2002).
exposure of 6 days to dinoseb concentrations of 15–30µg/gdry soil seems
to stimulate the weight, lipid,
protein content of the exposed organisms. This stimulation cannot easily
be explained by the specific mode of action of dinoseb, but is probably
a general stress reaction, a theory strengthened by the induction of
heat shock proteins. Over time dinoseb most likely affects energy
metabolism by inhibiting ATP synthesis. Dinoseb, much like triphenyltin
studied by Jager et al. (2004), could induce an increase in maintenance
costs, explaining the decrease of size, weight, and lipid content
observed after 21 days.
the protein content is equal or superior in exposed organisms compared
to the control. These results might indicate that defense and repair
mechanisms were activated (detoxification enzymes) (Staempfli et al.,
2002).These processes are costly in terms of energy and can be in part
responsible for the decrease of lipid reserves, and finally for an
increase in mortality.
biphasic concentration-response pattern observed could be linked to a
hormesis phenomenon characterized by a low-dose stimulation,
corresponding to a short exposure time in our case, and a high-dose
inhibition, corresponding to a long exposure time. This phenomenon is
usually observed in a low dose zone, below the no observed effect levels
(Calabrese, 2005), but in this case it appears at an exposure
concentration that induces death 10 days later. It is not clear whether
this phenomenon can be considered as hormesis.
effect on adult survival was observed after 28 days at the hightest
tested concentration of 20 µg/kg soil dw (NOEC);
of 17 µg dinoseb/kg soil dw was obtained for a survival endpoint in a
mini Petri dish (growth test);
of 14.4 µg dinoseb/kg soil dw in the reproduction test.
of weight, length, and lipid content in Folsomia candida have been
observed at a dinoseb concentration of 25 µg/g dry soil after 21 days of
exposure. Growth (weight and length), lipid content, and survival are
affected to a similar degree. The decrease of growth, lipid and protein
content are not more sensitive parameters than mortality, all these
parameters being less sensitive than reproduction.
an increase in the number of eggs, the weight and the lipid and protein
content is observed after 6 days of exposure to dinoseb. If we consider
this stimulation as an early warning signal, it can be can concluded
that these parameters are more predictive of toxicity than mortality and
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